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Methods in Molecular Biology 1247

Mónica V. Cunha João Inácio Editors

Veterinary Infection Biology: Molecular Diagnostics and High-Throughput Strategies

METHODS IN MOLECULAR BIOLOGY

Series Editor John M. Walker School of Life Sciences University of Hertfordshire Hatfield, Hertfordshire, AL10 9AB, UK

For further volumes: http://www.springer.com/series/7651

Veterinary Infection Biology: Molecular Diagnostics and High-Throughput Strategies Edited by

Mónica V. Cunha Instituto Nacional de Investigação Agrária e Veterinária, Lisbon, Portugal Centro de Biologia Ambiental, Faculdade de Ciências, Universidade de Lisbon, Lisbon, Portugal

João Inácio Instituto Nacional de Investigação Agrária e Veterinária, Lisbon, Portugal School of Pharmacy and Biomolecular Sciences, University of Brighton, United Kingdom

Editors Mónica V. Cunha Instituto Nacional de Investigação Agrária e Veterinária Lisbon, Portugal

João Inácio Instituto Nacional de Investigação Agrária e Veterinária Lisbon, Portugal

Centro de Biologia Ambiental Faculdade de Ciências, Universidade de Lisbon Lisbon, Portugal

School of Pharmacy and Biomolecular Sciences University of Brighton United Kingdom

ISSN 1064-3745 ISSN 1940-6029 (electronic) ISBN 978-1-4939-2003-7 ISBN 978-1-4939-2004-4 (eBook) DOI 10.1007/978-1-4939-2004-4 Springer New York Heidelberg Dordrecht London Library of Congress Control Number: 2014953901 © Springer Science+Business Media New York 2015 This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. Exempted from this legal reservation are brief excerpts in connection with reviews or scholarly analysis or material supplied specifically for the purpose of being entered and executed on a computer system, for exclusive use by the purchaser of the work.

Duplication of this publication or parts thereof is permitted only under the provisions of the Copyright Law of the Publisher’s location, in its current version, and permission for use must always be obtained from Springer. Permissions for use may be obtained through RightsLink at the Copyright Clearance Center. Violations are liable to prosecution under the respective Copyright Law. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use.

While the advice and information in this book are believed to be true and accurate at the date of publication, neither the authors nor the editors nor the publisher can accept any legal responsibility for any errors or omissions that may be made. The publisher makes no warranty, express or implied, with respect to the material contained herein.

Printed on acid-free paper Humana Press is a brand of Springer Springer is part of Springer Science+Business Media (www.springer.com)

Foreword Infectious diseases of domestic and wild animals have a tremendous impact on the economy and public health worldwide. Sixty percent of the pathogens that cause diseases in humans have their source in animals (whether domestic or wild), as do 75 % of emerging human diseases and 80 % of the pathogens that potentially could be used in bioterrorism. Animal diseases that are transmissible to humans (collectively known as zoonoses), such as avian influenza, rabies, bovine tuberculosis, and brucellosis, represent a very relevant public health threat worldwide, being imperative to prevent and/or control them. Likewise, pathogens that are not zoonotic but have a negative impact on livestock production, and the associated financial income, should not be overlooked. Some estimates suggest that world production of food animals is reduced by more than 20 % due to disease, which may be particularly important in developing countries where the unavailability of animal protein is socially impacting and inevitably exacerbates infectious diseases risk.

The World Organization for Animal Health (OIE) was created in 1924 with the aim of controlling the international spread of infectious animal diseases, but currently, over and above this original mission, the new mandate is “to improve animal health worldwide.” The OIE builds its expertise on a global network of Reference Laboratories and Collaborating Centers that play an essential role in prevention, detection, surveillance, and control of animal diseases worldwide. Reference Laboratories serve as centers of scientific and technical expertise for official listed diseases and have particular responsibility for carrying out confirmatory diagnostic tests for these diseases. Collaborating Centers are centers of expertise in key competences related to the management of a particular animal health issue, such as epidemiology, diagnosis, risk analysis, welfare, or veterinary training. This global network of excellence enables OIE to remain at the forefront of world veterinary scientific expertise and successfully carry out its activities, providing authoritative scientific opinions and advice on key topics, such as animal health and welfare and diagnostic techniques. Moreover, the OIE is involved in the preparation of standards on diagnostic assays and their official validation which are published in the OIE Terrestrial Manual and Aquatic Manual and updated annually by the World Assembly of the Delegates of the OIE (available from www.oie.int).

The achievements of biotechnology are significantly contributing to the development of novel, rapid, and powerful diagnostic assays for animal diseases, namely based on molecular approaches, such as standard and real-time polymerase chain reaction and isothermal nucleic acid amplification methods. The recent advent of high-throughput sequencing technologies is also boosting the prompt detection of all microbes in a sample, uncovering novel pathogens and significantly advancing veterinary diagnostic microbiology. Furthermore, the analysis of single nucleotide and large sequence polymorphisms in the genome of pathogens provides novel information on their traits, such as virulence and antimicrobial resistance determinants, and supports epidemiological studies. Outstandingly, biotechnology is enabling the development of diagnostic kits that can ultimately be used in point-ofdecision settings, away from centralized laboratories to assist in local decision-making, for instance during an outbreak. OIE is aware of the opportunities and challenges that these novel molecular-based approaches bring to veterinary diagnosis and several OIE Reference

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Foreword

Laboratories and Collaborating Centers are also dedicated to Research and Development activities on these fields. In this context, the present Springer’s book on the use of molecular biology strategies in veterinary laboratory practice showcases the versatility of the veterinarian profession in meeting the challenges posed by the continuous advance of science, namely in the biotechnology field. The editors have succeeded in bringing together an impressive group of renowned collaborating authors, primarily veterinary doctors and researchers of veterinary science, from institutions with established and prized expertise in the fields of molecular biology and veterinary diagnostics, including several OIE Reference Laboratories and Collaborating Centers. This book provides therefore an excellent and authoritative overview of molecular biology strategies for pathogen detection and characterization, along with the most recent technological innovations and their potential to reconstruct transmission chains and to disclose pathogen biology, evolution, and interaction with the host. Beyond the techniques, the book also discusses the integration of these new molecular approaches in the framework of veterinary practice and animal health management. In recognition of the challenge faced by the public and private components of Veterinary Services and all veterinarians to improve the animal health status worldwide, this book may aid veterinary laboratories in setting up molecular diagnosis valences and is a valuable resource to veterinary doctors and laboratory technicians, researchers, and students of veterinary medicine and science interested in knowing more about these challenging, but promising molecular strategies to unravel animal infectious diseases.

Director General World Organization for Animal Health (OIE)

Bernard Vallat

Preface Infectious diseases of animals represent a significant economic burden to economies worldwide and may raise relevant Public Health concerns. Molecular diagnostics has emerged as the fastest growing and trending segment of the in vitro diagnostics industry and seems also promising in the veterinary sector, providing novel and powerful tools that enable the fast and accurate diagnosis of animal diseases.

In this book, we provide an overview of molecular biology tools for pathogen detection and characterization, along with potential applications to disclose pathogen biology and interaction with the host. In Part I, biobanking, biosafety and good laboratory practices, biological specimen collection and processing, quality assurance and control, and validation of molecular diagnostic assays for veterinary use are concisely introduced as a basis for the molecular strategies enclosed in protocol sections. The significance and integration of molecular biology methods in the framework of veterinary practice are discussed, along with a SWOT analysis presenting our view on the advantages, limitations, opportunities and weaknesses, or challenges, of these approaches. The rationale is on the techniques and applications rather than the pathogen or disease models. Part II is dedicated to the molecular detection and identification of animal pathogens using a wide range of established techniques and covering, also, emerging diagnosis approaches. A brief overview of the advances on nanoscience and microfluidics is provided, including a discussion on how the recent developments in these research fields may be translated into field deployable biosensors.

Part III addresses genotyping tools for assessing the genetic landscape and epidemiology of pathogens. Part IV introduces integrative omics and high-throughput technologies as powerful research tools, yielding massive amounts of information that may ultimately improve the control and management of infectious diseases. We strived to span a broad range of molecular approaches, from simple and affordable to emerging and sophisticated, which we anticipate will be useful for veterinary doctors and laboratory technicians, researchers, and students of veterinary medicine and science.

Lisbon, Portugal

Mónica V. Cunha João Inácio

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Acknowledgements The editors were supported by Fundacão para a Ciência e Tecnologia (FCT, Portugal) through projects PTDC/CVT/111634/2009 (JI) and PTDC/CVT/117794/2010 and in the framework of Projecto 3599—Promover a Produção Científica e Desenvolvimento Tecnológico e a Constituição de Redes Temáticas (MVC). Alice Santos (INIAV IP, Lisbon, Portugal) is acknowledged for the cover image artwork. The authors of Chap. 17 are acknowledged for providing the fluorescence microscopy image used in the artwork.

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Contents Foreword . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Contributors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

PART I

v vii ix xv

INTRODUCING NUCLEIC ACID TESTING IN THE VETERINARY LABORATORY

1 Overview and Challenges of Molecular Technologies in the Veterinary Microbiology Laboratory . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Mónica V. Cunha and João Inácio 2 Significance and Integration of Molecular Diagnostics in the Framework of Veterinary Practice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Alicia Aranaz 3 Biosafety Principles and Practices for the Veterinary Diagnostic Laboratory . . .

Joseph Kozlovac and Beverly Schmitt 4 Veterinary Biobank Facility: Development and Management for Diagnostic and Research Purposes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Tina Lombardo, Silvia Dotti, Riccardo Villa, Stefano Cinotti, and Maura Ferrari 5 Biological Specimen Collection and Processing for Molecular Analysis. . . . . . .

Susan Sanchez and Holly Sellers 6 Validation of Molecular Diagnostic Assays and Quality Assurance and Control in the Veterinary Laboratory . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Nick Saunders and Ian Sharp

PART II

3

19 31

43

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MOLECULAR DETECTION AND IDENTIFICATION OF ANIMAL PATHOGENS IN LABORATORIAL SETTINGS

7 Molecular Approaches to Recognize Relevant and Emerging Infectious Diseases in Animals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Fredrik Granberg, Oskar E. Karlsson, Mikael Leijon, Lihong Liu, and Sándor Belák 8 Real-Time Reverse Transcriptase PCR for the Detection of Bluetongue Virus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Carrie Batten, Lorraine Frost, and Chris Oura 9 Nested and Multiplex Real-Time PCR Using Dual-Labeled Probes: Detecting and Discriminating Mycobacterium tuberculosis Complex Members in Cultures and Animal Tissues . . . . . . . . . . . . . . . . . . . . .

Pedro Costa, Isabel Couto, Miguel Viveiros, and João Inácio

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10 A Real-Time PCR Assay for the Diagnosis of Gastrointestinal Nematode Infections of Small Ruminants. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Florian Roeber, Aaron R. Jex, and Robin B. Gasser 11 Improved Detection of Mycobacterium bovis in Bovine Tissues Using Immunomagnetic Separation Approaches . . . . . . . . . . . . . . . . . . . . . . .

Irene R. Grant and Linda D. Stewart 12 Detection of Fish Pathogens by Loop-Mediated Isothermal Amplification (LAMP) Technique . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Hatem Soliman, Mona Saleh, and Mansour El-Matbouli 13 Direct Detection of Theileria annulata in Bovine Blood Samples Using Standard and Isothermal DNA Amplification Approaches . . . . . . . . . . . . . . . .

Jacinto Gomes and João Inácio 14 Reverse Line Blot Hybridization with Species-Specific Oligonucleotide Probes: Application to Piroplasm Detection. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Ana Hurtado 15 DNA Microarray-Based Detection of Multiple Pathogens: Mycoplasma spp. and Chlamydia spp.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Christiane Schnee and Konrad Sachse 16 In Situ Hybridization with Labeled Probes: Assessment of African Swine Fever Virus in Formalin-Fixed Paraffin-Embedded Tissues . . . . . . . . . . . . . . . . . . . .

Maria Ballester and Fernando Rodríguez 17 Fluorescence In Situ Hybridization for the Tissue Detection of Bacterial Pathogens Associated with Porcine Infections . . . . . . . . . . . . . . . . . . . . . . . . .

Henrik Elvang Jensen, Louise Kruse Jensen, Kristiane Barington, Susanne Elisabeth Pors, Thomas Bjarnsholt, and Mette Boye 18 Identification of Animal Pasteurellaceae by MALDI-TOF Mass Spectrometry .

Joachim Frey and Peter Kuhnert 19 Gold Nanoparticles as a Potential Tool for Diagnosis of Fish Diseases . . . . . . .

Mona Saleh, Hatem Soliman, and Mansour El-Matbouli 20 Nucleic-Acid Testing, New Platforms and Nanotechnology for Point-of-Decision Diagnosis of Animal Pathogens . . . . . . . . . . . . . . . . . . .

Fernando Teles and Luís Fonseca

PART III

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MOLECULAR PROFILING OF VETERINARY RELEVANT MICROBIAL PATHOGENS

21 Molecular Typing Tools: From Pattern Recognition to Genome-Based Algorithms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Konrad Sachse and Petra Moebius 22 Characterization of Campylobacter jejuni and Campylobacter coli Genotypes in Poultry Flocks by Restriction Fragment Length Polymorphism (RFLP) Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Ana Cláudia Carreira and Mónica V. Cunha 23 Pulsed-Field Gel Electrophoresis (PFGE): Application in Population Structure Studies of Bovine Mastitis-Causing Streptococci. . . . . . . . . . . . . . . .

Ilda Santos-Sanches, Lélia Chambel, and Rogério Tenreiro

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Contents

24 Multiple-Locus Variable-Number Tandem Repeat (VNTR) Analysis (MLVA) Using Multiplex PCR and Multicolor Capillary Electrophoresis: Application to the Genotyping of Brucella Species . . . . . . . . .

Giuliano Garofolo 25 Multilocus Sequence Typing (MLST): Markers for the Traceability of Pathogenic Leptospira Strains . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Ahmed Ahmed, Ana S. Ferreira, and Rudy A. Hartskeerl 26 Single-Nucleotide Polymorphism Discrimination Using High-Resolution Melting Analysis for the Genotyping of Bacillus anthracis . . . . . . . . . . . . . . . .

Sylviane Derzelle 27 Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) Analysis of Members of the Mycobacterium tuberculosis Complex . . . . . . . . . . .

Ana Botelho, Ana Canto, Célia Leão, and Mónica V. Cunha 28 Rapid Microarray-Based Genotyping of Chlamydia spp. Strains from Clinical Tissue Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Konrad Sachse and Anke Ruettger 29 Multiplexed Genotyping of Bacillus anthracis by Luminex xMap Suspension Array. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Simon Thierry and Sylviane Derzelle

PART IV

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INTEGRATIVE OMICS AND HIGH-THROUGHPUT PLATFORMS TO UNRAVEL THE BIOLOGY OF PATHOGENS AND THEIR INTERACTION WITH THE HOST

30 Next-Generation Sequencing in Veterinary Medicine: How Can the Massive Amount of Information Arising from High-Throughput Technologies Improve Diagnosis, Control, and Management of Infectious Diseases? . . . . . .

Steven Van Borm, Sándor Belák, Graham Freimanis, Alice Fusaro, Fredrik Granberg, Dirk Höper, Donald P. King, Isabella Monne, Richard Orton, and Toon Rosseel 31 Impact of Next-Generation Technologies on Exploring Socioeconomically Important Parasites and Developing New Interventions . . . . . . . . . . . . . . . . .

Cinzia Cantacessi, Andreas Hofmann, Bronwyn E. Campbell, and Robin B. Gasser 32 Functional Genomics of Tick Vectors Challenged with the Cattle Parasite Babesia bigemina . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Ana Domingos, Sandra Antunes, Margarita Villar, and José de la Fuente 33 Metagenomic Approaches to Disclose Disease-Associated Pathogens: Detection of Viral Pathogens in Honeybees. . . . . . . . . . . . . . . . . . . . . . . . . . .

Fredrik Granberg, Oskar E. Karlsson, and Sándor Belák 34 Proteomics Characterization of Tick-Host-Pathogen Interactions . . . . . . . . . .

Marina Popara, Margarita Villar, and José de la Fuente Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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491 513 529

Contributors AHMED AHMED • WHO/FAO/OIE and National Collaborating Centre for Reference and Research on Leptospirosis, Department of Biomedical Research, Royal Tropical Institute (KIT), Amsterdam, The Netherlands SANDRA ANTUNES • Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Lisbon, Portugal ALICIA ARANAZ • Departamento de Sanidad Animal, Facultad de Veterinaria, Universidad Complutense de Madrid, Madrid, Spain MARIA BALLESTER • Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, Bellaterra (Cerdanyola del Vallès), Spain; Centre de Recerca en Agrigenòmica (CRAG), Consorci CSIC-IRTAUAB-UB, Bellaterra, Spain; Departament de Ciència Animal i dels Aliments, Facultat de Veterinària, Universitat Autònoma de Barcelona, Bellaterra, Spain KRISTIANE BARINGTON • Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark CARRIE BATTEN • The Pirbright Institute, Surrey, UK SÁNDOR BELÁK • OIE Collaborating Centre for the Biotechnology-based Diagnosis of Infectious Diseases in Veterinary Medicine, Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden; Department of Biomedical Sciences and Veterinary Public Health (BVF), Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden; The National Veterinary Institute (SVA), Uppsala, Sweden THOMAS BJARNSHOLT • Department of International Health, Immunology and Microbiology, Costerton Biofilm Center, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark; Department of Clinical Microbiology, Copenhagen University Hospital, Copenhagen, Denmark STEVEN VAN BORM • Veterinary and Agrochemical Research Center, Brussels, Belgium ANA BOTELHO • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal METTE BOYE • National Veterinary Institute, Technical University of Denmark, Kongens Lyngby, Denmark BRONWYN E. CAMPBELL • Department of Veterinary Science, The University of Melbourne, Parkville, VIC, Australia CINZIA CANTACESSI • Department of Veterinary Science, The University of Melbourne, Parkville, VIC, Australia ANA CANTO • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal ANA CLÁUDIA CARREIRA • iMed.UL—Research Institute for Medicines and Pharmaceutical Sciences, Nanomedicine & Drug Delivery Systems Group, Faculdade de Farmácia, Universidade de Lisboa, Lisbon, Portugal; Instituto Nacional de Investigação Agrária e Veterinária, I.P. (INIAV, I.P.), Lisbon, Portugal

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Contributors

LÉLIA CHAMBEL • Centro de Biodiversidade, Genómica Integrativa e Funcional, Faculdade de Ciências, Universidade de Lisboa, Lisbon, Portugal STEFANO CINOTTI • Cell Culture CentreA, Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (IZSLER), Brescia, Italy PEDRO COSTA • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal; Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Lisbon, Portugal ISABEL COUTO • Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Lisbon, Portugal MÓNICA V. CUNHA • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal; Centro de Biologia Ambiental, Faculdade de Ciências, Universidade de Lisbon, Lisbon, Portugal SYLVIANE DERZELLE • Bacterial Zoonosis Unit, Animal Health Laboratory, University Paris-Est, Anses, Maisons-Alfort, France ANA DOMINGOS • Centro de Malária e Outras Doenças Tropicais; Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Lisbon, Portugal SILVIA DOTTI • Cell Culture Centre, Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (IZSLER), Brescia, Italy MANSOUR EL-MATBOULI • Clinical Division of Fish Medicine, University of Veterinary Medicine, Vienna, Austria MAURA FERRARI • Cell Culture Centre, Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (IZSLER), Brescia, Italy ANA S. FERREIRA • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal LUÍS FONSECA • Department of Bioengineering, Instituto Superior Técnico (IST), Institute for Biotechnology and Bioengineering (IBB), Centre for Biological and Chemical Engineering, Lisbon, Portugal GRAHAM FREIMANIS • The Pirbright Institute, Pirbright, UK JOACHIM FREY • Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Bern, Bern, Switzerland LORRAINE FROST • The Pirbright Institute, Surrey, UK JOSÉ DE LA FUENTE • SaBio, Instituto de Investigación en Recursos Cinegéticos IREC, UCLM-JCCM, Ciudad Real, Spain; Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK, USA ALICE FUSARO • Istituto Zooprofilattico Sperimentale delle Venezie, Padova, Italy GIULIANO GAROFOLO • Istituto Zooprofilattico Sperimentale dell’Abruzzo e del Molise “G. Caporale”—National and OIE Reference Laboratory for Brucellosis, Teramo, Italy ROBIN B. GASSER • Department of Veterinary and Agricultural Sciences, The University of Melbourne, Parkville, VIC, Australia; Institute of Parasitology and Tropical Veterinary Medicine, Berlin, Germany JACINTO GOMES • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal FREDRIK GRANBERG • OIE Collaborating Centre for the Biotechnology-based Diagnosis of Infectious Diseases in Veterinary Medicine, Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden; Department of Biomedical Sciences and Veterinary

Contributors

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Public Health (BVF), Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden IRENE R. GRANT • Institute for Global Food Security, School of Biological Sciences, Queen’s University Belfast, Northern Ireland, UK RUDY A. HARTSKEERL • WHO/FAO/OIE and National Collaborating Centre for Reference and Research on Leptospirosis, Department of Biomedical Research, Royal Tropical Institute (KIT), Amsterdam, The Netherlands ANDREAS HOFMANN • Structural Chemistry Program, Eskitis Institute, Griffith University, Brisbane, QLD, Australia DIRK HÖPER • Friedrich Loeffler Institut—Federal Research Institute for Animal Health, Riems (Greifswald), Germany ANA HURTADO • Department of Animal Health, NEIKER—Instituto Vasco de Investigación y Desarrollo Agrario, Bizkaia, Spain JOÃO INÁCIO • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal; School of Pharmacy and Biomolecular Sciences, University of Brighton, Brighton, UK HENRIK ELVANG JENSEN • Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark LOUISE KRUSE JENSEN • Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark AARON R. JEX • Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Parkville, VIC, Australia OSKAR E. KARLSSON • OIE Collaborating Centre for the Biotechnology-based Diagnosis of Infectious Diseases in Veterinary Medicine, Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden; Department of Biomedical Sciences and Veterinary Public Health (BVF), Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden DONALD P. KING • The Pirbright Institute, Pirbright, UK JOSEPH KOZLOVAC • United States Department of Agriculture (USDA), Agricultural Research Service (ARS), Office of National Programs, Beltsville, MD, USA PETER KUHNERT • Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Bern, Bern, Switzerland CÉLIA LEÃO • Instituto Nacional de Investigação Agrária e Veterinária IP, Lisbon, Portugal MIKAEL LEIJON • OIE Collaborating Centre for the Biotechnology-based Diagnosis of Infectious Diseases in Veterinary Medicine, Uppsala, Sweden; The National Veterinary Institute (SVA), Uppsala, Sweden LIHONG LIU • OIE Collaborating Centre for the Biotechnology-based Diagnosis of Infectious Diseases in Veterinary Medicine, Uppsala, Sweden; The National Veterinary Institute (SVA), Uppsala, Sweden TINA LOMBARDO • Cell Culture Centre, Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (IZSLER), Brescia, Italy PETRA MOEBIUS • Institute of Molecular Pathogenesis at Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Jena, Germany ISABELLA MONNE • Istituto Zooprofilattico Sperimentale delle Venezie, Padova, Italy RICHARD ORTON • Institute of Biodiversity, Animal Health and Comparative Medicine, University of Glasgow, Glasgow, UK

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Contributors

CHRIS OURA • School of Veterinary Medicine, University of the West Indies, St. Augustine, Trinidad and Tobago MARINA POPARA • SaBio, Instituto de Investigación en Recursos Cinegéticos IREC, UCLM-JCCM, Ciudad Real, Spain SUSANNE ELISABETH PORS • Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark FERNANDO RODRIGUEZ • Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, Bellaterra (Cerdanyola del Vallès), Spain FLORIAN ROEBER • Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Parkville, VIC, Australia; AusDiagnostics Pty. Ltd., Beaconsfield, NSW, Australia TOON ROSSEEL • Veterinary and Agrochemical Research Center, Brussels, Belgium ANKE RUETTGER • Institute of Molecular Pathogenesis at Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Jena, Germany KONRAD SACHSE • Institute of Molecular Pathogenesis at Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Jena, Germany MONA SALEH • Clinical Division of Fish Medicine, University of Veterinary Medicine, Vienna, Austria SUSAN SANCHEZ • Athens Veterinary Diagnostic Laboratory, College of Veterinary Medicine, The University of Georgia, Athens, GA, USA ILDA SANTOS-SANCHES • Centro de Recursos Microbiológicos, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Lisbon, Portugal NICK SAUNDERS • Public Health England (PHE), Microbiology Services Colindale, London, UK BEVERLY SCHMITT • United States Department of Agriculture (USDA), Animal and Plant Health Inspection Service, National Veterinary Services Laboratories, Ames, Iowa, USA CHRISTIANE SCHNEE • Institute of Molecular Pathogenesis at Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Jena, Germany HOLLY SELLERS • Poultry Diagnostic and Research Center, College of Veterinary Medicine, The University of Georgia, Athens, GA, USA IAN SHARP • Public Health England (PHE), Microbiology Services Colindale, London, UK HATEM SOLIMAN • Clinical Division of Fish Medicine, University of Veterinary Medicine, Vienna, Austria; Fish Medicine and Managements, Department of Animal Medicine, Faculty of Veterinary Medicine, Assiut University, Assiut, Egypt LINDA D. STEWART • Institute for Global Food Security, School of Biological Sciences, Queen’s University Belfast, Northern Ireland, UK FERNANDO TELES • Laboratory of Mycology/Microbiology Unit and Centre for Malaria and Tropical Diseases (CMDT), Institute of Hygiene and Tropical Medicine (IHMT), Universidade Nova de Lisboa (UNL), Lisbon, Portugal ROGÉRIO TENREIRO • Centro de Biodiversidade, Genómica Integrativa e Funcional, Faculdade de Ciências, Universidade de Lisboa, Lisbon, Portugal

Contributors

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SIMON THIERRY • Bacterial Zoonosis Unit, Animal Health Laboratory, University Paris-Est, Anses, Maisons-Alfort, France BERNARD VALLAT • World Organization for Animal Health (OIE), Paris, France RICCARDO VILLA • Cell Culture Centre, Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (IZSLER), Brescia, Italy MARGARITA VILLAR • SaBio, Instituto de Investigación en Recursos Cinegéticos IREC, UCLM-JCCM, Ciudad Real, Spain MIGUEL VIVEIROS • Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Lisbon, Portugal

Part I Introducing Nucleic Acid Testing in the Veterinary Laboratory

Chapter 1 Overview and Challenges of Molecular Technologies in the Veterinary Microbiology Laboratory Mónica V. Cunha and João Inácio Abstract Terrestrial, aquatic, and aerial animals, either domestic or wild, humans, and plants all face similar health threats caused by infectious agents. Multifaceted anthropic pressure caused by an increasingly growing and resource-demanding human population has affected biodiversity at all scales, from the DNA molecule to the pathogen, to the ecosystem level, leading to species declines and extinctions and, also, to host-pathogen coevolution processes.

Technological developments over the last century have also led to quantic jumps in laboratorial testing that have highly impacted animal health and welfare, ameliorated animal management and animal trade, safeguarded public health, and ultimately helped to “secure” biodiversity. In particular, the field of molecular diagnostics experienced tremendous technical progresses over the last two decades that significantly have contributed to our ability to study microbial pathogens in the clinical and research laboratories.

This chapter highlights the strengths, weaknesses, opportunities, and threats (or challenges) of molecular technologies in the framework of a veterinary microbiology laboratory, in view of the latest advances.

Key words Molecular diagnostics, Veterinary microbiology, Nucleic acid testing, Animal health, Animal welfare, Public health, Biodiversity, Validation, In-house, Molecular assays, Quality assurance, Quality control

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Introduction Active and passive surveillance of infectious diseases in farmed and wild animals enable the detection of (re)emergent pathogens and the monitoring of endemic and/or zoonotic diseases and, ultimately, back up stakeholders, supporting national and international policies aimed at the protection of animal health and welfare, food security and public health, and the trade of animals and animal products.

During recent years, the world has witnessed the emergence or reemergence of a number of major animal diseases such as footand-mouth disease (FMD), bluetongue, classical swine fever (CSF), and avian influenza, which are emblematic examples of the overwhelming economic, social, and public health impacts that animal diseases may cause. Effective surveillance and control of animal

Mónica V. Cunha and João Inácio (eds.), Veterinary Infection Biology: Molecular Diagnostics and High-Throughput Strategies, Methods in Molecular Biology, vol. 1247, DOI 10.1007/978-1-4939-2004-4_1, © Springer Science+Business Media New York 2015

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Mónica V. Cunha and João Inácio

diseases require a rapid and accurate detection of the etiological agent by the use of effective diagnostic tests. Veterinary microbiology laboratories play a fundamental role in this context, the main purposes being the detection, identification, and characterization of any pathogenic organisms present in a diverse range of biological samples, such as tissues, blood, urine, and other fluids collected from suspect animals. The characterization of the microorganisms may include drug susceptibility testing and genotyping of the isolates for epidemiological purposes, which is essential to track sources and routes of infection, as well as for pathogen population studies.

The direct observation by optical microscopy of biological samples, usually stained by different processes, may give a first indication of the presence of a microorganism. Histopathological examination of hematoxylin-eosin-stained paraffin-embedded tissues may also give indications on the infectious disease and the putative agent, thus potentially complementing the diagnostic algorithm. However, the confirmation of an infection usually involves the isolation of the causative agent(s), followed by identification through additional phenotypic tests. Also widely used are immunological-based diagnostic methods relying on the specific binding between antigens and antibodies. These methods allow detecting the presence of antibodies produced in response to an antigen, or the presence of an antigen, through various experimental formats, such as agglutination tests, immunofluorescence, and ELISA. All these laboratory diagnostic approaches are based on the analyses of phenotypic characteristics of the microorganisms and can be designated as traditional, classical, or conventional methods. In contrast, the socalled molecular diagnostics methods detect or analyze, in the majority of the cases, genotypic characteristics of the microorganisms, such as the presence of certain genes or their respective polymorphisms. In this context, it is possible to identify the microorganisms at different taxonomic levels, from domain to species, and even down to intraspecific levels, through the analysis of genes or other genome regions. Moreover, relevant information can be obtained about pathogenic microorganisms by the analysis of molecular determinants encoding virulence factors (e.g., toxins) or conferring antimicrobial resistance. Importantly, in this field are the availability, appropriateness, and accuracy of diagnostic and phenotypic characterization tests to avoid selective pressure from unnecessary or erroneous antimicrobial use during therapy.

The field of molecular diagnostics experienced tremendous technological developments in the last two decades that have contributed significantly to our ability to study microbial pathogens in the clinical laboratory. Molecular methods have thus been taking an increasingly important role, supporting, complementing, and, in some cases, even replacing the conventional methods. This chapter highlights some of the strengths, weaknesses, opportunities, and threats (or challenges) of molecular technologies designed for the diagnosis of infectious diseases in animals.

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