Mycotoxins and disruption of vaccination efficacy in swine

MYCOTOXINS AND DISRUPTION of vaccination efficacy IN SWINE

Assist. Prof. Panagiotis Tassis Assistant Professor of Swine Medicine and Reproduction, Clinic of Farm Animals, School of Veterinary Medicine, Aristotle University of ThessalonĂ­ki, Greece

1

Pig farm vaccination programs are a major

Therefore, proper vaccine selection

preventive tool for a wide range of diseases and

and proper implementation are

syndromes affecting swine. They have colossal importance in terms of herd health and productivity, as well as from a financial standpoint.

the basis for the construction of a herd immune status that will counteract antigenic pressure during different productive stages.

As already discussed in the respective literature, mycotoxins seem to play an important role in disrupting this

EFFECTS OF MYCOTOXINS ON THE SWINE IMMUNE SYSTEM

major preventive health tool1.

In our previous technical article regarding the effects of major mycotoxins on the immune system of swine and

↓VACCINATION EFFICACY

cellular and molecular mechanisms involved, it had been reported that the health and economic impact of mycotoxins on the immune defense system of pigs is significant.

Three major outcomes of these effects on the swine immune system, herd health and productivity have been described2: Increased susceptibility to infectious diseases

↑ SUSCEPTIBILITY TO INFECTIOUS DISEASES

REACTIVATION OF CHRONIC INFECTIONS

Reactivation of chronic infections Decreased vaccination efficacy The present article will focus on the main mycotoxins affecting swine and extensively contaminate crops worldwide3.

The current knowledge on the effects of aflatoxins (AFs), fumonisins (FBs and mainly FB1) deoxynivalenol (DON), zearalenone (ZEN), ochratoxin A (OTA) and T-2 toxin, on the immune response after sensitization or vaccination in swine will be presented.

Emphasis will be put on studies with pigs and vaccines against swine pathogens.

2

Swine vaccines and vaccinal immunity The use of veterinary vaccines in swine production is a disease prevention tool that has been used

Porcine Reproductive and Respiratory Syndrome (PRRS) Porcine Circovirus 2 -associated diseases (PCV2-AD)

by swine farmers worldwide in a

Aujeszky’s disease (PRV)

variety of production systems.

Parvovirus infection (PPV)

It is still implemented in every conventional pig production system.

In the past few decades, facts in the field of novel vaccine production have changed rapidly.

The scientific field of swine vaccine

Swine influenza (SIV) Enzootic pneumonia (Mycoplasma hyopneumoniae) Pleuropneumonia (App – Actinobacillus Pleuropneumoniae) Glasser’s disease (Haemophilus parasuis) Atrophic Rhinitis (Pasteurella multocida ± Bordetella bronchiseptica)

development is a rapidly evolving

Erysipelas (Erysipelothrix rhusiopathiae)

research and innovation field.

Leptospirosis (Leptospira spp.)

Major swine diseases that can be prevented or controlled at field level with the use of commercially available vaccines, as part of a veterinary health management programme, include:

Escherichia coli infections Clostridium spp. infections Ileitis (Lawsonia intracellularis) Classical Swine Fever

Salmonella spp. and others

In cases such as the recently introduced in the European region African Swine Fever, there aren’t any commercial vaccines available so far. However respective research and development efforts are under way4.

3

Apart from typical intramuscular vaccination

Swine vaccines are usually either “dead”

against one pathogen, innovations of vaccine

(inactivated) or “live” (attenuated) and

technology in the past decades have resulted

can be used in different production stages

in the production of intradermal vaccines,

in the breeding stock and/or in suckling,

intranasal vaccines, as well as vaccines against

weaned or growing pigs, depending on

more than one pathogen, reaching up to three

the vaccine, its pathogenic target and

swine pathogens in one vaccine up today.

suggested administration programme.

ADMINISTRATION ROUTE Intramuscular Intradermal Intranasal

NUMBER OF PATHOGENS 1 2 3

VACCINE CLASSIFICATION TYPE OF VACCINE Inactivated (“dead”) Attenuated (“live”)

PRODUCTION STAGE Breeding stock Suckling piglets Weaned piglets Growing pigs

Vaccination success will result in a timely and appropriate humoral response to counteract the specific pathogen in the most susceptible population and production stage at an acceptable extent.

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Vaccine success includes: Reduction of infected animals and susceptibility to infection against a specific pathogen Reduced animals as pathogencarriers (reduced pathogen spreading in terms of time and microbial load) Improved herd immunity levels

However, limited attention has been given to the impact of mycotoxin contaminated feed as a reason for vaccine failure under field conditions.

Vaccine failure is an issue of great concern due to the severe health and productivity impact on farms. Many reasons for such failure have been described so far, including environmental and management reasons, such as:

Nevertheless, recent research data demonstrates the ability of a number of mycotoxins to affect vaccine-induced humoral response with significant impact on health and productivity results.

Improper vaccine storage, handling and administration Incorrect timing of vaccination and others.

5

Effects of Trichothecenes on vaccinal immunity DON H

DON immunostimulatory

O

O OH

HO

O HO

or immunosuppressing

DON

H

O

effects depend on the dose, frequency and duration of exposure5.

O

Apoptosis of lymphocytes

OH HO

O HO

It has been suggested that high doses of DON (greater than 10 μM), cause apoptosis of lymphocytes, resulting in immunosuppression, increased susceptibility to infection, reactivation of latent infections and reduced vaccine efficiency .

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↓ Vaccine efficacy

Immunosuppression ↑ Susceptibility to infections

Reactivation of latent infections

After DON exposure, inhibition of immune response has been demonstrated after porcine parvovirus vaccination in rats8.

Further three studies using ovalbumin (OVA) immunization suggested that DON affects anti-OVA

According to a previous review7, trichothecenes may generate an “immune evasion” environment that allows pathogens to escape host and vaccine immune defenses.

immunoglobulins response9-11.

1. In the first study (2.2–2.5 mg DON/kg feed, weaned pigs for 9 weeks), DON increased OVA-specific IgA and IgG, whilst

2. In a second study, 3.5 times greater

a biphasic effect of the toxin

levels of anti-OVA IgG titers in

on lymphocyte proliferation

comparison to control animals (3.5 mg

after antigen stimulation

DON/kg feed for 42 days) two weeks

(upregulation on 21st day post

after the first OVA immunization (day

exposure and down-regulation on

7 of the study) were demonstrated.

35th to 49th day post exposure) were reported, along with lower expression of both TGF-β and IFN-γ mRNA expression levels9.

6

3. In the latter study, seven days

Moreover, after feeding 1.8

Quite similarly in the study of

after a second OVA immunization

or 4.7 mg DON/kg feed (pigs

(day 21 of the study) anti-OVA

over 25.3 kg body weight at the

IgG levels were similar

start of the trial) a significant

between groups (DON-fed

dose-dependent reduction in

animals vs. control animals).

secondary antibody response to

Gutzwiller et al.16, pigs were fed 3.2 mg DON and 0.06 mg ZEN, or 2.1 mg DON and 0.25 mg ZEN/ kg diet and received PPV vaccination (one-tenth of the recommended dose).

Anti-OVA IgA levels in that study were similar between the two

tetanus toxoid was present when compared to the control group14.

In that study mycotoxin exposure

trial groups up to one week

Furthermore, after feeding a

did not affect antibody production,

prior to the end of the study

mixture of 1.0 mg DON /kg and

probably due to administration of

period, and then reduced in

250Îźg ZEN/kg contaminated

low vaccine dose and the short

the DON-treated animals .

feed and PRV double vaccination,

time interval between vaccination

PRV antibody titers were

and antibodies determination.

10

According to a recent study11 in

significantly decreased 14 days

which pigs were immunized with

after booster vaccination15.

OVA, PRV, swine fever and porcine circoviruses vaccines, ingestion of feed contaminated with 1.0 and 3.0 mg DON /kg feed reduced the concentration of serum porcine circoviruses antibody titer in pigs, whereas serum OVA antibody titer levels were not affected.

Nevertheless, it should be mentioned that inhibition of IFN-Îł (as suggested previously9) and Toll-like Receptors (TLR) expression, assists pathogens escaping host and vaccine immune defenses12.

In a 28-day feeding study with 0.15, or 1.5, or 3 mg DON/kg feed13 and subsequent immunization with sheep red blood cells, delayed peak titers were observed (one week later) in DON-fed animals, when compared with control animals.

7

According to a series of studies

In an in vitro study (permissive

Moreover, in another study with

on the effects of DON on the

cells infected with PRRSV were

pigs, DON increased the severity

immune response against PRRS

treated with 140–280 ng/

of the viral infection in the

and PCV2 viruses, very interesting

ml DON) by the same research

presence of porcine circovirus

group , a similar observation was

type 2 (PCV2) virus20.

findings have been presented

.

17-20

Major findings included that ingestion of DON contaminated

reported i.e. replication of PRRSV was significantly inhibited.

Results showed that viremia and lung viral load tended

feed can decrease the immune

to be higher in animals

response against PRRSV

ingesting DON contaminated

and influence the course of PRRSV infection in pigs.

In the study of Savard et al.17, piglets received DON- naturally DON vs PRRS & PCV2 VACCINATION

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contaminated diets (2.5 and 3.5 mg/kg) and were then inoculated with PRRSV.

In vivo effects of DON ingestion supported a negative effect of the toxin on PRRSV-specific humoral responses (DON at 2.5 mg/kg significantly decreased PRRSV specific humoral responses), as well as amplification of PRRSVattributed negative effects such as those on weight gain, lung lesions, and mortality.

However, such an effect did not associate with a significant increase in viral replication, since DON ingestion resulted in a decrease of PPRSV replication.

diet at 2.5 mg/kg (pigs were

In the latter study it was demonstrated that the reduction

inoculated with PCV2b virus).

of viral replication could be

However, DON had no

attributed to a DON-induced

significant effect on clinical

pro-inflammatory cytokine

manifestation of PCV2-AD.

environment that promoted activation of apoptosis, which is an

Authors of the latter study

important host defense mechanism,

supported that DON has neither in vitro nor in vivo clear potentiating effects in the development of PCV2 infection despite slight increases in viral replication.

as it interrupts viral replication and eliminates virus-infected cells12,21.

Furthermore, it has been proved that DON can decrease the replication of the attenuated PRRSV vaccine strain in

Nevertheless, it can be

vaccinated pigs and their antibody

concluded that DON exposure

response to the vaccine .

may hamper the acquisition

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of vaccine-induced protective Such significant finding is

immune responses.

consistent with results from a study with DON-exposed mice vaccinated with inactivated PPV, which demonstrated disruption of the immune response to the vaccine through modulation of specific cytokines and chemokines22.

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T-2 TOXIN

T-2 toxin is reported to

Pigs fed 1.324 or 2.102

be immunotoxic, through

mg T-2 toxin/kg exhibited

its cytotoxic, apoptotic

reduced anti-OVA antibody

or immunosuppressive

production on day 21 without

attributes .

significant alteration to specific

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lymphocyte proliferation26.

A study with necrotic enteritis B (NEB) vaccination of pigs that received 5 mg T-2/kg feed, resulted in significantly reduced NEB antibody

Apoptosis

levels in T-2-exposed animals24.

Immunosuppression due to T-2 has

Cytotoxicity

been also observed in another feeding

Immunosuppression

study with pigs, (0.5-3.0 mg T-2/ kg feed), in which animals were immunized with horse globulin.

TOXIN T-2

Results suggested reduction of anti-horse globulin antibodies synthesis, whereas a dose

IMMUNOTOXICITY

dependent depletion of lymphoid elements in the thymus and spleen, was also reported25.

After OVA immunization, subclinical doses of T-2 toxin induced an early and transient increase of total IgA plasma concentration but a decrease in the anti-OVA IgG titer.

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Effects of Fumonisins on vaccinal immunity

FUMONISIN B1

Disruption of sphingolipid biosynthesis FBs competitively inhibit ceramide synthases (CerS), a group of key enzymes in the biosynthesis of ceramide and more complex sphingolipids, resulting in the

Accumulation of sphinganine & sphingosine

disruption of sphingolipid metabolism, whilst they have

Ceramide synthase

been also linked with impairment of innate and acquired immune response, including reduction of specific antibody response during vaccination27,28.

level of IL-10 were observed

Alterations were statistically

in a sex-related manner, thus

significant for animals

Previous studies have supported

proving the immunosuppressive

receiving FB-contaminated

that FB1 modifies the Th1/Th2

effects of the toxin.

feed, and more pronounced in animals that recived

(T-helper 1/T-helper 2) cytokine balance in pigs similar to an

Such differences in the

the combined mycotoxins-

impaired humoral response27,29.

specific immune response

contaminated diet.

were observed only in male

In vivo exposure (28 days) of weanling piglets to feed contaminated with 8 mg FB1/ kg significantly decreased the expression of IL-4 mRNA (IL-4 is a Th2 cytokine involved in the humoral response) by porcine whole blood cells and diminished the specific antibody titer after vaccination against Mycoplasma agalactiae27.

pigs, but not female ones29.

An increase of specific IgA was In a study by Grenier et al.30, pigs received a diet contaminated with

reported for animals receiving DON-contaminated diet, but not

either DON (3 mg/kg) or FB (6 mg/

for those that received combined

kg) or both toxins and immunized

DON and FB, possibly due to FB

twice with OVA.

interference at the intestinal level

Ingestion of diets contaminated with DON or FB individually

through its action on sphingolipids.

At the same time, reduced

or in combination altered

lymphocyte proliferation upon

In a quite similar study with FB1

immunoglobulins production

OVA stimulation was demonstrated

(8mg FB1/kg feed), significantly

after OVA immunization

in the animals receiving any of the

decreased specific antibody

and reduced anti-OVA IgG

three contaminated diets (DON,

levels after vaccination against

plasma concentration.

FB, or combined DON and FB).

Mycoplasma agalactiae, as well as the mRNA expression

10

The humoral immune response

Both, F4-specific IgM and IgA

Quite similarly, pigs fed low

was significantly disturbed, with a

antibody secreting cells were

levels of FB-contaminated feed

strong decrease in antibodies levels

reduced after FB1-exposure

(2 mg FB1/kg contaminated

at days 21 and 35 after vaccination,

and the authors suggested

culture material/day for 5

in pigs exposed to 0.5 mg OTA/

that FB1 could interfere with

weeks) and vaccinated with PRV

kg feed and/or 10 mg FB1/kg feed

the induction phase of the

vaccine showed absence of an

for three months, and vaccinated

immune response through

FB-attributed significant effect

against Aujeszky’s disease (Suid

reduction of in vivo antigen

on PRV antibody titers34.

Herpesvirus 1 [SuHV1]) .

presenting cells maturation.

31

That antibody disruption

On the other hand, few particular

was detected in animals

studies have shown the absence of

fed both mycotoxins, either

FB-attributed significant effects on

alone or in combination.

immune response after vaccination.

Nevertheless, the majority

In another study with piglets that

Exposure of piglets to FB1-

were orally exposed to a low dose of FB1 (1 mg FB1/kg body weight)

contaminated feed for up to 4 months (1, 5, and 10 mg FB1/kg feed) did

immunosuppressive properties,

for 10 days, a longer shedding of

not affect significantly their antibody

alters the cytokine profile

F4(+) enterotoxigenic Escherichia

titers against Aujeszky’s disease33.

and reduces the specific

coli (ETEC) following infection and lower induction of the antigenspecific immune response following oral immunization, were presented32.

of studies and respective findings support that FB1 has

antibody response built during a vaccination protocol.

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Effects of Zearalenone on vaccinal immunity

ZEN

ZEN has significant estrogenic

(intraperitoneally), revealed that ZEN, with or without

potency in swine but has also been

immune challenge, can decrease immunoglobulins

suggested as an immunotoxic

in serum and cytokines in lymphoid organs36.

compound35.

A study with pigs that received ZEN (dietary Few studies have investigated ZEN

levels of 1.1 to 3.2 mg/kg feed for 18 days) and

effects on humoral immune

a swine fever live vaccine, demonstrated that

response after vaccination with

specific antibody titers in the group treated with

a commercial vaccine or other

ZEN (2.0 and 3.2 mg/kg) were significantly lower

type of immunization in pigs.

18 days after immunization in comparison with the control group, in a dose-dependent manner.

Nevertheless, the effect of estrogens on the immune system

Levels of IgM showed a trend of decreasing

have received attention due to

linearly with increased levels of ZEN, indicating

their immunomodulatory activity

that ZEN (3.2 mg/ kg) inhibited humoral

on cell-mediated responses

immunity in piglets, whilst it was also suggested

and antibody production .

that ZEN may affect protein metabolism37.

35

A study performed with ZEN in rats

Additionally, it has been discussed that effects

(mycotoxin administered via gavage

of ZEN on humoral immune response could be

at dosages of 0, 1, 5, and 30 mg/

related to receptor-specific effects, since ZEN is

kg for 36 days) and subsequent

an agonist toward estrogen receptors α (ERα) and a

inactivated PPV vaccine administration

mixed agonist-antagonist of ERβ, with possible full antagonism of the ERβ expressed in B cells35.

12

Effects of Aflatoxins on vaccinal immunity

For more than half century,

Previous studies with pigs treated with

the detrimental effects of

AFs showed contradicting results.

AFLATOXIN B1

AFs on vaccinal response have been reported.

It has been shown that AFB1 interferes with the development of acquired immunity in swine following erysipelas vaccination with bacterin preparation (a suspension of killed bacteria) of E. rhusiopathiae and increases the severity of infection with E. rhusiopathiae38.

In a previous study that included

Joens et al.40 reported significantly lower hemagglutination titers against Treponema hyodysenteriae (name used at present: Brachyspira hyodysenteriae as swine dysentery causative agent), whilst AF ingestion did not alter humoral response of weanling pigs to sheep red blood cells41 or to Erysipelothrix rhusiopathiae42 in other studies.

In that later study, authors supported that AFB1 exposure does not result in significant modulation of the humoral immune response, whilst it can induce IgA increase but not at statistically significant levels.

ingestion of low doses of AFs (140 and 280 ppb for 4 weeks) a tendency

Moreover, after immunization of

towards reduced immune response

pigs with OVA and concurrent

Findings of another investigation in

against Mycoplasma agalactiae

AFB1 exposure (385 μg AFB1/

mice44 on the involvement of AFB1

(280-ppb-treated group)

kg feed; 867 μg AFB1/kg feed, or

in Swine Influenza Virus (SIV)

was observed39.

1807 μg AFB1/kg feed) absence of

replication in vitro and in vivo,

major effect on humoral immunity

supported that 10–40μg/kg of AFB1

(concentrations of total IgA, IgG

in vivo promotes SIV replication, inflammation and lung damage by activating TLR4-NFkB signaling.

and IgM and specific anti-OVA IgG), but impaired lymphocyte activation was reported43.

13

Effects of Ochratoxin A on vaccinal immunity

OCHRATOXIN A

OTA has a a well described signi icant nephrotoxic mode of action in swine, whilst it has been suggested as a compound that can affect immune response in swine.

It has been reported that immunosuppression is the first expressed toxic effect of OTA that may become evident clinically

As reported in the FB section, alterations in humoral immune response were reported also in an in vivo study with pigs (500 Îźg OTA/ kg feed for 3 months with or without 10 mg FB1/ kg feed), in which a strong decrease in antibody titer was observed after immunization against Morbus Aujesky (PRV)31.

before nephropathy45.

A possible synergistic action of OTA and FB1 on immunosuppression in pigs could be discussed.

In a study with 1 OTA/kg feed provided to swine for up to three weeks, animals were immunized against Salmonella choleraesuis haemorrhagic diarrhea45.

Results proved OTA-attributed immunosuppression (reduced mean antibody titer on day 21 post immunization) and delayed response to immunization.

Moreover, increased susceptibility to infectious agents (Brachyspira

hyodysenteriae and Campylobacter coli infections) was observed.

14

Remarks and conclusions as regards field conditions Taken together, a large number of studies have demonstrated the

Taking into account that a farm vaccination programme is of colossal importance in terms

negative effects of the previously

of disease prevention, the effects of mycotoxins

mentioned mycotoxins on the humoral

should be taken into consideration.

response after sensitization or vaccination.

A vast majority of significant mycotoxins for swine have shown potential to induce a clear negative effect on immune response against various swine pathogens after vaccination in pigs.

However, it is important to remember that, under field

At the field level, diagnostic investigation of reduced vaccine efficacy cases could include feed mycotoxicological analysis, particularly when reduced vaccine efficacy is correlated with other clinical signs of mycotoxicosis or has occurred at a subsequent time interval after alterations in feed raw materials or feed production.

It should be highlighted that the presence of mycotoxins in the feed may lead to a breakdown in vaccinal immunity and to the occurrence of disease even in properly vaccinated flocks2.

conditions, such mycotoxins concurrently contaminate pig feed. Therefore, the immune system of pigs receives pressure from more than one mycotoxin, that could result in various interactions as regards immune response after vaccination.

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REFERENCES 1. Oswald, I.P.; Marin, D.E.; Bouhet, S.; Pinton, P.; Taranu, I.; Accensi, F. Immunotoxicological risk of mycotoxins for domestic animals.

Food Addit Contam. 2005, 22, 354-60. doi: 10.1080/02652030500058320.

2. Pierron, A.; Alassane-Kpembi, I.; Oswald, I.P. Impact of mycotoxin on immune response and consequences for pig health. Anim. Nutr.

2016, 2, 63-68. doi:10.1016/j.aninu.2016.03.001.

3. Gruber-Dorninger, C.; Jenkins, T.; Schatzmayr, G. Global Mycotoxin Occurrence in Feed: A Ten-Year Survey. Toxins 2019, 11, 375, doi:10.3390/toxins11070375.

4. Barasona, J.A., Gallardo, C., Cadenas-Fernández, E., Jurado, C., Rivera, B., Rodríguez-Bertos, A., Arias, M., Sánchez-Vizcaíno, J.M. First Oral Vaccination of Eurasian Wild Boar Against African Swine Fever Virus Genotype II. Front. Vet. Sci. 2019, 6, 137. doi: 10.3389/ fvets.2019.00137.

5. Pestka JJ, Zhou HR, Moon Y, Chung YJ. Cellular and molecular mechanisms for immune modulation by deoxynivalenol and other trichothecenes: unraveling a paradox. Toxicol. Lett., 2004, 153, 61–73. doi: 10.1016/j.toxlet.2004.04.023.

6. Maresca, M. From the gut to the brain: journey and pathophysiological effects of the food-associated trichothecene mycotoxin deoxynivalenol. Toxins, 2013, 5(4), 784–820. doi: 10.3390/toxins5040784.

7. Wu, Q, Wu W, Franca TCC, Jacevic V, Wang X, Kuca K. Immune Evasion, a Potential Mechanism of Trichothecenes: New Insights into Negative Immune Regulations. Int J Mol Sci. 2018, 19, 3307. doi: 10.3390/ijms19113307.

8. Choi, B.K., Cho, J.H., Jeong, S.H., Shin, H.S., Son, S.W., Yeo, Y.K., Kang, H.G. Zearalenone affects immune-related parameters in lymphoid organs and serum of rats vaccinated with porcine parvovirus vaccine. Toxicol Res. 2012, 28, 279-88. doi: 10.5487/ TR.2012.28.4.279. PMID: 24278621; PMCID: PMC3834426.

9. Pinton, P.; Accensi, F.; Beauchamp, E.; Cossalter, A-M.; Callu, P.; Grosjean, F.; Oswald, I.P. Ingestion of deoxynivalenol (DON) contaminated feed alters the pig vaccinal immune responses. Toxicol. Lett. 2008, 177, 215-222. doi:10.1016/j.toxlet.2008.01.015.

10. Lessard, M.; Savard, C.; Deschene, K.; Lauzon, K.; Pinilla, V.A.; Gagnon CA.; Lapointe, J.; Guay, F.; Chorfi, Y. Impact of deoxynivalenol (DON) contaminated feed on intestinal integrity and immune response in swine. Food Chem Toxicol. 2015, 80, 7-16.

11. Zhang, L., Ma, R., Zhu, M.-X., Zhang, N.-Y., Liu, X.-L., Wang, Y.-W., Qin, T., Zheng, L.-Y. Liu, Q., Zhang, W.-P., Karrow, N. A., Sun, L.-H. Effect of deoxynivalenol on the porcine acquired immune response and potential remediation by a novel modified HSCAS adsorbent. Food Chem Toxicol, 2020, 138, 11187. https://doi.org/10.1016/j.fct.2020.111187.

12. Wu, Q.H.; Wang, X.; Nepovimova, E.; Miron, A.; Liu, Q.Y.; Wang, Y.; Su, D.X.; Yang, H.L.; Li, L.; Kuca, K. Trichothecenes: Immunomodulatory effects, mechanisms, and anti-cancer potential. Arch. Toxicol. 2017, 91, 3737–3785.

13. Rotter, B.A.; Thompson, B.K.; Lessard, M.; Trenholm, H.L.; Tryphonas, H. Influence of low-level exposure to Fusarium mycotoxins on selected immunological and hematological parameters in young swine. Fundam. Appl. Toxicol. 1994, 23, 117–124.

14. Overnes, G.; Matre, T.; Sivertsen, T.; Larsen, H.J.; Langseth, W.; Reitan, L.J.; Jansen, J.H.. Effects of diets with graded levels of naturally deoxynivalenol-contaminated oats on immune response in growing pigs. Zentralbl Veterinarmed A. 1997, 44, 539–550.

15. Cheng YH, Weng CF, Chen BJ, Chang MH. Toxicity of different Fusarium mycotoxins on growth performance, immune responses and efficacy of a mycotoxin degrading enzyme in pigs. Anim Res 2006, 55, 579-90. https://doi.org/10.1051/animres:2006032.

16. Gutzwiller, A.; Czegledi, L.; Stoll, P.; Bruckner, L. Effects of Fusarium toxins on growth, humoral immune response and internal organs in weaner pigs, and the efficacy of apple pomace as an antidote. J. Anim. Physiol. Anim. Nutr. 2007, 91, 432–438.

17. Savard C, Pinilla V, Provost C, Gagnon CA, Chorfi Y. In vivo effect of deoxynivalenol (DON) naturally contaminated feed on porcine reproductive and respiratory syndrome virus (PRRSV) infection. Vet Microbiol 2014a, 174, 419–26. doi:10.1016/j.vetmic.2014.10.019.

18. Savard C, Pinilla V, Provost C, Segura M, Gagnon CA, Chorfi Y. In vitro effect of deoxynivalenol (DON) mycotoxin on porcine reproductive and respiratory syndrome virus replication. Food Chem Toxicol 2014b, 65, 219–26. doi:10.1016/j.fct.2013.12.043.

19. Savard C, Gagnon CA, Chorfi Y. Deoxynivalenol (DON) naturally contaminated feed impairs the immune response induced by porcine reproductive and respiratory syndrome virus (PRRSV) live attenuated vaccine. Vaccine 2015a, 33, 3881–6. doi:10.1016/j. vaccine.2015.06.069.

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20. Savard C, Provost C, Alvarez F, Pinilla V, Music N, Jacques M, et al. Effect of deoxynivalenol (DON) mycotoxin on in vivo and in vitro porcine circovirus type 2 infections. Vet Microbiol 2015b, 176, 257–67. doi:10.1016/j.vetmic.2015.02.004.

21. Thomson, B.J. Viruses and apoptosis. Int J Exp Pathol, 2001, 82, 65–76.

22. Choi, B.K., Jeong, S.H., Cho J.H., Shin, H.S., Son, S.W., Yeo, Y.K., Kang, H.G. Effects of oral deoxynivalenol exposure on immune-related parameters in lymphoid organs and serum of mice vaccinated with porcine parvovirus vaccine. Mycotoxin Res 2013, 29:185–192.

23. Bondy, G.S.; Pestka, J.J. Immunomodulation by fungal toxins. J. Toxicol. Environ. Health. B. Crit. Rev. 2000, 3, 109-143.

24. Rafai, P., Tuboly S. Effect of T-2 Toxin on Adrenocortical Function and Immune Response in Growing Pigs. Zbl. Vet. Med. B, 1982, 29, 558-565.

25. Rafai, P.; Tuboly, S.; Bata, A.; Tilly, P.; Vanyi, A.; Papp, Z.; Jakab, L. Tury, E. Effect of various levels of T-2 toxin in the immune system of growing pigs. Vet. Rec. 1995, 136, 511-514.

26. Meissonnier GM, Laffitte J, Raymond I, Benoit E, Cossalter AM, Pinton P, Bertin, G.; Oswald, I.P.; Galtier, P. Subclinical doses of T-2 toxin impair acquired immune response and liver cytochrome P450 in pigs. Toxicol. 2008a, 247, 46-54.

27. Taranu, I.; Marin, D.E.; Bouhet, S.; Pascale, F.; Bailly, J.D.; Miller, J.D.; Pinton, P.; Oswald, I.P. Mycotoxin fumonisin B1 alters the cytokine profile and decreases the vaccinal antibody titer in pigs. Toxicol Sci 2005, 84, 301-7.

28. EFSA CONTAM Panel (EFSA Panel on Contaminants in the Food Chain), Knutsen H-K, Alexander J, Barregard L, Bignami M, et al.

Scientific opinion on the risks for animal health related to the presence of fumonisins, their modified forms and hidden forms in feed. EFSA J. 2018, 16, 5242, 144 pp. https://doi.org/10.2903/j.efsa.2018.5242.

29. Marin, D.E.; Taranu, I.; Pascale, F.; Lionide, A.; Burlacu, R.; Bailly, J.-D.; Oswald, I.P. Sex-related differences in the immune response of weanling piglets exposed to low doses of fumonisin extract. Br. J. Nutr., 2006, 95, 1185-1192. doi: 10.1079/BJN20061773.

30. Grenier, B.; Loureiro-Bracarense, A.P.; Lucioli, J.; Pacheco, G.D.; Cossalter, A.M.; Moll, W.D.; Schatzmayr, G.; Oswald, I.P. Individual and combined effects of subclinical doses of deoxynivalenol and fumonisins in piglets. Mol Nutr Food Res 2011, 55, 761-71.

31. Stoev, S.D.; Gundasheva, D.; Zarkov, I.; Mircheva, T.; Zapryanova, D.; Denev, S.; Mitev, Y.; Daskalov, H.; Dutton, M.; Mwanza, M.; Schneider, Y.J. Experimental mycotoxic nephropathy in pigs provoked by a mouldy diet containing ochratoxin A and fumonisin B1.

Experim. Toxicol. Pathol. 2012, 64, 733–741.

32. Devriendt, B.; Gallois, M.; Verdonck, F.; Wache, Y.; Bimczok, D.; Oswald, I.P.; Goddeeris B.M.; Cox E. The food contaminant fumonisin B1 reduces the maturation of porcine CD11R1+ intestinal antigen presenting cells and antigen-specific immune responses, leading to a prolonged intestinal ETEC infection. Vet. Res. 2009, 40, 40. doi:10.1051/vetres/2009023.

33. Tornyos, G., Kovacs, M., Rusvai, M., Horn, P., Fodor, J., Kovacs, F. Effect of dietary fumonisin B1 on certain immune parameters of weaned pigs. Acta Vet. Hung. 2003, 51, 171–179.

34. Gumprecht, L.A., Peavey, C., Zuckerman, F., Rottinghaus, G., Haschek, W., Wollenberg G. Effects of fumonisin on specific and nonspecific immunity in pigs after pseudorabies vaccination. Vet. Pathol. 1997, 34, 519.

35. Hueza, I.M.; Raspantini, P.C.; Raspantini, L.E.; Latorre, A.O.; Górniak, S.L. Zearalenone, an estrogenic mycotoxin, is an immunotoxic compound. Toxins 2014, 6, 1080-95.

36. Choi, B.K.; Cho, J.H.; Jeong, S.H.; Shin, H.S. Zearalenone affects immune-related parameters in lymphoid organs and serum of rats vaccinated with porcine Parvovirus vaccine. Toxicol. Res. 2012, 28, 279–288.

37. Yang L, Yang W, Feng Q, Huang L, Zhang G, Liu F, Jiang S, Yang Z. Effects of purified zearalenone on selected immunological measurements of blood in post-weaning gilts. Anim Nutr. 2016, 2, 142-148. doi: 10.1016/j.aninu.2016.04.008.

38. Cysewski, S.J.; Wood, R.L.; Pier, A.C.; Baetz, A.L. Effects of aflatoxin on the development of acquired immunity to swine erysipelas. Am J Vet Res 1978, 39, 445-8.

39. Marin, D.E.; Taranu, I.; Bunaciu, P.R.; Pascale, F.; Tudor, D.S.; Avram, N.; Sarca, M.; Cureu, I.; Criste, R.D.; Suta, V.; Oswald, I.P. Changes in performance, blood parameters, humoral and cellular immune response in weanling piglets exposed to low doses of aflatoxin. J Anim. Sci. 2002, 80, 1250–1257.

40. Joens, L.A., Pier, A.C., Cutlip, R.C. Effects of aflatoxin consumption on the clinical course of swine dysentery. Am. J. Vet. Res 1981, 42,1170–1172.

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41. van Heugten, E., Spears, J.W., Coffey, M.T., Kegley, E.B., Qureshi, M.A. The effect of methionine and aflatoxin on immune function in weanling pigs. J. Anim. Sci. 1994, 72, 658–664.

42. Pananagala, V.S., Giambrone, J.J., Diener, U.L., Davis, N.D., Hoerr, F.J., Mitra, A., Schultz, R.D., Wilt, G.R. Effects of aflatoxin on the growth performance and immune responses of weanling swine. Am. J. Vet. Res. 1986, 47, 2062–2067.

43. Meissonnier, G.M.; Pinton, P.; Laffitte, J.; Cossalter, A.M.; Gong, Y.Y.; Wild, C.P.; Bertin, G.; Galtier, P.; Oswald, I.P. Immunotoxicity of aflatoxin B1: impairment of the cell-mediated response to vaccine antigen and modulation of cytokine expression. Toxicol Appl Pharmacol 2008b, 231, 142-149.

44. Sun, Y.; Su, J.; Liu, Z.; Liu, D.; Gan, F.; Chen, X.; Huang, K. Aflatoxin B1 Promotes Influenza Replication and Increases Virus Related Lung Damage via Activation of TLR4 Signaling. Front Immunol. 2018, 9, 2297. doi: 10.3389/fimmu.2018.02297.

45. Stoev, S.D.; Goundasheva, D.; Mirtcheva, T.; Mantle, P.G. Susceptibility to secondary bacterial infections in growing pigs as an early response in ochratoxicosis. Experim. Toxicol. Pathol. 2000, 52, 287-296. doi: 10.1016/s0940-2993(00)80049-4.

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