The impact of mycotoxins on the immune status of ruminants
Consequences on disease susceptibility andvaccine efficacy

Numerous investigations have shown that, once ingested, mycotoxin-contaminated feed can induce harmful effects, such as hepatorenal toxicity, reproductive, cardiac, and neurotoxicity, as well as immunotoxicity in animals (Fang et al., 2022).

When livestock ingests one or more mycotoxins, the effect on health can be acute, showing evident signs of disease or even causing death.

However, acute manifestation of mycotoxicosis is rare under farm conditions (Vieira et al., 2014).

The effects of mycotoxin ingestion are mainly chronic, leading to hidden disorders with reduced ingestion, productivity, and fertility (Fink-Gremmels, 2008). Such effects cause severe economic losses due to:

• ⇰ Clinical changes in animal growth
• ⇰ Reduction in feed intake or complete anorexia
• ⇰ Alteration in nutrient absorption and metabolism
• ⇰ Effects on the endocrine system
• ⇰ Suppression of the immune system

(Oswald et al., 2005; Storm et al., 2014a)

The immunotoxic effects of MYCOTOXINS in ruminants

Mycotoxins have been proved to have immunosuppressive effects depending on:

• The mycotoxin(s) involved
• The concentration
• The parameter studied

For example, aflatoxin B₁ (AFB₁) has immunotoxic consequences, such as the disruption of innate and acquired/adaptive immunity (Meissonnier et al., 2008; Mohsenzadeh et al., 2016).

In ruminants, mycotoxins have been confirmed to induce oxidative stress, hypoxia, and immunosuppression.

Additionally, emerging evidence shows that mycotoxins have the potential to induce cellular senescence, which is involved in their immunomodulatory effects (You et al., 2023).

Penicillium-derived toxins

P. roqueforti and P. paneum produce several secondary metabolites with immunosuppressive, antibacterial, and other not well-defined toxicological effects for animals (Storm et al., 2008; Storm et al., 2014b).

Monascus ruber-derived toxins

Based on in vitro results, immunotoxic effects of citrinin have been documented at very high doses (Stec et al., 2008).

Fusarium-derived toxins

Fusarium-derived toxins, mainly deoxynivalenol (DON) and Fumonisin B₁ (FB₁), can drastically alter the defense mechanisms in the intestine, reducing epithelial integrity, cell proliferation and mucus production or increasing intestinal permeability, as well as the production of immunoglobulins and cytokines (Antonissen et al., 2014).

For example, A. fumigatus produces several mycotoxins, including gliotoxin and tremorgens that are toxic to cattle.

⇰ Gliotoxin, an immune suppressant, has been present in animals infected with A. fumigatus (Bauer et al., 1989).

In an insect model, the role of gliotoxin in increasing the virulence of A. fumigatus has been demonstrated A. fumigatus (Reeves et al., 2004).

Several metabolites from Fusarium fungi, collectively known as Fusarium toxins, have shown deleterious effects in humans and livestock by targeting the immune and hepatic systems (Bondy and Pestka, 2000b; Trenholm et al., 1984).

The impact of mycotoxins on the ruminant immune system

In ruminants, AFB₁, ochratoxin A (OTA), and DON are the three mycotoxins that have received the most scholarly attention and have been tested most routinely in clinical settings.

These mycotoxins have been found not only to suppress immune responses but also to induce inflammation and even increase susceptibility to pathogens (Sun et al., 2023a).

It is theorized that, with in the development of mycosis, mycotoxins produced by the invading fungi can suppress immunity, therefore increasing their infectivity (Whitlow and Hagler Jr, 2010).

Leukocytes and differential leukocytic count

In a 3-week experimental case of mycotoxicosis in steers, there was leukocytosis with lymphocytosis, monocytosis, and neutropenia. However, 1.7 mg of DON and 3.5 mg of fumonisins (FUM) per kg of total ration were insufficient to cause significant cytotoxic effects on circulating mononuclear leukocytes in these animals (Roberts et al., 2021b).

The same authors added that the phagocytic capacity of granulocytes from all the steers was substantially reduced by week 5, whereas monocyte activity began to decline even earlier in the study.

In a study with control-fed steers, CD8+ lymphocytes decreased during the first two weeks of the treatment period, while treatment-fed animals (DON and FUM) had significantly higher CD4-CD8+ lymphocyte populations during week 2 (Roberts et al., 2021b).

It has been concluded that bovine cells are more sensitive to DON than those from other livestock species (Novak et al., 2018). However, their effect on body condition or growth status was not been formally evaluated (Pestka, 2008).

It has been suggested that the CD4:CD8 ratio could be a candidate biomarker of early DON exposure in beef cattle, which may be attenuated by co-occurring fumonisin toxicity (Taranu et al., 2010).

RNA-Seq analysis has revealed 217 differentially expressed genes between treatment- and control-fed steers, with mycotoxin exposure broadly leading to down-regulation of annotated features.

⇰ Downregulated genes from treatment-fed steers indicates that iron utilization may be adversely affected by DON/ FUM exposure (Roberts et al., 2021b).

Previous studies have shown alterations in cellular oxygen transport mechanisms by in vitro administration of fumonisins (Osweiler et al., 1993; Yin et al., 1996).

In an experimental study, mycotoxins have shown to reduce phagocytic activity in finishing-stage steers, regardless of prior mycotoxin exposure (Roberts et al., 2021a).

Inflammatory Response, Neutrophil Chemotaxis and Immunoglobulins

In an experimental study in finishing steers, exposure to DON/FUM induced upregulation of inflammatory response times at days 7 and 21, along with neutrophil chemotaxis times at day 7 (Roberts et al., 2021a).

The same authors found that early mycotoxin exposure was accompanied with a numerical increase in neutrophil abundance, but at the same time, functional assays showed a slight decrease in granulocyte phagocytosis and oxidative burst. Additionally, immunoglobulin-related pathways were enriched in animals exposed to DON/FUM on day 21.

The effects of DON on laboratory animal immunoglobulin levels have been previously reviewed (Bondy and Pestka, 2000a), showing alterations in immunoglobulin production in a class-dependent manner. However, disruptions to antibody production are an important nonlinear toxicological outcome in cattle (Dänicke et al., 2018).

DIA and DAVID analysis tools revealed the effects of dietary mycotoxin treatments on the beef cattle transcriptome, showing consistent inhibition of focal adhesion, ECM signaling and PI3K-Akt signaling pathways (Roberts et al., 2021a).

In vitro studies have demonstrated a suppressive effect of aflatoxins (AF) on inflammatory cytokine levels in cattle (Kurtz and Czuprynski, 1992).

In pigs, mycotoxin exposure inhibits lymphocyte proliferation and alters cytokine production.

Specifically, AFB₁ increases the synthesis of IFN-gamma, a Th1 cytokine involved in the cell mediated immune response, and decreases IL-4 synthesis, a Th2 cytokine involved in the humoral response (Pierron et al., 2016).

Significant changes in some coagulation factors in the extrinsic pathway in the intoxicated lambs and prothrombin time determination could be used as an indicator of aflatoxicosis in lambs (Fernández et al., 1995).

DON, nivalenol (NIV), zearalenone (ZEN), and FB₁ have been proven to modulate the secretory mucins MUC5AC and MUC5B, as well as total mucin-like glycoprotein secretion, all of which are essential components of host mucosal immunity (Wan et al., 2014).

Susceptibility to infection and failure of vaccination

Mycotoxin exposure can affect the infection severity of some pathogens, including bacteria, viruses, and parasites. Their specific mechanisms of action include three aspects:

• ⇰ Mycotoxin exposure directly promotes the proliferation of pathogenic microorganisms.
• ⇰ Mycotoxins induce toxicity, destroying the integrity of the mucosal barrier and promoting an inflammatory response, thereby increasing the susceptibility of the host to infections.
• ⇰ Mycotoxins reduce the activity of some specific immune cells and induce immune suppression, resulting in reduced host resistance.
• (Sun et al., 2023a)

Mycotoxin-associated increased susceptibility to infection has been documented in experimental and natural conditions.

In calves, exposure to AFs has shown to increase susceptibility to Shiga toxin-producing Escherichia coli (STEC) (Baines et al., 2013a).

Alteration of lymphocyte proliferation and cytokine production may explain vaccine failures that have been observed in vivo.

⇰ The presence of mycotoxins in the feed may lead to a decline in vaccinal immunity and to the occurrence of disease even in properly vaccinated flocks (Pierron et al., 2016).

Synergistic effects between viruses and mycotoxins have been extensively studied in swine (Gan et al., 2022; Gan et al., 2018) and the outcomes of the ingestion of mycotoxin-contaminated feed are increased susceptibility to infectious diseases, reactivation of chronic infection and a decreased vaccine efficacy (Pierron et al., 2016).

Exposing calves to 1–3 ppb of AF and 50–350 ppb of FUM facilitated STEC-associated outbreaks. In addition, in vitro inclusion of 0.02 ppb of AF in the growth media of STECs resulted in higher cytotoxin production and cytotoxicity, highlighting the role of mycotoxins in STEC pathogenesis (Baines et al., 2013b).

In other studies, cattle exposed to mycotoxin mixtures were colonized by two or more STECs, suggesting that mycotoxins facilitate co-infection (Baines et al., 2011a; Baines et al., 2011b).

In lambs, experimental aflatoxicosis revealed that albumin and alfa-globulin levels were lower for intoxicated lambs than in the control group. Although beta-globulin concentrations did not change, increases in gamma-globulins levels in dosed lambs were observed throughout the experiment.

⇰ These results suggest that AF cause a failure in the lambs’ acquired immunity by decreasing antibody production and altering serum proteins (Fernández et al., 1997).

However, in another experiment, results indicate that a decline in the average daily gain was the most sensitive indicator of aflatoxicosis in lambs, showing that the altered immune response could render the animals more susceptible to infectious diseases (Fernández et al., 2000).

An experiment carried out by Dzidic et al. (2010) indicated that sheep fed 300 mg of mycophenolic acid/sheep/day from contaminated silage did not show any immunodepression effects

Although mycotoxins have been confirmed to modulate the effects of bacteria within animals and increase their virulence, it has also been suggested that strains of L. acidophilus CIP 76.13T and L. delbrueckii subsp. bulgaricus CIP 101027T may be included in feed to reduce mycotoxin contamination (Ragoubi et al., 2021).

As mycotoxin exposure can be detrimental to certain intestinal microbial populations, the use of beneficial bacteria can alleviate these effects. For example, Lactobacillus is a critical genus for detoxifying OTA in vivo (Guerre, 2020; Jin et al., 2021; Sun et al., 2023b).

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