Vol 80 No 3 November 2026

ISSN 3021-2529 | VOLUME 80 | EDITION 03 | NOVEMBER 2026

MEDCONN □ DIAGN

STICS

DIAGNOSTICS FOR MEDICAL CONNECTION

Editor Lisa Cambridge, NZCS DipQA B.ApplManagement, MNZIMLS, NZIMLS, Rangiora

New Zealand Journal of

Deputy Editors Michael Legge, PhD MRSB FIBMS FNZIMLS FFSc(RCPA), University of Otago, Dunedin Holly Perry, DipMLS MAppSc(Hons) PhD MNZIMLS, University of Otago

Medical Laboratory Science

Emeritus Editor Rob Siebers, PGCertPH FNZIC FNZIMLS FRSB HonFNZAP, Wellington Editorial Board Paul Austin, MSc(Hons) DipMLT MNZIMLS, LabPlus, Auckland Jillian Broadbent, FNZIMLS, NZIMLS, Rangiora Julie Creighton, DipMLS, FNZIMLS, Canterbury Health Laboratories, Christchurch Lauren Eddington, DipGradSci, BMLSc, MSc, Awanui Laboratories, Dunedin Chris Kendrick, GradDipSci MSc MNZIMLS, Massey University, Palmerston North Craig Mabbett, BMLSc PGDipHSM, LabCare Pathology, New Plymouth Mohd. Shahid, MBBS MD PhD FNZIMLS, PGDipHSM, Arabian Gulf University, Bahrain Terry Taylor, BSc DipMLS MNZIMLS, Southern Community Laboratories, Dunedin Dr Donna Rudd, BappSci, PhD, MAIMS, CCS, FFSc (RCPA), James Cook University, Australia Formatting Sharon Tozer, AT DipBusStud, Executive Office NZIMLS, Rangiora About the Journal The New Zealand Journal of Medical Laboratory Science (the Journal) is the official publication of the New Zealand Institute of Medical Laboratory Science (NZIMLS). The Journal is peer reviewed and publishes original and review articles, case studies, technical communications, and letters to the Editor on all subjects pertaining to the practice of medical laboratory science. The Journal is open access (www.nzimls.org.nz/nzimlsjournal) and is published three times per year in March, July, and November. Hard copies are circulated to all NZIMLS members and universities and research units in New Zealand and overseas. Current circulation is about 2,800 copies per issue. Printed by Spectrum Print, Christchurch, using environmentally responsible paper from elemental chlorine free, third‑party certified pulp sourced from well‑managed and legally harvested forests. Manufactured under Spectrum’s Toitū Diamond Environmental Management Certification. The Journal is indexed by CINAHL, EMBASE, SCOPUS, Informit, Thomson Gale, EBSCO and Biosis Citation Index, and the Journal Editors are members of the World Association of Medical Editors (www.wame.org). Brief instructions to authors The Journal accepts original submissions from anyone and anywhere. Comprehensive instructions can be found on the NZIMLS website (www.nzimls.org.nz/instructions-to-authors. html). All submissions will undergo single-blind peer review and possibly plagiarism checking with iThenticate™ software. If accepted for publication, copyright is vested in the author(s) under terms of the Creative Commons Attribution License (www. creativecommons.org/licenses/by/2.5/legalcode). The authors are responsible for the scientific content and views. Opinions expressed in the Journal are not necessarily those of the Editors, Editorial Board, or Council of the NZIMLS. Journal cover © Kelly Craig, Medlab Central, Palmerston North Advertising and subscription Advertisement bookings and enquiries should be addressed to the NZIMLS Executive Officer, Sharon Tozer: sharon@nzimls.org.nz. Phone +64 3 313 4761.

Volume 80 Number 3 November 2026 ISSN 1171-0195 80th Anniersary Guest Editorials My experiences and the NZIMLS. Tony Mace............................................................................122 A different journey through medical laboratory science. Warren Dellow...............................................................122-123 Reviews Review on current β-thalassaemia therapeutic treatments and future research. Navana Mclachlan ........................................................127-131 Original articles Cephalosporin and fluoroquinolone-resistant gram-negative bacteria in the gut microbiota of healthy humans in Bahrain. Khaled Ahmed Alnaimi, Mohd Shadab, Ronni Mol Joji, Abdulrahman Yousif Ismael, Ali Ebrahim Almahmeed and Mohammad Shahid ......................................................132-137 Prevalence and determinants of Chlamydia trachomatis infection among HIV-infected patients and pregnant women in Auchi and environs, Nigeria. Favour Osazuwa, Osayande Osabuohien and Innocent Godfrey Iyare..............................................................................138-142 Role of Decorin and its relationship with biochemical variables in the serum of patients with osteoarthritis Sarah A Younus and Zahraa MA Hamodat...................144-150 Case Studies Benign acute childhood myositis (BACM): a case study from a rural hospital. Rebecca Busch ............................................................151-153 When APTT refuses to correct high-titre factor VIII inhibitor inducing false positive lupus anticoagulant and spurious factor IX deficiency: a case of assay interference. Hari Priya Raghvan, Wee Shiang Yui, Caroline Ho Siew Ling, Ahmad Izzudin Mustaffa, Ehram Jamian and Indhira Subbiah ............................................................154-156 Book reviews Other scientists: routes to innovation in biology, edited by Oren Harman and Michael R. Dietrich. Reviewed by Michael Legge................................................ 158 Invisible evidence in forensics in New Zealand by Bill O’Brian. Reviewed by Michael Legge................................................ 158 What we inherit: how technologies and old myths are shaping our genomic future by Sam Trejo and Daphne O Martschenko. Reviewed by Michael Legge................................................ 158 Conference proceedings Barrie Edwards & Rod Kennedy scholarship conference report. Savannah Young ................................................................. 156

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Fiji Institute of Medical Laborator Science Annual Seminar Kirsten Birrel and Leah Pringle .....................................161-162 Regular features Advertisers........................................................................... 119 Christmas Quiz .............................................................179-182 In this issue...................................................................120-121 Journal questionnaire ...................................................177-178 Microbiology SIG 2026 report .............................................. 166 NICE Weekend 2026 report ................................................ 165 NZIMLS calendar................................................................. 142

NZIMLS Annual General Meeting Minutes ..................... 159-160 Obituary: Walter Wilson ......................................................... 167 Otago BMLSc student research project abstracts, semester 1, 2026 ............................................................ 168-173 Pacific Way ..................................................................... 174-176 Reviews of interest ................................................................ 137 Reprint, Love and Death, Tristan Jones, NEJM .................... 157 Retiring colleagues: Ross Hewett.......................................... 121 Science digest ................................................................ 163-164 TH Pullar address ........................................................... 124-126

Advertisers in this issue: Abacus DX ...................................................................................................................................................................................119 Helena Laboratories .............................................................................................................................................. Inside front cover

In this issue In the final instalment of our series celebrating 80-years of the NZIMLS, we feature reflections on laboratory and professional life from two esteemed NZIMLS Life Members, Warren Dellow and Tony Mace. We are pleased to reprint Julie Creighton’s outstanding TH Pullar Address presented at the Wellington ASM in August; “Try not to have a good time, this is supposed to be educational”. Thalassaemia is a group of inherited haemoglobin disorders that impose a substantial global health burden, and while its genetic basis is well established, increasing evidence highlights the role of epigenetic regulation in modulating disease severity and as a target for therapy. Navana McLachlan, a student and Medical Laboratory Technician from the Haematology Department at Medlab Central in Palmerston North, conducted a literature review on current β-thalassaemia therapeutic treatments and future research. Focusing on three therapeutic approaches for transfusion-dependent β-thalassaemia (TDT); iron chelation therapy, CRISPR–Cas9 gene editing targeting the BCL11A enhancer, and allogeneic hematopoietic stem cell transplantation (HSCT), McLachlan highlights the contrast between supportive, curative, and emerging molecular therapies, emphasising the need for safer, more accessible treatments that target the underlying genetic and epigenetic mechanisms of thalassaemia. The gut microbiota is one of the densest microbial ecosystems on earth. Although it has many positive effects on health, the high population mass of microbes in this ecosystem can result in the horizontal spread of antimicrobial resistance (AMR) genes to potentially harmful bacteria. Professor Shahid and colleagues from the College of Medicine & Health Sciences, Arabian Gulf University, Bahrain conducted a study to analyse the presence of cephalosporin and fluoroquinolone-resistant gram-negative bacteria in the gut microbiota from 29 healthy patients to assess the growing threat of antibiotics resistance within the population and globally. A total of 92 isolates were found with 84.7% identified as gram-negative bacteria. Antimicrobial susceptibility testing showed that bacteria exhibited the highest resistance against ampicillin (64%), followed by cefazolin (45%). Resistance was lowest towards imipenem and meropenem, at 5% and 3% respectively. Genotypic analysis revealed 52% of isolates carried the blaCTX-M-15. gene. This study raises findings that warrant further examination, as the discovery of antibiotic resistance genes harboured within the gut of volunteers with no history of antibiotic exposure holds concerning implications for public health and its burden to effectively treat infectious diseases. Osazuwa and fellow researchers from Edo State University, Nigeria investigated the prevalence and determinants of Chlamydia trachomatis infection among HIV-infected patients and pregnant women. C. trachomatis infection has been associated with the increased risk of human immunodeficiency virus (HIV), as Chlamydia triggers an inflammatory response that draws CD4+ T-cells to the site of the infection, leading to an increase in immune cell recruitment, particularly CD4+ T cells, which are targeted by HIV. The dual burden of HIV and Chlamydia trachomatis co-infection presents significant clinical and public health challenges, especially among vulnerable groups such as pregnant women. Using CD4+T cell count and PCR methods of 190 women comprising of 95 HIV-infected patients and 95 pregnant HIV-negative women. Results showed C. trachomatis was significantly higher in HIV-infected subjects (21.1%) than compared to pregnant HIV-negative women (5.3%). Additionally, a high prevalence (13.2%) of asymptomatic infection was shown across the study group. These findings highlight the need for routine screening, education and control programs in the Auchi and surrounding areas, particularly for at-risk groups and individuals of all ages. Osteoarthritis classified as a chronic degenerative joint disease characterised by cartilage loss, pain, and limited mobility and leading to significant health problems and physical disability

that negatively impacts an individual’s quality of life. Leucinerich small proteoglycans, specifically Decorin, play a vital role in maintaining the integrity of cartilage and the extracellular matrix. Younis and Hamodat from the University of Mosul and Ninevah (respectively) in Iraq, investigated the role of Decorin and its relationship with parathyroid hormone (PTH), vitamin D, calcium, phosphorus, and body mass index (BMI) in the serum of patients with osteoarthritis using ELISA and spectrophotometry methods and statistical analysis. Data demonstrated a distinct positive correlation between serum Decorin levels and the progressive stages of joint degeneration, highly significant differences between the patient and control groups. Results confirmed that elevated Decorin and parathyroid hormone levels, compared to low vitamin D and calcium levels, provide a diagnostic picture that accurately reflects the severity of joint tissue degeneration and mineral imbalance in affected individuals. With further investigation these biomarkers could be integrated into clinical evaluation practises to improve disease grading and facilitate therapeutic intervention in the treatment of osteoarthritis. Medical Laboratory Scientist, Rebecca Busch from CDHB, New Zealand reports four benign acute childhood myositis (BACM) cases seen in a rural hospital in the winter of 2025. BACM is a rare, self-limiting, characterised by the sudden onset of lower extremity muscle pain following a viral infection. Laboratory investigations included respiratory viral panels, creatine kinase (CK) measurements and complete blood count. All cases tested positive for Influenza B and had markedly elevated CK levels. In one patient CK levels were monitored and returned to near normal levels with eight days. There are no clear guidelines for treatment or management of this condition, all patients were managed conservatively, discharged home with instructions to rest, use paracetamol and hydrate and all made a full recovery. While these cases were rapidly identified and managed, previous studies show that many cases of BACM are not recognised by clinicians and may be misdiagnosed as more serious conditions and unnecessary testing and treatment. Monitoring CK levels is recommended to assist in confirming that the muscle injury has fully resolved. Acquired haemophilia A (AHA) due to autoantibodies against factor VIII (FVIII) is a very rare condition with severe bleeding and high mortality. Raghvan and colleagues from Ministry of Health in Malaysia report a patient with spontaneous bruising and isolated prolongation of activated partial thromboplastin time (aPTT), which did not correct in mixing studies. Lupus anticoagulant (LA) testing also demonstrated an abnormal partial thromboplastin time. These preliminary results led to suspicion of concomitant lupus anticoagulant (LA) and factor assays revealed a very low FVIII level (<1%, reference 50%-1005), accompanied by spuriously low FIX (7%, reference 50%150%). Further investigation identified a high-titre factor VIII inhibitor, and the patient was treated with Factor VIIa, however the patient developed intracranial bleeding and succumbed to her condition. This unique case of acquired haemophilia A causing interference with LA and FIX assays illustrates the need of careful clinical correlation and specific assay selection for accurate diagnosis. Savannah Young, NZIMLS recipient of the Barrie Edwards & Rod Kennedy Scholarship, reports on her attendance at the Blood and Regional Congress of the ISBT in Perth, held in October 2025. The Journal is excited to again been given permission from the New England Journal of Medicine, to reprint Tristan Jones perspective article entitled; Love and Death, published in N Engl J Med 2026; 395(4): 321-323. We announce the retirement of Ross Hewett a long-serving NZIMLS life member. The NZIMLS and profession thank him for his services and wish him all the best in his well-earned retirement. The NZIMLS farewells founding member, Walter Wilson, who

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sadly passed away this year and we pay tribute to his personal warmth and lasting contribution to the profession. Our resident bibliophile, Dr Michael Legge reviews three books; ‘Other scientists: routes to innovation in biology’, edited by Oren Harman and Michael R. Dietrich; ‘Invisible evidence in forensics in New Zealand’ by Bill O’Brien; ‘What we inherit: how technologies and old myths are shaping our genomic future’ by Sam Trejo and Daphne O Martschenko. Regular features include Pacific Pathology Training Centre (PPTC) updates in the Pacific Way, CPD Questionnaire, meeting reports, Otago University BMLSc student research abstracts from Semester 1, the Science Digest and Recent Reviews. The successful Christmas Quiz is back with more fun and prizes for

the most successful laboratory teams. 2026 marked 80-years of the Journal and the Institute. This milestone anniversary is typically symbolised by an Oak tree, representing strength, wisdom, endurance and legacy. On behalf of the NZIMLS and Journal, thank you to all our authors, contributors, Editorial Board members, subject-matter-experts, reviewers, NZIMLS Executive & Editorial Office, Editorial team and the continued support of members and Council members, past and present for their amazing and dedicated work within the profession. Slàinte mhath* - Lisa Cambridge, Editor * “Good health” (Scottish Gaelic), pronunciation: slanj-a-va

Retirement Announcement Ross Hewett: a career of leadership in Medical Laboratory Science Ross Hewett’s career in medical laboratory science spans clinical practice, international laboratory management, education, industry, professional leadership and conference presentation. Beginning as a trainee in 1971, he developed broad experience in New Zealand and overseas and became an active contributor to the profession through the New Zealand Institute of Medical Laboratory Technology, now the New Zealand Institute of Medical Laboratory Science (NZIMLS). Training and Early Career Ross began as a Trainee Medical Laboratory Technologist at Palmerston North Hospital in January 1971, the same year he joined the NZIMLT. He completed his basic training certificate in 1974, followed by the Part 2 Chemical Pathology examination in 1975 and Part 3 in 1976. International Experience In April 1977, Ross travelled to the United Kingdom and worked for 18 months at J. S. Pathology Ltd, 81 Harley Street, where he served as second-in-charge in Haematology. At the end of 1978, he moved to Jeddah, Saudi Arabia, to become Laboratory Manager at Dr S. Fakeeh Hospital. There, he established a complete laboratory from the ground up. He concluded this role in 1981 and returned to New Zealand that September, following six months travelling through Europe in a camper van with his wife Jill and their baby, who had been born in Saudi Arabia. Teaching, Clinical Practice and Industry On returning home, Ross worked at the Central Institute of Technology in Upper Hutt, teaching medical laboratory science students. He subsequently joined Hutt Hospital, where he worked from 1982 until the end of 1983. In 1984, he moved to Auckland and began a long association with Boehringer Mannheim and Roche, remaining with the organisation until 2000. Professional Leadership and Continuing Education Ross presented the T. H. Puller in 1998 and has shared his knowledge at many conferences and special interest group meetings throughout his career. His commitment to reflective professional practice was also evident in his completion of a Postgraduate Diploma in Health Science in Healthcare Ethics at Auckland University of Technology. He has played a prominent role in convening major professional gatherings, including the combined New Zealand and Australian South Pacific Congress at Auckland’s SkyCity Convention Centre in August 2007, the 2010 NZIMLS Scientific Meeting in the Bay of Islands, the 2013 NZIMLS North Island Seminar in Auckland, and the 2015 South Pacific Congress in Auckland.

NZIMLS Leadership and Service • • • • • • • • •

Joined the NZIMLS in 1971. Elected Region 1 representative in 2003. Elected Secretary/Treasurer in 2004. Served as NZIMLS President from 2014 to 2017. Awarded Life Membership at the 2014 Dunedin Conference. Delivered the 1998 T. H. Pullar Address at the Palmerston North Conference, titled Trends, Technologies and the Medical Laboratory Scientist. Hosted several Biochemistry Special Interest Group meetings. Regularly presented at Annual Scientific Meetings. Played a pivotal role in establishing the NZIMLS accounts system in 2004. By introducing an updated chart of accounts and regular financial reporting, he supported the development of the NZIMLS administration and Executive Office as it operates today.

A Lasting Contribution Across laboratory practice, management, teaching, industry and professional service, Ross has demonstrated a sustained commitment to medical laboratory science. His career reflects both the expanding scope of the profession and the importance of sharing knowledge, building professional communities and supporting continuing development. We wish Ross all the best in his well-earned retirement. Contributed by: Sharon Tozer, CEO, NZIMLS

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80th ANNIVERSARY GUEST EDITORIALS

My experiences and the NZIMLS Tony Mace

I would like to start by saying what an honour and privilege to be able to write as a guest on what the NZIMLS has meant to me over the years as a trainee, staff technologist, technologist in charge and then as a charge Medical Laboratory Scientist, over 43 years in the medical industry. My journey with the Institute started when I joined the laboratory at Tauranga Hospital as a trainee technologist in the mid to late 1970’s. I was a late bloomer as I was already married and had a child, a house and a mortgage. This was because I had been working before as a science technician for the then National Health Institute in Wellington and had gained a NZCS (Biology). So, when joining I was informed that I needed a NZCS (Medical Biology) qualification from the then NZIMLT (New Zealand Institute of Medical Laboratory Technology) before being able to go straight into doing ‘O’ and ‘A’ level exams. So back to school, doing all the medical side of the certificate over another 3-year period. Travelling to Hamilton Polytechnic on day release and doing night classes at this stage of life, plus working and family life was a big task, but being young and enthusiastic I did achieve my proper qualification to carry on. Finally qualified with a Diploma in Medical Technology, I also joined the Institute and so began a close liaison with them and my involvement. At the beginning of this career, the NZIMLT was a ghost who required a subscription for some things I was unaware of through my own naivety. Subscription rates were around $30 a year back then and I was amazed that work paid this, how fortunate was I. So, who was this mysterious group that required money on a regular basis and what were they doing for me? I think time has changed that and people in the profession now certainly have more information. It was in the eighties when I had become a Technologist in charge at Waipukurau Hospital lab that I became more aware of what the NZIMLT was about as industrial relations and a wage freeze for 3 years eventually led to industrial action by laboratory workers in 1985/86 and consequently the splitting of the NZIMLT into the professional and educational side and the separate formation of the New Zealand Laboratory Workers Union in 1988. I became interested when I heard that they would like qualified people to help with setting QTA exams. This started my involvement actively with the NZIMLT. So, for three years I set

the QTA general exams learning that this ghost was becoming more real and getting to know some of the people who were on the committee and its workings. I started to go to conferences and have a closer liaison with Blood Bank at Palmerston North Hospital and meet up with them on a regular basis and was present for the first few years of the NICE weekend, which was like the forefront of the Special Interest Groups (SIG). Moving to Hamilton in the early nineties to help start a new laboratory, my relationship with the Institute became even more important as it helped with collegiate and professional help and I also took on Biochemistry SIG convenor for two years as I wanted to give something back to the profession that had helped me realise that professional growth was now becoming an essential part of our industry and the Institute was (and still is) a vital part of who we are. When Hamilton hosted the annual conference, I was part of the team and because of that involvement I thought that I could be of more help if I joined the committee as Region 2 Representative. In 1997 I became Region 2 Rep and joined the Institute committee. We had had the name change from NZIMLT to NZIMLS (New Zealand Institute of Medical Laboratory Science). This ghost had now turned into a living being of likeminded people who had the interests of the industry at heart and for our profession and its education. Under excellent leadership we managed to turn our financial problems round keep the journal going from four publications to three and started to put in protocols for running conferences, SIG’s, polling the industry to ascertain what works and what doesn’t, running the exams with tighter financial control, setting up a website. Serving on the committee for 5-6 years was a highlight of my career in being able to help in an ever-changing world. This was why I believe in what the Institute is, it is for the people, their professional development, education, and ultimately for the patient for whom we do this work, for they deserve a gold standard service and nothing less, and because of the Institute, that is what they receive.

AUTHOR INFORMATION

Tony Mace, (retired) DipMLT, Life MNZIMLS, Hamilton, New Zealand. Email: tonym52@me.com

A different journey through medical laboratory science Warren Dellow In late January 1974 I started work in the Nelson Hospital Laboratory as a trainee medical laboratory technologist. Two weeks after starting, knowing almost nothing about how medical laboratories functioned, I was off to the Central Institute of Technology in Heretaunga to start my studies towards NZCS (paramedical). After this three-month block course, we returned home to start learning about what really happened in medical laboratories. Similar length block courses were held each year, before sitting the final NZCS examinations. After attaining NZCS (paramedical), I embarked on studies in Part II biochemistry. A good choice, as the department was undergoing significant change, with the introduction of automation and new technology. After successfully passing my Part II examinations I was emboldened to continue with Part III studies. Early in that year we were advised by hospital management that of the remaining trainees who started in 1974, only two, or at most three, would be employed as medical technologists, once they had passed the examinations required for registration.

Feeling vulnerable, and with a baby expected in June, I started looking for alternative employment options. Eventually I heard about a possible position in Christchurch at Pearson Laboratory. After interviews, I was offered the position of acting 2IC in biochemistry, with the position to be confirmed on passing my Part III examinations and gaining registration. I was also able to negotiate access to the Christchurch Hospital biochemistry Part III lecture programme along with work experience in their specialised departments such as toxicology, steroid laboratory etc. A year after I moved to Christchurch, Jim Le Grice was employed as Head of Biochemistry. We worked well together and assumed different responsibilities for different areas of the laboratory. When I started at Pearson’s, the laboratory required upgrading in almost all areas with substantial investment required in automated equipment to meet a very rapidly growing workload. Fortunately, the owners supported these requirements and over a three-year period we fully transformed the department into a high throughput laboratory where the major focus was to process

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and report patient test results as quickly as possible. In 1982 I changed direction and started work for a commercial laboratory supply company. This led to an opportunity to form my own company, (Med-Bio Limited) in 1985. While completing my Part III training at Christchurch Hospital I came to understand how little I really knew about the specialist areas of my chosen subject, compared to those who worked in those areas full time. I realised, that to really gain competency, I needed to continue to learn. In the five years following registration as a medical laboratory technologist, I had limited opportunity for further study. However, once I was my own boss that responsibility lay entirely with me. I seized the opportunity and completed short courses, both in New Zealand and overseas, in topics related to our companY’S work, but also in subjects that just interested me. I continued to do that for the remainder of my working life. By the early 1980’s I was also aware that many of my colleagues in laboratories had very limited opportunities for continuing education. After leaving Pearson laboratory I remained close friends with Jim Le Grice and others within the profession. Over a few beers we hatched a plan to resurrect the South Island Seminar. The first meeting of the new era was held at Mariua Springs, close to the centre of the South Island. That first meeting was light on attendee numbers, light on papers but a good time was had by all participants, especially those who stayed on for the evening meal and a soak in the hot pools. From that very first

meeting of the new era, South Island Seminar has gone from strength to strength. Today, some 40 years on, the meeting is still very well attended with a high standard of scientific presentations. Med-Bio had agreed to underwrite the meeting financially (not required thankfully), and our involvement was locked in for more than 20 years. We also supported the meeting with a generous prize for the best presentation. At many of these meetings I also presented, along with some of my staff members. Our focus was to present new information on methods and technologies that many attendees may not have been aware of. Once SIG meetings become established, we continued to support some of those in much the same way as we supported the South Island seminar. On reflection, I had a very successful career in Medical Laboratory Science. I attribute much of this success to my personal commitment to continuing my own education. This was an investment of my own time and money, and I regard this as having been a worthy investment in my future. It is my belief that our personal investment in ongoing education pays dividends for furthering our careers and maintaining our employability within our chosen profession.

AUTHOR INFORMATION

Warren Dellow, (Retired) Life MNZIMLS, HFNZSP Email: wdellow@xtra.co.nz

Advertisement extracted from the Journal of the New Zealand Association of Bacteriologists, Vol. 1, October 1946, No. 3 New Zealand Journal of Medical Laboratory Science 123

TH PULLAR MEMORIAL ADDRESS

Try not to have a good time… this is supposed to be educational Julie Creighton

INTRODUCTION

It was certainly both a privilege and a responsibility to open the 2026 NZIMLS Annual Scientific Meeting with the TH Pullar Memorial address, and I thank the Council for considering me worthy. I am acutely aware that many exceptional laboratory professionals have gone before me. This address honours the legacy of Dr Thomas Henry Pullar; recognising him as a pioneer of New Zealand pathology. Dr Pullar was born in New Zealand in 1907, educated in the United Kingdom, then returned to New Zealand, working as a pathologist in Palmerston North from 1937. Sadly, he passed away at only 59. For many years he was involved in the development of professional laboratory standards, and he played a key role in the formation of the NZ Association of Bacteriologists (which is now the NZIMLS). He was a champion of medical laboratory technologists, deeply committed to their training and welfare, including conditions of employment, preparing syllabi for examinations, supervising examinations and introducing new educational training schemes. Dr Pullar certainly paved the way for the future of our profession! And talking about the future… what a terrific conference title this year! 80 years since the formation of the New Zealand Association of Bacteriologists is a remarkable milestone. The profession has come a long way from the days of manual everything, very rudimentary organism identification, handwritten results and mouth pipetting! And, sadly, I remember them all! We now have automated analysers processing thousands of samples a day, molecular diagnostics and some impressive digital systems. We can look forward to artificial intelligence, pathogen genomics and personalised medicine reshaping laboratory diagnostics. However, I will leave some of these topics for other presenters at this conference. The importance of enjoying education What I would like to focus on is education – what you can learn along the way if you are curious, engaged and enjoying it. Education is something I have always been passionate about – either self-education or passing on my knowledge. I recently counted that I have done nearly 100 PowerPoint presentations. This photo (Figure 1) shows me on holiday – well actually my deck chair - with a stack of journal papers at the end. I often get teased about it at work, but I always take away a pile of journal papers with me on holiday, and I read them while travelling in the car, at the airport, on the plane or sitting by the pool! I really enjoy it!

Figure 1. On holiday – well actually my deck chair - with a stack of journal papers at the end Which leads me to the title of my talk which is based on a quote by Charles Schulz, from the Peanuts comic strip. Try not to have a good time. this is supposed to be educational! The line is attributed to Lucy, who liked to drain the fun out of everything.

Obviously, this is the opposite of good teaching wisdom and certainly does not align with the beginnings of my career. Enjoying what I did and what I learned was what attracted me to medical laboratory science and what keeps me motivated – and has hopefully motivated other scientists over the years. My career started after I left school, not sure where I was heading, but contemplating something in the medical field, or maybe accounting. I applied for a lab assistant role at Dunedin Public Hospital, in Microbiology. On day one, I was taken under the wing of another lab assistant, Anita Wilson, who has ‘mothered’ many new staff over the years and remains a dear friend. Initially I was employed in the media room. In those days we made all our own media, and the kitchen ladies did all the cleaning and sterilising. My father was horrified that I had a job surrounded by glassware! I was quite a clumsy child – you could always rely on me to get stuck up a tree, trip over, break a bone, stand on glass etc. I did drop a fair few test tubes at the beginning, and there was an incident involving a large cardboard carton of cream cakes at the main entrance of Dunedin Hospital, at the busiest time of the day, which turned into an embarrassing and expensive event! My most memorable incident involved a flask of MacConkey Agar and a poorly designed lab coat. I was sitting at the media room table, swirling the flask, to cool it down before pouring it into Petrie dishes, when I cracked it against the table edge. 1L of warm agar spilled onto my woollen skirt. I had no choice but to take it off, hand wash it and dry it in the hot air oven. I spent the rest of the day in my poorly designed lab coat and cowboy boots. If that wasn’t bad enough, when it was home time, I discovered that my skirt had shrunk! I managed to squeeze in to it; walked like a Geisha girl to the bus stop, but was a bit late, so I had to leap up onto the bus resulting in a very large rip up the back seam! I don’t remember the rest of story – obviously too traumatised, but that was certainly one way to learn a lesson! Those early days in the lab were a lot of fun, with numerous water fights and lots of socialising. With four new trainee scientists each year, there was a big pool of young staff. One of the highlights was the Christmas party – held in the Haematology lab. We’d clear off the benches, leaving a little bit of bench for the person still working making blood films, spread out some brown paper, then get out the drinks and nibbles. Obviously minimal Health & Safety back then! The evening always included numerous playing of Snoopy’s Christmas, while we formed a long chain and danced our way through the whole lab. However, there was education as well. I was fortunate to join the training scientists for lectures. I found the theory fascinating and decided that I would stay at this job and study towards becoming a Qualified Technical Assistant (QTA). After I passed, I wanted to go on and become a scientist. Unfortunately, the ‘rule’ at the time was that assistants were just not capable of advancing to the next level! This was despite that I had achieved top marks in my QTA exam, done a chemistry paper in the evenings, and shown aptitude on the bench. Fortunately, the Microbiology second-in-charge, Ron Fyfe, advocated strongly on my behalf and managed to change the minds of the Chief Scientists. Hence, I began my career as a clinical laboratory scientist. I completed my training, ending with O and A levels in Microbiology. Overseas experience I then headed off to London. My first locum position was at King’s College, a liver transplant specialist hospital. I was involved with a study investigating the use of aztreonam (a broadspectrum beta-lactam antibiotic) to prevent hospital acquired infections in patients with fulminant hepatic failure. A tour of the ward presented me with a big surprise – I had expected to see modern facilities; however, it was more Florence Nightingale era!

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A distressing aspect was seeing the study patients who were mainly young girls, all with bloated faces and yellow eyes and skin due to their liver failure following an attempted suicide by paracetamol overdose. It was a harrowing experience. Sadly, many of the patients died, so I took on the MRSA screening bench – despite having never seen MRSA before! This was in the era before chromogenic media, or staphylococcus slide latex and way before MALDI-TOF for identification. But there was a lot of MRSA in London hospitals at the time, so I learnt quickly! After 6 months at King’s College, I moved to St Thomas’s Hospital – right across the river from Parliament House and Big Ben. Every day, I’d leave work and I’d never get sick of admiring that view. I was at St Thomas’s for more than 3 years, advancing to a senior role. During that period, we went to South Africa for nine months. While in Durban I had a short contract at the Research Institute for Tropical Diseases, looking into antimalarial drug resistance. A highlight was going on a field trip to Jozini, a small military town near the Mozambique border. We collected mosquitoes and visited local clinics and the hospital to collect patient’s blood. The hospital was very basic, with most patients spending their days outside on the grass, drips attached! This was a real eye opener for me! Domestic and wild animals were abundant, with goats, cows and donkeys a constant road hazard, and snakes, hippos and crocodiles not difficult to spot! The photo (Figure 2) shows typical village life, with the women fetching water and doing their washing in the nearby river. The bridge was destroyed in a severe storm more than a year prior but had still not been repaired.

Figure 2. Jozini, a small military town near the Mozambique border. After 5 years away, now married and with a young baby, we returned to New Zealand. Soon after arrival I accepted a role at a Christchurch-based Diagnostics company, MedBio Ltd, who were the New Zealand agents for bioMèrieux Diagnostics and DAKO immunohistochemistry products. For the first time I was on the commercial side of science, and I learnt a lot during this period. In 2000 it was time to get back to the laboratory when a scientist role became available at Canterbury Health Labs. 26 years later, I’m still there and still loving it. Developing antimicrobial resistance passion It would be fair to say that antimicrobial resistance (AMR) has developed into a major passion. Right from the beginning I was interested in AMR, particularly enzymes found in Gram-negative bacteria (Figure 3). When I first started at CHL, ExtendedSpectrum β-lactamases (ESBLs) were rare and caused great excitement when we found one! Soon they became more frequent and these days isolation of ESBLs is very common, with endemic spread in the community. In 2008 we found our first carbapenemase producer – only the second reported in NZ but relatively few were isolated until 2024 when numbers across New Zealand increased dramatically (1). The World Health Organisation (WHO) lists these types of organisms as critical priority pathogens, and it is vital that laboratories can rapidly and accurately detect them. Consequently, AMR detection and screening is a large component of what I do in microbiology. I am very thankful that

I have been supported by the Clinical Microbiologists along the way (Professors David Murdoch and Anja Werno initially) and given a lot of freedom to experiment, to acquire isolates from ESR, to ‘play’ with different resistant isolates, to compare new products, and to develop and refine our methods. I’m always trying to improve what we do. Thank you to everyone who has sent me their difficult isolates over the years.

Figure 3. Gram-negative bacteria In 2014 we were proudly the first laboratory in New Zealand to introduce European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines for antimicrobial testing and interpretation. This was a move away from CLSI, the Americanbased system. In 2017 a group of microbiologists and scientists formed the NZ National Antimicrobial Susceptibility Testing Committee (NZ NAC), with the aim to provide expert advice to NZ diagnostic laboratories. The committee has done a lot of work over nearly a decade, including a guideline for reporting antimicrobials; recommendations for the minimum requirements for laboratory detection of carbapenemase-producing Enterobacterales; education bugs; various susceptibility methodology studies and newsletters (2). We can now say that every clinical microbiology lab in New Zealand is standardised with EUCAST guidelines. AMR interesting finds AMR is an ever-increasing clinical threat and it’s important to recognise the changing epidemiology and complex resistance mechanisms now evolving, especially in carbapenemase producing Enterobacterales (3). Continual learning, being curious about unusual results and careful investigation are essential to the steady accumulation of knowledge. I have used this knowledge to uncover some interesting NZ-first or world-first finds along the way. One case occurred at the end of 2015. A 20-year-old male NZ tourist suffered electrical burns and head trauma following a live-wire shock in Romania. He spent six months in a Bucharest hospital before being flown to Christchurch Hospital. On admission he was put into isolation with contact precautions and multiple screening samples were taken, including rectal swab, five electrical exit-wound sites, urine and tracheal aspirate. The bench was covered in plates growing so many resistant organisms, and I remember feeling quite overwhelmed! It is important to note that up until this point our laboratory had only isolated eight carbapenemase producing organisms between 2008 and 2015 – all from patients with overseas acquisition, except for another NZ first involving transmission of NDM-5producing K. pneumoniae among four patients at Christchurch Hospital (4). A paper that you’d really rather not be publishing! From our patient we recovered a total of 15 carbapenem resistant organism, yielding seven distinct organism/carbapenemase combinations. Globally this was the highest number reported from a single patient. In addition, there were two isolates that had not previously been found in NZ, an ESBL-producing Pseudomonas aeruginosa and an ACC-1-producing Proteus mirabilis. I have not found either of these resistance mechanisms since (5). Another case was in 2017 when we received a urine from a female patient with a spinal cord injury and a history of recurrent urinary tract infections (UTI). She had travelled to Thailand four

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months prior to her current UTI. The culture grew an ESBLproducing E. coli which was also resistant to colistin. Resistance to colistin was rare and of concern because colistin was among a diminishing group of antimicrobial agents considered last resort for the treatment of MDRO infections. Plasmid-mediated colistin resistance due to mcr-1 had only recently been discovered had not been reported in NZ before. I sent the isolate to ESR (PHF) and clever scientists performed whole genome sequencing and discovered that there were in fact two colistin resistant gene types (mcr-1 and mcr-3) located on two different plasmids. Globally, cooccurrence of mcr-1 and mcr-3, from a single organism, had only been reported twice before. In addition, the mcr-3 gene and the CTX-M-55 (ESBL gene) were found within a uniquely arranged InFII plasmid. This was a New Zealand first and a rare global find (6). More recently a 49-year-old woman presented at Christchurch Hospital emergency department with shortness of breath and underlying asthma and immunosuppression. Importantly, she had no history of overseas travel. Her sputum grew Serratia marcescens with an unusual susceptibility profile, including high-level resistance to meropenem but susceptible to extendedspectrum cephalosporins. GeneXpert PCR (detects the five most common carbapenemase enzymes) was negative, but a biochemical test, Carba NP, confirmed a carbapenemase. WGS revealed Serratia marcescens enzyme SME-2, which was not only a first for New Zealand, but rarely reported worldwide and our isolate included a unique gene sequence (7). Travel and education Combining my loves of travel and teaching, I was invited to go to Vietnam in 2008 and 2009. We were based in a city called Quy Nhon, essentially a fishing village, but home to a leprosy hospital and the Provincial Hospital. The trips were funded by the NZ Vietnam Health Trust, of which Graham Paltridge was a long-time associate (Figure 4). The focus of my trips was on reproductive and sexual health. My presentations had to be translated into Vietnamese, so I had to be very careful with my dialogue, especially considering the subject matter. The health system was very basic. The hospital even had wards for administration of natural/herbal medicine treatments. There was no such thing as hospital meals and patients had to be fed by their family. Many families from the outlying villages couldn’t afford accommodation locally so we would find them sleeping around the hospital grounds or stairwells. It was certainly a very interesting experience, but also lots of fun and the food was fantastic!

Figure 4. New Zealand Vietnam Health Trust with Graham Paltridge, a long-time associate. I have been fortunate to attend many international conferences over the years. I have been supported by CHL management, but also by the NZIMLS with a Barrie Edwards & Rod Kennedy scholarship and with best paper or speaker awards. I have also benefited from a CDHB quality award and some industry sponsorship. I’m very grateful for all this support. One of my first times attending a major international conference, was the European Society of Clinical Microbiology and Infectious

Diseases. (ESCMID) in London, 2012. I was so excited! These are massive conferences and some of my AMR heroes were going to be there. One globally renowned French researcher was due to give a presentation, so I sat up the front, eagerly awaiting his talk, with visions of a movie star. (No google images back then!) However, he ended up looking like a cross between Dobby and Lucius Malfoy from Harry Potter! Never mind, his talk was excellent, and I have read many of his papers over the years. To finish on a personal ‘hobby’ note, I like to combine my loves of travel, bacteria and yeast into what I’ll call ‘wine education’. I have been fortunate to experience wine in many amazing locations, often combined with conference travel, and to share these experiences with family and friends (Figure 5). It’s a big wine world and there is still so much to explore! So, Lucy, I did have a good time, AND it was educational! A final thanks to the NZIMLS, and my awesome colleagues who have supported me over the years. And to everyone attending remember to be curious, to make new connections and to have fun and enjoy what you are learning!

Figure 5. Wine education

AUTHOR INFORMATION

Julie Creighton, DipMLS. FNZIMLS, Technical Lead and Clinical Lecturer 1,2 Department of Microbiology, University of Otago, New Zealand Microbiology, Canterbury Health Laboratories and West Coast District Laboratory, Christchurch, New Zealand Email: Julie.Creighton@cdhb.health.nz

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REFERENCES

1. PHF Science Antimicrobial Resistance Reports. [Available from: https://www.phfscience.nz/expertise/ infectious-disease/antimicrobial-resistance/] Accessed September 2026. 2. New Zealand National Antimicrobial Susceptibility Testing Committee, NZ NAC [Available from: https://www.nzmn.org. nz/nz-nac] Accessed September 2026. 3. Macesic N, Harris PN, Mo Y, Gomez-Simmonds A. Carbapenemases: epidemiology, detection and management in a changing global landscape. JAC Antimicrob Resist 2026; 8(3): d lag099. 4. Howard JC, Creighton J, Heffernan H, Werno A. Evidence of transmission of an NDM-5-producing Klebsiella pneumoniae in a healthcare facility in New Zealand. J Antimicrob Chemother 2016; 72: 949-951. 5. Creighton J, Heffernan H, Howard JC. Isolation of seven distinct carbapenemase-producing Gram-negative organisms from a single patient. J Antimicrob Chemother 2017; 72: 317319. 6. Creighton J, Anderson T, Howard JC, Dyet K, Ren X, Freeman JT. Co-occurrence of mcr-1 and mcr-3 genes in a single Escherichia coli in New Zealand. J Antimicrob Chemother 2019; 74: 3113-3116. 7. Creighton J, Howard JC, Anderson T. Serratia marcescens enzyme SME-2 isolated from sputum in New Zealand. JAC Antimicrob Resist 2023; 5(6): dlad126.

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REVIEW ARTICLE

Review on current β-thalassaemia therapeutic treatments and future research Navana McLachlan ABSTRACT

Thalassaemia is a group of inherited haemoglobin disorders characterised by defective globin chain synthesis, resulting in ineffective erythropoiesis, chronic anaemia, and significant clinical variability. While its genetic basis is well established, increasing evidence highlights the role of epigenetic regulation in modulating disease severity and as a target for therapy. This literature review examines three key therapeutic approaches for transfusion‑dependent β‑thalassaemia: iron chelation therapy, CRISPR–Cas9 gene editing targeting the BCL11A enhancer, and allogeneic hematopoietic stem cell transplantation (HSCT). Iron chelation remains essential for managing transfusion-induced iron overload, although current agents are limited by toxicity, adherence challenges, and an inability to address the underlying molecular defect. In contrast, CRISPR–Cas9-mediated disruption of BCL11A demonstrates significant potential by reactivating fetal haemoglobin (HbF) production, thereby restoring globin chain balance, though concerns regarding long-term safety and accessibility remain. HSCT is the only widely established curative option, with high success rates in well-matched, younger patients. Still, it is associated with risks such as graft-versushost disease and limited donor availability. Collectively, these approaches highlight the contrast between supportive, curative, and emerging molecular therapies, emphasising the need for safer, more accessible treatments that target the underlying genetic and epigenetic mechanisms of thalassaemia. Keywords: Thalassaemia, iron chelation therapy, CRISPR–Cas9, allogeneic haematopoietic stem cell transplantation, haemoglobin disorders. NZ J Med Lab Sci 2026; 80(3): 127:131

INTRODUCTION

Thalassaemia comprises a group of inherited haemoglobin disorders characterised by reduced or absent synthesis of globin chains, resulting in ineffective erythropoiesis and chronic anaemia. Haemoglobin is composed of protein subunits known as globin chains, primarily α, β, γ, and δ, whose expression is tightly regulated throughout human development (1). The genes encoding these globin proteins are organised into two distinct clusters: the α-globin locus on chromosome 16, containing HBA1 and HBA2, and the β-globin locus on chromosome 11, which includes HBB and several other developmentally regulated globin genes (1). Mutations affecting these loci disrupt globin production, leading to an imbalance in globin chain synthesis. The accumulation of excess unmatched globin chains damages erythroid precursor cells, leading to membrane instability, oxidative stress, and premature red blood cell destruction (1). Although the genetic basis of thalassaemia has been well established for decades, increasing evidence suggests that regulatory mechanisms controlling gene expression also contribute to disease severity and may represent important therapeutic targets (2). Clinically, thalassaemia presents with considerable phenotypic variability. Mild carrier states are often asymptomatic and may only be identified through routine haematological screening. In contrast, severe forms such as haemoglobin H disease and β-thalassaemia major are associated with profound anaemia and require lifelong clinical management (3). Treatment frequently involves regular blood transfusions combined with iron chelation therapy to prevent iron overload (4). Disease severity largely reflects the degree of imbalance between α and β-like globin chains. When excess globin chains accumulate within developing erythroid cells, they precipitate, damaging the cell membrane and leading to ineffective erythropoiesis and compensatory expansion of the bone marrow. Individuals carrying similar genetic mutations often display markedly different clinical outcomes, highlighting the importance of additional regulatory mechanisms in influencing disease expression (5). Thalassaemia represents one of the most common inherited blood disorders worldwide, with approximately 1.5% of the global population carrying a thalassaemia-associated mutation (6). Over recent decades, global migration has altered the epidemiology of the disease, increasing its prevalence in regions where it was historically uncommon and placing new demands on healthcare systems that may lack established screening and management frameworks.

This literature review will examine three key therapeutic strategies for transfusion-dependent β-thalassaemia, focusing on both current clinical management and emerging curative approaches. Specifically, it will evaluate the role of iron chelation therapy in mitigating transfusion-induced iron overload, assess advances in CRISPR–Cas9-mediated gene editing targeting the BCL11A enhancer to reactivate fetal haemoglobin production, and explore the efficacy and limitations of allogeneic haematopoietic stem cell transplantation as a curative treatment. Through critical analysis of these approaches, this review aims to highlight their respective benefits, limitations, and implications for the future development of more effective and accessible therapies.

REVIEW

Iron chelation Iron chelation therapy remains a central component of clinical management for patients with transfusion-dependent β-thalassaemia (TDT). Regular blood transfusions are required to maintain adequate haemoglobin levels and suppress ineffective erythropoiesis; however, repeated transfusions inevitably lead to progressive iron accumulation. Because humans lack a physiological mechanism for active iron excretion, excess iron accumulates in vital organs, including the liver, heart, and endocrine glands, leading to severe complications such as cardiomyopathy, liver dysfunction, and endocrine disorders (13). Consequently, the administration of iron-chelating agents is essential to prevent iron overload and improve long-term survival in patients with thalassaemia major. Several iron-chelating agents are currently used in clinical practice, including deferoxamine (DFO), deferiprone (DFP), and deferasirox (DFX) (7), each with distinct pharmacological properties and limitations. Deferoxamine, the oldest available chelator, is a hexadentate molecule produced by Streptomyces pilosus that binds ferric iron in a 1:1 ratio. Once bound, the iron– DFO complex is excreted primarily via the kidneys, while unbound DFO is eliminated through the biliary system. Importantly, DFO can directly remove iron from myocardial tissue, making it particularly effective at preventing cardiac complications associated with iron overload (7). Despite its efficacy, DFO therapy is limited by the requirement for parenteral administration, typically through prolonged subcutaneous infusion. This route of administration can significantly reduce patient adherence, particularly in paediatric populations. Furthermore, DFO has been associated with several adverse effects, including renal dysfunction, gastrointestinal disturbances, growth retardation, and dose-

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dependent ototoxicity and ophthalmic toxicity, indicating that long-term use may carry substantial clinical risks. In response to these limitations, orally administered iron chelators have been developed to improve patient compliance. Deferiprone, first introduced in the 1980s, is a bidentate lipophilic molecule that forms a stable 3:1 complex with iron and is rapidly absorbed following oral administration (8). Unlike DFO, deferiprone can penetrate iron-overloaded cells, including cardiomyocytes, facilitating the removal of intracellular iron and its transfer to circulating transferrin for excretion in the urine. Clinical studies have demonstrated that deferiprone can significantly reduce systemic iron stores and may be more effective than subcutaneous DFO in removing cardiac iron and improving cardiac function (9). However, the use of deferiprone is limited by potentially severe adverse effects, including agranulocytosis. Agranulocytosis is considered an unpredictable and potentially life-threatening reaction that necessitates regular haematological monitoring during treatment. Although clinical trials have reported no fatalities associated with agranulocytosis, the overall incidence of agranulocytosis, 1.5%; 1.1/100 patient‐ years of drug exposure, raises ongoing concerns regarding the safety of this therapy, particularly in paediatric patients (10). The mechanisms underlying deferiprone-induced neutropenia remain poorly understood, although in vitro studies suggest a possible toxic effect on myelopoiesis (11). As a result, the risk– benefit balance of deferiprone therapy continues to be debated in the literature. Deferasirox is a more recent development in iron chelation therapy and was designed to provide a once-daily oral treatment to improve patient adherence. Deferasirox is a lipophilic tridentate molecule that binds iron to form a stable complex, which is primarily excreted via the biliary system. Clinical studies comparing deferasirox with deferoxamine have demonstrated comparable efficacy in reducing iron burden, although some trials have suggested that suboptimal dosing may influence treatment outcomes (12). Deferasirox has generally been associated with improved treatment compliance due to its oral administration and convenient dosing schedule. However, adverse effects, including renal impairment, hepatic dysfunction, and gastrointestinal bleeding, have been reported, and in rare cases these complications have been fatal. Additionally, cases of Fanconi syndrome have been observed in a small proportion of patients, particularly among children and adolescents, indicating that careful monitoring of renal function is required during treatment (7). Combination chelation therapy has also been investigated as a strategy to enhance iron removal in patients with severe iron overload. The most extensively studied combination involves deferoxamine and deferiprone. These agents exhibit complementary pharmacokinetic properties, with deferiprone mobilising intracellular iron and transferring it to deferoxamine, which has a higher affinity for iron and facilitates its excretion (12). Studies suggest that this synergistic interaction may enhance the removal of hepatic and cardiac iron, leading to improved cardiac function and the reversal of iron-related endocrine complications (4). However, the safety profile of combination therapy appears similar to that observed with individual chelators, and further research is required to determine optimal treatment regimens. Despite the clear clinical importance of iron chelation therapy, several limitations remain in the current evidence base. Retrospective studies evaluating chelation therapies in patients with thalassaemia major often report a wide range of adverse effects, including gastrointestinal disturbances, elevated liver enzymes, and auditory or visual impairment (4). However, the interpretation of these findings is complicated by the underlying complications of thalassaemia itself, making it difficult to determine whether these effects are directly attributable to the chelating agents or to the disease process. Furthermore, many studies involve relatively small and heterogeneous patient populations, limiting the ability to establish clear associations

between treatment regimens, ferritin levels, and treatment discontinuation rates (7-10). Importantly, iron chelation therapy addresses a major complication of transfusion-dependent thalassaemia but does not directly correct the underlying defect in globin chain synthesis. As a result, patients remain dependent on lifelong transfusion therapy and continued chelation treatment. This limitation highlights a significant gap in current treatment strategies and underscores the need for therapies that target the disease’s primary molecular mechanisms. Future research should therefore aim to improve the safety and efficacy of existing chelating agents while also investigating alternative therapeutic strategies that target the underlying genetic and epigenetic mechanisms of thalassaemia. Larger, more homogeneous clinical studies are required to characterise long-term treatment outcomes better and to clarify the relationship between iron burden, treatment response, and adverse effects. In addition, emerging molecular therapies that modify globin gene regulation, such as gene editing and epigenetic modulation, may ultimately reduce reliance on transfusion and chelation therapy by addressing the fundamental causes of globin imbalance. CRISPR–Cas9 Gene Editing Targeting the BCL11A Enhancer Recent advances in gene-editing technologies have enabled the development of therapeutic strategies to modify the epigenetic regulation of globin gene expression in β-thalassaemia. One of the most promising approaches involves CRISPR–Cas9 mediated disruption of regulatory elements controlling the transcription factor BCL11A, a key repressor of fetal γ-globin expression in adult erythroid cells. The developmental switch from fetal haemoglobin (HbF, α₂γ₂) to adult haemoglobin (HbA) is largely regulated by BCL11A, which suppresses γ-globin transcription following birth (14). Experimental evidence has demonstrated that inhibition or reduction of BCL11A expression can reactivate fetal haemoglobin production, thereby compensating for the absence or dysfunction of β-globin chains in disorders such as β-thalassaemia and sickle cell disease (15). Preclinical studies have demonstrated that targeted disruption of erythroid-specific regulatory elements within the BCL11A enhancer can effectively increase HbF expression in haematopoietic stem and progenitor cells (HSPCs). Using CRISPR–Cas9 ribonucleoprotein complexes, Wu et al. (17) showed that cleavage at a GATA1-binding site within the +58 BCL11A erythroid enhancer resulted in a substantial reduction in BCL11A expression and the induction of γ-globin production. In edited CD34⁺ HSPCs derived from patients with β-thalassaemia, γ-globin expression increased significantly, restoring globin chain balance and improving erythroid maturation. Importantly, these edited stem cells retained their ability to engraft and differentiate in vivo, indicating that modification of the BCL11A enhancer does not appear to impair haematopoietic stem cell function (16). These findings support the hypothesis that disruption of erythroid-specific enhancer regions can selectively reactivate fetal haemoglobin without affecting BCL11A activity in nonerythroid tissues. Clinical translation of this approach has been investigated in the CLIMB-THAL-111 trial using CTX001, an autologous CRISPR–Cas9 gene-edited stem cell therapy (15). In this study, haematopoietic stem cells were collected from a patient with transfusion-dependent β-thalassaemia (TDT), edited ex vivo using a CRISPR–Cas9 system targeting the BCL11A enhancer, and reinfused following busulfan myeloablation. The edited cells demonstrated an allelic editing frequency of approximately 68.9%, resulting in substantial increases in fetal haemoglobin production. HbF levels increased from 0.3 g/dL (3.0g/L) pretreatment to 13.1 g/dL (131g/L) by 12 months post-infusion, while F-cell expression rose from 10.1% to 99.7% by six months and remained stable during the 18-month observation period. These results suggest that BCL11A enhancer disruption can effectively mimic the benign phenotype observed in hereditary persistence

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of fetal haemoglobin, where elevated HbF levels compensate for defective β-globin production. Despite these promising outcomes, several limitations and uncertainties remain. Although early clinical data demonstrate sustained HbF induction, the long-term safety and durability of CRISPR-mediated editing in haematopoietic stem cells remain to be fully established. Adverse events reported during the trial included neutropenia, pneumonia, abdominal pain, and venoocclusive disease. However, many of these complications were associated with the myeloablative conditioning regimen rather than with the gene-editing procedure itself (15). Additional cases of severe complications, including hemophagocytic lymphohistiocytosis and acute respiratory distress syndrome, have also been reported in ongoing trials, highlighting the need for continued monitoring of treatment safety. Furthermore, discrepancies exist between experimental models and clinical outcomes regarding editing efficiency and HbF induction levels. While preclinical studies often report editing efficiencies exceeding 80% in cultured HSPCs (17), clinical applications may demonstrate greater variability due to differences in patient genotype, stem cell quality, and editing efficiency. These inconsistencies suggest that additional regulatory factors may influence globin gene expression beyond BCL11A alone. Indeed, earlier studies have demonstrated that globin switching is controlled by complex interactions between cis-regulatory elements and trans-acting transcription factors, with BCL11A representing only one component of a broader regulatory network (14). Several important gaps in knowledge remain to be addressed. The long-term genomic stability of edited haematopoietic stem cells remains uncertain, particularly regarding the potential accumulation of off-target mutations and clonal expansion. In addition, further research is required to clarify the broader epigenetic mechanisms through which BCL11A interacts with chromatin remodelling complexes and histone modification pathways to silence γ-globin expression. Understanding these regulatory networks may enable the identification of additional therapeutic targets to enhance fetal haemoglobin induction. Future investigations should therefore focus on improving editing efficiency, reducing reliance on toxic myeloablative conditioning regimens, and exploring alternative epigenetic targets within the globin regulatory network. Advances in gene-editing precision, including base editing and prime editing technologies, may provide safer, more controlled methods for modifying globin gene regulation. Expanding knowledge of the epigenetic mechanisms that control globin switching will be essential for developing nextgeneration therapies to achieve durable and safe correction of β-globin disorders. Allogeneic haematopoietic stem cell transplant Allogeneic haematopoietic stem cell transplantation (HSCT) remains the only widely established curative therapy for transfusion-dependent β-thalassaemia (TDT). However, treatment outcomes vary considerably depending on patient characteristics, disease severity, and the presence of preexisting organ damage. Gaziev, J et al. (19) demonstrated encouraging engraftment outcomes in paediatric populations, particularly when an HLA-matched sibling donor is available. In a cohort of 60 patients with a mean age of approximately 7 years, successful engraftment occurred in nearly all recipients, with median neutrophil and platelet recovery times of approximately 20 days (18). Despite these promising results, complications remained evident, as several patients developed chronic graftversus-host disease (GVHD), and transplant-related mortality was reported within the early post-transplant period. These findings suggest that although HSCT can achieve sustained donor engraftment and transfusion independence in the majority of cases, transplant-related morbidity continues to represent a significant clinical challenge. Outcomes are particularly variable in high-risk patients, including those with advanced disease or pre-existing organ dysfunction

caused by chronic transfusion therapy and iron overload. Such patients, commonly classified as Class 3, historically demonstrate higher rates of graft rejection and transplant-related mortality (18). Nevertheless, attempts to improve outcomes in this population have led to the development of modified conditioning protocols. For example, Gaziev et al. (19) reported that an intensified conditioning regimen incorporating pre-transplant cytoreduction significantly reduced graft rejection rates and improved thalassaemia-free survival, achieving approximately 92% survival compared with 73% using earlier protocols. These findings highlight the importance of optimising conditioning strategies to enhance engraftment success in high-risk patients without substantially increasing treatment-related toxicity. Although emerging therapies such as gene replacement therapy have shown promise as potential curative approaches, their clinical implementation remains limited. Gene therapy must typically be administered before the development of severe iron overload or major organ damage and is currently restricted by high treatment costs and limited access to specialised healthcare services (20). Consequently, only a small proportion of patients globally can access this treatment, emphasising the continued reliance on HSCT as the primary curative option for many individuals with TDT. These limitations further underscore the importance of improving HSCT outcomes through enhanced pretransplant management, including optimisation of transfusion protocols, aggressive iron chelation therapy, and earlier identification of suitable transplant candidates. Although research is limited, a long-term study (21) provides further evidence for the benefits of HSCT in this patient population. It evaluated patients more than 20 years after transplantation and found that survivors reported health-related quality-of-life outcomes broadly comparable to those of the general population. Furthermore, transplanted patients demonstrated improved quality of life compared with individuals receiving conventional transfusion-based therapy. Importantly, the study suggested that patients transplanted at a younger age, particularly before 15 years, were more likely to achieve favourable long-term physical and psychosocial outcomes. These findings support the growing consensus that early transplantation, before irreversible organ damage develops, may significantly improve both survival and long-term quality of life. However, despite these encouraging outcomes, long-term complications remain an important clinical consideration. Chronic GVHD continues to represent a major source of late morbidity following allogeneic transplantation and may result in persistent complications, including endocrine dysfunction, growth retardation in paediatric patients, and long-term organ impairment. Nevertheless, survivorship studies indicate that outcomes are generally favourable for patients who successfully recover from the early post-transplant period. For instance, La Nasa et al. (21) reported that more than 90% of patients who survive the first two years after allogeneic HSCT become longterm survivors. Despite these positive findings, the literature highlights a significant gap in understanding the long-term effects of newer conditioning regimens and evolving transplant strategies. Consequently, further longitudinal research is required to characterise late complications better and optimise long-term survivorship care in patients undergoing HSCT for thalassaemia. What does this mean for New Zealand? Collectively, thalassaemia represents one of the most common monogenic disorders worldwide and is particularly prevalent among people of African, South Asian, Southeast Asian, and Mediterranean ancestry (22). This is increasingly relevant to New Zealand as the country’s population becomes more ethnically diverse, leading to a greater likelihood of individuals and families affected by thalassaemia. Statistics New Zealand, using data from Te Whatu Ora – Health New Zealand, recorded 277 hospitalisations for thalassaemia across all age groups (0-85+ years) in the year ended June 2023 (Figure 1) (23). In New Zealand medical laboratories, screening for thalassaemia

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and other haemoglobinopathies is routinely performed upon clinician’s request or when abnormalities are identified during HbA1c testing, enabling the referral of individuals with suspected haemoglobin variants for further investigation and diagnosis.

These data demonstrate that thalassaemia is an existing health concern in New Zealand and highlight the importance of awareness, early diagnosis, appropriate laboratory screening, and ongoing clinical management.

Figure 1. Number of hospitalisations with a diagnosis of thalassaemia in New Zealand by age group, year ended June 2023 - source: Statistics New Zealand using Te Whatu Ora – Health New Zealand hospitalisation data (23).

CONCLUSION

Thalassaemia continues to impose a substantial global health burden, reflecting its genetic basis, lifelong clinical management, and associated complications. While regular blood transfusions combined with iron chelation therapy have significantly improved survival, these approaches remain fundamentally supportive, addressing the consequences of globin imbalance rather than its underlying cause. Iron chelation is indispensable for preventing transfusion-induced iron overload; however, its lifelong nature, associated toxicities, and reliance on patient adherence limit its capacity to alter disease progression. As such, it highlights a critical gap between disease management and definitive treatment. In contrast, emerging molecular therapies aim to directly target the pathophysiology of thalassaemia by restoring the balance of globin chains. CRISPR–Cas9-mediated gene editing of regulatory elements such as the BCL11A enhancer represents a paradigm shift, offering the potential to reactivate foetal haemoglobin and functionally compensate for defective β-globin production. Early clinical data demonstrate substantial and sustained increases in HbF levels, with some patients achieving transfusion independence. However, the requirement for myeloablative conditioning, variability in editing efficiency, and unresolved concerns regarding long-term genomic safety underscore that this approach, while promising, remains in an early stage of clinical translation. These limitations also reflect the broader complexity of globin gene regulation, suggesting that targeting a single regulatory factor may not fully account for interindividual variability in treatment response. Allogeneic hematopoietic stem cell transplantation is the only widely established curative therapy, effectively replacing the defective hematopoietic system. Its success, particularly in younger patients with well-matched donors, demonstrates the feasibility of achieving long-term disease resolution.

Nevertheless, its clinical application is constrained by donor availability, transplant-related morbidity, and the risk of graftversus-host disease. Compared with gene editing, HSCT offers proven curative potential but lacks scalability and carries significant procedural risks, highlighting a trade-off between clinical maturity and accessibility. Collectively, these therapeutic strategies illustrate a continuum from supportive care to curative intent, each associated with distinct advantages and limitations. Iron chelation remains essential but non-curative; HSCT provides established but limited curative potential; and gene editing offers a highly targeted yet still evolving solution. The future of thalassaemia treatment will depend on integrating these approaches with advances in epigenetic understanding, improving the precision and safety of gene-editing technologies, and expanding access to curative therapies. Ultimately, achieving durable, safe, and globally accessible treatments will be critical to transforming thalassaemia from a chronic, life-limiting disorder into a universally manageable and potentially curable condition.

ACKNOWLEDGMENTS

I would like to acknowledge Massey University for providing access to academic databases, journal articles, and research resources that made this review possible, and the support and guidance provided by my lecturers throughout the development of this review. In addition, I would like to thank the Haematology Department at Medlab Central for their continued support and encouragement throughout my studies. Their professional guidance and ongoing support have been greatly appreciated.

AUTHOR INFORMATION

Navana McLachlan BSc, PGCertScTech Haematology Department, Medlab Central, Palmerston North, New Zealand

New Zealand Journal of Medical Laboratory Science 130

Email: navanamclachlan@icloud.com

REFERENCES 1.

Traeger-Synodinos J, Vrettou C, Sofocleous C, et al. Impact of α-Globin gene expression and α-Globin modifiers on the phenotype of β-thalassemia and other hemoglobinopathies: implications for patient management. Int J Mol Sci; 2024; 25(6): 3400. 2. Mettananda S. Genetic and epigenetic therapies for β-thalassaemia by altering the expression of α-globin genes. Front Genome Ed 2021; 3: 752278. 3. Bunyaratvej A, Sahaphong S, Bhamarapravati N, Wasi P. Quantitative changes of red blood cell shapes in relation to clinical features in β-thalassemia/HbE disease. Am J Clin Pathol 1985; 83(5): 555-559. 4. Uygun V, Kurtoglu E. Iron-chelation therapy with oral chelators in patients with thalassemia major. Hematology 2013; 18(1): 50-55. 5. Bank A. Understanding globin regulation in beta-thalassemia: it's as simple as alpha, beta, gamma, delta. J Clin Invest 2005; 115(6): 1470–1473. https://doi.org/10.1172/JCI25398. 6. Rao E, Kumar Chandraker S, Misha Singh M, Kumar R. Global distribution of β-thalassemia mutations: an update. Gene 2024; 896: 148022. https://doi.org/10.1016/j. gene.2023.148022 7. Adramerina A, Economou M. Challenges of iron chelation in thalassemic children. Thalass Rep 2024; 14(1): 1-9. https:// doi.org/10.3390/thalassrep14010001 8. Hider RC, Hoffbrand AV. The role of deferiprone in iron chelation. N Engl J Med 2018; 379(22): 2140–2150. https:// doi.org/10.1056/NEJMra1800219. 9. Brittenham GM, Griffith PM, Nienhuis AW, et al. Efficacy of deferoxamine in preventing complications of iron overload in patients with thalassemia major. N Engl J Med 1994; 331(9): 567-573. 10. Tricta F, Uetrecht J, Galanello R, et al. Deferiprone-induced agranulocytosis: 20 years of clinical observations. AM J Hematol 2016; 91(10): 1026–1031. https://doi.org/10.1002/ ajh.24479 11. Fioredda F, Forni GL. Deferiprone and idiosyncrasic neutropenia: light and shadow. Blood Adv 2024; 8(21): 56515652. https://doi.org/10.1182/bloodadvances.2024013479 12. Lal A, Porter J, Sweeters N, et al. Combined chelation therapy with deferasirox and deferoxamine in thalassemia. Blood Cells Mol Dis 2013; 50(2): 99-104.

13. Wallace DF. The regulation of iron absorption and homeostasis. Clin Biochem Rev 2016; 37(2): 51–62. 14. Sankaran VG, Orkin SH. The switch from fetal to adult hemoglobin. Cold Spring Harb Perspect Med 2013; 3(1): a011643. https://doi.org/10.1101/cshperspect.a011643 15. Frangoul H, Altshuler D, Cappellini MD, et al. CRISPR-Cas9 gene editing for sickle cell disease and β-thalassemia. N J Engl J Med 2021; 384(3): 252-260. 16. Wilber A, Hargrove PW, Kim YS, et al. Therapeutic levels of fetal hemoglobin in erythroid progeny of β-thalassemic CD34+ cells after lentiviral vector-mediated gene transfer. Blood 2011; 117(10): 2817–2826. 17. Wu Y, Zeng J, Roscoe BP, Liu P, et al. Highly efficient therapeutic gene editing of human hematopoietic stem cells. Nat Med 2019; 25(5): 776–783. https://doi.org/10.1038/ s41591-019-0401-y. 18. Bernardo ME, Piras E, Vacca A, et al. Allogeneic hematopoietic stem cell transplantation in thalassemia major: results of a reduced-toxicity conditioning regimen based on the use of treosulfan. Blood 2012; 120(2): 473– 476. https://doi.org/10.1182/blood-2012-04-423822 19. Gaziev J, Isgrò A, Sodani P, et al. Optimal outcomes in young class 3 patients with thalassemia undergoing HLA-identical sibling bone marrow transplantation. Transplantation 2016; 100(4): 925–932. 20. Wingard JR, Majhail NS, Brazauskas R, et al. Long-term survival and late deaths after allogeneic hematopoietic cell transplantation. J Clin Oncol 2011; 29(16): 2230–2239. https://doi.org/10.1200/JCO.2010.33.7212 21. La Nasa G, Caocci G, Efficace F, et al. (). Long-term healthrelated quality of life evaluated more than 20 years after hematopoietic stem cell transplantation for thalassemia. Blood 2013; 122(13): 2262–2270. 22. Li CK. New trend in the epidemiology of thalassaemia. Best Pract Res Clin Obstet Gynaecol 2017; 39: 16-26. https://doi. org/10.1016/j.bpobgyn.2016.10.013. 23. Health New Zealand – Te Whatu Ora. National minimum dataset: publicly funded hospital discharges by recorded external cause by age and sex 2023. [Available from: https:// tewhatuora.shinyapps.io/hospitals-web-tool]. Accessed September 2026. Copyright: © 2026 The author(s). This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

New Zealand Journal of Medical Laboratory Science 131

ORIGINAL ARTICLE

Cephalosporin and fluoroquinolone-resistant Gram-negative bacteria in the gut microbiota of healthy humans in Bahrain Khaled Ahmed Alnaimi, Mohd Shadab, Ronni Mol Joji, Abdulrahman Yousif Ismael, Ali Ebrahim Almahmeed and Mohammad Shahid

ABSTRACT

Objectives: The human gut harbours antibiotic resistance genes that can spread through various means. The study aimed to analyse the presence of fluoroquinolone and cephalosporin resistance genes in gram-negative bacteria isolated from stool samples of the healthy human population in Bahrain. Methods: Stool samples were collected from 29 healthy volunteers and cultured on cystine lactose electrolyte deficient agar to isolate enteric gram-negative bacilli. Initial isolate identification was by standard microbiological techniques. Representative isolates were confirmed by using the VITEK 2 Compact automated system. Antimicrobial susceptibility testing of all the isolates was performed using the disk diffusion method. Isolates exhibiting phenotypic resistance were further examined via polymerase chain reaction to detect the presence of genes linked to cephalosporin and fluoroquinolone resistance (blaCTX-M, blaTEM, blaSHV, qnrA, qnrB, and qnrS). Results: A total of 92 isolates were yielded from the stool cultures, with 78 (84.7%) identified as gram-negative bacteria. Escherichia coli accounted for 56.4% of the gram-negative isolates. Klebsiella pneumoniae and Klebsiella oxytoca made up 27% and 6.4% of the isolates, respectively. Other isolates included Enterobacter cloacae complex, Morganella morganii, Brevundimonas species, and Citrobacter koseri. Antimicrobial susceptibility testing unveiled the highest level of resistance against ampicillin (64%), followed by cefazolin (45%). Resistance was lowest towards imipenem and meropenem, at 5% and 3% respectively. Genotypic analysis revealed that 52% carried blaCTX-M-15. Among blaCTX-M–negative isolates, blaSHV (84%) was more common. Of 23 fluoroquinolone-resistant isolates, qnrS predominated (78%). Conclusion: The study’s findings revealed a high prevalence of blaCTX-M-15 and qnrS, indicating a pressing concern for public health. If antibiotic resistance continues to escalate within the population in the future, clinical services may face an overwhelming surge in cases of difficult-to-treat infections. Addressing the growing threat of antibiotic resistance requires a collaborative approach to ensure future effectiveness. Keywords: Antibiotic resistance, cephalosporin, fluoroquinolone, Gram-negative gut bacteria, healthy human. NZ J Med Lab Sci 2026; 80(3): 132:137

INTRODUCTION

The gut microbiota is one of the densest microbial ecosystems on earth. Although it has many positive effects on health, the high population mass of microbes in this ecosystem can result in the horizontal spread of antimicrobial resistance (AMR) genes to potentially harmful bacteria. In the earlier few decades, most studies on antibiotic susceptibility have used Escherichia coli as an indicator bacterium from the microbiome. Investigations using this method have significantly increased our knowledge of the prevalence, distribution, and related risk of AMR (1). Antibiotic resistance is a major global health and economic issue due to the rise of new resistance mechanisms. Overuse of antibiotics is a key driver of rising resistance worldwide. Lack of regulation around antibiotic access further exacerbates resistance in some countries. The World Health Organization (WHO) has stated that resistance spreads more where antibiotics can be obtained without a prescription for human and animal consumption (2). Common infections such as, urinary tract infections, foodborne infections and lung infections are becoming harder to treat with standard and combined antibiotics due to increasing resistance. These microbes have co-evolved with humans and play a crucial role in a multitude of activities to keep them healthy (3). The human gut resistome is a diverse set of genes and genetic material that confers antibiotic resistance. Factors like diet, travel and antibiotic use impact gut resistome. Moreover, geographical regions also play a role in antibiotic resistance (3). In the Gulf Cooperation Council countries (GCC), antibiotic resistance is rising alarmingly, especially during the peak travel seasons. The prevalence of multidrug resistance is also increasing which increases the prolonged hospitalisation and complications. Most GCC countries like United Arab Emirates (UAE), Qatar, Bahrain and Saudi Arabia attract tourists. Significant travel escalates acquiring and transmitting resistant genes (4). With one of the largest and most varied populations of foreigners, the UAE is one of the most popular commercial and tourism hubs (5). AMR development in the GCC has been linked to several factors, including a high rate of inappropriate antibiotic use (6), the widespread dispensing of antibiotics without a prescription at community pharmacies (7), the use of antibiotics as growth promoters in agricultural settings, sometimes without proper supervision (7), a strong inward migration pattern and mobile

populations (8). While antibiotic abuse and overuse are the primary causes of resistance, the fast transmission of antibioticresistant genes and bacteria throughout the world's ecosystems, including human populations, animals, and environments, also play a significant role in the emergence and spread of antibiotic resistance (9). The complete knowledge of the consequences of antibiotic resistance on human health is limited due to the paucity of quantitative data on such a pathway (10). This study was conducted to explore the presence of fluoroquinolone and cephalosporin resistance genes through molecular analysis in healthy human participants from diverse ethnic backgrounds residing in Bahrain for most of the time. Additionally, we also sequenced representative resistance amplicons (blaCTX-M & qnrS) to verify the specific gene types.

MATERIALS AND METHODS

This study was approved by the Research and Ethics Committee, College of Medicine and Medical Sciences, Arabian Gulf University (E38-PI-1-22). Stool samples were collected from healthy volunteers (January 2022-February 2022) after taking informed consent. The main inclusion criteria were healthy adults not currently on antibiotics or in the past three months. Exclusion criteria were antibiotic use in the past three months or known acute or chronic diseases requiring medication. Stool samples were inoculated on cystine lactose electrolyte deficient agar (CLED) for culture and incubated at 37°C for 24-hours. Isolates were sub-cultured for purity based on colony morphology. These were further identified by colony characteristics, Gram-staining and biochemical tests. Among 29 healthy human stool samples collected, 78 were Gram-negative bacteria. Biomerieux VITEK 2 Compact system was used to identify isolates that eluded manual identification methods. Due to limited resources, not all isolates could be identified using the Biomerieux VITEK 2 Compact system. Using the Kirby-Bauer disk diffusion method, antibiotic sensitivity testing (AST) was carried out. The results were read as per Clinical and Laboratory Standard Institute (CLSI) guidelines, 2021. The following antibiotic discs were used for AST: ampicillin (10 µg), ampicillin-sulbactam (10/10 µg), piperacillin-tazobactam (100/10 µg), cefazolin (30 µg), cefepime (30 µg), ceftriaxone (30 µg), ceftazidime (30 µg), cefoxitin (30 µg) imipenem (10

New Zealand Journal of Medical Laboratory Science 132

µg), meropenem (10 µg), gentamicin (10 µg), amikacin (30 µg), ciprofloxacin (5 µg), levofloxacin (5 µg), chloramphenicol (30 µg). In this study, gram-negative isolates that exhibited resistance to cephalosporins, and fluoroquinolones, were analysed using PCR screening to identify specific resistance markers. First, all gram-negative isolates showing cephalosporins resistance were screened for extended spectrum beta-lactamases gene (blaCTX-M, blaTEM and blaSHV). Initially the isolates were screened with primers designed for universal amplification of CTX-M-encoding genes. In house known blaCTX-M positive K. pneumoniae strain was taken as the positive control in this experiment. Among these isolates, if any lacked the blaCTX-M gene, they were further screened for the presence of blaTEM and blaSHV. Four selected Gram-negative isolates carrying blaCTX-M gene were amplified again with ISEcp1/ P2D primers to amplify the whole blaCTX-M gene. For gram-negative isolates resistant to Ciprofloxacin and Levofloxacin, PCR screening was performed to detect the presence of the qnrA, qnrB, and qnrS genes. The PCR cycles were performed with annealing temperatures specified in Table 1. Six representative amplicons were sequenced (four blaCTX-M positive and two qnrS positive genes). Online BLASTn tool (http://www.ncbi.nlm.nih.gov/BLAST/) was used to analyse the similarity of our nucleotide sequences. Further, multiple sequence alignment was done by Clustal Omega tool (https:// www.ebi.ac.uk/Tools/msa/clustalo/) and Jalview software.

RESULTS

Bacterial isolates A total of 29 stool samples were collected from healthy human volunteers. Among them 5 were Bahraini and 24 were nonBahraini volunteers, with 28 males and one female. NonBahraini volunteers were from India, Pakistan, Bangladesh, and Philippines as shown in Figure 1. Among the 78 GNB analysed, 44 (48%) were E. coli, followed by 21 (23%) K. pneumoniae among others (Figure 1). Antibiotic susceptibility testing The antibiotic susceptibility testing showed that bacteria exhibited the highest level of resistance to ampicillin, with a prevalence of 64% among the tested isolates. Following closely behind, cefazolin displayed a resistance rate of 45%, while ciprofloxacin exhibited resistance in 21% of the isolates. Gentamicin showed a lower resistance rate of 10%. In contrast, imipenem and meropenem demonstrated relatively low resistance rates of 5% and 3%, respectively. However, it is important to note that resistance to these carbapenem antibiotics was observed in a few isolates only. Two E. coli, one K. pneumoniae, and one M. morganii were found to be resistant to imipenem. Additionally, one E. coli and one M. morganii were resistant to meropenem. Table 2. encompasses a range of different antibiotics and their corresponding resistance rates among the tested isolates.

Molecular detection of antibiotic resistance markers Conventional PCR was used to screen for the presence of resistance markers in all isolates that displayed phenotypic resistance to the corresponding antibiotics. Detection of blaCTX-M All isolates (n=52) that showed resistance to ampicillin and/or other antibiotics in the cephalosporin class of drugs were tested by PCR to detect the blaCTX-M gene. Of these, 27 (52%) were positive and 25 (48%) were negative (Figure. 1). The 27 isolates that tested positive included 14 E. coli, 9 K. pneumoniae, 2 M. morganii, 1 K. oxytoca, and 1 Citrobacter koseri (Figure 1; Supplementary Figure 1). Detection of blaTEM, and blaSHV All the isolates that tested negative for blaCTX-M (n=25) were screened for blaTEM and blaSHV genes. These 25 isolates comprise of E. coli (n=14), K. pneumoniae (n=7), K. oxytoca (n=2), and Enterobacter cloacae complex (n=2). Of these, 12% were positive for blaTEM, while 88% tested negative. For blaSHV, 84% tested positive while 16% were negative. Nine isolates of E. coli, 7 K. pneumoniae, 1 each of K. oxytoca, and Enterobacter cloacae complex were positive for blaSHV and negative for blaTEM. One Enterobacter cloacae complex and three E. coli isolates were negative for both blaTEM, and blaSHV. Two E. coli isolates, and one K. oxytoca were positive for both blaTEM and blaSHV. Detection of fluoroquinolone resistance genes: qnrA, qnrB, and qnrS The presence of qnr genes was determined by conventional PCR on 23 isolates that showed resistance to ciprofloxacin and levofloxacin. For each isolate distinct primer sets were utilized to amplify the qnrA, qnrB, and qnrS genes (Table 1). 18 of the 23 (78%) isolates were tested qnrS positive (Figure 1), whereas 5 (22%) were found negative. Nine E. coli, eight K. pneumoniae, and one K. oxytoca were found harbouring qnrS gene among the 18 positive isolates. On the contrary, two isolates, one K. pneumoniae, and one K. oxytoca were positive with qnrB gene (Figure 1). And no isolates showed harbouring a qnrA gene. The prevalence of ARGs in all Gram-negative isolates is shown in Figure 1. Sequencing of ISEcp1/P2D and qnrS blaCTX-M gene is carried by ISEcp1 insertion sequences. To sequence the full gene region, we reamplified 4 representative blaCTX-M positive DNA samples with ISEcp1/P2D primer set, and the amplicons were sent for sequencing. All the amplicons selected here, were related to E.coli. To confirm the fluoroquinolone resistance genes, two qnrS positive amplicons were also sent for sequencing. Total 6 representative amplicons were sent for sequencing to know the precise gene type (Geno

Figure 1. Distribution of samples, bacterial isolates and ARGs. New Zealand Journal of Medical Laboratory Science 133

Screen Lab (Campus Institute Pasteur de, France). Sequence analysis of blaCTX-M BLAST results showed 100% identity and 0.0 E-value aligning with blaCTX-M gene. Thus, ISEcp1/P2D product was confirmed containing and matching with the complete gene sequence of blaCTX-M gene inserted between the known insertion sequences. Further, multiple sequence alignment (MSA) was done using the reference sequence of blaCTX-M-15 gene (accession number NG_048935.1) from https://www.ncbi.nlm.nih.gov/pathogens/ refgene/ which confirmed all of the nucleotide sequences matching with blaCTX-M-15 subtype (11). Supplementary Figure 2 shows the MSA and Figure 2 shows a neighbour joining tree created by Jal view software showing clonal relationship of our DNA sequences with blaCTX-M-15 gene.

All the sequences were submitted to the NCBI GenBank which can be accessed by the following accession numbers OP852367, OP852368, OP852369, OP852370 for blaCTX-M-15 and OP852371, OP852372 for qnrS at https://www.ncbi.nlm.nih.gov/genbank/

Sequence analysis of qnrS gene The partial sequences of the qnr gene were obtained through screening primers. Upon similarity analysis using BLAST, these sequences were confirmed to be the qnrS gene associated with fluoroquinolone resistance. By aligning our qnrS sequences with reference alleles from NCBI refseq (https://www.ncbi. nlm.nih.gov/pathogens/refgene/), we found 100% identity and query coverage with eight qnrS subtypes: qnrS1, qnrS4, qnrS8, qnrS10, qnrS11, qnrS13, qnrS14, and qnrS15. This suggests that our isolates may harbour variants of any of these qnrS alleles. Unfortunately, due to the partial gene amplification, we couldn’t predict the specific gene subtype. The similarity pattern can be seen with NCBI MSA Viewer, version 1.25.0 in Supplementary Table 1. Primers used for detection of antibiotic resistance genes. Figure 3. Primer (target gene)

Sequence (5’ to 3’)

Amplicon size

Annealing temperature

Reference

Primers used for detection of cephalosporin resistance genes blaCTX-M ISEcp1 (F) P2D (R) blaTEM

F-ATG TGC AGY ACC AGT AAR GT R- TGG GTR AAR TAR GTS ACC AGA

593 bp

50 °C

(29)

F- AAA AAT GAT TGA AAG GTG GT R- CAG CGC TTT TGC CGT CTA AG

~1100 bp

55 °C

(30)

F- KAC AAT AAC CCT GRT AAA TGC R- AGT ATA TAT GAG TAA ACT TGG

936 bp

58 °C

(29)

58 °C

(29)

F- TTT ATC GGC CYT CAC TCA AGG 930 bp R- GCT CGC GGC CGG ATA ACG Primers used for detection of fluoroquinolone resistance genes blaSHV

qnrA

F-CAG CAA GAG GAT TTC TCA CG R-AAT CCG GCA GCA CTA TTA CTC22

630 bp

54 °C

(31)

qnrB

F- GGC TGT CAG TTC TAT GAT CG R- GAG CAA CGA TGC CTG GTA G

488 bp

54 °C

(31)

qnrS

F- GCA AGT TCA TTG AAC AGG GT R- TCT AAA CCG TCG AGT TCG CGC

428 bp

54°C

(31)

Table 2: Antibiotic resistance pattern of gut GNB isolates

Antibiotics

E. coli (n=44) R (%)

K. pneumoniae (n=21) R (%)

Gram Negative Isolates* (n=13) R (%)

Ampicillin (10µg)

20 (45)

-

8 (62)

Ampicillin/ Sulbactam (10/10µg)

4 (9)

3 (14)

2 (15)

0

1 (5)

1 (8)

Piperacillin/ Tazobactam (100/10µg) Cefazolin (30µg)

21 (48)

7 (33)

8 (62)

Cefoxitin (30µg)

1 (2)

3 (14)

2 (15)

Ceftazidime (30µg)

6 (14)

0

1 (8)

Ceftriaxone (30µg)

15 (34)

1 (5)

4 (31)

Cefepime (30µg)

7 (16)

0

1 (8)

Gentamicin (10µg)

3 (7)

3 (14)

2 (15)

Amikacin (30µg)

2 (5)

0

1 (8)

Ciprofloxacin (5µg)

9 (20)

6 (29)

1 (8)

Levofloxacin (5µg)

6 (14)

1 (5)

1 (8)

Imipenem(10µg)

2 (5)

1 (5)

1 (8)

Meropenem (10µg)

1 (2)

0

1 (8)

Chloramphenicol (30µg) 2 (5) 2 (10) 1 (8) *Includes K.oxytoca, E.cloacae, M.morganii, C.koseri, Brevundimonas; R: resistant; - : not tested as they have intrinsic resistance New Zealand Journal of Medical Laboratory Science 134

Figure 2. Neighbour joining tree showing clonal relationship of our blaCTX-M harbouring isolates (K5.2, K6.2, K7.2 and K11.4) with blaCTX-M-15 reference gene (NG_048935.1).

DISCUSSION

There is a wide variety of bacteria present in the human gut, many of which are resistant to different types of antibiotics. Antibiotic-resistant genes can be transmitted to humans from other sources, such as animals. This makes the gut resistome, a critical component of the antimicrobial resistance challenge (12). This is the first study conducted in Bahrain focusing on the resistance of cephalosporins and fluoroquinolones, as well as the identification of specific resistance genes present in the gut GNB of healthy individuals. Among all the studied bacterial isolates blaCTX-M was tested positive in 27 (52%). blaCTX-M was found highest in E. coli strain in the present study. This finding is in line with the study published by Bindayna KM. et al. in 2022, which revealed that blaCTX-M is the most prevalent AMR-conferring gene in E. coli (13). A study from South India reported the presence of blaCTX-M (62/65, 95.3%), blaTEM (11/65, 16.9%) and blaSHV (1/65, 1.5%) in the healthy human gut. Hence presence of blaCTX-M E. coli is an important finding of this study because the high rate of blaCTX-M E. coli intestinal carriage may lead to the sudden occurrences of infections in a hospital setting (14). According to another study, blaCTX-M was more common in human gut isolates (13.89%; 5/36) compared to poultry and cattle (15). Most blaCTX-M genes detected were found within E. coli isolates, which is consistent with previous reports of this species being a primary reservoir for such resistance genes. Notably, blaCTX-M genes were also identified in K. pneumoniae, Citrobacter spp. and Pseudomonas spp. In the current study, the presence of the blaCTX-M-15 gene was detected among the participants. According to a study conducted in Venezuela found that the blaCTX-M-15 gene was present in all the ESBL E. coli strains in a group of healthy children (16). Another study in China reported that the CTX-M-9 group was the most detected (74.5%), followed by the CTX-M-1 group (29.1%)(17). This potentially suggests about the widespread dissemination of these resistance elements among clinically significant gram-negative pathogens commonly implicated in healthcareassociated infections (15). A pilot investigation by scientists at the School of Medicine found that many antibiotic-resistant genes were present in the bacteria in the guts of healthy American children (18). In one Swedish study, the gut microbiome of 12 out of 18 Swedish students tested negative for bacterial isolates that produce extended-spectrum β-lactamase (ESBLs) prior to their travels, but they tested positive for ESBLs after arriving in India which is due to dissemination of antibiotic-resistant genes during travel (19). Another Dutch study examined the faecal carriage of several beta-lactamase genes in a population-based cohort of middle-aged and older residents of the community. The findings

revealed that blaTEM (53.0%) was the most often occurring beta-lactamase gene, followed by blaSHV (18.4%). Much less frequently (5.4%) found was blaCTX-M gene (20) which is in contrast to the present study findings. A study carried out on young Japanese individuals found that while 62.5% of them had a moderate detection of blaTEM, 21.9% showed modest detection of blaSHV. These findings indicate the likelihood of blaTEM and blaSHV spreading to some extent among gut bacteria in humans, particularly Enterobacteriaceae (21). For many years, fluoroquinolones were the go-to antibiotics for treating UTIs and other infections (22). However, due to their overuse, these antibiotics have developed several disturbing side effects such as tendinitis, neurotoxicity, and the emergence of Clostridium difficile infections (23). It is worth noting that fluoroquinolone resistant pathogens, particularly fluoroquinolone resistant E. coli, responsible for extra-intestinal infections, have seen a significant increase in the past two decades (24). During the study, it was found that K. pneumoniae and E. coli were resistant to fluoroquinolone. Nine isolates of E. coli and eight isolates of K. pneumoniae were tested positive for the qnrS gene. Except for two isolates that carried the qnrB gene, none of the other isolates had qnrA or qnrB. This finding supports the research conducted by Xiang Chen et al. (2012), which demonstrated the prevalence of qnr gene in humans, animals, and the environment. Moreover, the research study revealed that qnrS was present in diarrheal E. coli and human commensals, whereas qnrA and qnrB were not found (25). A study on healthy Dutch individuals found that 17.4% carried the ß-lactamase encoding blaCTX-M gene, 5.4% had qnrB, and none tested positive for the qnrS gene (26). According to the study, conducted in the Netherlands there is a strong correlation between international travel and high frequencies of ESBL-encoding gene blaCTX-M and quinolone resistance-encoding genes qnrB and qnrS acquisition. Before traveling, the prevalence of these genes was 9.0%, 6.6%, and 8.2% respectively. However, following travel, the prevalence of these genes significantly increased to 33.6%, 36.9%, and 55.7%, respectively(27). A recent study from Russia has reported the presence of MDR E. coli strains alongside rising fluoroquinolones and cephalosporins resistance in the gut microbiota of young children. The findings highlighted that for higher levels of resistance, particularly against ciprofloxacin and levofloxacin, a more complex mutational pattern was evident. At least three QRDR mutations—S83L and D87N/Y in gyrA, together with S80I in parC—were consistently observed, indicating that stepwise accumulation of chromosomal mutations is critical for achieving clinically significant resistance (28). The presence of numerous antibiotic resistance genes among the healthy volunteers in the study is indeed a cause for concern

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and merits serious consideration. As antibiotic resistance genes spread, the burden of antibiotic resistance increases, which indirectly affects human health and well-being. It can result in limited treatment options, thus increasing the rate of morbidity and mortality. This study had some limitations. First, the sample size was modest - a larger cohort may have provided additional insights. Second, participants' diversity was limited, as viewpoints from various ethnic/cultural backgrounds were underrepresented. Third, gender balance was impacted by the inability to recruit more female volunteers. Finally, screening of beta-lactamase resistance genes was narrowed to blaTEM and blaSHV in blaCTX-M negative isolates only, constrained by available resources. We could not even screen for other antibiotic-resistant genes due to resource constraints.

CONCLUSION

As this is the first study in this region focusing on gut bacteria in relation to commonly used drug classes such as cephalosporins and fluoroquinolones, it raises thought-provoking findings that warrant deeper examination. The discovery of antibiotic resistance genes harboured within the gut of volunteers with no history of antibiotic exposure holds concerning implications. If such resistance were to disseminate more broadly among community members and within local healthcare settings, it could significantly drive the burden on clinical services attempting to treat infectious diseases. Given the potential public health impacts suggested by these preliminary results, expanded investigation of this issue is strongly recommended. As resistance patterns evolve, maintaining awareness of the breadth of organisms and resistance genes involved will be important for public health decision-making. With continued diligent research and openminded assessment of evidence as it emerges, we can work to comprehend resistance dynamics on a deeper level. This will better empower efforts to safeguard treatment effectiveness through prudent stewardship and preventive strategies.

DATA AVAILABILITY STATEMENT

The datasets presented in this study can be accessed online at; https://www.ncbi.nlm.nih.gov/genbank/. Accession numbers can be found below: OP852367, OP852368, OP852369, OP852370, OP852371, OP852372. Supplementary figures available at https://www.nzimls.org.nz/ recent-journals

ACKNOWLEDGMENTS

This research was conducted within the framework of HH Sheikh Jaber Al-Ahmad Al-Sabah Academic Chair of Microbiology & Immunology at the Arabian Gulf University. All authors acknowledge Genoscreen Lab, France for providing sequence facility support.

CONFLICT OF INTEREST

The authors declare no potential conflict of interest.

AI DISCLOSURE STATEMENT

The authors used an AI-based language model (ChatGPT (GPT5.3), developed by OpenAI) to assist with grammar correction and language refinement. The authors reviewed and edited all content and take full responsibility for the final manuscript. Date/ Year of Use and Access: November 2025.

AUTHOR INFORMATION

Khaled Ahmed Alnaimi, MLM, Quality Control Analyst1 Mohd Shadab, MSc, Technologist1 Ronni Mol Joji, MD, PhD, Assistant Professor1 Abdulrahman Yousif Ismael, PhD, Vice President for Administrative Affairs / Dean of Student Affairs, Associate Professor1 Ali Ebrahim Almahmeed, MSc, Specialist1 Mohammad Shahid, MBBS, MD, PhD, FNZIMLS, PGDipHSM, Professor and Chairperson1

Department of Microbiology, Immunology and Infectious Diseases, College of Medicine and Health Sciences, Arabian Gulf University, Kingdom of Bahrain

1

Corresponding Author Professor Mohammad Shahid, Department of Microbiology, Immunology and Infectious Diseases, College of Medicine & Health Sciences, Arabian Gulf University, Manama, Bahrain. Email: mohammeds@agu.edu.bh

REFERENCES 1.

Penders J, Stobberingh E, Savelkoul P, Wolffs P. The human microbiome as a reservoir of antimicrobial resistance. Front Microbiol 2013; 4:87. 2. World Health Organisation. Antibiotic resistance 2020 [Available from: https://www.who.int/news-room/fact-sheets/ detail/antibiotic-resistance. (Accessed 22 October 2023). 3. Singh S, Verma N, Taneja N. The human gut resistome: Current concepts & future prospects. Indian J Med Res. 2019; 150(4): 345-358. 4. Pagani L, Dell'Amico E, Migliavacca R, et al. Multiple CTXM-type extended-spectrum beta-lactamases in nosocomial isolates of enterobacteriaceae from a hospital in northern Italy. J Clin Micro 2003; 41(9): 4264-4269. 5. Sonnevend Á, Abdulrazzaq N, Ghazawi A, et al. The first nationwide surveillance of carbapenem-resistant Enterobacterales in the United Arab Emirates – increased association of klebsiella pneumoniae CC14 clone with Emirati patients. Int J Infect Dis 2022; 120: 103-112. 6. Butt AA, Navasero CS, Thomas B, et al. Antibiotic prescription patterns for upper respiratory tract infections in the outpatient Qatari population in the private sector. Int J Infect Dis 2017; 55: 20-23. 7. Balkhy HH, Assiri AM, Al Mousa H, et al. The strategic plan for combating antimicrobial resistance in Gulf Cooperation Council States. J Infect Public Health 2016; 9(4): 375-385. 8. Le Quesne WJF, Baker-Austin C, Verner-Jeffreys DW, et al. Antimicrobial resistance in the Gulf Cooperation Council region: A proposed framework to assess threats, impacts and mitigation measures associated with AMR in the marine and aquatic environment. Environ Int 2018; 121: 1003-1010. 9. Hernando-Amado S, Coque TM, Baquero F, Martínez JL. Defining and combating antibiotic resistance from One Health and Global Health perspectives. Nat Microbiol 2019; 4(9): 1432-1442. 10. Zhou Z, Shuai X, Lin Z, et al. Association between particulate matter (PM)2.5 air pollution and clinical antibiotic resistance: a global analysis. Lancet Planet Health 2023; 7(8): e649-e659. 11. Waterhouse AM, Procter JB, Martin DM, et al. Jalview Version 2--a multiple sequence alignment editor and analysis workbench. Bioinformatics 2009; 25(9): 1189-1191. 12. American Society for Microbiology. The Gut resistome and the spread of antimicrobial resistance 2023 [Available from: https://asm.org/articles/2022/june/the-gut-resistome-andthe-spread-of-antimicrobial] (Accessed 9 November 2023). 13. Bindayna KM, Joji RM, Ezzat H, Jahrami HA. Antibioticresistance genes in E. coli strains in GCC Countries: a meta-analysis. Saudi J Med Med Sci 2022; 10(1): 1-11. 14. George EA, Sankar S, Jesudasan MV, et al. Molecular characterization of CTX-M type extended spectrum beta lactamase producing E. coli isolated from humans and the environment. Indian J Med Microbiol 2015; 33: S73-S79. 15. Tayyaba U AS, Singh A, Shahid M. Gut resistome with special reference to beta-lactamase-producers from human, poultry, and cattle from North-Indian region: a step towards “One Health” approach. NZ J Med Lab Sci 2022; 76(1): 1825. 16. Araque M, Labrador I. Prevalence of fecal carriage of CTX-M-15 beta-lactamase-producing escherichia coli in healthy children from a rural Andean village in Venezuela. Osong Public Health Res Perspect 2018; 9(1): 9-15. 17. Li B, Sun JY, Liu QZ, et al. High prevalence of CTX-M β-lactamases in faecal escherichia coli strains from healthy humans in Fuzhou, China. Scand J Infect Dis 2011; 43(3): 170-174.

New Zealand Journal of Medical Laboratory Science 136

18. Washington University in St. Louis. Gut microbes in healthy kids carry antibiotic-resistance genes. 2014 [Available from: https://outlook.wustl.edu/2014/apr/microbes/] 19. Bengtsson-Palme J, Angelin M, Huss M, et al. The human gut microbiome as a transporter of antibiotic resistance genes between continents. Antimicrob Agents Chemother 2015; 59(10): 6551-6560. 20. Mulder M, Arp PP, Kiefte-de Jong JC, et al. Prevalence of and risk factors for extended-spectrum beta-lactamase genes carriership in a population-based cohort of middle-aged and elderly. Int J Antimicrob Agents 2021; 58(3): 106388. 21. Ohashi Y, Fujisawa T. Detection of antibiotic resistance genes in the feces of young adult Japanese. Biosci Microbiota, Food Health 2017; 36(4): 151-154 22. Bratsman A, Mathias K, Laubscher R, et al. Outpatient fluoroquinolone prescribing patterns before and after US FDA boxed warning. Pharmacoepidemiol Drug Saf 2020; 29(6): 701-707. 23. Barlam TF, Cosgrove SE, Abbo LM, et al. Implementing an antibiotic stewardship program: guidelines by the Infectious Diseases Society of America and the Society for Healthcare Epidemiology of America. Clin Infect Dis 2016; 62(10): e51-e77. 24. Spellberg B, Doi Y. Editor's choice: the rise of fluoroquinoloneresistant escherichia coli in the community: scarier than we thought. Journal Infect Dis 2015; 212(12): 1853-1855. 25. Chen X, Zhang W, Pan W, et al. Prevalence of qnr, aac(6')Ib-cr, qepA, and oqxAB in escherichia coli isolates from humans, animals, and the environment. Antimicrob Agents Chemother 2012; 56(6): 3423-3427.

26. Malin JJ, von Wintersdorff CJH, Penders J, et al. Longitudinal fluctuations of common antimicrobial resistance genes in the gut microbiomes of healthy Dutch individuals. Int J Antimicrob Agents 2023; 61(3): 106716. 27. von Wintersdorff CJ, Penders J, Stobberingh EE, et al. High rates of antimicrobial drug resistance gene acquisition after international travel, The Netherlands. Emerg Infect Dis 2014; 20(4): 649-657. 28. Suzhaeva L, Egorova S, Polev D, et al. Fluoroquinolone resistance patterns in multidrug-resistant escherichia coli from the gut microbiota of young children. Antibiotics (Basel) 2026; 15(2): 140. 29. Shahid M. Citrobacter spp. simultaneously harboring blaCTX-M, blaTEM, blaSHV, blaampC, and insertion sequences IS26 and orf513: an evolutionary phenomenon of recent concern for antibiotic resistance. J Clin Microbiol 2010; 48(5): 1833-1838. 30. Ensor VM, Shahid M, Evans JT, Hawkey PM. Occurrence, prevalence and genetic environment of CTX-M betalactamases in enterobacteriaceae from Indian hospitals. J Antimicrob Chemother 2006; 58(6): 1260-1263. 31. Ciesielczuk H, Hornsey M, Choi V, et al. Development and evaluation of a multiplex PCR for eight plasmid-mediated quinolone-resistance determinants. J Med Microbiol 2013; 62(Pt 12): 1823-1827. Copyright: © 2026 The author(s). This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

REVIEWS OF INTEREST The reviews below can be accessed for their Abstracts, and “Open Access” is indicated where applicable. Unfortunately, the NZIMLS cannot provide full access due to copyright restrictions. However, full access may be available through various institution agreements with publishers. Any feedback on the Reviews can be sent to: editor@nzimls.org.nz. 1.

Suryawanshi R, Sharme, S etc al. Cortisol as a stress marker: analytical perspectives and challenges. Clin Chim Acta 2026; 15:121008. doi: 10.1016/j.cca.2026.121008. 2. Dey A, Gres V, Nelipovich A et al. Breaking barriers: how cytomegaloviruses manipulate myeloid cells to invade tissues. EMBO Reports 2026; 27(9): 2160-2179. doi: 10.1038/s44319-026-00763-y. [Open Access] 3. Rocca B, Ten Cate H. Antidotes for anticoagulation reversal. N Eng J Med 2026; 394(22): 2235-2254. 4. Wu R, Zachou G, Michael A. Diagnostic and prognostic performance of circulating biomarkers in ovarian cancer. Academia Oncol 2026; 3(1): doi: 10.20935/Acad Onco8189. [Open Access]. 5. Baker RE, Stamper AR, Burrows HA. Climate changes and infectious diseases. Nat Med 2026; 32(5): 1634-1645. 6. Bayani A, Mohammadi Z, Harrison P et al. Conventional and novel platelet parameters: clinical and laboratory approaches. Clin Chim Acta 2026; 15: 589. doi: 10.1016/j. cca.2026.120994. 7. Maqsood Q, Khan MU, Fatima T et al. Recent insights into breast cancer: molecular pathways, epigenetic regulation and emerging targeted therapies. Breast Cancer (Auckl) 2025; 13:19. doi: 10.1771//1782234251355663. [Open Access] 8. Rai N, Abdel-Wahab, Zhang L. Molecular and therapeutic frontiers in anemia therapy. J Clin Invest 2026; 136(10): e203401. doi: 10.1172/JCI1203401. [Open Access] 9. Liu Y. 100 years of the theory of the gene. EMBO J 2026; 27: 2517-2521. [Open Access] 10. Mantzouratou A, Mania A. Embryo ploidy correction: an update on mechanisms and insights from mosaic embryo

transfer. Hum Reprod 2026; 41(3): 319-331. doi: 10.1093/ humrep/deaf254. 11. Muller K, Gillard J, Typas A et al. Antibiotic-induced gut microbiome dysbiosis: risks and strategies for mitigation. EMBO J 2026; 45(11): 3642-3654. doi: 10.1038/s44318026-00796-y. [Open Access] 12. Hauser SL, Advances in multiple sclerosis. N Eng J Med 2026; 395(1): 54-68. doi: 10.1056/NEJMra2501195. 13. Yee A, Smith J, Kumar R et al. et al. Towards implementation of precision medicine biomarkers in early detection and prognostication of prostate cancer. Cancer Metas Rev 2026; 45(3): 50. doi: 10.1007/s1055-026-10357-8. [Open Access] 14. Kristiansen K. Digital diagnostic pathology between promise and professional erosion: artificial intelligence and the future of diagnostic sovereignty. J Clin Pathol 2026; 79(8): 523525. 5. doi: 136/jcp.2026-210728. [Open Access] 15. Mann SP, Savulescu J, Earp BD. When to create embryos or organoids for research. J Med Ethics 2026; 52(7): 459-464. [Open Access] 16. Teixeira AP, Aubel D, Fussengger M. Next generation programmable cell therapies for precision medicine. Nature Rev Genet 2026; 27(8): 585-602. doi: 10.1038/s41576-02600945-3. 17. Rakha OE, Rakha EA. Beyond root cause analysis: a practical systems engineering approach to incident investigation in histopathology. J Clin Pathol 2026; 79(10): 649-658. doi: /10.1136/jcp-2026-10875. 18. Williams DA, Clauw DJ. Fibromyalgia. N Eng J Med 2026; 395(3): 267-277. 19. Rugge M, Fraschini M, Orvito E et al. Patient safety in AIpowered diagnostic pathology. J Clin Pathol 2026; 79(4): 271-277. 20. Aronson NE, Musa AM, Satoskar AR. Leishmaniasis. N Eng J Med 2026; 394(20): 2026-2039. Prepared by: Michael Legge

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ORIGINAL ARTICLE

Prevalence and determinants of Chlamydia trachomatis infection among HIVinfected patients and pregnant women in Auchi and environs, Nigeria Favour Osazuwa, Osayande Osabuohien and Innocent Godfrey Iyare

ABSTRACT

Aim: Chlamydia trachomatis is the commonest bacterial sexually transmitted infection and known to increase the risk of the acquisition of human immunodeficiency virus (HIV). The aim of this study is to determine the prevalence and determinants of Chlamydia trachomatis using polymerase chain reaction among HIV-infected females and pregnant women in Auchi and environs. Methods: This cross-sectional descriptive study was carried out between January and July 2025 and included one-hundred and ninety subjects (95 HIV-infected women and 95 pregnant HIV-negative women). Endocervical swabs were collected for analysis of Chlamydia trachomatis DNA using polymerase chain reaction (PCR). Self-administered questionnaires were collected for information on participant socio-demographics, sexual history and sexual risk behaviours. Results: Overall, 25/190 (13.2%) subjects were Chlamydia trachomatis positive. Chlamydia trachomatis was significantly higher in HIV-infected subjects (21.1%) compared to pregnant HIV-negative women (5.3%) (Chi square 10.363, p value = 0.0013). Age of participants in both groups did not influence the distribution pattern of Chlamydia trachomatis. HIV-infected subjects with CD4+ T cells ≤ 200 cells/uL were significantly more infected, with a relative risk of 33% (OR: 0.1889, RR: 0.33, p = 0.031). Other significant risk factors of Chlamydia trachomatis infection were an early age at first sexual debut, a history of sexually transmitted infection (STI), employment and marital status. Conclusions: Asymptomatic infection with Chlamydia. trachomatis is highly prevalent among study subjects in Auchi and environs, routine screening among at-risk individuals and education on risk factors is however encouraged. Key words: Chlamydia trachomatis, human immunodeficiency virus (HIV), pregnant women, PCR, prevalence, people living with HIV (PLWH). NZ J Med Lab Sci 2026; 80(3): 138:142

INTRODUCTION

Chlamydia trachomatis (C. trachomatis) is an obligate intracellular parasite that replicates within the cytoplasm of host cells (1, 2). C. trachomatis infection is the most common bacterial sexually transmitted disease and more common in young people (3,4). Risk factors of Chlamydia infection includes, multiple sexual partners, use of oral contraceptives, and failure to use barrier methods of contraception (5). Others include a history of sexually transmitted diseases (STDs), cervical ectopy, low socioeconomic status, HIV sero-positivity, and seroconversion (5). Chlamydia trachomatis belongs to the genus Chlamydiae (6). C. trachomatis has 15 immunotypes (7). Clinical symptoms in women include vaginal discharge, dysuria, induced endocervical bleeding, irregular menstruation or intermenstrual bleeding and dyspareunia (8). Adverse outcomes associated with C. trachomatis in pregnancy include premature rupture of membranes, postpartum endometrial inflammation, and low birth weight, also chlamydial infection in pregnant women can enhance the risk of preterm labour, and perinatal death (9, 10). C. trachomatis vertical transmission can cause neonatal inclusion conjunctivitis and/or pneumonia (11). C. trachomatis also increases the risk of human immunodeficiency virus (HIV), because Chlamydia triggers an inflammatory response that draws CD4+ T-cells to the site of the infection (12). The presence of Chlamydia can lead to an increase in immune cell recruitment, particularly CD4+ T cells, which are targeted by HIV (12). This can provide more opportunities for HIV to infect these cells. The dual burden of HIV and Chlamydia trachomatis co-infection presents significant clinical and public health challenges, especially among vulnerable groups such as pregnant women. In semi-urban and rural settings like Auchi, where healthcare infrastructure may be limited, the lack of routine screening and reliable epidemiological data further compounds the problem. Consequently, the actual burden and dynamics of Chlamydia infections among HIV patients and pregnant women remain under-researched and poorly addressed, hindering effective intervention strategies. The aim of this study is to investigate the prevalence of Chlamydia trachomatis among HIV-infected patients and pregnant women in Auchi and environs.

METHODS

Study Design This cross-sectional descriptive study was done among HIVinfected female patients and pregnant women (aged 18 to 70 years) in Auchi and environs. Study participants were recruited from Edo State University Teaching Hospital (EDSUTH) Auchi, Edo State, Nigeria. Structured questionnaires were used to obtain socio-demographic and obstetric information such as educational level, occupation, socio-economic status, gravidity, gestational age, parity status, number of sexual partners in the past six months, history of STD, age, latest CD4+ count (for HIV patients). The sample size for this study was determined with the Cochran formula. While the minimum sample size was 138; we recruited 190 participants to increase the power of the study. Ethical Consideration Ethical approval was obtained from Health Research Ethics Committee (HREC) EDSUTH Auchi with the approval numberEDSUREC25/044. Informed consent was obtained from all the participating subjects following the standards of human experimentation and with the Helsinki Declaration of 1975, as revised in 2000. This was done via informed consent forms duly completed by all participants. Sample Collection All participant’s endocervical swabs were collected by a trained Clinician or Nurse. Swabs were placed in a sterile tube containing 1mL of phosphate-buffered saline (PBS) and transported on ice to the molecular laboratory within two hours of collection. Samples were stored at -70°C until DNA extraction. Sample Processing CD4+ T-cell count estimation CD4+ T-cell count was determined by flow cytometry (Partec, Germany), following the manufacturer’s instructions. 20µl of whole blood was placed in a Partec tube and 20µL of CD4+ T-cell monoclonal antibodies were added. The mixture was then incubated in the dark for 15 minutes at room temperature after which 800µL of buffer was added. The tube was then analysed

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using a flow cytometer to count and determine sample CD4+ T-cell values. DNA extraction Genomic DNA was extracted from endocervical swab samples using the QIAamp DNA Mini Kit (Qiagen, Hilden, Germany). Stepwise protocols were followed as recommended by the manufacturer’s instructions. Real-time PCR Real time PCR was used for the positive detection of Chlamydia trachomatis. Extracted DNA was amplified by PCR. Specific primers included one pair of oligonucleotide primers specific for a conserved 144 region of the Chlamydia trachomatis gene coding for the major outer membrane protein (MOMP). The sequences from 5ʹ to 3ʹ of the forward and reverse oligonucleotide primers are ‘TGA ACC AAG CCT TAT GAT CGA CGG A’ and TCT TCG AYT TTA CCT TTA GAT TGA’ respectively. In vitro DNA amplification was performed in a 0.5mL thin-walled microfuge tubes containing PCR premix, by adding 1µL each of forward and reverse primers, 1µL of DNA template and 17µL of molecular grade water. Samples were then placed in a Biometra T3 thermocycle (Jena, Germany) and subjected to the following reaction conditions: Pre-Denaturation at 95°C for 5min, Denaturation at 94°C for 1 min, annealing at 52°C for 1 min, extension at 72°C for 1 min, 25 cycles and final extension at 72°C for 7min. A positive control (C. trachomatis) and a negative reagent control (distilled water) were included in the PCR run. Statistical analysis Data was analysed using SPSS v.21. Association between sociodemographic characteristics was determined using ANOVA, Chi square and significance were inferred if p< 0.05.

RESULTS

A total of 25 out of 190 subjects sampled were positive for Chlamydia trachomatis and given a prevalence of 13.2% (Table 1). The prevalence of C. trachomatis was found to be significantly higher among HIV-infected subjects (21.1%), compared to asymptomatic pregnant women (5.3%) (Table 1). Age-related distribution of Chlamydia trachomatis among HIV– infected subjects showed C. trachomatis was numerically higher among those in the range ≥ 36 years but was not statistically significant (p=0.227). CD4+ T cell count levels was a significant predictor of a positive C. trachomatis assay among HIV-infected subjects, the percentage relative risk of C. trachomatis acquisition at CD4+ cell count <200 was 33%. C. trachomatis prevalence was found to be statistically higher among HIV-infected subjects with low CD4+ T cell counts (Table 2). The association between participant’s sexual behaviour and C. trachomatis infection was investigated. Having a history of sexually transmitted disease/infection and age at first at sexual debut (coitarche) were significant cofounders contributing to acquisition of C. trachomatis (Table 3). The number of sexual partners was not associated with C. trachomatis acquisition (p=0.4022). The socio-demographic characteristics of the study population were analysed with respect to their PCR results for C. trachomatis. Among the pregnant women, only employment status had a significant association with being positive for C. trachoma (p< 0.001) (Table 4). Marital status, parity status and educational status were not associated with infection with C. trachomatis. Among the HIV-infected subjects, the analysis of their socio-demographic characteristics/parameters found out that only marital status had association with C. trachomatis, HIVinfected subjects who reported to be single (unmarried) had a significantly higher prevalence (p< 0.001) (Table 5). Age was not a significant factor (p= 0.211), in the prevalence and distribution of C. trachomatis among pregnant women in our study area (Table 6).

Table 1. Prevalence of Chlamydia trachomatis among the study population Chlamydia trachomatis status

Subjects

Positive (13.2%)

Chi square

p-value

20 (21.1)

10.363

0.0013*

5 (5.3)

-

-

HIV-infected CT Positive (n=95) Pregnant CT Positive (n=95) *Statistically significant (p< 0.05) Table 2. Association between Chlamydia trachomatis infection and CD4+ cell counts among HIV-infected subjects CD4+ Cell Counts < 200 (n=22)

C. trachomatis (+)

95% CI

RR

OR

p-value

2

0.039, 0.900

0.33

0.1889

0.031*

-

-

-

≥ 200 (n=53) 18 *Statistically significant; RR: Relative Risk; OR: Odd Ratio

Table 3. Association between C. trachomatis positivity and sexual behaviours of all study participants Sexual Behaviours Age at first sexual debut ≤ 14 (n=29) 15-19 (n=97) 20-25 (n=58) > 25 (n=6) History of STI Yes (n=53) No (n=137) Total *Statistically significant (p< 0.05)

Chlamydia trachomatis Positive (%)

p value

15 (60.0) 7 (28.0) 3 (12.0) 0

<0.001*

23 (92.0 2 (8.0) 25

<0.001*

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Table 4. Association between socio-demographic characteristics and Chlamydia trachomatis among pregnant women Parameter

Number studied (%)

Employment status Employed 63(66.3) Unemployed 30(31.6) Apprentice 2(2.1) Marital status Married 43(45.3) Single 29(30.5) Divorced 23(24.2) Education Primary 16(16.8) Secondary 54(56.8) Tertiary 25(26.3) Parity Primigravidae 72(75.8) Multigravida 23(24.2) Total * Statistically significant (p<0.05)

C. trachomatis Positive (%)

Chi square

p-value

1(20) 2(40) 2(40)

18.2345

<0.001*

1(20) 2(40) 2(40)

1.4005

0.4964

2(40) 1(20) 2(40)

3.3164

0.1904

3(60) 2(40) 5

0.6316

0.4268

Table 5. Association between socio-demographic characteristics and Chlamydia trachomatis among HIV-infected subjects Parameter

Frequency (%)

Employment status Employed 49(51.5) Unemployed 37(37.9) Apprentice 9(9.5) Marital status Married 61(64.2) Single / Divorced 11(11.6) Widowed 23(24.2) Education Primary 12(12.6) Secondary 53(55.8) Tertiary 30(31.6) Total 95 * Statistically significant (p<0.5)

C. trachomatis Positive (%)

Chi square

p-value

9(45) 10(50) 1(5)

1.0159

0.6017

7(35) 8(40) 5(25)

10.465

0.0054*

2(10) 13(65) 5(25) 20

0.3508

0.8391

Table 6: Effect of age on the prevalence of Chlamydia trachomatis among the studied population Age (years) HIV-infected patients 18-24 25-30 31-55 ≥ 36 Pregnant women 18-24 25-30 31-55 ≥ 36 ns Not statistically significant

DISCUSSION

C. trachomatis Positive (n)

C. trachomatis Negative (n)

1 1 1 17 2 1 1 1

In this study, the overall prevalence of C. trachomatis infection, assessed by PCR was 13.2%. This reported prevalence report is close to other study reports of 9.6% and 7.3% reported in Kano (13) and Ibadan (14) respectively. Higher prevalence of C. trachomatis (34%) was reported in Eket et al, (15). These observed differences in C. trachomatis prevalence shows that its distribution is population specific. The prevalence of C. trachomatis in HIV-infected subjects was 21.1%, while amongst pregnant women it was 5.3%, statistically, a significant difference was found (p = 0.0013). This shows that genital Chlamydia asymptomatic carriage was higher among HIV-infected subjects than among pregnant women in our study area. Generally, asymptomatic Chlamydia infection has been

Chi square

p-value

1 3 18 53

4.3361

0.227ns

50 31 7 3

4.5119

0.211ns

documented to be more commonly found among HIV-infected and STI clinic patients than pregnant women and any other subject groups (16). Similar studies among HIV-infected individuals within Nigeria have produced varying Chlamydia prevalence reports, ranging from the very low 1.7% reported in Owerri (17), to the moderate to high prevalence of 12.7% in Port-Harcourt, Rivers State (18). In 2025, the mean prevalence of Chlamydia among adults in Sub-Saharan African region had been estimated to be 6.9% (19). The incidence of C. trachomatis found in this study and from other Nigerian studies have been found to be higher than those of developed counties. The current overall prevalence of C. trachomatis among at risk individuals in USA is placed at 5% (20). These differences might be due to level of awareness,

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qualitative and organised screening programs in developed countries. In this study, age was not a significant risk factor in the acquisition of C. trachomatis in our both subject groups, although some studies found a strong association between age and C. trachomatis (21, 22). It could be argued to avoid age stratification when organising screening for at risk individuals in our study area. In other words, all age groups of HIV-infected patients and pregnant women are capable of being infected with C. trachomatis in Auchi and environs, and as such there should be age-related priority group in Chlamydia and STI screening and control programs. The prevalence of C. trachomatis among the HIV-infected subjects was found to be significantly higher in subjects with CD4+ T cell count of ≤200. It has been documented that immunosuppression due to HIV may lead to acquisition of C. trachomatis and increases in Chlamydial disease conditions such as pelvic inflammatory disease (PID) in those infected (23). Lower CD4+ has been linked to increased risk of acquisition of C. trachomatis in HIV infected individuals (24). Some relationships have been hypothesized between HIV and C. trachomatis, majorly that the invasive intracellular pathogenesis of Chlamydia can cause substantial damage to the genital epithelial layer which may facilitate HIV infection (25). Also, immunological changes due to HIV infection may favour C. trachomatis (26). Furthermore, epidemiological studies have shown that untreated genital C. trachomatis infection can lead to an increased risk for heterosexual acquisition of HIV (27). We advocate routine screening for genital Chlamydia be encouraged in various antenatal clinics, STI clinics and general outpatient wards in Nigeria. Sexual behaviours and sexual risk histories were collected and their association with genital Chlamydia was investigated. Sexual debut (Coitarche) at age ≤ 14 and a history of STIs were found to be significant risk factors or Chlamydia acquisition. Risky sexual behaviours are primary cofounders and are reported to account for majority of acquisition of C. trachomatis, other sexually transmitted disease and HIV (28). It is advised that any individual with symptomatic STI must ensure proper diagnosis and treatment to prevent the risk of acquisition of other preventable diseases. Currently, there are no public education programs tied to sexual behaviours on a mass scale in our study region and the introduction of adequate education could help to mitigate or reduce infection with C. trachomatis. The socio-demographic characteristics found to be associated with asymptomatic Chlamydia trachomatis infection among the pregnant women were employment status as the unemployed among the subjects were found to have higher Chlamydia prevalence. It has been shown that socio-economic status which can be reflected from their employment status influences their sexual health and preponderance among study subjects in this area. Single (unmarried) and divorced HIV-infected patients were more infected, this may stem from risky sexual behaviours engaged in by this set of subjects, when they have multiple sexual partners. This finding can be corroborated with the works of Ogedengbe et al. (28), where it was found that C. trachomatis infections were higher among unmarried HIV-infected subjects and those with multiple sexual partners.

genital Chlamydia infection. Common risk factors of Chlamydia acquisition were risky sexual behaviours, unemployment among pregnant women, unmarried and divorced among HIV-infected subjects. This study was a one-centre study, future studies may provide better prevalence data by recruiting subjects from many healthcare providing centres, this will provide a more realistic prevalence data of C. trachomatis for our region.

CONCLUSION

6.

This study shows a high prevalence (13.2%) of asymptomatic infection with Chlamydia trachomatis in Auchi and environs. It clearly shows that C. trachomatis infection is more prevalent in HIV-infected females when compared to HIV-uninfected pregnant women. This study did not find any age-related priority in designing screening and control programs; however, this study encourages testing for Chlamydia among all age groups. It was also found that, HIV-infected subjects with CD4+ T cell count ≤ 200 cells/mL had a higher prevalence of asymptomatic

ACKNOWLEDGEMENTS

Sincere thanks to all study participants for providing consent and participation in this study. Special thanks to Mrs Aminat Daud, for assistance with laboratory analysis.

AUTHOR INFORMATION

Favour Osazuwa, PhD, Senior Lecturer Medical Laboratory Science1 Osabuohien Osayande, BMLS, Graduate Scholar1 Innocent Godfrey Iyare, PhD, Senior Lecturer and Head of Department1 Department of Medical Laboratory Science, Faculty of Applied Health Sciences, Edo State University, Edo State, Nigeria. 1

Corresponding author: Dr Favour Osazuwa, Department of Medical Laboratory Science, Faculty of Applied Health Sciences, Edo State University, Edo State, Nigeria. Email: favourdesires@gmail.com Article Preprint Osazuma F and Osayande O. Prevalence of Chlamydia trachomatis among HIV-infected patients and pregnant women in Auchi and environs. 28 October 2025. Version.1 Available at Research Square. https://doi.org/10.21203/rs.3.rs-7633218/v1

REFERENCES 1.

2.

3.

4.

5.

7.

8.

Omosigho PO, Ajide TE, Izevbuwa O E, et al. Seroprevalence of Chlamydia trachomatis and associated risk factors among HIV-positive women in North Central Nigeria. Infez Med 2024; 32(1): 52–59. Egbe NE, Bakori BF, Abdulsalam I, et al. Molecular screening for Chlamydia trachomatis among pregnant women attending antenatal clinics in Kaduna metropolis, Kaduna State, Nigeria. The Nigerian Health Journal 2017; 24(4): 1746-1752. World Health Organization. Prevalence and incidence of selected sexually transmitted infections: chlamydia trachomatis, neisseria gonorrhoeae, syphilis, and trichomonas vaginalis: methods and results used by WHO to generate 2005 estimates. Geneva: WHO 2011. [Available from www.who.int.] Acessed, August 15, 2025. Centers for Disease Control and Prevention. Sexually transmitted infections treatment guidelines, 2021. Atlanta, GA: US Department of Health and Human Services. 2021. [Available from www.CDC.com] Accessed July 5, 2025. Rashidi BH., Chamani-Tabriz L, Haghollahi F, et al. Effects of chlamydia trachomatis infection on fertility: a case-control study. J Reprod Infertil 2013, 14(2): 67–72. Arif ED. Chlamydiosis in human. One Health Triad. Faisalabad, Pakistan Unique Scientific Publishers 2023; 7379. Eashwernath RP. Prevalence of Chlamydia trachomatis infection among women of reproductive age group attending the gynaecology department in a tertiary care centre. (Doctoral dissertation). Rajiv Gandhi University of Health Sciences, India. 2017. Muhammad HID, Jatau ED, Olonitola OS, et al. Association of chlamydia serology with HIV in Nigerian women. NJBAS

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2016; 24(2): 15-22. Silva MJP, Florêncio MA,Gabiatti GL. Et al. Perinatal morbidity and mortality associated with chlamydial infection: a meta-analysis study. Braz J Infect Dis 2011; 15(6): 533– 539. 10. Osazuwa F, Ifueko OM. Seroprevalence of sexually transmitted diseases (herpes, chlamydia, and syphilis) in pregnant women in Warri, Nigeria. CHRISMED J Health Res 2017; 4(2): 155-156. 11. Metallinou D, Nanou C, Sarantaki A, et al. Chlamydial infection. Primary Health Care. IntechOpen, London 2021; 8(6): 81-89. http://doi.org/10.5772/intechopen.9650. 12. O’Connell CM, Ferone M E. Chlamydia trachomatis genital infections. Microb Cell 2016; 3(9): 390-403. 13. Nwankwo EO, Magaji SN. Prevalence of chlamydia trachomatis among patients attending infertility and STI clinic in Kano, North Western Nigeria. Afr Health Sci 2014; 14: 672-678. 14. Ajani TA, Oluwasola TAO, Anaedobe CG, et al. Correlates of genital chlamydia trachomatis in a cohort of women in Ibadan, Nigeria. Int J Reprod Contracept Obstet Gynecol 2017; 6(9): 12-16. 15. Agbulu BF, Ekuma AE, Umoh SG, Adie SE. Prevalence of Chlamydia trachomatis colonization against asymptomatic pregnant women in Eket and its environs, Akwa Ibom State, Nigeria. World J Appl Sci Technol 2025; 17(1): 202-225. 16. Bhattar S, Bhalla P, Chadha S, et al. Chlamydia trachomatis infection in HIV-infected women: need for screening by a sensitive and specific test. Infect Dis Obstet Gynecol 2013; 2013: 960769. 17. Ohanu EC, Kone JB, Ezeh GL. Prevalence and association of chlamydia and HIV infections among symptomatic and asymptomatic patients in Owerri, Nigeria. Int J Pathog Res 2025; 14(6): 137-143. 18. Lawson S, Ogbonnie E, Ikpeama RA, et al. Seroprevalence of chlamydia trachomatis among HIV patients and healthy female volunteers in Port-Harcourt, Rivers State, Nigeria. Int J Pathog Res 2023; 12(6): 136-141. 9.

19. Michalow J, Hall L, Rowley J, et al. Prevalence of chlamydia, gonorrhoea, and trichomoniasis among male and female general populations in sub-Saharan Africa from 2000 to 2024: a systematic review and meta-regression analysis. EClinicalMedicine 2025; 83: 103210. 20. Centers for Disease Control and Prevention. Chlamydia screening among STI patients. MMWR 2020; 69: 312-318. 21. Keshinro B, Crowell TA, Nowak RG, et al. High prevalence of HIV, chlamydia, and gonorrhoea among men who have sex with men and transgender women attending trusted community centres in Abuja and Lagos, Nigeria. Intl J AIDS Soc 2016; 19(1): 2127–2129. 22. Twarhima R, Rema JO. Cross-sectional determinants of STIs among men who have sex with men and transgender women in Kigali, Rwanda. Sex Transm Infect 2022; 105: 342–365. 23. Soper DE. Pelvic inflammatory disease. Obstet Gynecol 2010; 116: 419-428. 24. Wodarz D, Hammer DH. Infection dynamics in HIV-specific CD4 T cells: Does a CD4 T cell boost the host or the virus? Math Biosci 2007; 20991): 14-29. 25. Altes HK, Wodarz D, Jansen VA. The dual role of T helper cells in the infection dynamics of HIV and their importance for vaccination. J Theor Biol 2020; 214(4): 633–646. 26. Joyee AC, Thyagarajan SP, Riddy E, et al. Chlamydia infection in patients: its relation to HIV infection. Indian J Med Microbiol 2005; 23(1): 37-40. 27. Oharume IM. Knowledge, sexual behaviours, and risk perception of sexually transmitted infections among students of the Polytechnic Ibadan, Oyo State. Afr Health Sci 2020; 20(1): 39-44. 28. Ogedengbe S, Agbah MI, Omosigho PO. Seroprevalence of chlamydia trachomatis among HIV positive women in Bida, North Central Nigeria. Int J of Epid Infect 2020; 10: 23-28. Copyright: © 2026 The author(s). This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

NZIMLS CALENDAR 2027 Council meetings

Venue

Agenda items to office

25-26 February 2027

Christchurch

12 February 2027

27-28 May 2027

Christchurch

14 May 2027

1616-1 August 2027

Brisbane

August 2027

25-26 November 2027

Christchurch

12 November 2027

Examiner/Moderator Workshop 8 May 2027

Jetpark Hotel Auckland

Seminars (Dates subject to change) 20 February 2027

Haematology SIG

15 May 2027

Biochemistry SIG

18-20 Augu 2027 18-

South Pacific Congress

sharon@nzimls.org.nz Venue/Convenor Jetpark Auckland Mala.govender@awanuilabs.co.nz Jetpark Auckland Lukleo82@gmail.com / natasha.dutt@awanuilabs.co.nz Brisbane Convention Centre Sharon@nzimls.org.nz

New Zealand Journal of Medical Laboratory Science 142

The NZIMLS original Certificate of Incorporation, 9 April 1946

New Zealand Journal of Medical Laboratory Science 143

ORIGINAL ARTICLE

Role of Decorin and its relationship with biochemical variables in the serum of patients with osteoarthritis Sarah A Younus and Zahraa MA Hamodat ABSTRACT

Background: Osteoarthritis is a chronic joint disease characterised by cartilage loss, pain, and limited mobility. Leucine-rich small proteoglycans, specifically Decorin, play a vital role in maintaining the integrity of cartilage and the extracellular matrix. This study aimed to determine serum Decorin levels in patients with osteoarthritis and their relationship to parathyroid hormone (PTH), vitamin D, calcium, phosphorus, and body mass index (BMI). Methods: The study included 90 patients and 90 healthy individuals (males and females) aged 35 to 60 years. Patients were classified into two groups, based on clinical examination and radiographic findings: a mild‑symptom group and a severe‑symptom group. Decorin, parathyroid hormone and vitamin D levels were measured using ELISA, mineral levels were estimated using spectrophotometry. Results: The results showed a significant increase in Decorin levels in patients with osteoarthritis compared to the control group (34.68 ± 11.83 µg/L) and this increase correlated with disease severity. Decorin levels were higher in patients with severe osteoarthritis (41.87 ± 5.89 µg/L) compared to patients with mild osteoarthritis (22.85 ± 4.77 µg/L). The results also showed a strong positive correlation between Decorin and parathyroid hormone (PTH), body mass index and a strong negative correlation with vitamin D, calcium, and phosphorus, compared to healthy individuals. ROC curve analysis revealed high diagnostic accuracy (p ≤ 0.001) for all biomarkers. Decorin and vitamin D showed the highest discriminatory power (area under the ROC curve): 1.000 and 0.999, respectively, followed by parathyroid hormone (0.967) and calcium (0.895). Conclusions: These results confirm the high reliability of these biomarkers as sensitive tools for detecting joint degeneration and mineral imbalance and highlighted the potential of Decorin as a valuable biomarker for assessing arthritis severity and monitoring treatment effectiveness. Keywords: Decorin (DCN), osteoarthritis (OA), parathyroid hormone (PTH), Vitamin D, calcium, phosphorus. NZ J Med Lab Sci 2026; 80(3): 144:150

INTRODUCTION

Osteoarthritis (OA) is classified as a chronic degenerative joint disease that leads to significant health problems and physical disability, negatively impacting patients’ quality of life (1). Clinically, it is characterised by the loss of articular cartilage and pain during movement, which in advanced stages can lead to complete immobility (2-4). Statistically, OA is the second most common rheumatic disease after rheumatoid arthritis in clinical practice (5), with prevalence rates reaching tens of millions in the United States alone. Globally, estimates suggest that approximately 300 million people exhibit radiographic evidence of the disease, whether through overt symptoms or incidental findings (6). Although the primary cause of the disease is attributed to the progressive loss of articular cartilage, the modern understanding of the pathophysiology of OA emphasises the involvement of multiple tissue components within the synovial joint, including subchondral bone, synovial membrane, ligaments, and the muscles that stabilise the joint (7). As the condition progresses, the biological balance is disrupted, favouring the deterioration of joint tissues. This leads to synovitis, subchondral bone remodelling, and the formation of osteophytes, which manifest clinically as pain, stiffness, joint cracking, and reduced mobility (8). OA is diagnosed by combining clinical examination with radiographic findings (9), and its progression is classified into different stages based on the severity of the condition (9). Literature studies indicate that women have higher prevalence rates, particularly after age 50 (years) (10-12), and obesity is identified as a significant risk factor that increases the likelihood of developing osteoarthritis symptoms (13). Decorin (DCN) is classified within the small leucine-rich proteoglycans (SLRPs) family (14). These proteins are found in abundance in cartilage tissue, with DCN being one of the most abundant components of this family in this region (15). DCN possesses dual biological properties, including anti-fibrotic and anti-inflammatory activities (16). It also plays a pivotal role in fibrillogenesis, regulating fibre structure and determining fibre diameters to ensure tissue stability (17). In addition to its structural role, DCN is clinically associated with several inflammatory diseases, such as osteoarthritis, as well as various medical conditions including Ehlers-Danlos

syndrome, corneal diseases, and muscular dystrophy (18). Vitamin D is vital for regulating calcium levels and supporting bone health. Parathyroid hormone (PTH) works synergistically with vitamin D, playing a pivotal role in mineral metabolism and maintaining calcium and phosphate balance, thus ensuring overall bone health. Studies have shown that this balance is crucial, particularly in conditions related to bone regeneration, such as osteoporosis, a condition that shares several pathophysiological pathways with osteoarthritis (19). Furthermore, the association of these factors with bone microstructure in older adults underscores their importance and essential role in understanding and managing conditions like osteoarthritis (20). This study focuses on exploring the complex interaction and close correlation between these biomarkers in predicting osteoarthritis, opening new horizons for enhancing clinical management protocols and improving treatment efficiency for this condition.

MATERIALS AND METHODS

Ethical Approval Ethical approval for this study was obtained from the Scientific and Research Ethics Committee of the Nineveh Health Directorate (Session No. 212, December 4, 2024), and the research project was registered under reference number (2024228). Formal and prior written consent was obtained from all volunteer participants before the sample collection process began, with participants being informed of the research objectives and assured of the complete confidentiality of their personal data and anonymity. Population Study The study included 180 participants divided into two age- and sex-equivalent groups. The patient group consisted of 90 individuals with osteoarthritis (65 females and 25 males) aged between 35 and 60 years, who were clinically and radiologically with Kellgren Lawrence classification including 2 grades, (34 patients were grade II mild and 56 patients were grade IV severe) under the supervision of a specialist physician. The control group comprised 90 individuals who appeared healthy and without any joint pain or symptoms such as morning stiffness or bone cracking. Patients with other health conditions such as diabetes, immune disorders, cancer, kidney disease, thyroid disease, or

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those who had undergone previous knee surgery were excluded from the study. Sample collection took place at the Al-Salam Teaching Hospital in Mosul from September 5, 2024, to May 1, 2025, after obtaining formal ethical consent and written informed consent from all participants. Sample Collection The sample collection procedure involved drawing approximately 5 mL of venous blood from each participant. The collected blood was transferred into gel tubes and allowed to stand at room temperature for ten minutes to facilitate clotting. Subsequently, centrifugation at 3,000xg for five minutes to separate the serum. The isolated serum was then aliquoted into Eppendorf tubes and immediately stored at -80°C to maintain the stability of biochemical properties prior to laboratory investigations. Various Parameters Estimation Decorin was determined using enzyme-linked immunosorbent assay ELISA based on the sandwich method, using kit number RE2319H, according to the manufacturer’s instructions. Vitamin D, Parathyroid hormone (PTH): quantified using commercially available enzyme-linked immunosorbent assay ELISA kits (Sun Long) using kit number EL0320Ge and SL1342Hu respectively, according to the manufacturer’s instructions. Calcium levels and phosphorus levels in serum were measured using a BIOLABO test kit of French origin (Biolabs). Body mass index (BMI) was measured by dividing weight by the square of height according to the following equation: BMI (kg/m2) = weight (kg)/height (m2). Statistical analysis The data were analysed using (SPSS) Version 26. The results are expressed as mean ± standard deviation (SD); an independent t-test was used to compare the two groups. ANOVA test employed the differentiation between the two groups. Pearson’s correlation coefficient was also used to explore the relationship between the Decorin level and the variables studied. p-values of <0.05 were considered statistically significant (21).

RESULTS

Decorin levels in the participants’ serum showed a very high and significant increase (p ≤0.0001) in osteoarthritis patients, with levels (34.68 ± 11.83 µg/L) compared to the control group (8.72 ± 3.13µg/L). A clear effect of age on Decorin concentration was also observed. The older age groups showed a significantly higher (p ≤0.0001) increase in patients (38.06 ± 11.14 µg/L) and healthy individuals (9.51 ± 2.94 µg/L) compared to the younger age groups, which recorded (23.58 ± 5.60 µg/L) for patients and (6.67 ± 2.68 µg/L) for healthy individuals. Regarding gender variation, a significant increase (p ≤0.0001) was observed in healthy females (10.48 ± 1.56 µg/L) compared to males (4.41 ± 1.27 µg/L). This was even more pronounced in the patient group, where the Decorin level in female patients was significantly higher (p ≤0.0001), reaching (38.47 ± 11.52 µg/L) compared to male patients (24.83 ± 5.01 µg/L). Body mass index (BMI) was directly correlated with Decorin levels; in the healthy group, levels were significantly higher (p ≤0.0001) in overweight individuals (8.93 ± 3.05 µg/L) compared to normal weight (7.23±2.86 µg/L) and reached their highest levels in obese individuals (10.54 ± 2.80 µg/L) with a very high significant increase (p ≤0.0001). Similarly, in the osteoarthritis patient group, a significant increase (p ≤0.0001) was recorded for overweight individuals (29.37 ± 10.49 µg/L), while obese patients showed the most significant increase (p ≤0.0001) at a level of (37.61 ± 11.57 µg/L), confirming the close relationship between obesity and Decorin concentration in Table 1. Results in Table 2 show a very high and significant increase (p ≤0.0001) in the level of body mass index (BMI) in osteoarthritis patients (29.46 ± 2.54 kg/ m2) compared to healthy people (25.73 ± 3.14 kg/m2). There was a highly significant decrease (p ≤ 0.0001) in the level of calcium in osteoarthritis patients

(1.84 ± 0.22 mmol/L) compared to healthy people (2.20 ± 0.16 mmol/L). Highly significant decrease (p ≤0.0001) in the vitamin D levels was observed in OA patients (35.74 ± 15.95 nmol/L) compared to healthy participants (83.97±10.53 nmol/L). There was observed a highly significant increase (p ≤ 0.0001) in the parathyroid hormone level in osteoarthritis patients (14.38 ± 5.50 pmol/L) compared to healthy people (3.83 ± 0.40 pmol/L) and a highly significant increase (p ≤0.0001) in the level of phosphorus in osteoarthritis patients (1.46 ± 0.40 mmol/L) compared to healthy people (1.24 ± 0.15 mmol/L). The results in Table 3 show a highly statistically significant increase (p ≤ 0.001) in Decorin levels in patients with mild cases (22.85 ± 4.77 µg/L) compared to healthy individuals (8.72 ± 3.13 µg/L), with a further significant increase in severe cases (41.87 ± 5.89 µg/L). Regarding calcium levels, a highly statistically significant decrease (p ≤ 0.0001) was observed in patients with severe cases (1.69 ± 0.11 mmol/L) compared to both healthy individuals (2.20 ± 0.16 mmol/L) and those with mild cases (2.09 ± 0.07 mmol/L). In contrast, phosphorus levels showed a statistically significant increase (p ≤0.001) in patients with severe cases (1.76 ± 0.40 pmol/L). Parathyroid hormone (PTH) levels in patients with osteoarthritis (OA) were also significantly higher (p ≤ 0.001), rising from (3.83 ± 0.40 pmol/L) in healthy individuals to (9.60 ± 3.82 pmol/L) in mild cases, and reaching (17.29 ± 4.19 pmol/L) in severe cases. Conversely, vitamin D levels showed a gradual and highly significant decrease (p ≤ 0.0001) associated with disease severity, decreasing from (83.97 ± 10.53 nmol/L) in healthy individuals to (55.61 ± 3.42 nmol/L) in mild cases, and reaching their lowest levels in patients with severe cases (23.69 ± 3.67 nmol/L). compared to the control group (1.24 ± 0.15 mmol/L). Furthermore, parathyroid hormone (PTH) levels in osteoarthritis (OA) patients showed a significant increase (p ≤ 0.001), rising from (3.83 ± 0.40 pmol/L) in healthy individuals to (9.60 ± 3.82 pmol/L) in mild cases, and reaching their lowest levels in patients with severe cases (23.69 ± 3.67 nmol/L).and it showed a high significant decrease (p ≤ 0.001) in the severe case compared to the mild case. Table 3 showed that there was high significant (p ≤ 0.001) in BMI levels among the study groups. OA patients with mild cases exhibited a significantly higher BMI (28.22 ± 2.93 kg/m2) compared to healthy people (25.73 ± 3.14 kg/m2). Similarly, a substantial and highly significant increase (p ≤0.0001) was observed in severe cases (30.22 ± 1.95 kg/m2) when compared to both the healthy control group and those with mild disease severity, indicating a progressive increase in body mass index (BMI) associated with the progression of osteoarthritis. Effect of Decorin level and its correlation with some biochemical variables and body mass index in control and osteoarthritis patients. The results, as shown in Table 4, showed that the relationship of Decorin with each of the element’s calcium (r = -0.044; p ≤0.046), phosphorus (r = 0.172; p ≤0.105), parathyroid hormone (r = 0.382; p ≤0.02) in the control case showed no significant relationship with each of the elements, and the relationship was negative, and showed a high significant relationship with both calcium (r = -0.790; p ≤0.0001) and phosphorus (r = 0.644; p ≤ 0.0001) in osteoarthritis patients case and it was positive. On the contrary, it showed no significant relationship with parathyroid hormone (r = 0.105; p ≤ 0.324) in healthy control, while parathyroid hormone (r = 0.825; p ≤0.0001) there was a high significant relationship in osteoarthritis patients. In vitamin D, showed a significant relationship with healthy control (r = 0.211; p ≤0.046) and very high significant (r = -0.825; p ≤0.0001) in osteoarthritis patients. While there is no significant between body mass index in healthy control was showed (r = -0.019; p ≤0.862), While very high significant in osteoarthritis patients was show (r = 0.604; p ≤0.0001) it is that the effect of Decorin is as the severity of the disease progresses compared with healthy control. The results of the receiver operating characteristic (ROC) curve analysis showed good diagnostic efficacy for the studied

New Zealand Journal of Medical Laboratory Science 145

biological variables in differentiating between the study groups. The area under the curve (AUC) was calculated, and optimal cutoff points were determined to ensure the highest levels of sensitivity and specificity. The results, shown in Figure 1, Figure 2, and Table 5, reveal that all variables achieved high statistical significance (p ≤ 0.001). Decorin led the results with an AUC of 1.000 at a cutoff point of 45.74 µg/L, followed by vitamin D with an AUC of 0.999 at a cutoff point of 12.41 nmol/L. PTH and Calcium also demonstrated excellent discriminatory power, with AUCs of 0.967 and 0.895 at cutoff points of (18.06 pmol/L) and (1.75 mmol/L), respectively. Furthermore, the BMI demonstrated remarkable efficiency, with an area under the curve (AUC) of 0.794 at a

cutoff of 2.50. Although BMI showed a sensitivity of 78.6% and a specificity of 75.8%, its predictive accuracy was lower compared to biochemical marker compared with Decorin and parathyroid hormone level. These results confirm that elevated Decorin and parathyroid hormone levels, compared to low vitamin D and calcium levels, provide a comprehensive diagnostic picture that accurately reflects the severity of joint tissue degeneration and mineral imbalance in affected individuals. Achieving 100% specificity at the cutoff points for parathyroid hormone and vitamin D further strengthens the reliability of these markers as effective diagnostic tools that contribute to the accurate detection of the condition and support the clinical diagnostic pathway.

Table 1. Characteristics of serum Decorin level in healthy controls and osteoarthritis patients in relation to age, gender and BMI categories Decorin (µg/L) Controls, n= 90

Patients, n=90

8.72 ± 3.13

34.68 ± 11.83

Decorin level (µg/L) according to age Controls

Patients

(35-45), n= 25

(46-60), n= 65

(35-45), n= 21

(46-60), n= 69

6.67 ± 2.68

9.51 ± 2.94

23.58 ± 5.60

38.06 ± 11.14

Decorin level (µg/L) according to gender Controls

Patients

Female, n= 64

Male, n= 26

Female, n= 65

Male, n= 25

10.48 ± 1.56

4.41 ± 1.27

38.47 ± 11.52

24.83 ± 5.01

Decorin level (µg/L) according to BMI* Normal weight, n= 26 7.23 ± 2.86

Controls Overweight, n= 48 8.93 ± 3.05

Obese, n= 16 10.54 ± 2.80

Normal weight n= 0 0

Patients Overweight n= 32 29.37 ± 10.49

Obese n= 58 37.61 ± 11.57

*BMI categories are defined as: Normal weight (18.5-24.9 kg/m2), Overweight (25-29.9 kg/m2) and Obese (≥ 30 kg/m2).

Table 2. Level of biochemical parameters and BMI in osteoarthritis patients compared to controls

Parathyroid hormone (PTH) (pmol/L)

Controls n = 90 3.83 ± 0.40

Patients n = 90 14.38 ± 5.50 ***

Vitamin D (nmol/L)

83.97 ± 10.53

35.74 ± 15.95 ***

Variable

Calcium (Ca) (mmol/L)

2.20 ± 0.16

1.84 ± 0.22 ***

Phosphorus (P) (mmol/L)

1.24 ± 0.15

1.46 ± 0.40 ***

BMI (kg/m2)

25.73 ± 3.14

29.46 ± 2.54 ***

PTH – Parathyroid hormone; Ca – Calcium; P – Phosphorus; BMI – Body Mass Index. Values are expressed as (mean ± SD) ***p ≤ 0.0001 highly significant difference.

Table 3. The effect of severity disease on the Decorin and variables studied for osteoarthritis patients

Decorin (µg/L)

Controls n = 90 8.72 ± 3.13

Mild n = 34 22.85 ± 4.77

Severe n = 56 41.87 ± 5.89

Parathyroid hormone (PTH) (pmol/L)

3.83 ± 0.40

9.60 ± 3.82

17.29 ± 4.19

Vitamin D (nmol/L)

83.97 ± 10.53

55.61 ± 3.42

23.69 ± 3.67

Calcium (Ca) (mmol/L)

2.20 ± 0.16

2.09 ± 0.07

1.69 ± 0.11

Phosphorus (P) (mmol/L)

1.24 ± 0.15

0.97 ± 0.15

1.76 ± 0.08

BMI (kg/m )

25.73 ± 3.14

28.22 ± 2.93

30.22 ± 1.95

Variable

2

PTH – Parathyroid hormone; Ca – Calcium; P – Phosphorus; BMI – Body Mass Index; values are expressed as (mean ± SD) New Zealand Journal of Medical Laboratory Science 146

Table 4. The correlation of the Decorin level and some biochemical variables and body mass index in control and Osteoarthritis patients Decorin, Pearson correlation (r); p-value

Biochemical variable

Controls

Patients

Parathyroid hormone (PTH) (pmol/L)

0.105; 0.324

0.825***; 0.0001

Vitamin D (nmol/L)

0.211*; 0.046

-0.825*** ;0.0001

Calcium (Ca) (mmol/L)

-0.044; 0.680

-0.790 ***; 0.0001

Phosphorus (P) (mmol/L)

0.172; 0.105

0.644***; 0.0001

BMI (kg/m2)

-0.019; 0.862

0.604 ***; 0.0001

*Correlation is significant at the p ≤0.05 level. ***Correlation is significant at the p ≤ 0.0001 level. PTH – Parathyroid hormone; Ca – Calcium; P – Phosphorous; BMI – Body Mass Index; p <0.05 statistically significant Table 5. Area under the curve (AUC), cut-off values, sensitivity, and specificity of serum Decorin and biochemical parameters in Osteoarthritis patients Cut-off Sensitivity Specificity Variable AUC p-value Value (%) (%) Serum Decorin (DCN) (µg/L)

45.74

53.6

100

1.000

< 0.001

Vitamin D (nmol/L)

12.41

40.0

100

0.999

< 0.001

Parathyroid hormone (PTH ) (pmol/L)

18.06

57.1

98.4

0.967

< 0.001

Calcium (Ca) (mmol/L)

1.75

37.8

98.9

0.895

< 0.005

BMI (kg/m2)

2.50

78.6

75.8

0.794

< 0.001

DCN – Decorin; PTH – Parathyroid hormone; Ca – calcium; BMI– body mass index; p <0.05 statistically significant

Figure 1. ROC curve analysis for Decorin, Parathyroid hormone levels and body mass index in serum osteoarthritis patients

Figure 2. ROC curve analysis for Decorin , Calcium and Vitamin D levels in serum osteoarthritis patients New Zealand Journal of Medical Laboratory Science 147

DISCUSSION

The study results demonstrated a distinct positive correlation between serum Decorin levels and the progressive stages of joint degeneration, highly significant differences between the patient and control groups. A clear variation in Decorin concentrations was observed according to the Kellgren-Lawrence (KL) classification; the highest levels were recorded in patients with severe osteoarthritis compared to those with early-stage disease, while the lowest levels were observed in the healthy control group. This substantial increase in severe cases can be explained by the extensive loss and degradation of cartilage extracellular matrix tissue (22). These findings closely align with the study conducted by Ozlar (23), who investigated individuals with knee osteoarthritis and reported a significant higher level of serum Decorin in patients than in healthy controls, indicating a direct positive correlation with clinical severity indicators. Furthermore, the investigation by Bock et al (24). Supported these outcomes by examining articular cartilage samples, which demonstrated a marked increase in leucine-rich proteoglycans, specifically identifying elevated levels of Decorin and biglycan in affected tissues. In contrast, a study by Manfort et al (25) reported a lower baseline, showing no statistically significant differences in Decorin expressions when the early stages of osteoarthritis joint disease comparing to healthy controls. Another study found that Decorin levels in patients with osteoarthritis were significantly higher than in healthy individuals (26). To fully capture the clinical context, serum Decorin was evaluated against established risk factors such as age, gender, obesity and disease severity. The progression of osteoarthritis is heavily driven by cellular aging, with advanced age being one of the primary risk factors for mechanical joint degradation (27). As cartilage tissue undergoes senescent modifications, the compensatory release of non-collagenous small leucine-rich proteoglycans like Decorin rises significantly to counteract the ongoing structural breakdown. This phenomenon is particularly pronounced in aging populations, where osteoarthritis stands as a leading cause of global physical disability (28,29). The number of osteoarthritis cases or primary knee replacement surgeries globally are increasing due to aging populations (30). At this stage, a localised increase in Decorin proteoglycan levels is observed, which can be explained by the assumption that the cells in the cartilage, as a result of aging, begin to produce larger quantities of these proteins in response to aging, or perhaps it is a result of the loss of other structural proteins such as collagen, which changes the composition of the tissue (31,32). Females often experience greater pain, stiffness, and reduced physical capacity compared to males, although radiographic assessment scores at diagnosis can be similar in both sexes (33). It has been noted that the concentration of Decorin levels in women is more susceptible than in men, and this study is consistent with Kellgren-Lawrence grading (34). Furthermore, obesity is recognised as one of the critical risk factors in patients with osteoarthritis, especially in knees (35). It is well established that excess weight and obesity increase the risk of osteoarthritis by increasing the mechanical load on the joints (36). However, abnormally excessive mechanical loading leads to cartilage imbalance and deformation of the joint's natural shape, which stimulates and accelerates the development of osteoarthritis in the knee (37). This pathophysiological mechanism is strongly supported by an experimental model of chronic knee compression, which found a progressive decrease in cartilage thickness and cellular density, accompanied by pronounced histological deterioration and an increase in subchondral bone thickness (38). Another study utilising magnetic resonance imaging (MRI) on a different biological model of periodic knee loading for two weeks revealed bone marrow lesions, bone resorption, and a significant increase in microscopic tissue damage associated with increased loading (39). Our study observed a significant decrease in vitamin D levels and calcium concentration in patients with osteoarthritis

compared to healthy individuals. Conversely, the study noted elevated parathyroid hormone (PTH) levels in the patient group compared to healthy individuals. Our findings are also consistent with a study conducted on postmenopausal women with osteoporosis, which indicated that vitamin D deficiency and impaired parathyroid hormone response negatively impact bone regeneration, thus highlighting the importance of maintaining adequate levels to support the skeletal system and reduce the risk of general mortality (40). These results reinforce the overall picture presented by global research on the role of vitamin D in the development of osteoarthritis. The low levels observed in our study align with international trends and systematic reviews indicating the prevalence of vitamin D deficiency among knee osteoarthritis patients in various communities (41). The observed elevation in parathyroid hormone in our patients also explains the close inverse relationship with vitamin D and clarifies how this deficiency contributes to the acceleration of the disease by affecting mineral and bone metabolism. This underscores the importance of therapeutic intervention to ensure adequate levels of these elements in the body to mitigate the effects of osteoarthritis (42). Scientific research has demonstrated an inverse relationship between serum parathyroid hormone levels and the risk of developing osteoarthritis, with a particular focus on knee osteoarthritis. A Mendelian randomisation study highlighted this relationship, suggesting a potential protective role for parathyroid hormone against disease progression. However, the same study did not establish a direct causal effect of vitamin D levels on the development of osteoarthritis, reflecting the complexity of these biological connections and the need for further investigation into the mechanisms by which parathyroid hormone affects joints (43). In contrast, our study results indicate that vitamin D deficiency is directly associated with the exacerbation of knee osteoarthritis. The study indicates that individuals with this deficiency face an increased risk of joint deterioration, underscoring the importance of maintaining adequate vitamin D levels to promote joint health and slow disease progression (44). Our findings are supported by longitudinal data from the Osteoarthritis Initiative (OA), which demonstrated that low vitamin D levels significantly increase the risk of knee osteoarthritis progression over four years, with this risk tripling in individuals with both low vitamin D and elevated parathyroid hormone levels (40). This is in agreement with a previous study (45) on patients with severe osteoarthritis, where they observed a significant increase in parathyroid hormone levels in patients compared to healthy controls, alongside a significant decrease in vitamin D levels in the same patients compared to healthy individuals, which strongly strengthens the reliability of our current findings. Furthermore, a cross-sectional study of patients with earlystage osteoarthritis (EOA) revealed that these patients had significantly lower vitamin D levels compared to matched control groups. Clinically, low vitamin D levels were associated with increased pain, worsening functional disability and anxiety, as well as a decline in physical performance indicators such as walking speed and the ability to transition from a seated to a standing position (46). These findings underscore the vital importance of both vitamin D and parathyroid hormone in the management of osteoarthritis, as their interrelationship suggests that maintaining their levels within normal limits may constitute a protective mechanism against joint deterioration. The vitamin D and parathyroid hormone not only regulate calcium balance in the body but also plays a broader role in metabolic pathways closely related to the osteoarthritis bioenvironment (40,42).

CONCLUSION

This study demonstrates that serum Decorin levels are significantly elevated in osteoarthritis patients and directly correlate with clinical disease severity, highlighting its potential as a reliable biomarker for structural joint deterioration. Furthermore, the progression of osteoarthritis is closely linked to

New Zealand Journal of Medical Laboratory Science 148

metabolic disturbances, characterised by vitamin D deficiency, hypocalcaemia, and secondary elevation of parathyroid hormone. Collectively, these findings underscore the importance of monitoring the Decorin, vitamin D and parathyroid hormone to assess arthritis progression. Integrating these biomarkers into clinical evaluation could enhance disease grading and facilitate targeted therapeutic interventions to mitigate joint degeneration. We recommend further studies with larger sample sizes to confirm the role of Decorin in determining osteoarthritis grade.

CONFLICT OF INTEREST

The authors confirm that they have no competing interests, financial or otherwise, that could be perceived as influencing the objectivity of this work.

FUNDING

This research received no outside support or specific funding from any agency in the public, commercial, or not-for-profit sectors.

ACKNOWLEDGMENTS

The authors express their sincere gratitude to all who contributed to the completion of this research. They also extend special thanks to the Department of Chemistry, College of Science, University of Mosul, for their ongoing support throughout the research process.

AUTHOR INFORMATION

Sarah A Younus1,2 Zahraa Mohammed Ali Ahmed Hamodat, PhD, Professor1 Department of Chemistry and Biochemistry, College of Medicine, University of Ninevah, Mosul, Iraq 2 Department of Chemistry, College of Science, University of Mosul, Mosul, Iraq

1

Corresponding author: Zahraa MA Hamodat, Department of Chemistry, College of Science, University of Mosul, Iraq Email: zahraahamodat@uomosul.edu.iq

REFERENCES 1. 2.

3. 4. 5.

6. 7. 8. 9.

Karimi SB, Awalkhail F, Walizada S. Knee osteoarthritis: assessment of quality of life in these patients. Sci Educ 2024; 5(3): 105-110. Cao Y, Winzenberg T, Nguo K, et al. Association between serum levels of 25-hydroxyvitamin D and osteoarthritis: a systematic review. Rheumatology (Oxford). 2013; 52(7): 1323-1334. Mermerci Başkan B, Yurdakul FG, Aydın E, et al. Effect of vitamin D levels on radiographic knee osteoarthritis and functional status. Turk J Phys Med Rehabil. 2017; 64(1): 1-7. Hamodat ZMA, Khaleel NF. Study on some biochemical markers for women affected with joint disease. Biochem Cell Arch 2020; 20(2): 5540-5543. Mohammed A, Alshamarri T, et al. A comparison of risk factors for osteo- and rheumatoid arthritis using NHANES data. Prev Med Rep 2020; 20:101242. doi: 10.1016/j. pmedr.2020.101242. Allen KD, Thoma LM, Golightly YM. Epidemiology of osteoarthritis. Osteoarthritis Cartilage 2022; 30(2):184-195. Sharma AR, Jagga S, Lee SS, Nam JS. Interplay between cartilage and subchondral bone contributing to pathogenesis of osteoarthritis. Int J Mol Sci 2013; 14(10): 19805-19830. Salvo SG. Musculoskeletal pathologies, disorders, and injuries. Mosby's Pathology for Massage Professionals. Elsevier, EvolveTM, 5th Edition. 2021; 544 Jang S, Lee K, Ju JH. Recent updates of diagnosis, pathophysiology, and treatment on osteoarthritis of the knee. Int J Mol Sci 2021; 22(5): 2619.

10. Wallace IJ, Worthington S, Felson DT et al. Knee osteoarthritis has doubled in prevalence since the mid-20th century. Proc Natl Acad Sci USA 2017; 114(35): 9332-9336. 11. Allen KD, Thoma LM, Golightly YM. Epidemiology of osteoarthritis. Osteoarthritis Cartilage 2022; 30(2): 184-195. 12. Litwic A, Edwards MH, Dennison EM, CooperC. Epidemiology and burden of osteoarthritis. Br Med Bull 2013; 105: 185-199. 13. Godziuk K, Hawker GA. Obesity and body mass index: Past and future considerations in osteoarthritis research. Osteoarthritis Cartilage 2024; 32(4): 452-459. 14. Claus Rh, Elke S, Horst R, et al. The glycosaminoglycan chain of decorin plays an important role in collagen fibril formation at the early stages of fibrillogenesis. FEBS J 2007; 274(16): 4246–4255. doi:10.1111/j.1742-4658.2007.05951.x. 15. Dipon KM, Christopher X, Gabriel JP, et al. Decorin suppresses tumor lymphangiogenesis: a mechanism to curtail cancer progression. Proc Natl Acad Sci U S A. 2024; 4(18): e2317760121. 16. Lourdes AR,Ramón C,Juan MD. Proteoglycans in articular cartilage and their contribution to chondral injury and repair mechanisms. Int J Mol Sci. 2023; 24(13): 10824. doi: 10.3390/ijms241310824. 17. Goncalo B, Antti S, Pekka Y, et al. Soluble biglycan: a potential mediator of cartilage degradation in osteoarthritis. Barreto et al. Arthritis Res Ther 2015; 17: 379. doi:10.1186/ s13075-015-0902-0. 18. Han B, Li Q, Wang C, et al. Differentiated activities of decorin and biglycan in the progression of post-traumatic osteoarthritis. Osteoarthritis Cartilage 2021; 29(8):11811192. doi: 10.1016/j.joca.2021.03.019. 19. Sahota O, Mundey MK, San P, et al. The relationship between vitamin D and parathyroid hormone: calcium homeostasis, bone turnover, and bone mineral density in postmenopausal women with established osteoporosis. Bone 2004; 35: 312-319. 20. Bobillier A, Wagner P, Whittier DE, et al. Association of vitamin D and parathyroid hormone status with the agingrelated decline of bone microarchitecture in older men: the prospective structure of aging men’s bones (STRAMBO) study. J Bone Miner Res. 2022; 37: 1903-1914. 21. Hammodat, Z, Layla AM, Biochemical studies on synovial fluid and serum from rheumatoid arthritis patients. Rafidain Journal of Science 2018 27(3): 2018-2055. doi: 10.33899/ rjs.2018.159385. 22. Zhu J, Li Y, Shen W, et al. Relationships between transforming growth factor-beta1, myostatin, and decorin: Implications for skeletal muscle fibrosis. J Biol Chem 2007; 282(35): 25852-25863. 23. Ozler K. Relationship between increased serum & synovial fluid decorin levels & knee osteoarthritis. Indian J Med Res 2021; 153(4): 453-458. 24. Bock HC, Michaeli P, Bode C, et al. The small proteoglycans decorin and biglycan in human articular cartilage of latestage osteoarthritis. Osteoarthritis Cartilage, 2001; 9(7): 654-663. doi: 10.1053/joca.2001.0420. 25. Monfort J, Tardif G, Reboul P, et al. Degradation of small leucine-rich repeat proteoglycans by matrix metalloprotease-13: identification of a new biglycan cleavage site. Arthritis Res Ther 2006; 8: R26. 26. Hegazy SM, Mohamed MS, Elnaggar BM, AboZaid SY. Level of decorin as a small leucin rich proteoglycanin knee osteoarthritis patients. JRAM 2023; 4(1): 23-29. doi: 10.21608/jram.2022.140359.1169. 27. Kulkarni P, Srivastava V, Tootsi K, et al. Synovial fluid in knee osteoarthritis extends proinflammatory niche for macrophage polarization. Cells 2022; 11(24): 4115. doi: 10.3390/cells11244115. 28. Popper HR, Fliegel BE, Elliott DM, Su AW. Surgical management of traumatic meniscus injuries. Pathophysiol 2023; 30: 618–629.

New Zealand Journal of Medical Laboratory Science 149

29. Kuczyński N, Boś J, Białoskórska K, et al. The meniscus: basic science and therapeutic approaches. J Clin Med 2025; 14(6): 2020. doi: 10.3390/jcm14062020. 30. Ozeki N, Koga H, Sekiya I. Degenerative meniscus in knee osteoarthritis: from pathology to treatment. Life (Basel) 2022; 12(4): 603. doi: 10.3390/life12040603. 31. Loeser RF. Aging processes and the development of osteoarthritis. Curr Opin Rheumatol 2013; 25(1): 108-113. doi: 10.1097/BOR.0b013e32835a9428. 32. Pauli C, Grogan SP, Patil S. Macroscopic and histopathologic analysis of human knee menisci in aging and osteoarthritis. Osteoarthritis Cartilage 2011; 19(9) :1132–1141. doi: 10.1016/j.joca.2011.05.008. 33. Ratan R, Basak M, Pal D, et al. Females with knee osteoarthritis have greater pain and reduced physical function but similar radiological grading at presentation: a cross-sectional observational study. Indian J Orthop 2025; 59(4): 549-557. doi: 10.1007/s43465.025.01365.x. 34. Cho HJ, Chang CB, Yoo JH, et al. Gender differences in the correlation between symptom and radiographic severity in patients with knee osteoarthritis. Clin Orthop Relat Res 2010; 468(7): 1749-1758. 35. Chen L, Zheng JJY, Li G, et al. Pathogenesis and clinical management of obesity-related knee osteoarthritis: impact of mechanical loading. J Orthop Translat 2020; 24: 66–75. doi: 10.1016/j.jot.2020.05.001. 36. Shumnalieva R, Kotov G, Monov S. Obesity-related knee osteoarthritis. current concepts. Life (Basel) 2023; 13(8): 1650. doi: 10.3390/life13081650. 37. Raud B, Gay C, Guiguet-Auclair C, et al. Level of obesity is directly associated with the clinical and functional consequences of knee osteoarthritis. Sci Rep 2020; 10(1): 3601. doi: 10.1038/s41598.020.60587.1. 38. Roemhild ML, Beynnon BD, Gauthier AE, et al. Chronic in vivo load alteration induces degenerative changes in the rat tibiofemoral joint. Osteoarthr Cartilage 2013; 21: 346–357. doi: 10.1016/j.joca.2012.10.014.

39. Matheny JB, Goff MG, Pownder SL, et al. An in vivo model of a mechanically-induced bone marrow lesion. J Biomech 2017; 64: 258–261. 40. Sahota O, Mundey MK, San P, et al. The relationship between vitamin D and parathyroid hormone: calcium homeostasis, bone turnover, and bone mineral density in postmenopausal women with established osteoporosis. Bone 2004; 35(1): 312-319. 41. Annamalai R, Sujhithra A, Danis Vijay D. Association between vitamin D and knee osteoarthritis in Indian population: A systematic review and meta-analysis. J Clin Orthop Trauma 2023; 46: 102278. 42. Fang FZ, Jeffrey BD, Grace HL, et al. Vitamin D deficiency is associated with progression of knee osteoarthritis. J Nutr 2014; 144(12): 2002-2008. doi: 10.3945/jn.114.193227. 43. Qu Z, Yang F, Yan Y, et al. A Mendelian randomization study on the role of serum parathyroid hormone and 25-hydroxy vitamin D in osteoarthritis. Osteoarthritis Cartilage 2021; 29(9): 1282-1290. 44. Zhang FF, Driban JB, Lo GH, et al. Vitamin D deficiency is associated with progression of knee osteoarthritis. J Nutr 2014; 144(12): 2002-2008. 45. Hamodat ZMA, Al-Ashou K.O. Study of cartilage oligomeric matrix protein (COMP) level in serum and synovial fluid for osteoarthritis patients and its relationship level of vitamin D, parathyroid hormone and disease severity. Biochem Cell Arch 2021; 21(Suppl 1): 2111-2122. 46. Alabajos-Cea A, Herrero-Manley L, Suso-Martí L, et al. The role of vitamin D in early knee osteoarthritis and its relationship with their physical and psychological status. Nutrients 2021; 13(11): 4035. Copyright: © 2026 The author(s). This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

Advertisements extracted from the Journal of the New Zealand Association of Bacteriologists, Vol. 1, October 1946, No. 3 New Zealand Journal of Medical Laboratory Science 150

CASE STUDY

Benign acute childhood myositis (BACM): a case study from a rural hospital Rebecca Busch ABSTRACT

Benign acute childhood myositis (BACM) is a rare, self-limiting, sudden onset of lower extremity muscle pain that usually occurs after a viral infection. It often presents with difficulty walking due to severe calf pain. Symptoms of BACM are usually limited to muscle weakness but can rarely progress to rhabdomyolysis and renal failure if not managed correctly. Due to the limited number of cases of Benign acute childhood myositis, there are no clear guidelines for treatment or management of the condition. This case study will focus on four children seen at a small rural hospital over three weeks in winter 2025. All four children had calf pain following a respiratory illness in the previous five days, and two could not bear weight on their legs at all. A respiratory viral panel was positive for influenza B, and the initial creatine kinase (CK) was elevated in all four children. Following a paediatric consultation via phone, a diagnosis of benign acute childhood myositis was made. All patients were managed conservatively and discharged home with clear instructions for supportive care, including paracetamol and adequate hydration. Caregivers were advised to return if symptoms worsened or if there was a change in urine colour, which could suggest myoglobinuria. Only one of the four children was followed until creatine kinase returned to normal levels. Keywords: Benign acute childhood myositis (BACM), Viral myositis, creatine kinase (CK), Rhabdomyolysis, Upper Respiratory Infection (URI). NZ J Med Lab Sci 2026; 80(3): 151:153

INTRODUCTION

Benign acute childhood myositis (BACM) was first described and identified as a distinct syndrome in 1957 by the Swedish paediatrician Ake Lundbery (1). Lundberg originally named the condition “myalgia cruris epidemica” based on a study of 74 cases of calf pain in children following a viral upper respiratory infection, which was published in Acta Paediatrica in 1957 (1). BACM is also known as “influenza-associated myositis”, “viral myositis” and “acute myositis” (2,3). It is a self-limiting inflammatory condition of skeletal muscle that usually occurs after a viral infection. The incidence of BACM is approximately 2.69 cases per 100,000 (4), making it a rare clinical syndrome. Although most commonly associated with viral pathogens, especially Influenza B, bacterial and rarely fungal infections have also been implicated (4). Most reported cases involve school–age children, with a median age of 8.3 years and a male-to-female ratio of 2:1 (5). BACM often begins with a viral upper respiratory tract infection with symptoms such as headaches, fever, cough and fatigue. A few days later, the patients describe muscle pain, usually in the legs, that typically lasts an average of 3 days (5). Often, patients refuse to walk or walk with a “Frankenstein gate”, a wide-based, stiff- legged gait due to severe calf pain (2,6). Diagnosis of the patient usually relies on a combination of clinical findings and investigations, including elevated serum creatine kinase, altered complete blood counts and elevated liver enzymes (3,7). Typically, CK levels peak in the first 2-5 days, then gradually decrease over the next 5-7 days (7,8). In most cases, patients can be effectively managed at home with paracetamol, rest, and good hydration with support from clinician’s if required (1,3). Ensuring proper hydration of the patient is key to avoiding the risk of myoglobinuria, which is muscle protein in the urine resulting from muscle breakdown (7,9).

CASE STUDY

Patient A, an 8-year-old girl presented to a rural emergency department in late July 2025 with a four-day history of fever, headache, and fatigue that had been managed at home with paracetamol. On the fifth day, her fever resolved, but she developed calf pain in both legs. By lunchtime, the pain had worsened to the point that she was unable to walk. She described the pain as “squeezy,” occasionally sharp, and radiating toward her feet. On examination, she had no joint, chest, or abdominal pain, no vomiting or diarrhoea, no rash, and was passing clear urine. Neurological examination of the lower limbs was normal, with “intact reflexes and sensation”. She was able to stand and walk on her tiptoes but became tearful when attempting to stand flat-footed. Given her recent upper respiratory illness, a viral

swab was performed and returned positive for Influenza B. Laboratory investigations demonstrated normal renal function, a low WBC with neutropenia, and a markedly elevated creatine kinase (CK) level. Urinalysis was unremarkable, with no evidence of myoglobinuria. The clinician’s impression was acute myositis, most likely secondary to a recent viral illness. The patient received a 500mL intravenous fluid bolus and was discharged home while a repeat CK sample was sent for analysis. Caregivers were advised to encourage adequate oral hydration and provide supportive measures, including paracetamol, warm compresses, warm baths and rest. They were instructed to return for medical review if pain worsened, urine output decreased, or new concerns arose. Repeat CK measurement and renal function testing were arranged for the following day. Creatine kinase (CK) levels were repeated twice more and showed a reduction to near-normal values eight days later (Figure 1). Renal function test remained within the reference range, and the CBC was within the reference range, and the neutropenia had corrected by day 8. Key laboratory parameters of the patient presented in Table 1. Five days later, Patient A’s 6-year-old brother (Patient B) presented to the same emergency room with similar symptoms, including calf pain following a recent respiratory illness. After a thorough examination, he was also diagnosed with viral-induced myositis caused by Influenza B. Key laboratory parameters of Patient B are shown in Table 2. Over the next two weeks, two more children, Patient C (Table 3), an 8-year-old female, and Patient D (Table 4), a 6-year-old male were seen at the same rural hospital. All of them experienced lower limb pain, elevated CK levels, neutropenia, and were diagnosed with Influenza B-related myositis. Each child was diagnosed with BACM and sent home with instructions to rest, hydrate and return if symptoms worsened. Only Patient A’s CK levels were monitored until they returned to normal. None of the children required readmission or any follow-up treatment.

Figure 1: Patient A CK levels over time (normal range 30-180 U/L)

New Zealand Journal of Medical Laboratory Science 151

Table 1: Complete blood counts (CBC) for Patient A over time Complete Blood Count

28/07/2025

29/07/2025

30/07/2025

04/08/2025

Ref Range

Haemoglobin (g/L)

138

142

150

143

113-145

Haematocrit ratio

0.42

0.42

0.45

0.41

0.33-0.42

MCV (fL)

83

82

82

81

74-87

MCH (pg)

27

28

28

28

24-29

Platelets x109/L

216

192

176

365

150-475

WBCx109/L

2.9

2.3

2.2

5.3

4.5-12

Neutrophils x109/L

1.2

0.8

0.6

1.8

1.5-8.0

Lymphocytes x109/L

1.4

1.2

1.3

2.7

1.4-5.7

Monocytes x109/L

0.32

0.23

0.16

0.40

0.3-1.0

0.09

Eosinophils x109/L

0.03

0.05

Basophils x109/L

0.01

0.01

Meta/myelocytes x109/L

0.37

0.0-1.0

0.02

0.0-0.2

0.01

0.0-0.06

Red = outside reference range Film comment (29/07/2025), Neutrophils show a left shift; Reactive lymphocytes +. Neutropenia – Please refer to health pathways for possible causes and appropriate investigations. 28/7/25: LFT AST=75 (10-50) otherwise normal Renal function normal on 29th/30th and 4th. Table 2: Patient B, complete blood counts (CBC) Complete Blood Count

Ref Range

Haemoglobin (g/L)

123

113-145

Haematocrit ratio

0.37

0.33-0.42

MCV (fL)

79

74-87

MCH (pg)

27

24-29

Platelets x10 /L

157

150-475

WBC x109/L

2.6

4.5-12

Neutrophils x10 /L

1.7

1.5-8.0

Lymphocytes x109/L

0.7

1.4-5.7

Monocytes x10 /L

0.19

0.3-1.0

9

9

9

Eosinophils x10 /L

0.04

0.0-1.0

Basophils x109/L Meta/myelocytes x109/L

0.01 0.01

0.0-0.2 0.0-0.06

9

Red = outside reference range, 06/08/2025, CK=270 (Reference Range, 30-180U/L) Film comment – Neutrophils show a left shift, Sodium 133mmol/L (135-145) crp 11mg/L (<5) Otherwise normal results Table 3: Patient C complete blood count (CBC) Complete Blood Count

5/8/2025

6/8/2025

Ref Range

Haemoglobin (g/L)

125

130

113-145

Haematocrit ratio

0.39

0.40

0.33-0.42

MCV (fL)

79

81

74-87

MCH (pg)

26

26

24-29

Platelets x109/L

138

144

150-475

WBC x109/L

2.6

3.1

4.5-12

Neutrophils x109/L

0.5

1.2

1.5-8.0

Lymphocytes x109/L

1.7

1.7

1.4-5.7

Monocytes x109/L

0.28

0.17

0.3-1.0

Eosinophils x109/L

0.02

0.03

0.0-1.0

Basophils x109/L

0.01

Meta/myelocytes x10 /L

0.0-0.2 0.0-0.06

9

Red = outside reference range, Ck -5/8/2025 1931, 6/8/2025 1995 Normal renal function (Reference Range, 30-180U/L) Film comment- Neutrophils show left shift; Reactive lymphocytes +. These changes are most likely reactive to infection/ inflammation. Neutropenia- Please refer to Health Pathways for possible causes and appropriate investigations.

New Zealand Journal of Medical Laboratory Science 152

Table 4: Patient D Complete Blood Count

13/8/2025

14/8/2025

15/8/2025

Ref Range

Haemoglobin (g/L)

131

121

131

113-145

Haematocrit ratio

0.37

0.34

0.37

0.33-0.42

MCV (fL)

78

78

78

74-87

MCH (pg)

27

28

27

24-29

Platelets x109/L

214

254

150

150-475

WBC x109/L

3.4

4.7

3.9

4.5-12

Neutrophils x109/L

0.5

0.7

1.3

1.5-8.0

Lymphocytes x109/L

2.0

2.8

1.7

1.4-5.7

Monocytes x109/L

0.34

0.43

0.20

0.3-1.0

Eosinophils x109/L

0.5

0.8

0.6

0.0-1.0

Basophils x109/L

0.02

0.02

0.02

0.0-0.2

0.01

0.0-0.06

Meta/myelocytes x109/L

Red = outside reference range, CK 13/8/2025 = 801 15/8/2025 144, Normal renal function (Reference Range, 30-180U/L)

DISCUSSION

This cluster of BACM cases reflects a pattern previously described in the literature, characterised by seasonal spikes of severe calf myalgia following upper respiratory infections during periods of increased influenza activity in the population (1,6). Previous studies suggest that BACM has a higher male-to-female ratio; however, in this cluster two of the four patients were female, and one case involved a sibling of a previously affected patient. Although these cases appear to have been rapidly identified and managed, previous studies show that many cases of BACM are not recognised by clinicians and may be misdiagnosed as more serious conditions (6). The sudden onset of bilateral calf pain observed in these patients can mimic several other conditions including Guillain–Barré syndrome, rhabdomyolysis, muscular dystrophy, septic arthritis, or osteomyelitis (6). Misdiagnosis of this condition may result in unnecessary hospital admissions, extensive laboratory testing and imaging, lumbar punctures, invasive procedures, and inappropriate treatments such as antibiotic therapy (6). However, in this age group, viral myositis is also the most common cause of rhabdomyolysis, a condition involving the breakdown of skeletal muscle that releases toxic intracellular components, including myoglobin, into the bloodstream and can lead to acute kidney injury (2). Although this complication is uncommon (approximately 3% of BACM cases) doctors must consider and exclude rhabdomyolysis before discharging a patient. This can be achieved through urine dipstick testing for haem and careful monitoring of renal function (2,5). None of the patients in this study demonstrated evidence of rhabdomyolysis. Although BACM is strongly associated with influenza viruses, the mechanisms by which viral infections lead to muscle involvement remain poorly understood (2). One proposed mechanism is direct viral invasion of muscle tissue, which results in muscle cell necrosis and subsequent elevation of creatine kinase (CK) levels (5,7). Developing muscle tissue in children may be more susceptible to this process; however, the precise immunological mechanisms remain unclear (5). Although these four cases represent a very small cluster of BACM, due to its rarity, it is important for medical staff to recognise these patterns in the community so that no future cases are misdiagnosed. Monitoring CK levels until they return to the normal reference range is recommended to ensure that muscle damage has resolved. However, only one child in this cluster was monitored after the initial diagnosis.

CONCLUSION

Benign acute childhood myositis (BACM) is an important but often under-recognised cause of acute bilateral calf pain in children following viral illness, particularly during periods of increased seasonal influenza activity. Despite its sometimesdramatic presentation, the condition is typically benign and selflimiting, with rapid recovery and normalisation of creatine kinase (CK) levels.

Monitoring CK levels can assist in confirming that the muscle injury has fully resolved. Early recognition of the characteristic clinical features of BACM can help clinicians distinguish BACM from more serious conditions and avoid unnecessary investigations, hospital admissions, and inappropriate treatments while still providing reassurance for patients and their families.

AUTHOR INFORMATION

Rebecca Busch, BMLSc, Medical Laboratory Scientist, Ashburton Hospital Laboratory, Ashburton, New Zealand Email: Rebecca.busch@tewhatuora.govt.nz

REFERENCES 1.

Lundberg A. Myalgia curis epidemica. Acta Pediatr 1957; 46: 18-31. 2. Brisca G, Mariani M, Pirlo D, et al. Management and outcome of benign acute childhood myositis in pediatric emergency department. Ital J Paediatr 2021; 47(1): 57. 3. Costa Azevedo A, Costa e Silva A, Juliana Silva C, et al. Benign acute childhood myositis: a 5-year retrospective study. Arch Pediatr 2022; 29(7): 490-493. 4. Alexander M, Kigotho A. Recurrent benign acute childhood myositis (BACM) and incidental generalised joint hypermobility (GJH): a case study and discussion. Med Clin Res 2024; 9(7): 01-05. 5. Huzior C, Chernick B, Nguyen L, Kumar B. Benign acute childhood myositis in a pediatric patient post influenza B infection. Cureus 2023; 15(12): e51171. 6. Sung-Won Choi, Hoon Ko. Benign acute childhood myositis: perplexing complication after acute viral pharyngitis. Korean J Fam Med 2018; 39(6): 375-378. 7. Altinok E, Yurdakul Erturk E, Kasar T, et al. Benign acute childhood myositis: evaluation of clinical and laboratory features. CSMJ 2025; 5: 25-30. 8. Pattiyakumbura T, Jayasundara J, Premarathne P, et al. Influenza B- associated acute myositis in a child in Sri Lanka: a case report. J Med Case Rep 2025; 19(1): 394. 9. Basheer N, Mneimneh S, Mariam R. Seven-digit creatine kinase in acute rhabdomyolysis in a child. Child Neurol Open 2016; 4: 1-4. 10. Koliou M, Hadjiloizou S, Ourani S, et al. A case of benign acute childhood myositis associated with influenza A (H1N1) virus infection. Clin Microbiol Infect 2010; 16:193-195. Copyright: © 2026 The author(s). This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

New Zealand Journal of Medical Laboratory Science 153

CASE STUDY

When APTT refuses to correct: High-titre factor VIII inhibitor inducing false positive lupus anticoagulant and spurious factor IX deficiency: a case study Hari Priya Raghvan, Wee Shiang Yui, Caroline Ho Siew Ling, Ahmad Izzudin Mustaffa, Ehram Jamian and Indhira Subbiah ABSTRACT Acquired haemophilia A (AHA) due to autoantibodies against factor VIII (FVIII) is a very rare condition with severe bleeding and high mortality. Herein, we describe a patient with spontaneous bruising and isolated prolongation of activated partial thromboplastin time (aPTT), which did not correct on mixing studies. Preliminary laboratory results led to suspicion of concomitant lupus anticoagulant (LA), and factor assays revealed a very low FVIII level accompanied by spuriously low FIX. A high-titre FVIII inhibitor was subsequently identified on further investigations, accounting for the laboratory abnormalities. Awareness of these assay limitations, together with careful clinical correlation, is crucial to prevent misdiagnosis and delays in initiating appropriate treatment for acquired haemophilia A. Keywords: acquired haemophilia (AHA), activated partial thromboplastin time (aPTT), lupus anticoagulant (LA), Factor VIII, FVIII inhibitor. NZ J Med Lab Sci 2026; 80(3): 154:156

INTRODUCTION

The isolated prolongation of the activated partial thromboplastin time (aPTT) is a frequent laboratory abnormality encountered in haemostasis practice, but it may be difficult to interpret due to the multitude of possible factors causing it, ranging from benign assay interference to life-threatening bleeding disorders (1). In clinical practice, circulating lupus anticoagulants, pharmacological interference from treatments such as unfractionated heparin or argatroban, or congenital and acquired coagulation factor deficiencies are common causes of an isolated prolongation of the aPTT (2). Typically, mixing tests that "corrects" indicate the possibility of factor deficiencies, whereas tests that "do not correct" indicate the presence of inhibitors (3). Acquired haemophilia A is caused by autoantibodies that bind and neutralize the activity of coagulation factor VIII (FVIII). Lupus anticoagulants (LAs) are autoantibodies that target phospholipids or protein-phospholipid complexes, which show up clinically as an elevated risk of thrombosis or fetal loss, in contrast to the haemorrhagic characteristics of FVIII inhibitors (4). We describe a case of isolated aPTT prolongation and no correction with mixing studies with presence of high-titre FVIII inhibitor generating assay interferences in this case report.

CASE REPORT

A 68-years-old female presented with right forearm painless mild bruising for one week duration. There was no history of trauma or any other bleeding manifestation. No constitutional symptoms noted. On examination, there were bruises seen on the right forearm but no swelling or evidence of bleeding in other areas. Patient denied family history of bleeding disorders or any haematological malignancy. Her past medical history only consisted of diabetes mellitus and hypertension on medication.

No previous history of thrombosis as well. She also denied any history of autoimmune disorders or recent infections. Upon admission, her haemoglobin level was 84g/L , white blood cells of 11x109/L and platelet of 283x109/L. Coagulation profile sent noted an isolated prolonged aPTT of 80 sec (reference range: 24-40 sec) with a normal prothrombin time (PT) of 12.7 sec (reference range: 11.7-15.3 sec). A mixing test was requested and were not corrected (54.3 sec at 0-minute, 70.3 sec following incubation at 37°C for 60 minutes). LA testing demonstrated an abnormal partial thromboplastin time – lupus anticoagulant (PTT-LA) (ratio 3.05, reference: <1.30) with PTT-LA confirmatory using hexagonal phase phospholipid neutralization test showing 9.9 sec (reference: <8 sec). Dilute Russell Viper Venom Test (DRVVT) method was negative. Anti Cardiolipin IgG/IgM and Anti-Beta2 Glycoprotein1 IgG/IgM were negative as well. Due to the abnormal aPTT mixing study with re‑prolongation at 60 minutes, along with the patient’s history of bleeding and weak LA positivity, further investigations were done to identify any other coagulation factor deficiency or factor inhibitor. Further testing using clot-based method for FVIII and FIX assays, identified a markedly low FVIII (<1%, reference: 50%150%) and low FIX (7%, reference: 50%-150%). FVIII inhibitor was detected and measured at 330 Bethesda Unit (Table 1) using Bethesda Assay. FIX inhibitor assay could not be done due to insufficient sample. Other blood investigations such as autoimmune panel were negative. A full blood picture did not reveal any evidence of haematological malignancy. Given the findings of high-titre FVIII inhibitor with clinical picture of bleeding, she was started on recombinant Factor VIIa for 2-3 doses. However, patient developed intracranial bleeding and succumbed to her condition.

Table 1. FVIII inhibitor Bethesda Assay (Control plasma: 96% factor activity) Factor activity (%)

Residual factor activity (%)

1:2

Dilution

1

2

1:5

2

4

1:10

3

6

1:20

3

6

1:50

4

8

1:100

6

12

1:200

16

32

1:500

35

71

1:1000 46 94 Inhibitor strength was calculated from residual factor activity versus inhibitor units. Residual activity nearest to 50% was taken (32%) and inhibitor level was calculated. Inhibitor in Bethesda Unit: 330 B.U. New Zealand Journal of Medical Laboratory Science 154

DISCUSSION

This uncommon combination of positive FVIII inhibitor and positive LA test in a single patient, highlights how difficult it can be to determine the cause of a prolonged aPTT. Both LA and an FVIII inhibitor may prolong the aPTT and interfere with assays intended to identify the other autoantibody. Interference from LA in PTT‑based assays can artifactually lower the measured FVIII activity level and lead to an incorrect diagnosis of haemophilia. FVIII inhibitor also can lead to false positive LA. This is usually seen with hexagonal phospholipid assay, but DRVVT method is not affected by this inhibitor (5). As seen in our case, DRVVT method was negative, thus, PTT-LA weak positivity could be likely due to interference by FVIII inhibitor. Chromogenic FVIII assay can also be done to be sure of low FVIII activity, however, our centre does not offer chromogenic assay (5). Another clue would be, LA usually do not show re-prolongation at 60 minutes in mixing studies compared to FVIII inhibitor as the latter is a time dependent inhibitor (4). As per our knowledge, there are very few similar cases published (less than 20) to date (4). An inhibitor targeting a specific coagulation factor can interfere with the assays of the other factors. While it selectively reduces its primary target, it may also cause an apparent decrease in other factor levels due to its effect on the factor-deficient plasma used in the test. A well-known example occurs with Factor VIII inhibitors, where all intrinsic factors may appear low. To clarify the results, performing serial dilutions will normalize the levels of the non-specifically reduced factors, while the genuinely targeted factor will remain low (6). As our tests are clot-based, interference by the high-titre of FVIII inhibitor is highly likely causing the low FIX level as well (6). This could be a reason for the apparent reduction of FIX in this patient. All these results were supposed to be repeated once patient has started on treatment, unfortunately, patient passed away during the process. On top of that, acquired FVIII inhibitor is more common than FIX inhibitor in our population. Published literature also indicates that the Bethesda assay is associated with many false-positive results (7). This complicated case highlights the need for detailed coagulation functional analyses and deeper understanding at various assays functionalities and their limitations. In our laboratory, further testing of a particular sample is driven by clinical findings. In our patient, the decision to test for LA was based on the need to interpret mixing studies particularly because LA may partly correct or incompletely inhibit the aPTT and can co-exist with other inhibitors. Ultimately, LA positivity was interpreted as an interfering phenomenon as the patient’s bleeding history, markedly low FVIII level with high- titre FVIII inhibitor with time-dependent mixing studies points towards a diagnosis of acquired haemophilia. Another differential diagnosis that can be considered is LA hypoprothrombinemia syndrome (LAHPS), where the Factor II will be low, causing bleeding in a LA positive patient. However, in this condition, PT will be prolonged as well which is not seen in our case (8). LAHPS is a rare condition and seen more frequently in paediatric patients, systemic lupus erythematous cases, or post-infectious state, none of which is applied in our case. Initiation of a suitable therapy was contingent upon the detection of the FVIII inhibitor. There are mainly three goal options: to stop bleeding, to eliminate the autoantibodies, and to identify the cause of bleeding if any (9). Treatment options in our centre are usually tailored to individual patient. Bypassing agent or recombinant FVIIa are used to stop the bleeding and immunosuppressive therapy to allow a remission from the disease (9). Subcutaneous haemorrhages are the most common bleeding symptoms of AHA, and they are frequently rather extensive. Fortunately, intracerebral haemorrhage is rare but often fatal (8). Life-threatening haemorrhage most commonly occurs within the first few weeks of presentation but may develop at any time in the absence of treatment (10). In the European Acquired

Haemophilia (EACH2) registry, bleeding at diagnosis was observed in 89% of cases (11). In the majority of cases, bleeding occurred spontaneously, while 25% of the cases were associated with trauma or invasive procedures (12). It should be noted that not all patients with AHA present with bleeding at diagnosis and some may have only mild haemorrhages with about 25% not requiring haemostatic treatment (12). Even low-titre inhibitors require constant vigilance, as their presence signals an ongoing risk of spontaneous haemorrhages anywhere in the body, with no reliable predictors for major or fatal bleeding events (9). In summary, this unique case of acquired haemophilia A causing interference with LA and FIX assays illustrates the need of careful clinical correlation and specific assay selection for accurate diagnosis. This is to assist in the management as this condition is considered an emergency in a haematology setting.

CONFLICTS OF INTEREST

The authors declare no conflict of interest.

ACKNOWLEDGEMENT

We would like to thank the Director of General of Health of Malaysia for his permission to publish this article. Our sincerest thanks to our scientific officer Mr Mohd Zamri and medical laboratory technologist Ms Vivian Kiu Wei Chee for assisting us in the technical aspects.

AUTHOR INFORMATION

Hari Priya Raghvan, MD (UCSI), DrPath, Haemotology1 Wee Shiang Yui, MBBS, DrPath, Haemotology1 Caroline Ho Siew Ling, MBBS, DrPath, Pathology1 Ahmad Izzudin Mustaffa, MBBS, General Physician & Internal Medicine Specialist2 Ehram Jamian, MD, MPath, Haemotology1 Indhira Subbiah, MBBS, DrPath, Haemotology1 Haematology Unit, Department of Pathology, Hospital Sultanah Aminah, Johor, Ministry of Health, Malaysia. 2 Clinical Haematology Unit, Department of Internal Medicine, Hospital Sultanah Aminah Johor, Ministry of Health, Malaysia. 1

Corresponding Author: Hari Priya Raghvan, Haematology Unit, Department of Pathology, Hospital Sultanah Aminah, Johor, Ministry of Health, Malaysia. Email: Priya_hari88@yahoo.com

REFERENCES 1. 2.

3.

4. 5. 6.

7. 8.

Santoro RC, Molinari AC, Leotta M, Martini T. Isolated prolongation of activated partial thromboplastin time: not just bleeding risk! Medicina 2023; 59(6): 1169. Rasmussen KL, Philips M, Tripodi A, Goetze JP. Unexpected, isolated activated partial thromboplastin time prolongation: a practical mini‐review. Eur J Haematol 2020; 104(6): 519525. Favaloro EJ. Coagulation mixing studies: utility, algorithmic strategies and limitations for lupus anticoagulant testing or follow up of abnormal coagulation tests. Am J Hematol 2020; 95(1): 117-128. Jacobs JW, Gisriel SD, Iyer K, Rinder HM. Concomitant factor VIII inhibitor and lupus anticoagulant in an asymptomatic patient. J Thromb Thrombolsis 2022; 53(4): 945-949. Adcock DM, Favaloro EJ. Pearls and pitfalls in factor inhibitor assays. Int J Lab Hemotol 2015; 37: 52-60. Anžej Doma S, Hillarp A, Pajič T, Andoljšek D, Černelč P, Preložnik Zupan I. Concurrent acquired inhibitors to factor VIII and IX, a laboratory artifact: a case report. Biochem Med 2016; 26(2): 279-284. Favaloro EJ, Verbruggen B, Miller CH. Laboratory testing for factor inhibitors. Haemophilia 2014; 20: 94-98. Gago L, Lourenço MH, Torres RP, et al. Lupus anticoagulant-

New Zealand Journal of Medical Laboratory Science 155

hypoprothrombinemia syndrome: a duality between thrombosis and hemorrhage. Cureus 2024; 16(11): e73149. 9. Marino R. Acquired hemophilia A: bleeding pattern and hemostatic therapeutic strategies. Medicina 2023; 59(10): 1739. 10. Ma AD, Carrizosa D. Acquired factor VIII inhibitors: pathophysiology and treatment. Hematology Am Soc Hematol Educ Program 2006; 432-437. 11. Knoebl P, Marco P, Baudo F, et al. Demographic and clinical data in acquired hemophilia A: results from the European Acquired Haemophilia Registry (EACH2). J Thromb Haemost 2012; 10(4): 622-631.

12. Baudo F, Collins P, Huth-Kühne A, et al. Management of bleeding in acquired hemophilia A: results from the European Acquired Haemophilia (EACH2) Registry. Blood 2012; 120(1): 39-46.

Copyright: © 2026 The author(s). This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

BARRIE EDWARDS & ROD KENNEDY SCHOLARSHIP RECIPIENT CONFERENCE REPORT

Savannah Young

CONFERENCE

Poster presentation at Blood 2025 and the 26th Regional Congress of the ISBT Perth, Australia, 26-29 October 2025. Blood 2025 Conference was combined with the 26th Regional Congress of the ISBT at the Perth Convention and Exhibition Centre, Australia, guaranteeing a bumper program. A highlight was the use of molecular techniques in transfusion medicine, not only for the resolution of blood groups, but for detection of cell-free DNA to predict fetal D-genotype, for prophylactic Anti-D immunoglobulin management. A predicted requirement for up to 20,000 less doses of prophylactic anti-D per annum are possible due to this testing in Australia. Another stand out was Sara Chiaretti’s presentation comparing cord blood derived cultured RBC and cord blood cells with adult RBC. It was highlighted that both cord RBC and cultured cord blood stem cells are promising candidates for neonatal transfusion as both have HbF (fetal haemoglobin) compositions similar to neonatal RBC, versus adult RBC. Australian transfusion laboratories were seeing increased effects of monoclonal antibody therapies such as Daratumumab (antiCD38) interfering with red cell antibody testing. A novel soluble CD38 product was trialled and found to successfully neutralize anti-CD38 with a short plasma incubation. Sessions by Jill Storry were great for brushing up on our serological “toolboxes” to solve complex antibody investigations and perfect for scientists newer to transfusion medicine. Finally, worldwide equity of safe blood and patient blood management systems were significant topics, ensuring the future blood supply is in safe hands. The poster I presented was titled Anti-M haemolytic disease of the fetus and newborn: an unexpected ene-M-y. Particularly rewarding was discussing the case with an author of a previous case study whom I had cited in my poster, as well as another who had already implemented the pre-treatment mentioned in their laboratory. I would like to extend my sincere thanks to the Barrie Edwards and Rod Kennedy scholarship, awarded by the NZIMLS, for funding this trip. As an early career transfusion scientist this scholarship provided immeasurable opportunity to grow my knowledge and confidence in presenting transfusion topics. Thanks to New Zealand Blood Service, Dr Dhana Gounder, Dr Ben Paterson and Janine Gundersen for the support in sharing this case study.

Barrie Edwards & Rod Kennedy Scholarship

Figure 1. Savannah Young, Poster presentation session

Figure 2. Iconic Western Australian Quokka

Correspondence: Savannah Young, MMLSc BMLSc, Medical Laboratory Scientist, New Zealand Blood Service. Email: Savannah.Young@nzblood.co.nz

The Barrie Edwards & Rod Kennedy scholarship is one of the most significant awards offered by the NZIMLS. The award provides support for attendance at an international or national scientific meeting up to a maximum value of $7,500.00.

Applicants must be current financial members of the NZIMLS and have been a financial member for at least two concurrent years prior to application. To be eligible applicants must make an oral or poster presentation as 1st author at their nominated scientific meeting.

Applications for this prestigious scholarship are invited from Fellows, Members and Associate Members of the NZIMLS.

For more information visit: https://www.nzimls.org.nz/scholarships

New Zealand Journal of Medical Laboratory Science 156

REPRINT

Love and Death Tristan Jones It’s December during my final year of residency, and I am ready to be done with intensive care units (ICUs). I am tired of the blood gases. I am tired of performing procedures I am not good at. I am tired of the sunless days that melt into weeks. But mostly I am tired of the death. I have recently completed a month each in the cardiac ICU and the pediatric ICU, and now am wrapping up a month in the medical ICU. I first became accustomed to death, in all its permanency, when I stalked these halls as an intern. At that time, in the spring of 2020, I cared for endless rows of intubated patients. I built bonds with families over days and weeks, but once a patient died, I learned I would barely see or hear anything more about them. The paperwork was signed, the body was whisked downstairs, and the good intern moved on to the next patient. Any relationship with the patient, any special compassion, was tossed like a disposable stethoscope. Somewhere in the recesses of my mind, a void would open. But voids, like empty beds and open lists, have a habit of being filled by the humdrum torrent of residency. This time through the ICU, the causes of illness are more diverse, but the death seems just as prevalent. I watched the slow failure of a young father’s bone marrow transplant. The nurse printed out the telemetry strip so his child could have a keepsake of his final heartbeat. I listened to a 20-something man with interstitial lung disease beg his mother, who had been standing vigil for days, to go home and rest. In his final act of love, he then asked to remove his face mask, so he could spare her witnessing his final suffering. And I responded to an elderly woman’s last request, as her lungs filled with fluid, for a breath of fresh air before the end. Though the modern hospital is equipped to provide many amenities, fresh air is not one of them. As a paltry substitute, I offered the imminent sun-rise. Motivated, she propped herself up facing eastward and sat alongside her son, watching in silence before gently drifting off. These were just a few of many deaths, and each leaves me with that small sense of emptiness. I fear I will not remember the patients I have lost, and I fear it is far too crass to try to keep track. Naturally, my holiday assignment is the medical ICU. Nights. Just like the month of day shifts I have just completed, but maybe with a few more codes and death exams. How lucky am I? On Christmas Eve, one of the new patients is Cassie, a 29-yearold woman admitted through the emergency department with skin yellow-bronze and lactate in the double digits. Between labored breaths, with tremulous voice, she tells her story. She was just diagnosed in October with metastatic lung cancer. It has taken a few weeks to get into care, and then a few more for staging. Her insurance has limited her ability to find doctors, and she struggles to find childcare. When she showed up to the oncology visit to discuss chemotherapy, she found out she was pregnant, so she was told to either terminate the pregnancy or postpone treatment. It is not clear what, if any, discussions were had regarding the viability of the pregnancy, let alone her own life expectancy. I cannot imagine the thoughts that filled her mind. What personal beliefs and expectations guided her? What feelings of love and guilt drove her? All I know is I don’t fault her for her subsequent hesitation; we are now 4 weeks past that visit, and she has yet to see an obstetrician. Her liver and kidneys are failing, we need consent and organ support quickly, and we are dealing with the complicating fact of pregnancy. Now, the β-hCG level is low, in the teens, and it has not budged in a month. It’s been a few years since I took my Step exams, but a jaunt to PubMed confirms my suspicion: a decent percentage of small-cell lung cancers can cause mild β-hCG elevations. A curbside consult with gynecology reassures us, the patient is not pregnant and care can proceed without any invasive testing. I delicately relay this information to Cassie. Though she’s shocked, her pained look gives way to relief. A guilt appears to be lifted. She is more resolute now; she would like to start chemotherapy this admission. Next I call her emergency contact, her boyfriend, Dion. He has been urging her to seek care for days. Still, he sounds a bit surprised when I tell him now is the time to start the 2-hour drive to our hospital; Cassie might not be responsive come daylight. When he shows up, right before shift change, we review Cassie’s

condition. He tells me of their life together, how they had met years ago, how she already had three small children, how he has become their stepfather in all but the official sense. He has just one question: Is there someone who can perform a wedding? I return in the evening on Christmas Day. The couple has selected a chaplain. The unit secretary has arranged some simple paper decorations. Two friends of the couple are waiting in the room. Just before midnight, after the other patients are tucked in, about a dozen staff wedge between the dialysis machine, IV poles, and external warmer around Cassie’s bed. Dion, at the head, clasps her hand through the entire ceremony. It is the sixth and final wedding I attend that year, and it is surprisingly routine. The chaplain, in his thick accent, starts by recognizing our role as a gathering of witnesses. He then reads the selected scripture (Book of Genesis). The sermon is an unexpectedly hermeneutic dissertation on the creation of Eve from Adam’s rib, a linguistically tenuous inter-pretation at best. I wonder how much Cassie will be able to serve her new husband as the preacher suggests. Is this really relevant? Ever heard of reading the room? But Cassie, between dozes, is all smiles. Dion is positively levitating. During this brief ceremony there is no mention of Cassie’s condition. For a moment, we suspend the reality written all over the room and simply witness two people’s love. The nurse plays Dion’s chosen recessional, and the staff files out. I swear you’re an angel Sent to this world What did I do right to deserve you, girl? I could stay here forever I’d be fine if we never Had to even leave this room1 On Boxing Day, I come in for signout and learn that Cassie died that afternoon. She was a wife for all of 15 hours. I don’t know what became of Dion or whether he was able to secure guardianship of his stepchildren. I just know that he and Cassie loved one another. The writer Stephen Bergman (better known as Samuel Shem) describes the privilege of medicine as bearing witness to the two great themes of human literature, love and death. When the death is so prominent, it can be easy to miss the love. Yet no other career offers such proximity to both. “How lucky we are,” he writes.2 Medical training, unlike death, is not forever. By the end of the week, I finish the last ICU shift I will ever work. I no longer fear that I will forget the patients I have lost, for other things are forever, too. Love for a partner. Love for stepchildren. A mother’s love. A love that fills voids. Identifying details have been changed to protect the patients’ privacy. ¹Virginia Commonwealth University Medi-cal Center, Richmond. This article was published on July 18, 2026, at NEJM.org and republished with their permission. N Engl J Med 2026; 395(4): 321-323.

REFERENCES 1. Brown K. Heaven. Written by Carter S, McGinn M, Rimes L. Nashville: RCA Rec-ords, 2017. 2. Shem S. Samuel Shem, 34 years after ‘The House of God.’ Washington, DC: Atlantic. November 28, 2012 (https://www.theatlantic.com/health/archive/2012/11/ samuel-shem34-years-after-the-house-of-god/265675/). DOI: 10.1056/NEJMp2516818 Copyright © 2026 Massachusetts Medical Society.

New Zealand Journal of Medical Laboratory Science 157

BOOK REVIEWS Reviewed by: Michael Legge, Deputy Editor

What we inherit: how new technologies and old myths are shaping our genomic future

Invisible evidence in forensics in New Zealand

Authors: Sam Trejo and Daphne O Martschenko, Princeton Press, USA, 2026 ISBN: 9780691237756

Author: Bill O’Brien, David Bateman Ltd, 2007 ISBN: 978-1-86953-676-3

The authors take the unusual position of investigating what genomics and genetic information cannot inform us about ourselves. The authors have developed the text around conversations between themselves for each of the nine chapters. Sam Trejo is a sociologist at Princeton University, USA and Daphne O Martschnkko is a bioethicist at Stanford University, USA. For each chapter they adopt adversarial positions based on their respective backgrounds and knowledge. This leads the reader into discussions relating to what is in our DNA and what are the myths of DNA for the inheritance of social factors. Some of the information is based on historic American society and its structure, and touches on the issues relating to gender, race (especially in the South) and social positions. They expertly consider how genes do not determine an individual’s future and the myths of race, gender and genetics are inherently connected. These considerations have been linked to social norms and in the final chapters they discuss future impacts of at-home genetic testing, and the increasing use of embryo polygenic testing in the USA to determine a child’s future. These link well with their previous discussions in the preceding chapters. The book is well written and clear, but it does take a little time for the reader to adjust to their writing style which works very well.

The author, Bill O’Brien, was a forensics expert who worked for the New Zealand Police before retiring. He has written on several topics and his book “22 Hours of Terror” about the Aramoana massacre was the basis of the 2006 film: “Out of the Blue”. In the present book, he presents a concise discourse on the forensic techniques being used and their applications. The book is well structured and has 17 individual chapters about specific forensic techniques and their background. The 17 chapters are well written, and the author explains the use of technologies in an easy-to-read style. Within each chapter the author provides mini-case studies (Forensic Files) of how a particular technique was used to detect a crime. There are no blood and gore in this book rather a clear explanation of how forensic science and technologies are used to detect crime. Including, the interesting and well-presented New Zealand case of Annette Bouwer, murdered by her clinician husband by manipulating her blood glucose and caught by Annette’s pathology results. An interesting aspect of this book is the occasional comments of where many TV series such as CSI are so very wrong with their forensic analysis. This prompted the reviewer to consider whether this is the book to watch any of the TV crime series to spot how many errors occur with the use of forensics but no CPD points!

Outsider scientists: routes to innovation in biology Edited by: Oren Harman and Michael R. Dietrich, University of Chicago Press, 2013 ISBN: 978-0-226-07840-3 This was an interesting book for anyone who thinks about how some things happen in science. It is a set of 18 essays written by different authors; most of whom knew the scientists they write about. There are certain exceptions such as Mendel and Pasteur. The two editors have published books previously in this general area but here they curate wide ranging essays about scientists from biology, physical sciences, mathematics and informatics. Many of the scientists discussed in this book were not acknowledged in their time or withstood serious criticism for their work and ideas by colleagues and in some cases the media. There are good examples of the well-known instance of “everyone knows this cannot be right” which is subsequently proven to be incorrect. Each essay provides background on the scientist’s work, how it fitted at the time and the reactions to their work. There is considerable individual style with the essays but there is a consistency in the messages about those considered “outsiders” in their time. The authors provide clarity on the value of the work the “outsider scientists” undertook and the ultimate contribution they made to their areas of science. A cautious word, the writing is particularly dense and perhaps not the best read after Christmas lunch.

NZJMLS 50th Anniversary Edition cover, 1996

New Zealand Journal of Medical Laboratory Science 158

Minutes of the 80th NZIMLS Annual General Meeting, Thursday 20 August 2026, 7:30am (breakfast) for a 7:45am start The President chaired the meeting, with approximately 30 members attending in person and five members attending online.

1.

Apologies Warren Dellow Moved: A Joseph Seconded: M Legge That the apologies be received. CARRIED

2.

Proxies (if any) The CEO received no proxies for this meeting.

3.

Minutes of Annual General Meeting dated 28 August 2025 (as circulated) Moved: A Joseph Seconded: R Hewett That the Minutes of the Annual General Meeting held 28 August 2025 be accepted. CARRIED

4.

Business arising from the Minutes NIL

5.

Remits as circulated. Moved: A Joseph Seconded: L Pringle That the Policy Decision No 1 (1971) and Policy Decision No 2 (1989) be accepted.

·

Policy Decision No 1 (1971): That all committees and meetings convened under the auspices of the New Zealand Institute of Medical Laboratory Science (Inc) be subject to a standard reference of parliamentary procedure and that this is a ‘A Guide for Meetings and Organisations’ by Renton.

·

Policy Decision No 2 (1989): That all persons wishing to undertake any examination offered by the Institute shall at the time of application and the taking of the examination be financial members of the Institute. CARRIED

6.

President’s Report Moved: A Joseph Seconded: S Tozer That the President’s Report be accepted as circulated in the Annual Report 2025/2026. T Barnett spoke to the President’s report. CARRIED

7.

Annual Report Moved: A Joseph Seconded: H Beall That the 2025/2026 Annual Report be accepted. CARRIED

8.

Financial Report Moved: A Joseph Seconded: R Hewett That the 2025/2026 Financial Report be accepted. CARRIED

9.

Updates to Constitution and Code of Ethics: • Use of nominals (Appendix 1) • Balance date (Appendix 2) Moved: A Joseph Seconded: T Barnett That the Use of nominals (Appendix 1) be accepted. A Joseph spoke to the use of post-nominals for the various types of NZIMLS membership. New Zealand Journal of Medical Laboratory Science 159

Proposed: That the following nominals be accepted for use by NZIMLS members: Associate: ANZIMLS Member: MNZIMLS Life Member: LMNZIMLS Honorary Member: HonNZIMLS Fellow: FNZIMLS CARRIED Moved: A Joseph Seconded: R Hewett That the Balance Date of 31 March (Appendix 2) be added to the Constitution and Code of Ethics. CARRIED 10.

SIG Policy document update re administrative fee. Increase fees by 15% including GST to cover ongoing expenses for SIG meetings from January 2027. Moved: A Joseph Seconded: K Birrell That the SIG Policy document update re administrative fee be accepted. CARRIED

11.

Council Honoraria Moved: A Joseph Seconded: R Siebers That no Council honoraria be made. CARRIED

12.

Auditor Moved: A Joseph Seconded: H Beall That the company of Martin Wakefield Ltd continue as the auditor of the NZIMLS financial accounts. CARRIED

13.

Annual PPTC Donation Moved: A Joseph Seconded: K Birrell That the Annual PPTC Donation be paid for 2026 at the rate of $8,000.00. CARRIED

14.

General Business – Strategic Priorities Moved: A Joseph Seconded: M Adriaansen That the NZIMLS Strategic Priorities 2026-2029 be accepted with the addition of the wording ‘Ti Tiriti’. CARRIED

15.

Region 5 representative Moved: A Joseph Seconded: H Beall That the Council decision to invite Tessy George to take the Region 5 representative role on Council due to the resignation of Leah Pringle be accepted. CARRIED

16.

NZIMLS AGM 2027 A Joseph announced that the 2027 AGM would be an online meeting, with the date to be confirmed by email to all members.

17.

2027 South Pacific Congress A Joesph announced that the 2027 South Pacific Congress will be held in Brisbane, at the Brisbane Convention Centre 18-20 August 2027.

There being no further business, the meeting closed at 8:10am

New Zealand Journal of Medical Laboratory Science 160

CONFERENCE REPORT

Fiji Institute of Medical Laboratory Science Annual Seminar 19-21 June 2026, Suva, Fiji Kirsten Birrell and Leah Pringle Kirsten Birrell (NZIMLS, Region 3 Representative) and Leah Pringle (NZIMLS, ex-Region 5 Representative) attended the FIMLS/AIMS Tropical Division Annual Seminar in Suva, Fiji, 1921 June 2026. The New Zealand Institute of Medical Laboratory Science Inc. (NZIMLS) was a sponsor of this event, and that sponsorship included sending two representatives as guest speakers to encourage professional development and learning between countries and also support the conference. This was a successful event with the attendees all highly motivated to listen and learn from the speakers who presented. The NZIMLS was recognised as an Event Partner, with both of us recognised and thanked as delegates, along with the NZIMLS logo displayed on the conference screens throughout the weekend with other sponsors/ partners. Thursday 18 June 2026 After an early start and some flight delays due to domestic fog, we flew from Auckland to Nadi and then onwards via road to Suva along the Coral Coast. The lush scenery, interestingly named snacks (Bongo Chips) and the 28°C temperatures announced our arrival in Nadi. Views along the Coral Coast were spectacular and a mongoose crossing the road was a bonus. After checking in to our room, we narrowly avoided seasoning our hotel dinner with a gecko who had made his home in the kettle. Mr Gecko was gently rehomed outside before we boiled the kettle, but we did check our shoes in the morning before putting them on, in case of stowaways. Friday 19 June 2026 The theme of the conference was Partnerships in the Pacific and aimed to promote collaboration and education across countries. To this end, there were speakers from the World Health Organisation, Fiji, Australia, New Zealand and Papua New Guinea. Over the three days we met representatives from Australia and Fiji, including James Cook University students and staff. Conversations ranged from “Where do you live? What does a laboratory look like in your country? How do you promote continuing professional development? And what registration bodies do you have for medical laboratory workers?” Fijian news outlets were present throughout the conference and delegates were asked to give interviews for Fiji One News: (https://www. facebook.com/people/Fiji-Institute-of-Medical-LaboratoryScience-FIMLS/61550658064136/) and it was excellent to see some media coverage for the profession. Friday’s schedule included; welcomes, introductions and photographs before the Open Expo in the afternoon, centring around introductions and time with product representatives. Afterwards, we jumped onboard with an Australian crew who had booked a tour of the local Colonial War Memorial Hospital. Visiting the Fijian laboratory was fascinating. Staff were enthusiastic, dedicated, highly skilled and knowledgeable and it was a pleasure to speak with them. The resilience of medical laboratory workers is the same across countries. We finished the day with a quick dip in our accommodation’s pool, followed by dinner at the Grand Pacific Hotel with delegates from Australia and Fiji, overlooking the ocean as the sun set. Saturday 20 June 2026 The main day for talks and when most attendees were expected. The first invited guest, The Honourable Penioni Ravunawa

(Assistant Minister for Health, Ministry of Health and Medical Services, Fiji), was formally welcomed and spoke on the hidden nature of medical laboratory science. Keynote speaker, Dr Tevita Qoriniasi (Acting Deputy Secretary - Public Health, Ministry of Health and Medical Services) spoke on the integral nature of laboratory scientists, focusing on the need for laboratory staff to perform research. Using an example, that research may be critical in assisting with early detection of outbreaks. There was a strong focus on the concept of “One Health” combining the needs of people, animals, earth and environment for the best possible health outcomes for all. Other speakers included: • Dr Sakiusa Baleivanualala: Fiji National University/Otago University, Antimicrobial resistance • Dr Philippe Guyant: Division for Pacific Technical Support – WHO, OneHealth in the Pacific • Leah Pringle: NZIMLS, Isolated raised APTT: causes and cases • Ailie Ross: James Cook University (Australia) Diagnosis of haematological malignancies in resource-limited settings • Kirsten Birrell: NZIMLS Equity, Te Tiriti o Waitangi and diagnostic medical laboratory research • Associate Professor Donna Rudd: James Cook University/ AIMS Tropical Division (Australia), Urinary nephrin for the detection of glomerular injury in pregnancy • Francisca Varpit: James Cook University (Papua New Guinea), Association between ABO groups and hepatitis virus infection • Neelambra Devi: Supervising Officer - Colonial War Memorial Hospital (Fiji), Evolution and current landscape of laboratory services at CWMH. Talks varied across a range of subjects with a research perspective and the day closed with panel discussions about “HIV diagnoses, testing, linkage to care, surveillance and collaborative public health response” and “Strengthening regional laboratory networks in the Pacific”. The conference dinner was held at the Grand Pacific Hotel with cultural performances by students from the Fiji National University and excellent food and beer. Some delegates stayed for the after-dinner clubbing, but we opted for an earlier night. Sunday 21 June 2026 The final half day of the conference included talks followed by breakout sessions for more in-depth and in-person learning. Most of these talks were from the students at James Cook University who had been on a short placement in Fiji as part of their medical laboratory science degree. They studied at the university in Fiji, went on lab tours and conducted an online case study that they then presented at the conference. The talks were interesting and well-informed. Talks included: • Khushi Nand: Scientist (Fiji), Amoebic liver infections at Lautoka Hospital • Nur Zyna: Sexual Reproductive Health and HIV Clinic, Ministry of Health, Fiji POCT and EQA, HIV and STI James Cook University Student Case Presentations: • Evelyn Waters; Carbapenem resistance in Fiji: Outbreak investigation • Rachel Beerwald; The acute febrile illness triad following a flood event • Hannah Ritchings; HIV and blood transfusion in Fiji • Jorja Lundie; AML in Fiji • Vivienne Gunn; Leptospirosis in Fiji

New Zealand Journal of Medical Laboratory Science 161

Leah attended the breakout session in haematology, a good overview of morphology, and interactive, using smartphone access to let people look at slides with Ailie Ross from James Cook University as she went through common red and white cell changes. Kirsten attended the Planetary Health panel discussion run by the Fijian Institute of Health Research, with a focus on Tadra Vanua (Tadra [Fiji] = to dream of, Vanua [Fiji/Vanuatu] or ‘Enua’ [Cook Islands] or Whenua [Māori] = people of/and place), moving beyond boundaries and looking holistically at health. This included a more in-depth dive into OneHealth and aspects of Fijian public health projects and initiatives, including; REFLECT (reducing heat within homes), RISE (Revitalising Informal Settlements and their Environments), and WISH (Watershed Interventions for Systems Health). The Biochemistry breakout session run by Associate Professor Donna Reid from James Cook University, was well received and attendee feedback was that it was interesting and informative. After the conference programme finished for the day we walked around Suva township before heading around the waterfront and through Thurston Gardens. Unfortunately, the Fiji Museum was closed, but we saw the Government Buildings before our dinner

with the Australian crew. Monday 22 June 2026 We had a predawn start with a three-hour taxi drive from Suva to Nadi Airport, followed by smooth flights to New Zealand and home. We are extremely grateful to the NZIMLS for providing us the opportunity to attend the FIMLS/AIMS Tropical Division Annual Seminar 2026. It was an amazing experience to attend an international conference, present talks for ongoing professional development for colleagues in the Pacific, and to represent New Zealand and the NZIMLS at a conference on collaboration in the Pacific, we donated some textbooks, which were gratefully received and we recommend continuing this practice in the future. The scenery was spectacular, the food amazing and the people welcoming. If anyone has another overseas conference needing speakers, do not hesitate to get in touch! Reported by: Kirsten Birrell and Leah Pringle (NZIMLS Regional Representatives)

Figure 1. Kirsten Birrell (NZIMLS Region 3 Representative) and Leah Pringle (NZIMLS ex-Region 5 Representative) and presenter awards.

Figure 2. FIMLS Conference Banners

Figure 3. Views of the Coral Coast, Fiji New Zealand Journal of Medical Laboratory Science 162

SCIENCE DIGEST Michael Legge Why DEET does not work – sometimes Mosquito repellents are essential to protect people from mosquito bites and one of the most widely used repellent is N,N-diethylmeta-toluamide (DEET). It is cheap to make, highly effective and its effects can last for up to five hours. How it may work was considered in 2008 when it was shown that DEET blocks mosquito sensory neurons, thereby confusing the mosquito. However, later research demonstrated a small proportion of mosquitos exposed to DEET that were insensitive to it and it was an inherited trait. In a new international collaboration (1) from France, Argentina and the USA, researchers tested the previous research and then went on to investigate DEET exposure by conditioning mosquitos to respond to various opportunities for access to warm blood. What was found was that those mosquitos exposed to DEET while feeding, had a significantly higher biting response when exposed to DEET again but exposure to DEET prior to feeding showed no biting response. The conclusion was that DEET does not affect mosquitos physiologically, if they sense DEET during the blood meal, then the mosquitos are more likely to bite people who smell of DEET and that mosquitos can learn and retain information. Formaldehyde risks in UK pathology laboratories Occupational exposure to formaldehyde is known to be associated with numerous short and long term deleterious health effects. As well as being a recognised carcinogen, formaldehyde exerts effects on the respiratory system, female reproductive and nervous systems. It has also been implicated in allergic reactions, asthma and cognitive impairment. Recently the European Union (EU) issued revised work formaldehyde exposure limits (long term 0.3ppm and short term 0.6ppm). In the current research undertaken in the UK (2) the authors obtained 12 months of formaldehyde airborne monitoring from histopathology laboratories in the UK, from 2024 to 2025. Of the 1,715,516 results obtained from 112 laboratories, formaldehyde monitoring was identified as infrequent (once weekly or less). The EU long term work exposure standard was exceeded regularly at 70% of sites and short-term exposure at 43% of sites monitored. Only 11% of sites monitored once daily or more. The authors also reported that the current UK formaldehyde work exposure limit is 2ppm for both long and short term exposure (approximately seven times higher than the EU limit). This research prompted workplace formaldehyde reviews by the UK Health and Safety Executive, the British Medical Association and the Royal College of Pathologists. The mummies curse: new risks on old legends Mummification of humans and animals has been undertaken for millennia with the oldest known human mummy dating back 16,000 years in China where bodies were ‘smoked’ over a slow burning fire. Over time the mummification process became more complex, using chemicals and physical processes to preserve the dead. The process and its outcomes have fascinated both scientific and public communities, as well as the film industry. Embalming fluid often contained a complex mix of heavy metals such as arsenic, mercury and lead, as well as oils and plant extracts. In more recent times a clandestine market has emerged for mummified remains and these are being traded on the internet in an unregulated market. Most international postal and courier services forbid the transmission of human remains. In the current collaboration (3) involving the UK, Italy and Australia, the authors monitored online sales posts for mummified human and animal remains, between 2015 and 2025. Nine countries including the USA (highest uptake), Europe and Australia identified a total of 128 on-line posts. They then undertook a comprehensive literature search of microorganisms associated with mummified remains and identified 18 species of bacteria including Protobacteria,

Actinobacteria and Firmicutes species, 21 fungi species including Basidiomycota, Zygomycota and Ascomycota species, and five yeast species. The literature search provided evidence of deaths associated with these microorganisms, many from Aspergillus spores, including the deaths of ten conservation scientists when a royal tomb was opened in Crete and probably the cause of death of Lord Carnarvon in 1923 following the opening of Tutankhamun’s tomb and not the mummies curse. The authors concluded that cultural considerations aside, not only are those owners of mummified remains at risk from infections but also the postal and courier workers who are unknowingly handling the packages. Authors urged for tighter controls and enforcement on the shipping of mummified remains. Hyperuricemia and cardiac multimorbidity Uric acid is an abundant antioxidant in the body and contributes to vascular homeostasis. Excessive uric acid, however, is associated with increased risks for coronary heart disease, type 2 diabetes and stroke via multiple metabolic pathways. In current research from the China (4) the authors used prospective data available through the UK Biobank and investigated the association of uric acid and metabolites with cardiometabolic disease (CMM), the evidence for progression to CMM and death, and the possible metabolic mechanisms that link the first two objectives. In total, data from 422,058 participants were analysed including follow-up data from the Biobank over a median time of 13.65 years. During that time 55,092 people developed cardiac metabolic disease (CMD) and 6,484 of these progressed to cardiac CMM and of these 2,234 died with CMM compared to those who did not have hyperuricaemia. In addition, the authors identified 126 differential metabolites associated with hyperuricacidaemia and constructed a metabolic score from these which had a strong correlation with CMD. This provided evidence of the complex interactions of hyperuricacidaemia with other metabolic pathways, and they conclude that effective management of elevated uric acid should be part of the strategy for mitigating the risks of CMM. Faecal calprotectin in older adults Faecal calprotectin is acknowledged as a useful marker for conditions such as inflammatory bowel disease and differentiation of irritable bowel syndrome. In addition, it may also be used as a marker for those undergoing endoscopy for colorectal cancer. In a study from the UK (5) the authors investigated the performance of calprotectin in identifying inflammatory bowel disease and the relevance of calprotectin in people aged >50years. Two groups were tested, 18 to 49 years old and >50 years old. All had faecal calprotectin assays followed by colonoscopy. In total there were 423 patients in the 18 to 49 age group and 246 who were >50 years old. There was no significant difference between the two groups and a calprotectin result of >50µg/g showed a high sensitivity for inflammatory bowel disease in both groups. The authors conclude that faecal calprotectin is a sensitive test for use with older adults and may help discriminate between inflammatory bowel disease and other gastrointestinal tract disorders. A blood test for Alzheimer’s edges closer While the development of Alzheimer’s and associated dementia disorders still avoids effective treatment. Many international research groups are investigating the possibility of blood biomarkers to use for early detection of these disorders and plan for timely intervention when necessary. Until now Alzheimer’s biomarkers necessitated the use of CSF or PET imaging. However, early investigations identified blood biomarkers associated with amyloid and tau pathology and neurodegeneration. Most of these associated investigations have been undertaken in clinical settings with patients already demonstrating cognitive disabilities.

New Zealand Journal of Medical Laboratory Science 163

New research from Sweden (6) used the Swedish National Study for Ageing and Care in a community-based setting. They included 2,148 adults aged 60 and over who have been followed for up to 16-years and the authors analysed six potential Alzheimer’s disease markers using blood samples. The markers were: amyloid-β-42/40 ratio, and the biomarkers p-tau181, p-tau217, total tau, NfL (neurofilament light chain) and GFAP (glial fibrillary acidic protein). The participants were clinically assessed over time but were all dementia free at the start of the research. Increased baseline levels of p-tau181, p-tau217, NfL ad GFAP had strong predictive values ranging from 72% to 82% over the 10-years for developing all-cause and Alzheimer’s dementia. The authors conclude that tailoring biomarker strategies would be crucial for counselling, care and intervention in communitybased settings.

2.

REFERENCES

6.

1.

Lazzari CR, De Luca D, Nally A et al. Associative learning switches DEET valence from aversive to appetitive in Aedes aegypti. J Exp Biol 2026; 229(10): jeb251935. doi: 10.1242/ jeb.251935.

3.

4.

5.

Plesa M, Yates RL. Under the microscope: formaldehyde exposure in National Health Service pathology departments in the United Kingdom. Occup Environ Health 2026; 83(3): 123-129. Squires K, Piombio-Mascali D, Urzi C, Huffer D. The real mummy’s curse: Health risks with the sale and trade of mummified remains. Int J Cultural Property 2026; 33: e1-23. doi: 10.1017/S094073912610093. [Open Access] Zhou Z, Wu Y, Chu J et al. Hyperuricemia, circulating metabolites, and the risk of cardiometabolic multimorbidity. Clin Chim Acta 2026; 18:593:121238. doi: 10.1016/j. cca.2026.121238. Perry RW, Foulser PFG, Zhang D et al. Evaluating the role of faecal calprotectin in older adults: a retrospective observational study. Brit J Gen Pract 2026; 76(263): e100-e107. doi: 10.3399/BJGP.2025.0169. Grande G, Valletta M, Rizzuto D et al Blood-based biomarkers of Alzheimer’s disease and incident dementia in the community. Nat Med 2026; 31(6): 2027-2035. [Open Access]

New Zealand Journal of Medical Laboratory Science 164

MEETING REPORT

NICE, Hamilton, June 2026 This year’s Transfusion Science Special Interest Group Weekend was held in June at the Distinction Hotel in Hamilton, bringing together 35 delegates from blood banks around the country. While attendance was lower than usual, partly due to competing conferences for transfusion scientists in 2026, the weekend still delivered a strong and engaging programme. Across the weekend, delegates presented 25 talks and 3 posters. Almost a third of attendees were first-time NICE participants, affectionately known as “NICE Virgins”, which is a positive sign of the event’s ongoing strength and appeal. The requirement for every attendee to present can be daunting, especially for those taking part for the first time. However, NICE continues to provide a welcoming and supportive environment, and a five-minute presentation is completed very quickly once you start. The calibre and variety of this year’s topics encouraged robust discussion across all sessions. With fewer presentations overall, there was more time for questions, answers, and valuable knowledge-sharing among delegates. Although our convenor, Prof. Holly Perry, was unable to join us this year, we look forward to welcoming her back in 2027. There was a good mix of NZBS and non-NZBS laboratories represented, with several smaller core laboratory and rural laboratory teams taking out a few prizes. Prize-winners and presentation topics: Best Overall Presentation, sponsored by Temprecord: Jo Armstrong, HNZ Waitemata — “What is a Purine Analogue?” • Best Poster, sponsored by Bio-Rad: Kristina Wheal, NZBS Christchurch — “I Need a Hero! A Case of a Pregnancy and an Operation 20 Years Later.” • NICEst Virgin – Best First-Time Presenter at NICE, sponsored by Grifols: Helen Doney, Medlab Whanganui — “Navigating Maternal Care from the Perspective of a Small Lab.”

•

• • • • •

Most Promising Transfusion Scientist, sponsored by Quidel Ortho: Jess Timms, NZBS Auckland — “Keeping It Flowing: The Blood Bank Behind Organ Transplants.” Sponsorship to Attend NICE 2027, sponsored by ParagonCare: Belinda Drummond, Awanui Labs Nelson — “Blood Grouping Anomalies.” Sponsored 4th Year BMLSc Transfusion Student, sponsored by Roche: Ashe Lim, AUT — “I Am Not KIDD-ing! Potential Naturally Occurring anti-Jka.” Highly Commended Presentations: Paul Lin and Juliet Jaggard, Waikato NZBS. Special Presentation – NZIMLS President’s Award: Raewyn Cameron, Pathlab Rotorua. Raewyn was awarded her pin by Ajesh at the Saturday evening dinner in recognition of her services to the profession and to NICE Weekend over many years.

Thank you!! As always, a massive miharo, awesome! To our sponsors for their continued support. Your contribution adds so much to the success of the weekend. Thank you also to Sharon, Fran, and Cathy at the NZIMLS office, who work so hard to make sure everything is organised before we arrive at the venue and who help keep Raewyn on track. A special mention goes to the scientists who put their hands-up, or, in some cases, were “voluntold” to serve as judges, chairpersons and general helpers throughout the weekend. The camaraderie of NICE Weekend never fails to warm my heart. We hope to see even more of you in 2027, when we hope to return to the Tron and continue sharing transfusion stories together. Reported by: Raewyn Cameron, Site Lead, Transfusion, Pathlab, Rotorua

Left to Right: Ashe Lim (Sponsored Student); Jess Timms (Most Promising Scientist); Kristina Wheal (Best Poster); Raewyn Cameron (TSSIG – NICE Coordinator); Helen Doney (NICEst Virgin); Robyn Roberts (TSSIG); Jo Armstrong (Best Presentation); Sarah Copedo (TSSIG); Belinda Drummond (Sponsored to NICE 2027); Juliet Jaggard (TSSIG); Diane Ritchie (TSSIG).

Raewyn Cameron with Belinda Drummond, ParagonCare sponsored NICE 2027 scholarship winner.

Ajesh Joseph, NZIMLS Treasurer, presents Raewyn Cameron with her 2026 President’s Award.

New Zealand Journal of Medical Laboratory Science 165

Microbiology Special Interest Group Meeting 27 June 2026, Tauranga We welcomed nearly 80 attendees to sunny Tauranga for this year’s Microbiology Special Interest Group (SIG) and yes, the sun did shine for us after some pretty wild weather. Unfortunately, the stormy weather further south resulted in some people pivoting to remote attendance and presentations, but thanks to the technology we had become familiar with during COVID times, we were able to continue relatively unscathed. The presentations throughout the day were interesting and varied, covering everything from parasites, mycology, novel antibiotics for unique clinical cases, unexpected outcomes to case studies, newly implemented methods and artificial intelligence. We had a magnificent guest speaker in Lizzie McKay who has grown up with Cystic Fibrosis and shared her story around what it was like growing up with the condition, undergoing a double lung transplant and the comparison between her journey and that of

Figure 1.

her twin brother who has travelled a different path with Trikafta. It was an insightful reminder for us all that the specimens we deal with daily all have a person at the other end and sometimes our results can have a significant impact on their management. Shannon Diggin's talk on small colony variant S.aureus saw her take the award for best presentation and Carly Williams was awarded runner-up with her talk on extra-intestinal Yersinia enterocolitica. Both winners were from Bay of Plenty Pathlab and both were first time presenters, so the future is bright with some enthusiastic young scientists taking part. I would like to extend a massive thank you to all those who took the time to prepare and present their talks to us. Without people like you we would not be able to organise these events which are important to our profession for knowledge sharing, growth and development.

Figure 2.

Shweta Shinde (left) from Ngaio Diagnostics, as one of our sponsors presented awards to Shannon Diggins, Best Presentation (Figure 1) and to Carly Williams, Runner up Best Presentation (Figure 2).

New Zealand Journal of Medical Laboratory Science 166

OBITUARY

Walter Joseph Wilson 1942-2026

Walter Wilson was a founding member of the New Zealand Institute of Medical Laboratory Science, and hugely influential in our profession. He would remain a member of the Institute for the remainder of his life, serving as Treasurer, Vice President and President from 1990‑1993. Walter’s influence and passion for education lives on in the special interest groups, whose establishment he championed. Walter began his career in the laboratory at Auckland Hospital in 1961. He managed the Coagulation Laboratory, appointed to this senior position at an early age, which was undoubtedly a testament to his technical and leadership skills. The years of Walter’s early career were among some of the pioneering days of medical laboratory science, and at Auckland Regional Blood Services (ARBS) Walter prepared antisera in a rabbit house, made blood agar, and worked on the development of both Factor VIII to treat haemophilia A, and RhD immunoglobulin to prevent haemolytic disease of the fetus and newborn. He would ultimately spend 34 years at Blood Services, many of these as Principal Technologist, in charge of the whole technical service. Walter was my manager at ARBS for the 10 years I spent there, and he was one of the best bosses I ever had; he was professional, kind, approachable and always willing to listen, with a truly open‑door policy for staff. One thing I remember about Walter was a notable day when a fire broke out in a side lab in which a researcher was extracting lipids with solvents. Walter discovered the fire and fought it single handed! In retrospect, we agreed that the building probably should have been evacuated and the fire service called, however it was a mark of the practical nature of the man! Amongst Walter’s notable achievements at ARBS was

tracking of blood and blood products. Walter travelled to Bangkok to advise and help set up the blood service there. In 1995 it was announced that there would be a national blood service in New Zealand and to manage this transition, the Ministry of Health set up the Blood Trust, with Walter as an Executive Officer. The role of the Executive Officer was to manage the transition from individual area health boards (AHB) to one national service. This was a huge undertaking, given that numerous AHBs each recruited and bled their own donors, in their own way with their own systems. New Zealand Blood Service (NZBS) became operational in 1998, following the work of the Blood Trust. With the opening of NZBS, the Blood Trust would be disestablished; Walter had effectively made himself redundant, a mark of his selfless work ethic and his belief in a national service. Not ready to stop working, Walter took up two new roles; Immunology Lecturer at Auckland University of Technology (AUT) and returning electoral officer for national elections. Walter was a very popular lecturer at AUT and was held in high regard by students and colleagues alike. Walter met his wife Denise early in his career; they were married in 1973 and raised three sons. In his retirement, Walter enjoyed time with his family, including eight grandchildren. At Walter’s memorial service, we heard about his interests outside work; construction on the family home, hunting, boating, waterskiing, scuba diving and cooking. Walter was modest about his achievements. He had a huge service ethic which was evident in his work and which he carried into retirement where he volunteered for Citizen’s Advice Bureau. I will personally remember Walter’s warm friendship and generosity outside work; when we were expecting our child, Walter and Denise gave us several items of baby equipment. I will remember his gentle manner and infectious smile. Walter was made a Life Member of NZIMLS at the Annual Scientific Meeting in Invercargill in 1990. He gave the TH Pullar Address at the 1994 conference in Hamilton with the title; “Health reforms; one year on the track” and in 2023 gave the Jim Le Grice toast at the South Pacific Congress in Auckland. Walter will be hugely missed in all spheres of his professional and personal life.

Prepared by: Holly Perry, Deputy Editor

the transition from paper records to computerised

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Otago BMLSc student research project abstracts: Semester 1, 2026 MAIT cells and the mystery of tissue repair Madison Adsett and James Ussher Department of Microbiology and Immunology, University of Otago, Dunedin Objectives: Mucosal associated invariant T (MAIT) cells are a subset of T cells with innate-like properties. The MAIT cell T cell receptor (TCR) recognises 5-(2-oxopropylideneamino)-6-Dribitylaminouracil (5-OP-RU). MAIT cell stimulation can also be TCR-independent through the combination of IL-12 and IL-18. It was also discovered that when stimulated via the TCR, a tissue repair gene signature was enriched. The tissue repair genes were shown to be expressed at a gene and protein level. Functional evidence was shown through accelerating tissue repair. The underlying mechanism of tissue repair in MAIT cells remains elusive. However, it is known anti-inflammatory cytokines help transition the immune response from a protective to healing state. There are many anti-inflammatory cytokines, in particular IL-4, IL-10, and TGF-β. Methods: RNA sequencing of MAIT cells with different stimulation conditions was analysed for IL-4RA, IL-10RA, TGFBR2, and tissue repair genes (n= 7). Peripheral blood mononuclear cells (PBMC) from healthy donors (n= 6) were stained for receptors at rest or stimulated with 5-OP-RU or Escherichia coli for 6 hours. Flow cytometry staining was performed and acquired on the Cytek Aurora. Analysis of data was done on FlowJo and GraphPad Prism. Results: Gene expression differed with stimulation, with an enrichment in tissue repair genes. IL-4RA and IL-10RA receptor expression was confirmed with flow cytometry. Intracellular expression exceeded surface expression. Receptor expression was also affected by 5-OP-RU and E. coli stimulation. TGFBR2 expression could not be confirmed due to non-specific staining. Conclusion: Together, this demonstrates that MAIT cells express IL-4RA, IL-10RA, and potentially TGFBR2 and expression is modulated by activation. Future studies should investigate whether cytokines IL-4, IL-10, and TGF-β directly act on MAIT cells, and whether they modulate their tissue repair function.

The difference between the 16S rRNA sanger sequencing assay and the MinION nanopore sequencer in the identification of bacterial species Lilia Aknine1 and Megan Burton2 University of Otago, Dunedin and 2Awanui Labs, Wellington.

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Objectives: Bacterial infections require different treatments depending on the species involved. Quick, reliable bacterial identification is crucial for correct patient care. Most bacteria are identified through microbiological techniques; however phenotypically abnormal or fastidious bacteria cannot always be. Nucleic acid testing is then required for bacterial identification. The 16S Assay uses Sanger sequencing to read short sections of the 16S rRNA gene, while the MinION uses nanopore technology to read the entire genome. The 16S Assay is currently used; howe ver, it is time-consuming and labour-intensive. This report will investigate whether the Nanopore MinION identifies bacterial species correctly compared to the 16S rRNA Sanger Sequencing Assay. Methods: 4 gram-positive and 3 gram-negative bacterial isolates, difficult to identify using microbiological techniques, were recultured on blood agar. All sample nucleic acids were extracted via the Roche High Pure PCR Template Preparation Kit and then underwent the 16S Assay. All gram-negatives were extracted again using the MagNAPure 96 instrument, while gram-positives were extracted using the Quick-DNA Fungal/Bacterial Miniprep Kit. Samples were then run through the MinION protocol. The data was analysed using Applied Biosystems software and POx_POC bioinformatic pipeline for 16S Assay and MinION, respectively.

Results: Of the 7 samples, 5 were speciated with the same result from both sequencers, 1 was unable to be speciated by either sequencer, while 1 was speciated only by the MinION. Conclusion: The results show the MinION can identify bacterial species to the same degree or better than the 16S Assay. The MinION has the potential to be an alternative for the 16S Assay; however, the procedure would need further testing before integration into the laboratory. Reticulocyte haemoglobin as a screening tool for iron deficiency in pregnant women Josie Batt1 and Clare Berry2 University of Otago, Dunedin and Christchurch

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Awanui Laboratories,

Objectives: Iron deficiency is the most common deficiency worldwide, affecting 16.7% of people. During pregnancy iron stores are even more depleted due haemodilution and foetal growth. Consequently, this causes a higher risk of neonatal and maternal morbidity linked to preeclampsia, postpartum haemorrhage and preterm birth. As a result of this, monitoring iron during pregnancy is incredibly important. Currently pregnant women are screened for iron deficiency in their first and second trimesters by measuring iron biomarkers in the serum, however, these can be unreliable in the presence of inflammatory molecules. This study investigates a new parameter for screening iron deficiency, reticulocyte haemoglobin. Methods: This study was conducted by collecting 44 second trimester antenatal samples and sampling ferritin, CRP and haemoglobin to determine the initial iron status of each patient. Using a Sysmex XN-20 analyser, a complete blood count was then run with an added reticulocyte channel, allowing reticulocyte haemoglobin to be measured. Results: Reticulocyte haemoglobin declined with increasing iron deficiency and remained stable during inflammation, as opposed to ferritin which showed erratic results. The average reticulocyte haemoglobin in iron replete patients was 34pg, compared to 30.4pg in iron deficient patients. ROC analysis gave an AUC of 0.85, which indicates good diagnostic accuracy. Conclusion: Across varying degrees of iron status, reticulocyte haemoglobin demonstrated consistent behaviour, despite inflammation. This supports its diagnostic utility in being able to detect cases of iron deficiency early and accurately. NKX3.1 as a replacement marker for Prostate-Specific Antigen in prostate immunohistochemistry Dior Bowden1, Jordyn Wilkinson2 and Grace Wilshaw2 1 University of Otago, Dunedin and 2Pathlab, Bay of Plenty Objectives: At Pathlab, Bay of Plenty, prostate-specific antigen is currently one of the standard antibodies used in immunohistochemistry for prostate tissue. However, its limitations and sub-optimal performance have led the laboratory to investigate whether a different marker would be more effective. This study aimed to evaluate whether the newer marker NKX3.1 could serve as a suitable replacement for prostate-specific antigen in the investigations of prostate tissue. In addition, the study sought to identify the most effective staining protocol for NKX3.1 and determine which supplier between Cell Marque and BioCare provided optimal staining performance for use in the laboratory. Methods: Immunohistochemistry was performed on the Leica BOND-PRIME platform using a polymer 3,3′-diaminobenzidine detection system. Seven staining protocols incorporating varying epitope retrieval and antibody incubation times were evaluated using a control block containing prostate and testis tissue. NKX3.1 antibodies from Cell Marque and BioCare were compared for staining quality and specificity.

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Results: It was found that of the two companies, BioCare produced the most optimal staining results when using 20 minutes of Epitope Retrieval 2 and a 10-minute antibody staining protocol. In prostate tissue, NKX3.1 provided strong, crisp nuclear staining in most of the luminal epithelial cells lining prostate glands. In testis which served as a sensitivity marker and positive control, there was weak to moderate nuclear staining in dispersed spermatogonia of the seminiferous tubules, indicating optimal staining. Conclusion: Compared to prostate specific antigen staining, pathologists at Pathlab determined that NKX3.1 demonstrated superior performance using this method and could therefore be adopted for use in the laboratory to safely replace prostate specific antigen for investigation of prostate tissue. Comparison of three Kleihauer-Betke staining kits Used for fetomaternal haemorrhage detection.

films were selected, and 10 random non-flagged samples were also selected. For each patient sample, three blood films were made using the CellaVision HemaPrep® for validation, totalling 60 films. One of each triplicate was stained using the Aerospray® Pro 7152, the Siemens Hematek® 3000, and the Sysmex SP-10 in Whangarei. Manual film examination using light microscopy was done to compare the three different stainers. Results: There was some variation in staining across the three different methods. The Aerospray® produced inconsistent results with slides having excessive stain deposit and blurry patches. The Hematek® seems to stain leukocytes bluer but with clearer overall film definition. The SP-10 was used as a reference stainer and produced clear well stained results each time. Conclusion: The Hematek® 3000 demonstrated more reliable and accurate staining than the Aerospray® Pro 7152, with less maintenance health and safety concerns. Due to these results, the Hematek® 3000 has been validated for diagnostic use and is being used.

Liu Cao1, Ramesh Tiwari2, and Caity Anderson2 University of Otago, Dunedin and 2Canterbury Laboratories, Christchurch

Y chromosome microdeletion screening: moving from gel to fragment analysis

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Objectives: The Kleihauer-Betke test detects fetomaternal haemorrhage by identifying fetal red blood cells in maternal blood, guiding anti-D administration in RhD-negative mothers. Because interpretation depends on clear staining and distinction between fetal cells and adult ghost cells, variation between Kleihauer kits may affect result reliability. This project compares the Australian Biostain and Guest Medical kits with the CHL in-house method to identify a kit with easier reagent management while maintaining good staining quality and reliable interpretation. Methods: Ten spiked samples ranging from 0.1% to 5% fetal cells were stained with all three Kleihauer methods, producing 30 slides excluding controls. One slide was prepared for each method at each dilution, and 222 adult ghost cells were counted once per slide. Fifteen patient samples were stained with all three methods and assessed qualitatively for fetal cell visibility, adult ghosting, staining uniformity, background staining, intermediate cells, and ease of counting, with additional review by five CHL scientists. Results: Although the Guest Medical kit improved workflow and reagent management, it tended to produce lower counts in spiked samples. Some slides also showed overly pink adult ghost cells, which reduced the contrast between ghost cells, intermediate cells, and fetal cells, making interpretation less clear. The Guest Medical kit showed the best workflow efficiency, with shorter processing time and no reagent preparation required, demonstrating staining quality comparable to the CHL in-house method. The Australian Biostain kit showed greater variability, with less consistent staining and more intermediate cells. Conclusion: The Australian Biostain kit showed greater staining variation and more intermediate cells. Therefore, the CHL inhouse method remained the preferred option, as it provided clearer staining, more reliable interpretation and lower cost for routine Kleihauer testing. Validation of the Siemens Hematek® 3000 for haematology staining, and comparison to the Aerospray® Pro 7152 Isabel Gallagher1, Michael Herring2 and Teresa Travers2 University of Otago, Dunedin and 2Te Whatu Ora, Kaitaia

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Objectives: Blood film examination is a key part of haematology, necessary in identifying normal and abnormal morphology for diagnoses. A factor that determines the quality of the morphology is the staining technique and equipment used. In the Kaitaia laboratory, haematological staining was done using the Aerospray® Pro 7152, producing inconsistent and suboptimal results. This project aimed to validate a new Siemens Hematek® 3000 stainer to be able to implement it for diagnostic use in the Kaitaia laboratory. Methods: K2EDTA samples were collected from patients across the period of a week for full blood count analysis. 10 random samples that had been flagged by the Sysmex XN-1000 for blood

Rocel Katuin1 and Kylie Drake2 1 University of Otago, Dunedin, Laboratories, Christchurch.

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Canterbury

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Objectives: Fertility referrals for azoospermic or oligozoospermic men include screening for Y-chromosome microdeletions in azoospermia factor (AZF) regions a, b, and c. Detection of partial or full deletions is associated with male infertility. Screening at Canterbury Health Laboratories is currently gel-based, with up to eight patient samples per run. This capacity is proving unsustainable as fertility referrals increase; managing larger volumes requires transitioning to a high-throughput method. This study aimed to validate fragment analysis for screening of Y chromosome microdeletions. Methods: Twenty-two DNA samples (12 with deletions, 10 normal controls) were re-tested using fragment analysis on the ABI3500 genetic analyser (ThermoFisher). Amplification was performed via fluorescent PCR with each primer set (n=8) containing a FAM-labelled primer. Prior to multiplexing, primers were checked to determine length. Two multiplex panels (A and B) were then created based on the original gel mixes. Panel A included markers sY127, sY84, sY254, and SRY; Panel B included sY134, sY86, sY255, ZFY. Diluted PCR products were run with GeneScan™ 600LIZ dye size standard and HIDI Formamide (ThermoFisher) on an ABI3500. GeneMapper™ Software (ThermoFisher) was used for analysis. Results: AZF deletion results were concordant between methods in 21/22 samples. The non-concordant result was a QAP sample that was incorrectly genotyped by gel and correctly genotyped using fragment analysis. Samples tested for inter- and intra-run variability produced identical results each time. Conclusion: This investigation has demonstrated that fragment analysis on the ABI3500 is a reliable method for accurate AZF deletion screening. Transitioning methods offers improved efficiency, standardisation, and scalability while maintaining analytical confidence. Implementation of fragment analysis will support increased demand from rising fertility referrals and reduce operational burden. Optimisation of the INSM1 antibody for diagnostic use in neuroendocrine tumour identification at Awanui Labs, Canterbury Katie Kinnaird¹ and Rowena Hunter² ¹University of Otago, Dunedin and ²Awanui Labs, Canterbury Objectives: Immunohistochemistry is a vital tool in the histopathology department, particularly in the identification of specific cancerous biomarkers. Insulinoma-associated

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protein 1 (INSM1) is a transcription factor playing a key role in neuroendocrine cell differentiation, making it an effective neuroendocrine tumour marker. INSM1 is widely utilised at various laboratories; however, the optimal conditions for its use at Awanui Labs, Canterbury, have not yet been established. This study aimed to identify the optimal protocol for the INSM1 antibody. Methods: The investigation was conducted using two INSM1positive pancreatic tissue controls. Tissue sections were stained using the VENTANA BenchMark ULTRA PLUS Automated Immunohistochemistry Staining System. This protocol utilised a Roche Optiview DAB detection kit and involved manual antibody titrations. Various conditions were trialled, including multiple antibody dilutions, incubation times, and the addition of an amplification step. Slides were then analysed microscopically for clarity and strength of DAB staining, and the minimisation of background staining. The staining pattern is nuclear staining in the islets of normal pancreas. Results: Optimal conditions were found to be an INSM1 concentration of 1:50, with an amplification step, and a 36-minute incubation. These conditions produced the most effective nuclear staining of the pancreatic tissue, with minimal background staining, and were the favoured result when validated by an Awanui anatomical pathologist. Conclusion: These findings allow implementation of the INSM1 protocol at Awanui Labs, Canterbury, with optimal detection for the diagnosis of neuroendocrine tumours. This allows INSM1 to be utilised on-site, without reliance on alternative laboratory send aways, and can serve as an alternative to synaptophysin and chromogranin staining. The optimised conditions produce strong, specific INSM1 staining, informing reliability for its diagnostic use. Comparison between tube and card techniques for P1 phenotyping with Grifols Anti-P1 Mono-Type Dual antiserum Megan Lake1, Nicola Crampton2 University of Otago, Dunedin and 2New Zealand Blood Service, Christchurch 1

Objectives: Tube and microcolumn agglutination methods are common techniques used in blood banks for a variety of tests, including phenotyping. Red cell antigen phenotyping is routinely performed during antibody investigation to confirm antibody specificity. The Christchurch Hospital Blood Bank has recently begun using Grifols Anti-P1 Mono-Type Dual antiserum, for which there are two possible methods. This project aimed to determine if the tube or card technique would be more suitable. Methods: 44 samples were randomly selected from red cell units less than 14 days old. These samples were tested in batches alongside suitable control cells by tube and microcolumn techniques following the methods detailed in the Grifols Anti-P1 Mono-Type Dual package insert. Red cell agglutination strength was graded on a scale of 0-12 for the tube technique, and 0-4 for the card technique. Results were recorded in a spreadsheet in Microsoft Excel and analysed. Results: The key factors compared were cost, time, strength of agglutination, and ease of use. Both methods had strong positive reactions and were easy to perform and grade. Most results were strong positives – 10-12 in tube and 3-4 in card. Performing the P1 phenotyping with the card technique had an overall lower cost than the tube method, and the microcolumn reactions were subjectively easier to grade. Conclusion: There was not a clear conclusion as to whether one method was more suitable than the other as both proved to be acceptable for P1 phenotyping. It is likely that the method used will be determined by personal preference or other such factors. Laboratory staff should use whichever method is more

appropriate at the time of testing. Evaluation of NG-TEST® CARBA-5 for the detection of carbapenemase enzymes from clinical isolates Sonika Maisuriya1 and Casey O’Byrne2 Univerisity of Otago, Dunedin and 2Awanui Labs, Wellington Hospital Objectives: Beta lactams are the first-line antibiotic treatment of infections caused by Enterobacterales and Pseudomonas aeruginosa, however their efficacy has been challenged by beta lactamase production. Recently, beta lactamases with the ability to hydrolyse and inactivate carbapenem antibiotics (carbapenemases) have increased in prevalence and spread globally. This makes reliable and rapid identification critical. The aim of this study was to evaluate the performance of the NGTEST® CARBA-5 multiplex immunochromatographic assay for the identification of the most prevalent carbapenemase families (KPC, OXA-48, VIM, IMP, and NDM) present in bacterial culture. Methods: Twenty clinical bacterial isolates were selected among archived Enterobacterales and Pseudomonas aeruginosa identified by Cepheid Xpert® Carba-R at Awanui Labs, Wellington Hospital. These included a minimum of n= 2 carbapenemasepositive isolates per targeted enzyme, n=3 negatives, n=1 meropenem-resistant, and n= 2 isolates with multiple targets. Isolates were sub-cultured onto blood-agar plates from frozen stock thrice for organism recovery and colony isolation. Single colonies were mixed with the provided extraction buffer and loaded onto the test cassettes. The results were observed after 15 minutes. Results: The NG-TEST® CARBA-5 successfully detected and differentiated the presence of carbapenemases with 100% sensitivity and 100% specificity. Results were produced rapidly with minimal hands-on activity and a simple procedure. This study provides insight to the assay’s efficacy, however further investigation with a larger sample number and varied selective media would reflect an in-depth validation. Conclusion: The NG-TEST® CARBA-5 can accurately detect multiple prevalent carbapenemases present in a clinical bacterial isolate. It aids the rapid identification of patients with carbapenem antibiotic-resistance to guide therapeutic selection. It has high potential to be implemented into current routine diagnostic laboratory methods at Wellington Hospital. 1

Haematoxylin & eosin staining methods for digital pathology Zaria Muirhead1, Diane Kenwright2, Alicia Clark2, Shweta Mohandas2 1 University of Otago and 2Awanui Labs, Wellington Objectives: Haematoxylin and Eosin is central to histological diagnostic practice. However, there are no standardised protocols across laboratories. Inter-lab variation will become more significant as more laboratories adopt digital pathology, as image quality and consistency are key for accurate interpretation, and for application of artificial intelligence. This project aims to recommend an effective haematoxylin and eosin staining protocol for digital pathology. Methods: Seven haematoxylin formations, two eosin formulations, various staining methods, and section thicknesses were systematically trialled and evaluated. Colon, renal, and skin tissues were sectioned at 2-4μm and were manually stained using both progressive and regressive methods. A pathology registrar and a pathologist interpreted the slides according to the RCPAQAP staining criteria. Results: The best performing haematoxylin was Epredia 7211 with progressive staining of 2 minutes haematoxylin and 2 minutes eosin. 7211 was only effective when used with Epredia 7212, an alcoholic eosin. The ideal section thickness was 3μm, which improved definition. Trials to create darker staining only resulted in decreased quality.

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Conclusions: Leica Biosystems Aperio GT450DX digital scanners produced better image quality with 3μm sections stained for 2 minutes each of haematoxylin and eosin. Optimisation and standardisation of haematoxylin & eosin staining protocols is essential to support effective implementation of digital workflow and AI-assisted diagnostics. Verifying the use of serum samples for HIV-1 viral load assays on the Alinity M system Emma Paine1 and Jennifer Fahey2 1 University of Otago, Dunedin and 2Canterbury Health Laboratories Objectives: Human immunodeficiency virus 1 (HIV-1) is an RNA retrovirus that infects 40.8 million individuals globally. HIV-1 is the causative agent of acquired immunodeficiency syndrome (AIDS) and is associated with severe mortality and morbidity. HIV1 diagnosis relies on antibody detection and viral load assays to quantify the virus. HIV-1 viral load assays are performed on plasma samples. Serum samples may provide an effective alternative which eliminates the need for patients to return for plasma collection. This research project aims to verify the use of serum samples for HIV-1 viral load assays on the Alinity M System. Methods: 13 positive and 15 negative patient samples were retrieved from storage. A single donor provided plasma and serum samples to spike with an HIV-1 positive control for a dilution series. All samples were processed on the Alinity M molecular analyser, which utilises a real-time, reverse transcriptase polymerase chain reaction for the extraction, amplification and detection of HIV-1 genomic sequences and produces a numerical value to represent the quantity of viral copies/mL. Results: The positive samples revealed consistency across both plasma and serum viral loads. However, five samples from known HIV positive patients returned low-level positive results, such findings are consistent with occasional amplification events that can occur at extremely low viral concentrations that are common in HIV positive patients. Conclusion: Serum samples are a consistent and accurate alternative matrix for HIV-1 viral load assays on the Alinity M and can therefore be used for reflex viral load testing. Further investigation with a larger positive sample size and the use of true-negative samples should be performed to reinforce the results of this project. Investigating Ferumoxytol-induced ferroptosis in Δ133p53 isoforms Phoebe Pepper, Adriana Saraiva, Tania Slatter Department of Medical Laboratory Science, University of Otago, Dunedin Objectives: Brain tumours are an aggressive form of cancer and magnetic resonance imaging (MRI) scans are used for provisional diagnoses. Ferumoxytol is an ultrasmall superparamagnetic iron oxide nanoparticle and can be used off-label as an alternate MRIcontrast agent to gadolinium. It is currently on trial at Dunedin Hospital for brain tumour patients to determine if it can provide enhanced imaging for some patients. Initial trial results suggest ferumoxytol may also have potential therapeutic effects on brain tumours with some patients surviving longer than expected. A potential mechanism could be that ferumoxytol induces ironrelated cancer cell death, ferroptosis. Δ133p53 is an isoform of tumour suppressor p53, which has been shown to be increased in brain metastases. Initial evidence shows the isoform effects the amount of transferrin receptor at the cell surface, a protein involved in iron regulation. Methods: This project aimed to see if the distribution of proteins involved in iron regulation was altered with the expression of the Δ133p53 isoform using immunohistochemistry and immunofluorescence staining on brain and lung cancer cell lines expressing Δ133p53. Proteins investigated were, Divalent

Metal Ion Transporter 1 (DMT1), Transferrin Receptor (TFRC), Glutathione Peroxidase 4 (GPX4) and SLC40A1 (encodes Ferroportin). Results: Chromogenic immunohistochemistry revealed no difference in total amount of protein with Δ133p53 expression. A multiplexed immunofluorescence assay was optimised for future use to investigate cellular distribution of these proteins with Δ133p53 isoform expression. Further studies will need to be done to confirm whether these proteins are altered in different parts of the cell for example upregulated on the cell surface. Conclusions: This knowledge will contribute to further understanding if ferumoxytol can induce ferroptosis in tumours and if this could provide therapeutic effects of ferumoxytol towards improved patient survival. From sediment to structured tissue: the evolution of cell block preparation methods and the diagnostic value of the cell block Samantha Reyes1, Krish Pillay2 University of Otago, Dunedin and 2APS/LabPlus Auckland

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Objectives: Cell blocks have become an essential component of modern cytology, being utilised not only in cytopathological processing but also within histopathology and molecular diagnostics. However, cell blocks often have issues in terms of cellularity, DNA contamination, and standardisation, with an abundance of different cell block preparation techniques being described. This review aims to explore various cell block preparation techniques, discussing advantages and limitations within each preparation method, and to highlight the diagnostic significance of cell blocks for diagnosis and further ancillary testing. Methods: A literature review was undertaken to highlight the most used cell block preparation techniques in diagnostic laboratories, such as the plasma-thrombin and HistoGel method, as well as novel modifications and novel advances in preparation technology, like the Automated Cellient method. A case study was then followed to demonstrate the practical value of the cell block in the diagnosis and further testing of a tumour. Results: The literature review demonstrated that the selection of cell block preparation techniques is influenced not only by the unique advantages and limitations of each preparation method, but also by the budgets, space, and overall requirements of each diagnostic laboratory, and therefore no single preparation method can be identified as universally superior. Additionally, the case study highlighted the growing importance of cell blocks in the diagnosis of tumours and in opening the possibilities for personalised treatment pathways, as cell blocks provide minimally invasive and suitable material for various testing. Conclusion: Cell block preparations remain an important component of modern cytopathology. Despite the variability in preparation techniques, their diagnostic utility reinforces their importance in tumour diagnosis and possibilities within personalised medicine. Audit of cardiovascular genetic testing sent by LabPLUS, Auckland to Blueprint Genetics, Finland Emily Smith1, Rebecca Lopes2 and Mark Greenslade2 1 University of Otago, Dunedin and 2LabPLUS, Auckland Objectives: To review the diagnostic yield of cardiac genetic testing sent to Blueprint Genetics Finland by LabPLUS, Auckland. It is important to ensure that international send away of genetic testing is being referred appropriately, due to the clinical significance of potential diagnoses. Methods: 318 patients who had DNA sent to Blueprint Genetics from LabPLUS between 01/03/2025 and 28/02/2026 for a cardiovascular gene panel were included. Requests for whole exome sequencing were included only if the request form indicated an exclusively cardiovascular phenotype. Results: 51 patients received a diagnostic result (Pathogenic/ Likely Pathogenic, 16.29%). The highest diagnostic yield seen

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was in dilated cardiomyopathy referrals (45.10%, n = 28). Multiple variants were found in 10 patients, and results were pending for 5 patients by date of publication. Variants of uncertain significance (VUS) were found in 14.69% of our cohort, with the lowest rates of VUS detection seen in Māori and Pasifika populations. 14 variants were requested for reclassification using ACMG criteria, of which 2 were pending, 4 did not change classification, 6 variants were downgraded from Pathogenic/Likely Pathogenic to VUS and 1 was upgraded from VUS to Pathogenic. Conclusion: Cardiovascular referrals for overseas genetic testing yielded a diagnosis in 16.29%, with patients referred for dilated cardiomyopathy most frequently receiving a diagnosis. VUS results were returned for 14.69%, leading to significant clinical uncertainty and additional follow up. LabPLUS reclassification of reported variants changed the reported classification in 66.66%. Further research focused on improving gene panel content and variant classification to minimise VUS rates is encouraged. Expanding this study to include all genetic testing sent from New Zealand to Blueprint may also increase relevance and reduce inter-laboratory variation. Phenotypic characterisation and antimicrobial susceptibility profiling of Carbapenemase-Producing Enterobacterales Grace Suaalii,1 Kate Elgar,2 Kristin Dyet,3 Juliet Elvy.2,3 University of Otago, Dunedin; 2Awanui labs Dunedin, and 3 The New Zealand Institute for Public Health and Forensic Science (PHF Science), Wellington.

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Objectives: Carbapenemase-producing Enterobacterales are multidrug-resistant gram-negative pathogens associated with limited treatment options. Emerging resistance mechanisms beyond carbapenemase production highlight the need to better understand their impact on novel β-lactam agents. This study aimed to characterize the antimicrobial susceptibility profiles of genomically characterized carbapenemase-producing Enterobacterales isolates. It also assessed the impact of resistance mechanisms, including ftsI mutations, on novel β-lactam agents. Additionally, disc diffusion and gradient diffusion (E-test) methods for aztreonam-avibactam susceptibility testing were compared. Methods: 31 Enterobacterales isolates were analysed. Phenotypic susceptibility testing was performed using routine antimicrobial susceptibility panels, extended β-lactamase screening, and multidrug resistance profiling. Aztreonamavibactam susceptibility was assessed using both disc diffusion and E-test methods. Results were compared for concordance. Cefiderocol susceptibility was evaluated by disc diffusion. Carbapenemase production was confirmed using the modified carbapenem inactivation method. All isolates had been fully characterized prior, at PHF Science, using whole genome sequencing to identify species and detect genotypic resistance mechanisms, including ftsI mutations. Results: Most isolates exhibited multidrug-resistant phenotypes consistent with carbapenemase production. Notably, isolates harbouring ftsI gene mutations showed reduced susceptibility to aztreonam-avibactam compared with ftsI-negative isolates, which were all susceptible, supporting a possible association between ftsI mutations and decreased activity. Cefiderocol activity was variable among ftsI-positive isolates, whereas all ftsInegative isolates were susceptible. Conclusions: Combined phenotypic and genomic analysis improves understanding of resistance mechanisms and treatment options in carbapenemase-producing Enterobacterales. These findings suggest ftsI mutations may contribute to reduced susceptibility to aztreonam-avibactam and should be considered when interpreting susceptibility results. Strengthening surveillance and diagnostic capacity may improve detection of resistant organisms and inform appropriate antimicrobial use. Is this the Key? validation of a digital Ki67 algorithm across breast cancer Anna Wilson1, Stephen Fox2 and Cameron Snell2

University of Otago, Department of Medical Laboratory Science Cancer Evolution and Metastasis Program and, Department of Pathology, Peter MacCallum Cancer Centre, VIC, Australia

1 2

Objectives: Ki67 immunohistochemistry provides critical prognostic information in breast cancer but suffers from interobserver variability limiting its clinical utility. Digital image analysis offers an objective solution; however, commercial algorithms require rigorous analytical validation before clinical implementation. The Roche uPath Ki67 algorithm has the potential to improve standardisation, yet its performance relative to manual scoring and Australian laboratory accreditation standards remains unevaluated. This study aims to validate the Roche algorithm in an Australian pathology laboratory setting. Methods: Retrospectively, 94 cases of primary breast cancer were retrieved from archival formalin-fixed paraffin-embedded (FFPE) tissue at Peter MacCallum Cancer Centre. Cases were stained using the Ventana Ki67 (30-9) assay on the BenchMark ULTRA platform and digitised at 20x magnification using the VENTANA DP 200 whole slide scanner. Ki67 proliferation indices were assessed independently by three expert pathologists using semi-quantitative visual estimation, reporting the result to the nearest 10%, providing the reference standard for the algorithm. Digital Ki67 proliferation index was derived using the Roche uPath Ki67 image analysis algorithm within the Navify digital pathology software platform. Results: Pathologists showed good agreement (ICC = 0.858). Good reliability was observed with an ICC of 0.890 (CI: 0.793– 0.936) between the algorithm and manual reference standard. Bland-Altman analysis showed a mean bias of -5.78% (95% limits of agreement: -29.83% to 18.27%) between methods, indicating a tendency toward algorithmic underestimation. Conclusion: This study demonstrates a digital image analysis algorithm has the potential to reduce variability in Ki67 scoring and be used clinically as a standardised Ki67 digital scoring method. Evaluation of the cost effectiveness of prophylactic anti-D vs fetal Rh (D) genotyping in New Zealand Brandon Winter¹, Ruth Brookes², Janine Gundersen² ¹University of Otago, Dunedin, ²New Zealand Blood Service (NZBS), Palmerston North Blood Bank Scope: Royal Australian and New Zealand College of Obstetricians and Gynaecologists (RANZCOG) guidelines recommend administering anti‑D immunoglobulin prophylaxis to Rh(D)-negative mothers to prevent haemolytic disease of the fetus and newborn (HDFN). However, only approximately 60% of these pregnancies result in a Rh(D)‑positive fetus, meaning prophylaxis is not required. Fetal RhD genotyping offers a targeted approach to determine foetal Rh(D) presence which establishes anti-D requirements. This study evaluated the potential cost‑effectiveness of adopting molecular genotyping to determine Rh(D) prophylaxis requirements within New Zealand. Methods: A cross‑sectional analysis was conducted using laboratory report data compiled by senior scientists. Cord blood group and direct antiglobulin test (DAT) results were reviewed to determine whether anti‑D administration was clinically indicated based on neonatal Rh(D) status. This data was compared with actual anti‑D issues to identify clinically unwarranted administration. The cost of current practice was compared with projected costs of implementing routine foetal Rh(D) genotyping. Results: Analysis showed that the cost difference between current universal prophylaxis and a genotyping‑guided approach was minimal. Although genotyping would slightly increase overall expenditure, the difference was negligible relative to total programme costs. Conclusion: While the financial impact of adopting fetal Rh(D) genotyping appears minimal, transitioning to molecular testing may offer long‑term advantages. These include more targeted use of anti-D immunoglobulin, reduced pressure on supply, and decreased patient exposure to blood-derived products. The downstream effects of current practice, such as increased laboratory workload and costs associated with positive antibody

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screens, antibody identification, and cross-matching in patients who may have received unnecessary anti-D prophylaxis—should be further evaluated in future studies. Incorporating genotyping into standard practice warrants further consideration as part of future NZBS strategy. Effect of the use of enzyme Bromelase 30 during phenotyping of Jkb phenotype Timothy Wong1, Georgia Medland2 1 University of Otago, Dunedin and 2New Zealand Blood Service, Dunedin Hospital Blood Bank. Objectives: In transfusion science, patient blood phenotyping may be required to ensure safe transfusion practices and to manage specific medical conditions. Phenotyping helps identify compatible blood units that are unlikely to stimulate the production of new antibodies. The aim of this project was to determine the effect of Bromelase on Kidd phenotyping using the card technique. This was achieved by following the manufacturer's guidelines and comparing phenotyping results obtained with and without the use of Bromelase. Methods: Six donor red cell units with known Kidd phenotypes were used. Three units were homozygous for Jkᵇ, while the other three were heterozygous for Jkᵇ. A heterozygous positive control and a Jk-null negative control were selected from the antibody panel. The Grifols test procedure was followed, as both the reagent and the IAT cards used in this study were manufactured by Grifols. Two tests were performed on each sample: one with Bromelase and one without Bromelase. The results were then interpreted and recorded using a grading system ranging from 0 to 4+. Results: Results confirmed phenotyping performed with Bromelase produced highly consistent results, with all samples showing strong reactions. In comparison, the method performed without Bromelase produced weaker reactions across all samples, with particularly reduced reactivity observed in the heterozygous samples. Conclusions: Bromelase demonstrated a significant enhancement effect on Kidd phenotyping reactions. This enhancement is thought to occur through several mechanisms, including the reduction of the zeta potential on the red blood cell surface and the modification of membrane structures, which may improve the binding of anti-Jkᵇ antibodies. However, repeat testing and a larger sample size are required to increase the reliability and validity of these conclusions. Assessing platelet flagging accuracy of the Sysmex XN10 haematology analyser for identification of platelet abnormalities Genna Woodward1, Catherine Cahill2, and Shweta Kumar2 1 University of Otago, Dunedin and 2Awanui Labs, Invercargill Objectives: Accurate quantitation of platelets is essential for evaluation of patient bleeding and thrombotic risk. Factors that falsely reduce platelet counts such as giant platelets and EDTAmediated platelet clumping interfere with obtaining accurate measurements. The objective of this project is to assess accuracy of the Sysmex XN-10 haematology analyser platelet flagging system for investigation of pseudothrombocytopenia and platelet abnormalities.

Methods: 170 samples flagged by the Sysmex XN-10 for “PLT Clumps?”, “PLT Abn Distribution”, “PLT Abn Scattergram”, or thrombocytopenia were examined microscopically for corresponding morphology. Correlation between platelet flagging and morphology was evaluated by calculating the positive predictive value, negative predictive value, sensitivity, specificity and false omission rate for the “PLT Clumps?” and “PLT Abn Distribution” flags. Results: 58 samples flagged for “PLT Clumps?” and the flag had a positive predictive value of 98.3%, negative predictive value of 82.1%, specificity of 98.9%, sensitivity of 74% and false omission rate of 17.9% for presence of clumping. 33 samples flagged for “PLT Abn Distribution” with a positive predictive value of 84.8%, negative predictive value of 40.1%, specificity of 91.7%, sensitivity of 25.5% and false omission rate of 59.9% for presence of platelet size variation or clumping. Conclusion: There exists a variable but strong relationship between Sysmex XN-10 platelet flags and corresponding morphology. In combination with appropriate reflex to blood film examination protocol, the flagging system functions effectively for identification of platelet abnormalities that may impact patient outcomes. Preliminary evaluation of biochemical analyte stability in refrigerated lithium heparin plasma samples for add-on testing Valerie Yeo1, Hank Ploeg2 and Kareen Igot2 University of Otago, Dunedin1 and Canterbury Laboratories, Christchurch

Health

Objectives: Clinicians often request add-on biochemical testing after initial laboratory analysis, requiring prolonged sample storage. This pilot study provides a preliminary evaluation of analyte stability in refrigerated lithium heparin plasma samples stored on separated red cells, aiming to provide storage duration guidelines for the Core Biochemistry Laboratory at Canterbury Health Laboratories. Methods: Fifty-one plasma samples collected in lithium heparin tubes were analysed over seven days using the Beckman Coulter AU5800 and Beckman Coulter DXI9000. Samples were stored at 2-8°C and re-analysed on Days 5 and 7. Data were analysed in Microsoft Excel using measurement uncertainty (MU) and compared against the Royal College of Pathologists of Australasia Quality Assurance Programs (RCPAQAP) analytical performance specifications (APS). Results: Potassium, phosphate, and glucose showed marked instability, with frequent APS and MU breaches by Day 5. Some analytes, including C-reactive protein (CRP) and thyroidstimulating hormone (TSH), had MU breaches without APS breaches, indicating analytical change without clinically significant instability. ALP and total bilirubin maintained short-term stability with no breaches by Day 5. Instability increased by Day 7. Conclusion: The preliminary analysis suggests that several plasma analytes may maintain acceptable short-term stability for add-on testing under current laboratory conditions prior to Day 7, while phosphate, potassium, and glucose indicate clinically significant instability by Day 5. The combination of APS and measurement uncertainty provided a clinically relevant framework for preliminary analyte stability testing in routine laboratory practice.

University of Otago accepts QMLT as a degree entry qualification. Following discussions with the University of Otago an agreement has been reached that the QMLT discipline-based qualifications will be accepted for entry into the first year Diploma of Science programme. This is a full-time on-campus course. Successful completion of the Diploma qualifies students to apply for entry into the second-year full-time BSc programmes. Discussions are ongoing for access to other degree programmes using this new option for the QMLT. Full information about the Diploma of Science can be found at www.otago.ac.nz and search for “Diploma of Science”. University enrolments for 2027 have recently opened. Thisis a significant outcome and recognition for the QMLT programme and qualification offered by the NZIMLS. New Zealand Journal of Medical Laboratory Science 173

UPDATES

The Pacific Way Warm Pacific greetings to you all from the PPTC

Strengthening Pacific Laboratory Capacity, PPTC Courses Blood Transfusion Science Course, 8 June – 3 July 2026 (4 weeks) This course aimed to provide students with a comprehensive theoretical component and a series of practical workshops in the medical laboratory field of Blood Transfusion Science. As a result, students acquired sufficient knowledge and skill to work confidently in their home laboratories to provide quality diagnostic test results to clinicians using the medical laboratory services for patient management and better health outcomes. This course was delivered over four weeks at the Pacific Pathology Training Centre, based at the Wellington Hospital campus in Wellington, New Zealand, to medical laboratory personnel working in hospital laboratories (or similar) within the with Pacific region. This course included knowledge and practical experience in the following: • Transfusion transmitted diseases (including HIV, Syphilis, Hepatitis B and C) • Donor selection criteria and collection • Blood processing, Blood Transfusion Practise, ABO, Rh and other blood group systems • Blood group genetics and basic immunology • Preparation of Coombs control cells, technical methods • Antibody screen and identification • Compatibility testing • Haemolytic Disease of the Newborn • Transfusion reaction investigations and haemovigilance • Blood Bank quality management processes • Equipment maintenance • Organisation of a Blood Bank including emergency planning, major trauma cases, backup planning • Appropriate use of blood components in transfusion medicine • Referral laboratory network • Practical sessions, focusing on correct technique and fundamental basic procedure • Tours of NZBS Crossmatch laboratory and the Blood Donor Centre. The content of this course ensured that the practical needs of all students attending were met and that the practical sessions provided focused on repeated basic methodology, so as techniques were mastered to levels of excellence. Course Convenor Mrs Telesia Apikotoa (BAppSc, MLabQAMgt TMLA, MNZIMLS, RNZMLS) Table 1. Four students attended the Blood Transfusion Course Student Name

Laboratory

Country

Feofaaki Latu

Vaiola Hospital Lab Northern Provinces Hospital Lab, Santo Princess Margaret Hospital Lab CWM Hospital Lab

Tonga

Pina Sam Salafuga Tailoa Shalni Narayan

Vanuatu Tuvalu Fiji

Figure 1. Blood Transfusion Science Course 2026, Students and Laboratory Staff, Students and Laboratory Manager (Dan Gyles), Palmerston North, New Zealand Blood Service, Blood Bank, Wellington Hospital Clinical Biochemistry Course, 13 July – 7 August 2026 (4 weeks) A comprehensive theoretical component was provided to students in the diagnostic medical field of Clinical Biochemistry. Laboratory tours in the Wellington region were also incorporated into this course, as were visits by various suppliers (Roche, BioRad) of Biochemistry equipment and reagents who were invited to present to the students, current technological advances in terms of diagnostic Clinical Biochemistry. This training course was delivered over four weeks at the Pacific Pathology Training Centre, based at the Wellington Hospital campus in Wellington, New Zealand, to medical laboratory personnel working in hospital laboratories (or similar) within the Pacific region. Course Content and Objectives • An overview of analytical principles utilised by most analytical systems for the analysis of biochemistry tests, e.g., spectrophotometry, immunoassay, blood gas analyser and other biochemistry analysers. • An overview of diabetes in terms of pathology, diagnosis and laboratory testing • An overview of cardiac tests in health and disease focusing on myocardial Infarction, diagnosis and laboratory analysis • An overview of protein analysis, liver function tests and specialist biochemical analysis • An overview of endocrine tests such as tumour markers, thyroid function tests, fertility hormones, other hormones (e.g. PTH), and associated abnormalities. • A molecular approach to biochemical pathology • Organisation and effective management of the biochemistry laboratory • Discussion on quality indicators such as TAT • Reporting of urgent and critical results • Presentations from various biochemistry suppliers will be incorporated • Quality Control; IQC, and External Quality Assessment in the biochemistry laboratory • An overview of reference intervals and Uncertainty of Measurement (UoM)

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•

• • • • • •

Discussion of quality systems, Standard Operating Procedures, stock control and its management, result processing, audit trails, reducing error and process improvement An overview of the validation process for introducing new methods and analysers A review of staff competency/ personal records, quality manual essentials, documentation framework, auditing and quality review An overview of Laboratory Information Systems An opportunity to tour Wellington City, Lower Hutt, Wellington and Kenepuru Hospital laboratories A description of blood collection sets and vacutainer tubes and needle systems An introduction to IATA regulations and packaging techniques for biological substances

Course Convenor Filipo Faiga (BSc, Dip MLSc, MNZIMLS, RNZMLS), Biochemistry Specialist Consultant and EQA Programme Manager Table 2. Seven students who attended the Clinical Biochemistry Course Student Name

Laboratory

Country

Escar Avock

VNH Lab

Vanuatu

Malti Raju

Nadi Sub-Divisional Hospital Lab

Fiji

Tokelau

Tokelau

Lupe Peau

Tokelau

Tokelau

Farisha Ali

Tavua Sub-Divisional Hospital Lab

Fiji

MTII Hospital Lab, Savaii

Samoa

Yap Department of Health Services

FSM

Elisaia Reima

Meleane Tapuala Sylvia Gap

• •

•

• • • • • • • • •

•

Sample processing methods used in microbiology and molecular laboratories. Including equipment, consumables and supplies used in for microscopy, culture and analysis. Theoretical and practical aspects of current methods used in the isolation and identification of bacterial pathogens isolated from various microbiology specimens. PCR molecular testing theory and methodology. Emergence of automation and equipment used in microbiology and molecular PCR laboratories including maintenance of instruments used, in-house media production for bacterial isolation, and choosing the right resources to carry out microbiological testing. Suppliers may also present on instruments currently available in the world market Antimicrobial Susceptibility Testing of micro-organism pathogens using the CLSI and EUCAST guidelines Monitoring of resistance patterns for multi drug resistant bacterial pathogens Discussions on emerging and re-emerging bacterial and viral organisms likely to cause infectious diseases. Identification and diagnosis of pathogenic parasites Health and Safety in a molecular and microbiology laboratory, including disposing of infectious waste and rubbish. Reporting and presenting microbiology results to clinicians and reporting requirements and guidelines for PCR testing. The role of the microbiology laboratory in the surveillance of nosocomial infections and identification of infections of public health importance. Serological and other rapid methods for the identification of bacterial and viral diseases including Hepatitis A, B, and C, HIV and other STIs. Standard Operating Procedures and written documents in microbiology and molecular laboratories. Applying Laboratory Quality Management to diagnostic processes, including management of quality controls, documentation, training records and stock management. Understanding UN/ IATA shipping requirements for infectious substances affecting humans and animals, biological substances category B.

Course Convenors Russell Cole, Dip MLSc, MNZIMLS, RNZMLS and Emmanuel Marshall, BMLSc, MNZIMLS, RNZMLS Table 3. Eight students who attended the Medical MicrobiologyMolecular Course. Student Name

Laboratory

Country

Andrea Havea

Vaiola Hospital Lab

Tonga

Fane Masaurua

CWM Hospital Lab

Fiji

Figure 2. Clinical Biochemistry Students and PPTC Staff, 2026.

Jekope Daunivala

Microbiology Course, 7 September – 2 October 2026 (4 weeks) A comprehensive theoretical component and a series of practical workshops were provided to students in the diagnostic field of Medical Microbiology including procedures and molecular testing platforms. The purpose of this training was designed to equip students with sufficient knowledge to be able to work confidently and competently in their home laboratories and be able to provide quality diagnostic test results to clinicians using the medical laboratory services for patient management and better health outcomes. This training course was delivered over four weeks at the Pacific Pathology Training Centre, based at the Wellington Hospital campus in Wellington, New Zealand to medical laboratory personnel working in hospital laboratories (or similar) in the South Pacific region.

Joy Karen

Course Content and Objectives • An understanding of major bacterial groups and identification characteristics of pathogens and commensal flora.

Kalita Homasi Elisaia Reima Lupe Peau Diana Leteism

Labasa Divisional Hospital Lab Kosrae State Hospital Lab Princess Margaret Hospital Lab Tokelau Tokelau Port Moresby General Hospital Lab

UPCOMING COURSES

Fiji FSM Tuvalu Tokelau Tokelau PNG

Haematology Course, 26 October – 4 December 2026 (6 weeks) The PPTC will be providing a second six-week course in Foundational Haematology, blood cell identification and blood film interpretation and this will be delivered at the PPTC in Wellington. As a part of this course and apart from all the teaching/learning resources given during the 6-week programme, each student will receive

New Zealand Journal of Medical Laboratory Science 175

three new textbooks to return home with: Cells, a practical guide (Barbara Bain), Microscopic Haematology- a practical guide for the laboratory (Gillian Rozenberg), and Cases in Microscopic Haematology (Gillian Rozenberg). Course Convenor Philip Wakem (NZCS, Dip MLSc, MMLSc (Otago, NZ), MNZIMLS, RNZMLS), Haematology Specialist Consultant and Chief Executive Officer. All PPTC Wellington-based courses are NZQA-accredited and designed to provide high-quality, reputable training that strengthens diagnostic pathology services across the Pacific region. For further information about this course or available courses for 2026, please contact the PPTC Education Programme Manager: Emmanuel Marshall, Pacific Pathology Training Centre Email: emmanuel.marshall@pptc.org.nz or pptc@pptc.org.nz Website www.pptc.org.nz, phone: +64 4 389 6294

IN-COUNTRY LABORATORY SUPPORT UPDATES: REGIONAL CAPACITY BUILDING

Fiji Filipo Faiga, the PPTC’s Biochemistry Specialist and EQA Programme Manager travelled to Colonial War Memorial Hospital laboratory in Suva on the 11-12 May 2026 to meet with laboratory management, including the laboratory managers, pathologists, quality officers, technical leads, and laboratory staff, to review progress, discuss strategic service plans, and consider updates to laboratory services. During this visit, Filipo reviewed the laboratory’s performance in the External Quality Assurance (EQA) programme and provided technical advisory support regarding corrective and preventive actions where necessary. He also provided technical advice and guidance on quality improvement initiatives, including test methodologies, workflow optimisation, result reporting practices, and documentation standards. Filipo also assisted in the development, review, and strengthening of core laboratory documentation, including the Quality Manual, Personnel Manual, Specimen Management Manual, Health and Safety Manual, and individual departmental Standard Operating Procedures (SOPs). Solomon Islands Russell Cole, PPTC’s Laboratory Quality Manager and Microbiology Specialist travelled to the provincial laboratories to meet with the laboratory managers and senior laboratory department heads, to give them technical support in the diagnostic laboratory disciplines and help progress Quality Management principles throughout the scope of laboratory services available. Munda and Gizo sites have not been visited by PPTC in the last 5 years. Munda was the first of the Solomon Island’s container laboratories to be commissioned and operational since the Covid outbreak in 2020. Samoa Both Philip Wakem and Emmanual Marshall visited Samoa to oversee the installation of a new Leica tissue processor funded and donated by the NZ Ministry of Foreign Affairs and Trade through its partnership with the Pacific Pathology Training Centre, New Zealand. This essential new item of equipment will improve the quality and consistency of tissue processing, reduce unacceptable turnaround times due to the inefficiency of the current machines, performance in terms of its downtime and ultimately provide a safer working environment for laboratory personnel. It has been unfortunate that Histology services at the National Hospital in Apia have experienced significant compromise in quality and efficiency since the breakdown of the previous machine in October 2025. This disruption not only slowed dramatically the normal workflow placing considerable pressure

on the clinical laboratory but also had a direct impact on patient diagnoses and clinical disease management, delaying critical results and care decisions. The official handover ceremony was officiated by the Deputy Director General for Clinical and Hospital Services, Ms Safaato’a Fereti, who delivered a welcoming speech and thankyou to the NZ Government and the PPTC for the contribution made. This was followed by the official handover address from the Chief Executive Officer of PPTC, Philip Wakem.

Figure 3. The Leica tissue processor, visiting Dignitaries, PPTC and laboratory staff, Samoa. Vanuatu Philip Wakem and Filipo Faiga visited Vanuatu to meet with Laboratory Management, the laboratory’s quality officer, technical leads, and staff to discuss laboratory progress, and service plans, as well as conduct a follow up with laboratory management to be updated on previous PPTC recommendations from audits and visitations. Filipo also provided technical quality improvement advice with regards to test methods, workflow issues, reporting of results and documentation particularly in Biochemistry. Phil assessed the Haematology diagnostic processes of the laboratory and addressed issues that were identified as compromising to both quality progression and efficiency of the services provided. NZIMLS Annual Science Meeting, Wellington, 19 - 21 August 2026 Russell Cole, the PPTC’s Laboratory Quality Programme Manager presented on New Zealand’s influence in “lower middle-income countries”. Russell indicated that New Zealand has had significant input into the strengthening and development of Pacific Island country laboratory services. The term LMIC has been used across African, Asian, and Pacific Island Nations to collectively describe the stages of development that their medical laboratory services have currently reached. Many are still perusing accreditation standards and are working to strengthen isolated provincial laboratory services. His presentation investigated the impact made by the PPTC’s EQA programme, regular Pacific in-country visits, LQMS auditing programmes and the PPTC’s educational diploma lecture series with current references to PNG, Timor Leste, Nauru and the Solomon Islands.

CAN YOU HELP?

If any New Zealand medical laboratories have items of diagnostic instrumentation that have been recently upgraded or continue to be stored in the laboratory that are still very operational but are surplus to requirements, the PPTC would be most grateful if such items could be donated through its Centre to Pacific Island laboratories where there is an exceptional need. Pacific laboratories have very restricted budgets and often cannot afford to replace troublesome instrumentation that continues to breakdown and which is often discontinued because it is so outdated. Please contact: Philip Wakem, Chief Executive Officer Pacific Pathology Training Centre, Wellington, New Zealand E-mail: pptc@pptc.org.nz or phil@pptc.org.nz Tel: 64-4-389 6294 or 027 2305483

New Zealand Journal of Medical Laboratory Science 176

JOURNAL QUESTIONNAIRE Read the articles carefully as most questions require more than one answer. Answers are to be submitted through the NZIMLS website. Make sure you supply your correct email address and membership number, it is recommended that you write your answers in a word document and then cut and paste your answers on the website. You are reminded that to claim valid CPD points for successfully completing the journal questionnaire you must submit an individual entry. It must not be part of a consultative or group process. In addition, members who have successfully completed the journal questionnaire cannot then claim additional CPD points for reading the articles from which the questions were derived. The site will remain open until Friday 12 February 2027. You must get a minimum of eight questions correct per questionnaire to obtain 5 CPD points. The Editor sets the questions and the CPD Co-Ordinator, Jillian Broadbent, marks the answers. Direct any queries to her at cpd@nzimls.org.nz.

NOVEMBER 2026 QUESTIONNAIRE

ANSWERS JULY 2026 QUESTIONNAIRE

1.

What is thalassaemia comprised of? And What does it result in? How does the accumulation of excess unmatched globin chains affect erythroid precursor cells?

1.

2.

Blood transfusions combined with iron chelation therapy remain fundamentally supportive in thalassaemia disease management, addressing the consequences of globin imbalance rather than its underlying cause. In contrast, emerging molecular therapies aim to directly target the pathophysiology of thalassaemia. Name the gene-editing therapy, its targeted regulatory element and potential. Name a widely-established curative therapy.

How long do plastic sample bags used in the laboratory reportedly take to decompose in landfill and compared to what Artificial intelligence (AI) is transforming every sector of society, what is cited by Yanfang Hu’s article as the most compelling argument for AI in New Zealand healthcare laboratories? What is the ‘human-in-the-loop’ model? Growing workforce crisis in laboratory professions. The human in the loop model refers to automating repetitive, high-volume and low complexity tasks, AI enables laboratories to reallocate highly trained professional towards complex diagnostic interpretation, quality assurance and multidisciplinary collaboration as a decision support tool rather than an autonomous decision maker.

3.

What is the human gut resistome? What factors impact it? What other factors have been linked to antimicrobial resistance (AMR) emergence and spread in the Gulf Cooperation Countries (GCC)?

2.

4.

What concerns are implicated by the discovery of antibioticresistant genes harboured within the gut of volunteers with no history of antibiotic exposure? Why is maintaining awareness of resistance patterns, including the breadth of organisms and resistance genes involved considered important?

List the three laboratory disciplines currently using specific AI applications and instrumentation in New Zealand. As organisations continue to expand their lab AI automation, what is recommended to remain focused on? Haematology, Microbiology and Histopathology. Focus must remain on ethical integration, patient safety, and the protection of indigenous data rights.

3.

When was the first artificial intelligence-based software approved by Food and Drug Administration (FDA) in USA? What does approved application employ and for which health condition? April 2018, Application employs using fundus image analysis to help diagnose diabetic retinopathy.

4.

What challenges remain for AI in genetics? What can health systems do to address these challenges and allow for responsible uptake? Interpretability, data bias, equity and regulation. Healthcare systems must invest in infrastructure, training personnel and governance frameworks.

5.

Urothelial carcinoma in situ (UCIS) is a malignant flat intraepithelial lesion of the urinary tract and a high-risk feature in non-muscle-invasive bladder cancer. What nuclear features are classically used to diagnose UCIS? what associated atypia seen with benign conditions make its diagnosis challenging?

5.

What adverse outcomes are associated with Chlamydia trachomatis infection in pregnant women and vertical transmission to the neonate? How does the presence of C. trachomatis infection increase the risk of HIV?

6.

Decorin (DCN) is classified within the small leucine‑rich proteoglycans (SLRPs) family. Where are these proteins found? What dual biological properties does DCN possess? And what structural role does it play?

7.

What diseases and medical conditions is Decorin clinically associated with?

8.

What did Younis and Hamodat observe for vitamin D, calcium concentration and PTH levels in patients with osteoarthritis compared to that of healthy individuals in their study?

9.

What is benign acute childhood myositis (BACM) also known as? Although rare, what is the syndrome most commonly associated with? Describe symptoms observed with the onset of muscle pain.

10. In clinical practise, what are the common causes of the isolated prolongation of the activated partial thromboplastin time (aPPT)? What is indicated when mixing tests do not correct for aPTT? What was found to be generating assay interference in the case reported by Raghvan et al?

Nuclear enlargement, hyperchromasis, membrane irregularity and pleomorphism. Benign conditions such as inflammation, lithiasis, viral cytopathic changes and therapy induced changes. 6.

In the study conducted by Elalfy and colleagues what percentage of inaccurate UCIS diagnoses in their cases were reported with H&E staining? Triple immunostaining panels (TIP) were shown to be important tools for distinguishing UCIS from its mimics, however what are the potential problems of this method?

New Zealand Journal of Medical Laboratory Science 177

7.

8.

30% inaccuracy of cases using H&E. Test is costly and requires a complex pattern to be translated (various immunostaining panels showed variability and discrepancy in the results) Malaria in pregnancy can leave the woman and the fetus extremely vulnerable, so how does the malaria parasite interfere? What clinical symptoms can develop with severe malaria in pregnant women? The parasite interferes with transmission of vital substances through the placenta, often resulting in stillbirth, spontaneous abortion, still birth or low birth weight. Persons with severe malaria can develop a range of clinical symptoms including bleeding, shock, kidney and liver failure, central nervous system dysfunction, and coma. Based on reported studies, what are the factors responsible for increased susceptibility to malaria infection in primigravida (first pregnancy) females compared to multigravida (multiple pregnancies) females? How is an absence of malaria seen in multigravida pregnant females explained? However, what reasons are given by opposing studies reporting that the 3639 age group may be more susceptible to severe malaria? Inhibition of type 1 cytokine responses and cell-mediated immune responses are markedly suppressed in the first as opposed to subsequent pregnancies. Multigravida females

are presumably less affected due to immunological memory activation. Absence of malaria in multigravida women may be due to continual exposure to malaria and strength of immune status; however, this age group may also have decreased immune function or underlying health conditions, delayed diagnosis and treatment due to atypical symptoms or comorbidities making them more susceptible. 9.

What did levels of HbA2 in EDTA blood samples show when blood samples were stored under routine refrigeration? And what does this finding confirm? What would be the advantages? Samples remained largely stable over a 10-day period. Confirms stability and extends the timeframe to 10 days. Would allow flexibility for batching and minor delays without compromising accuracy.

10. How long do plastic sample bags used in the laboratory reportedly take to decompose in landfill compared to what other plastics? 10-20 years compared with 1,000 years for other plastics and 450 years for plastic bottles.

Conference proceedings extracted from the Journal of the New Zealand Association of Bacteriologists, Vol. 1, October 1946, No. 3

New Zealand Journal of Medical Laboratory Science 178

Laboratory Christmas Quiz: Light relief from CPD questions The Christmas quiz was very successful again last year and the NZIMLS Council has agreed to sponsor another quiz this year. As with last year it is a General Knowledge Quiz and the NZIMLS Council has agreed to award a single prize of $300 towards a laboratory Christmas function for the most correct answers. In addition, if there are two successful teams the NZIMLS Council has agreed to make two awards of $200. While individuals can also complete the quiz, the prize is only to a laboratory team entry to encourage teamwork. There are no CPD points for this quiz, and it is not discipline related. The NZIMLS Council have not participated in the question setting or have knowledge of the answers. An answer sheet is available on the NZIMLS website, and the quiz answers can be submitted electronically as for the CPD questions. The answers will be published on the NZIMLS website after the quiz closes, and the winner (s) is announced. Closing date for all answers is 7 December 2026, no late entries will be considered. Send all completed quiz answers with a Laboratory name, Laboratory Team name and Contact person to: mike.legge@nzimls.org.nz with subject heading: Christmas Quiz. Please indicate if the entry is a team or individual entry. No correspondence will be entered into relating to the quiz. 1.

Which fruit is used to make the syrup vincotto? i. Orange ii. Raspberry iii. Grapes iv. Strawberry

2.

A person who studies shells is known as? i. Calcologist ii. Conchologist iii. Chonchlosist iv. Calchonosist

3.

Which Renaissance artist created the Pieta? i. Leonardo da Vinci ii. Titan iii. Raphel iv. Michelangelo

4.

5.

6.

7.

8.

Which pastry is a hybrid of a croissant and a doughnut? i. Crookie ii. Cronut iii. Craffle iv. Cruftin In Greek mythology who was Zeus’s father? i. Kronus ii. Dionysus iii. Hermes iv. Poseidon The insecticide pyrethrum was derived from which of the following? i. Marigolds ii. Geraniums iii. Chrysanthemums iv. Nasturtiums

originally

The Titanic was built in which city shipyard? i. Belfast ii. Glasgow iii. Liverpool iv. Newcastle Which meat is used to make lardons? i. Lamb ii. Beef iii. Pork iv. Goat

9.

The ruins of the ancient Greek city Pergamon are found in which modern country? i. Crete ii. Lebanon iii. Turkey iv. Cyprus

of

10. The original traditional eyeliner was made from Kohl and comes from which element? i. Lead ii. Zinc iii. Tin iv. Antimony 11. The ancient Greek athlete, Pheidippes, is believed to be the originator of which modern Olympic sport? i. Pentathlon ii. Wrestling iii. Marathon iv. Boxing 12. Which red wine grape’s name is thought to mean “little blackbird” in French? i. Merlot ii. Syrah iii. Shiraz iv. Pinot noir 13. Which of the following did the Aztecs refer to as “love apple”? i. Guava ii. Tomato iii. Avocado iv. Papaya 14. Which of the following countries has three capital cities? i. Bolivia ii. Malaysia iii. Netherlands iv. South Africa 15. The Caprera Canyon is located in. i. Mediterranean Sea ii. Atlantic Ocean iii. Indian Ocean iv. Caribbean Sea 16. The 1995 film “Clueless” was based on which Jane Austin novel? i. Lady Susan ii. Persuasion iii. Mansfield Park iv. Emma

New Zealand Journal of Medical Laboratory Science 179

17. In the 13th century England, it was compulsory for all yeomen to learn what? i. Sword Fighting ii. Animal husbandry iii. Archery iv. Blacksmithing

27. The word “golf” is believed to come from which language? i. Basque ii. Norse iii. Breton iv. Celtic

18. A poltroon is what? i. Warrior ii. Jester iii. Coward iv. Ghost

28. New Amsterdam became which American city? i. Atlantic City ii. Detroit iii. Boston iv. New York

19. Which tribe brought about the end of the Western Roman Empire in 476 CE? i. Visigoths ii. Huns iii. Vandals iv. Hercule

29. The “carat” was originally used as a unit of measure for what? i. Grain ii. Gold iii. Gems iv. Silver

20. Which of the following animals’ drinks salt water to hydrate? i. Whales ii. Penguins iii. Turtles iv. Dolphins

30. Of the six major animal groups, which one still contains dinosaur heritage? i. Reptiles ii. Fish iii. Birds iv. Amphibians

21. The very first video to appear on YouTube was filmed where? i. Video shop ii. Zoo iii. Shopping Mall iv. Street 22. The traditional name for a group of cheetahs is what? i. Chitty ii. Clamour iii. Charge iv. Coalition

31. Where is the Hermitage museum? i. Houston ii. St Petersburg iii. Paris iv. Amsterdam

23. Where would you encounter gyrification? i. Athletics ii. Brain iii. Sailing iv. Geology 24. Which of the following words is the most recent? i. Ditzy ii. Doozy iii. Neft iv. Spiffy 25. Which of the following countries claims territories in both the Arctic and Antarctica? i. Canada ii. Norway iii. China iv. Russia 26. In the nursery rhyme what did the old woman who lived in a shoe feed her kids? i. Gruel ii. Bread crusts iii. Meat iv. Broth

32. Which is the oldest of the following words? i. Coupon ii. Logo iii. Ticket iv. Label 33. Which is the strongest piece on the Chess board? i. Pawn ii. Queen iii. Knight iv. King 34. What is the national fruit of both India and Pakistan? i. Pineapple ii. Guava iii. Mango iv. Papaya 35. What is a tolt? i. Tin object ii. Horse gait iii. A medieval dance iv. An animal den 36. Which famous person created the first Paddington Bear stuffed toy? i. Bill Bailey ii. Jeremy Clarkson iii. Stephen Fry iv. John Cleese

37. Which religion was responsible for creating the dog collar (Clerical Collar)? i. Roman Catholic ii. Methodist iii. Anglican iv. Presbyterian

47. The Milankovitch cycle is which of the following? i. A minor metabolic pathway ii. Timing of the moons orbit around the Earth iii. A biological rhythm linked to seasons iv. Changes in the Earth’s orbit around the sun

38. What is a peloton? i. Group of pelicans ii. Result of a mud wrestling competition iii. Group of cyclists iv. Fast Basque ball game

48. “Cool Runnings” is a true comedy film about a bob Sleigh team from which country? i. Jamaica ii. Nepal iii. Barbados iv. Bhutan

39. The Malabar Coast of India is associated with which spice? i. All spice ii. Garam masala iii. Coriander iv. Black pepper

49. The middle of the 4% of New Zealand above land is where? i. Blenheim ii. Nelson iii. Wellington iv. Greytown

40. Harold van Braunhut is famous for developing what? i. Sea monkeys ii. Dishwashers iii. Sunglasses iv. Rockets

50. Tea bags are believed to have been first used in which city? i. Boston ii. Calcutta iii. London iv. Delhi

41. Kakorrhaphiophobic is the fear of which of the following? i. Failure ii. Money iii. Fog iv. Marriage

51. Which was the only country to put a dog into space? i. China ii. America iii. Russia iv. India

42. In Snow White and the Seven Dwarfs what did the dwarfs mine? i. Diamonds and emeralds ii. Rubies and sapphires iii. Emeralds and sapphires iv. Diamonds and rubies

52. In which country was the four-wheel drive tractor invented? i. Australia ii. America iii. New Zealand iv. Britain

43. How long is the Ninety Mile Beach? i. 52 km ii. 144 km iii. 36 km iv. 88 km

53. When Napolean Bonaparte went to live in France, which was the language he spoke? i. Bretan ii. Corsican iii. Sicilian iv. Alsatian

44. If something is hebdomadal it occurs how frequently? i. Monthly ii. Weekly iii. Daily iv. Fortnightly 45. Where is Gozo located? i. Portugal ii. Indonesia iii. Malta iv. Central Africa 46. Which word is from Greenlandic? i. Anorak ii. Macintosh iii. Cagoule iv. Parka

54. The term “Brent Crude” originally referred to oil from where? i. Texas ii. North Sea iii. Venezuela iv. Kuwait 55. Which religion has the four noble truths? i. Christianity ii. Islam iii. Sikhism iv. Buddhism 56. Which planet is named after the Roman god of agriculture? i. Saturn ii. Jupiter iii. Earth iv. Neptune

57. Which country has never been beaten by Brazil in a soccer match? i. Denmark ii. Scotland iii. Norway iv. Ireland 58. In which year were the first Oscars awarded? i. 1946 ii. 1929 iii. 1934 iv. 1950

59. When was the first New Zealand fish and chip shop established? i. 1860 ii. 1903 iii. 1894 iv. 1910 60. Elizabeth Alexander is considered to be the world’s first female what? i. Radio astronomer ii. Economist iii. Neurosurgeon iv. Forensic scientist

Good Luck! The NZIMLS Executive Office will close for 2026 on Friday 18 December, reopening Monday 11 January 2027. We would like to thank our members for their continued support of their professional organisation, and wish you and your families a very safe and happy Christmas and a wonderful New Year.

Merry Christmas from (L to R) Sharon, Cathy, Fran and Jillian

New Zealand Journal of Medical Laboratory Science 182