Comparative Assessment of Different Yeast Cell Wall-Based Mycotoxin Adsorbents Using a Model- and Bioassay-Based In Vitro Approach.

Frequently reported occurrences of deoxynivalenol (DON), beauvericin (BEA), and, to a lesser extent, ochratoxin A (OTA) and citrinin (CIT) in ruminant feed or feedstuff could represent a significant concern regarding feed safety, animal health, and productivity. Inclusion of yeast cell wall-based mycotoxin adsorbents in animal feeds has been a common strategy to mitigate adverse effects of mycotoxins. In the present study, an in vitro approach combining adsorption isotherm models and bioassays was designed to assess the efficacy of yeast cell wall (YCW), yeast cell wall extract (YCWE), and a postbiotic yeast cell wall-based blend (PYCW) products at the inclusion rate of 0.5% (w/v) (ratio of adsorbent mass to buffer solution volume). The Hill's adsorption isotherm model was found to best describe the adsorption processes of DON, BEA, and CIT. Calculated binding potential for YCW and YCWE using the Hill's model exhibited the same ranking for mycotoxin adsorption, indicating that BEA had the highest adsorption rate, followed by DON and CIT, which was the least adsorbed. PYCW had the highest binding potential for BEA compared with YCW and YCWE. In contrast, the Freundlich isotherm model presented a good fit for OTA adsorption by all adsorbents and CIT adsorption by PYCW. Results indicated that YCW was the most efficacious for sequestering OTA, whereas YCWE was the least efficacious. PYCW showed greater efficacy at adsorbing OTA than CIT. All adsorbents exhibited high adsorption efficacy for BEA, with an overall percentage average of bound mycotoxin exceeding 60%, whereas moderate efficacies for the other mycotoxins were observed (up to 37%). Differences in adsorbent efficacy of each adsorbent significantly varied according to experimental concentrations tested for each given mycotoxin (p < 0.05). The cell viability results from the bioassay using a bovine mammary epithelial cell line (MAC-T) indicated that all tested adsorbents could potentially mitigate mycotoxin-related damage to bovine mammary epithelium. Results from our studies suggested that all tested adsorbents had the capacity to adsorb selected mycotoxins in vitro, which could support their use to mitigate their effects in vivo.

In-Vitro Cell Culture for Efficient Assessment of Mycotoxin Exposure, Toxicity and Risk Mitigation.

Mycotoxins are toxic secondary fungal metabolites that commonly contaminate crops and food by-products and thus, animal feed. Ingestion of mycotoxins can lead to mycotoxicosis in both animals and humans, and at subclinical concentrations may affect animal production and adulterate feed and animal by-products. Mycotoxicity mechanisms of action (MOA) are largely unknown, and co-contamination, which is often the case, raises the likelihood of mycotoxin interactions. Mitigation strategies for reducing the risk of mycotoxicity are diverse and may not necessarily provide protection against all mycotoxins. These factors, as well as the species-specific risk of toxicity, collectively make an assessment of exposure, toxicity, and risk mitigation very challenging and costly; thus, in-vitro cell culture models provide a useful tool for their initial assessment. Since ingestion is the most common route of mycotoxin exposure, the intestinal epithelial barrier comprised of epithelial cells (IECs) and immune cells such as macrophages, represents ground zero where mycotoxins are absorbed, biotransformed, and elicit toxicity. This article aims to review different in-vitro IEC or co-culture models that can be used for assessing mycotoxin exposure, toxicity, and risk mitigation, and their suitability and limitations for the safety assessment of animal foods and food by-products.

Assessment of yeast Saccharomyces cerevisiae component binding to Mycobacterium avium subspecies paratuberculosis using bovine epithelial cells.

BACKGROUND: Since yeast Saccharomyces cerevisiae and its components are being used for the prevention and treatment of enteric diseases in different species, they may also be useful for preventing Johne's disease, a chronic inflammatory bowel disease of ruminants caused by Mycobacterium avium spp. paratuberculosis (MAP). This study aimed to identify potential yeast derivatives that may be used to help prevent MAP infection. The adherence of mCherry-labeled MAP to bovine mammary epithelial cell line (MAC-T cells) and bovine primary epithelial cells (BECs) co-cultured with yeast cell wall components (CWCs) from four different yeast strains (A, B, C and D) and two forms of dead yeast from strain A was investigated. RESULTS: The CWCs from all four yeast strains and the other two forms of dead yeast from strain A reduced MAP adhesion to MAC-T cells and BECs in a concentration-dependent manner after 6-h of exposure, with the dead yeast having the greatest effect. CONCLUSIONS: The following in vitro binding studies suggest that dead yeast and its' CWCs may be useful for reducing risk of MAP infection.

Assessment of bovine mammary chemokine gene expression in response to lipopolysaccharide, lipotechoic acid + peptidoglycan, and CpG oligodeoxynucleotide 2135.

During intramammary infections pathogen associated molecular patterns (PAMPs) induce an inflammatory response, recognized clinically as mastitis. Recognition of PAMPs by mammary cells leads to the production of the pro-inflammatory cytokines, TNF-alpha and IL-1beta. These cytokines augment the secretion of various chemokines that are responsible for directing the host cellular immune response, and consequently the outcome of infection. Previous research has shown that gram-negative and gram-positive bacteria elicit different types of innate immune responses. The purpose of this study, therefore, was to characterize the expression of various chemokine genes in bovine mammary gland explants in response to lipopolysaccharide (LPS), peptidoglycan (PTG) combined with lipotechoic acid (LTA), and CpG oligodeoxynucleotide (CpG-ODN) 2135 representing gram-negative bacteria, gram-positive bacteria, and bacterial DNA, respectively, to determine if these PAMPs induce different chemokine gene expression patterns. Explants from 3 Holstein cows were cultured with 10 microg/mL of LPS, LTA + PTG, or CpG-ODN 2135 for 6 and 24 h. Total RNA was extracted and the expression of CXCL8, MCP-1, MCP-2, MCP-3, MIP1-alpha, and RANTES genes was measured by real-time polymerase chain reaction (RT-PCR). Lipopolysaccharide significantly induced MCP-1, MCP-2, and MCP-3 expression, and slightly increased CXCL8 gene expression. The combined PAMPs, LTA + PTG, on the other hand, significantly induced MCP-1 gene expression, and slightly increased MCP-3 expression. No significant expression differences for any of the chemokine genes were observed in explants stimulated with CpG-ODN 2135. These results demonstrate that PAMPs associated with different mastitis-causing pathogens induce chemokine-specific gene expression patterns that may contribute to different innate immune responses to bacteria.

Assessment of the ovine acute phase response and hepatic gene expression in response to Escherichia coli endotoxin.

Lipopolysaccharide (LPS), a bacterial membrane endotoxin, induces a systemic inflammatory response (IFR) through the activation of blood monocytes and hepatic kupffer cells. These cells secrete pro-inflammatory cytokines, which subsequently activate the hypothalamic-pituitary-adrenal axis (HPAA) to release cortisol, an anti-inflammatory hormone that regulates the IFR and subsequent immune response (IR). The intent of this study was to characterize the acute phase response in female sheep challenged systemically with a range of doses of Escherichia coli endotoxin. Yearling ewes were challenged with an i.v. bolus dose of LPS (0, 200, 400, 600 ng/kg BW) and the acute phase response assessed by measuring serum interleukin (IL)-6 and cortisol concentrations, and the febrile response over time. A follow-up liver biopsy study was performed to determine kinetic differences in the expression of eight candidate hepatic genes between LPS dose groups using real-time RT-PCR. The initial time trail did not follow a linear dose response relationship with respect to the febrile and HPAA response to LPS challenge. Serum IL-6 concentrations increased in the two highest treatment groups but did not correlate with the observed febrile and HPAA response. The expression of Toll-like receptor 4, CD14, IL-6, tumor necrosis factor-alpha, IL-1beta, macrophage migration inhibitory factor, 11-beta-hydroxysteroid dehydrogenase (HSD), and tachykinin precursor 1 hepatic genes was dependent on both the dose and the kinetics of the response to LPS.