Marion T. Ryan

Major Histocompatibility Complex DRB1 gene: its role in nematode resistance in Suffolk and Texel sheep breeds

A potential control strategy for nematode infection in sheep is the implementation of a breeding programme to select for genes associated with resistance. The Texel breed is more resistant to gastrointestinal nematode infection than the Suffolk breed, based on faecal egg count, and this difference should enable the identification of some of the genes responsible for resistance. The objective of this study was to determine if variation at the ovine MHC-DRB1 locus was associated with variation in faecal egg count in Suffolk and Texel sheep. Ovar-DRB1 alleles and faecal egg count were determined for Texel (n = 105) and Suffolk (n = 71) lambs. Eight Ovar-DRB1 alleles, including 1 previously unknown allele, were identified in the Texel breed by sequence-base-typing. Seven Ovar-DRB1 alleles were identified in the Suffolk breed. Two Ovar-DRB1 alleles were common to both breeds, but were among the least frequent in the Suffolk population. In the Suffolk breed 1 Ovar-DRB1 allele was associated with a decrease in faecal egg count and 2 alleles with an increase in faecal egg count. This locus accounted for 14% of the natural variation in faecal egg count in Suffolks. There was no evidence for an association between Ovar-DRB1 alleles and faecal egg count in the Texel breed and the Ovar-DRB1 locus accounted for only 3% of the phenotypic variation in faecal egg count. These results suggest that the Ovar-DRB1 gene plays an important role in resistance to nematode infection in the Suffolk breed. The difference in faecal egg counts between these breeds may be attributable in part to the different allele profile at the Ovar-DRB1 locus.

Effect of purified β-glucans derived from Laminaria digitata, Laminaria hyperborea and Saccharomyces cerevisiae on piglet performance, selected bacterial populations, volatile fatty acids and pro-inflammatory cytokines in the gastrointestinal tract of pigs.

β-Glucans have been identified as natural biomolecules with immunomodulatory activity. The first objective of the present study was to compare the effects of purified β-glucans derived from Laminaria digitata, L. hyperborea and Saccharomyces cerevisiae on piglet performance, selected bacterial populations and intestinal volatile fatty acid (VFA) production. The second aim was to compare the gene expression profiles of the markers of pro- and anti-inflammation in both unchallenged and lipopolysaccharide (LPS)-challenged ileal and colonic tissues. β-Glucans were included at 250 mg/kg in the diets. The β-glucans derived from L. hyperborea, L. digitata and S. cerevisiae all reduced the Enterobacteriaceae population (P<0·05) without influencing the lactobacilli and bifidobacteria populations (P>0·05) in the ileum and colon. There was a significant interaction between gastrointestinal region and β-glucan source in the expression of cytokine markers, IL-1α (<0·001), IL-10 (P<0·05), TNF-α (P<0·05) and IL-17A (P<0·001). β-Glucans did not stimulate any pro- or anti-inflammatory cytokine markers in the ileal epithelial cells. In contrast, the expression of a panel of pro- and anti-inflammatory cytokines (IL-1α, IL-10, TNF-α and IL-17A) was down-regulated in the colon following exposure to β-glucans from all the three sources. However, the data suggest that the soluble β-glucans derived from L. digitata may be acting via a different mechanism from the insoluble β-glucans derived from L. hyperborea and S. cerevisiae, as the VFA profile was different in the L. digitata-treated animals. There was an increase in IL-8 gene expression (P<0·05) in the gastrointestinal tract from the animals exposed to L. digitata following an LPS ex vivo challenge that was not evident in the other two treatment groups. In conclusion, β-glucans from both seaweed and yeast sources reduce Enterobacteriaceae counts and pro-inflammatory markers in the colon, though the mechanisms of action may be different between the soluble and insoluble fibre sources.

Development of Multiple-Locus Variable-Number Tandem-Repeat Analysis for the Molecular Subtyping of Enterobacter sakazakii

ABSTRACT The genomic content of Enterobacter sakazakii strain ATCC BAA-894 was analyzed for variable-number tandem repeats (VNTRs). In this study we report the development of a multiple-locus VNTR analysis (MLVA) strategy for the subtyping of E. sakazakii. The method is based on a GeneScan analysis of four VNTR loci labeled with multiple fluorescent dyes. This approach was applied to a collection of 112 isolates representing all 16 of the currently defined E. sakazakii biogroups. MLVA successfully discriminated among these isolates and compared favorably with pulsed-field gel electrophoresis. The method was relatively fast and easy to perform. The potential value of MLVA as an epidemiological tool is discussed.

Indices of gastrointestinal fermentation and manure emissions of growing-finishing pigs as influenced through singular or combined consumption of Lactobacillus plantarum and inulin.

Conventional dietary strategies to reduce pig manure odor may either be costly, or impede nutrient digestibility. Additionally, the response of manure odor to such measures may be variable, indicating a complex relationship between environmental pollutant and diet. We hypothesized that dietary Lactobacillus plantarum (LP), with or without the inclusion of a purified oligofructose (inulin), may reduce odor without compromising nutrient digestibility. An experiment with a 2 × 2 factorial arrangement of treatments was conducted to investigate effects of dietary inulin (0 and 12.5 g/kg) and LP (0 and 0.5 g/kg) on nutrient digestibility, indicators of gastrointestinal tract fermentation, select fecal bacteria, manure content, and ammonia and odor emissions of 28 growing-finishing pigs (60.3 kg; n = 7/treatment). Dietary treatments had no effect on nutrient digestibility. Dietary treatments containing inulin had decreased Enterobacteriaceae (8.60 vs. 9.67 log gene copy number/g fresh feces; P = 0.03) when compared with unsupplemented diets. There was an interaction between dietary inulin concentration and LP supplementation on estimates of fecal Clostridia (P = 0.01). Pigs offered diets containing both inulin and LP in combination had increased Clostridia when compared with those offered the control diet. However, there was no effect of either LP or inulin fecal Clostridia when offered singularly. An interaction was also noted where diets supplemented with LP or inulin only reduced odor (P = 0.01) compared with the control diet. However, there was no effect of LP on manure odor emissions when offered in combination with inulin. In summary, this study demonstrated that dietary supplementation with either exogenous LP or inulin reduces manure odor but not when offered in combination.

SNP variation in the promoter of the PRKAG3 gene and association with meat quality traits in pig

BackgroundThe PRKAG3 gene encodes the γ3 subunit of adenosine monophosphate activated protein kinase (AMPK), a protein that plays a key role in energy metabolism in skeletal muscle. Non-synonymous single nucleotide polymorphisms (SNPs) in this gene such as I199V are associated with important pork quality traits. The objective of this study was to investigate the relationship between gene expression of the PRKAG3 gene, SNP variation in the PRKAG3 promoter and meat quality phenotypes in pork.ResultsPRKAG3 gene expression was found to correlate with a number of traits relating to glycolytic potential (GP) and intramuscular fat (IMF) in three phenotypically diverse F1 crosses comprising of 31 Large White, 23 Duroc and 32 Pietrain sire breeds. The majority of associations were observed in the Large White cross. There was a significant association between genotype at the g.-311A>G locus and PRKAG3 gene expression in the Large White cross. In the same population, ten novel SNPs were identified within a 1.3 kb region spanning the promoter and from this three major haplotypes were inferred. Two tagging SNPs (g.-995A>G and g.-311A>G) characterised the haplotypes within the promoter region being studied. These two SNPs were subsequently genotyped in larger populations consisting of Large White (n = 98), Duroc (n = 99) and Pietrain (n = 98) purebreds. Four major haplotypes including promoter SNP’s g.-995A>G and g.-311A>G and I199V were inferred. In the Large White breed, HAP1 was associated with IMF% in the M. longissmus thoracis et lumborum (LTL) and driploss%. HAP2 was associated with IMFL% GP-influenced traits pH at 24 hr in LTL (pHULT), pH at 45 min in LTL (pH45LT) and pH at 45 min in the M. semimembranosus muscle (pH45SM). HAP3 was associated with driploss%, pHULT pH45LT and b* Minolta. In the Duroc breed, associations were observed between HAP1 and driploss% and pHUSM. No associations were observed with the remaining haplotypes (HAP2, HAP3 and HAP4) in the Duroc breed. The Pietrain breed was monomorphic in the promoter region. The I199V locus was associated with several GP-influenced traits across all three breeds and IMF% in the Large White and Pietrain breed. No significant difference in promoter function was observed for the three main promoter haplotypes when tested in vitro.ConclusionGene expression levels of the porcine PRKAG3 are associated with meat quality phenotypes relating to glycolytic potential and IMF% in the Large White breed, while SNP variation in the promoter region of the gene is associated with PRKAG3 gene expression and meat quality phenotypes.

Anti‐inflammatory effects of a casein hydrolysate and its peptide‐enriched fractions on TNFα‐challenged Caco‐2 cells and LPS‐challenged porcine colonic explants

Bioactive milk peptides are reported to illicit a range of physiological benefits and have been proposed as potential functional food ingredients. The objective of this study was to characterize the anti-inflammatory properties of sodium caseinate (NaCAS), its enzyme hydrolysate (EH) and peptide-enriched fractions (5 kDa retentate [R], 1 kDaR and 1 kDa permeate [P]), both in vitro using a Caco-2 cell line, and also ex vivo using a porcine colonic tissue explant system. Caco-2 cells were stimulated with tumour necrosis factor alpha (TNFα) and co-treated with casein hydrolysates for 24 h. Following this, interleukin (IL)-8 concentrations in the supernatant were measured using enzyme-linked immunosorbent assay. Porcine colonic tissue was stimulated with lipopolysaccharide and co-treated with casein hydrolysates for 3 h. The expression of a panel of inflammatory cytokines was measured using qPCR. While dexamethasone reduced the IL-8 concentration by 41.6%, the 1 kDaR and 1 kDaP fractions reduced IL-8 by 68.7% and 66.1%, respectively, relative to TNFα-stimulated Caco-2 cells (P < 0.05). In the ex vivo system, only the 1 kDaR fraction elicited a decrease inIL1-α,IL1-β,IL-8,TGF-β andIL-10 expression (P < 0.05). This study provides evidence that the bioactive peptides present in the 1 kDaR fraction of the NaCAS hydrolysate possess anti-inflammatory properties in vitro and ex vivo. Further in vivo analysis of the anti-inflammatory properties of the 1 kDaR is proposed.

Molecular characterization of Irish E. coli O157:H7 isolates of human, bovine, ovine and porcine origin.

Aims: To determine the degree of relatedness between isolates of Escherichia coli O157:H7 of human, bovine, ovine and porcine origin.

Effects of dietary supplementation with Laminaria hyperborea, Laminaria digitata, and Saccharomyces cerevisiae on the IL-17 pathway in the porcine colon

An unregulated T(h)17 inflammatory response has been highlighted as a major contributor to the underlying pathology of inflammatory bowel diseases (IBD) whereas regulatory T (T(REG)) cells) have been highlighted as pivotal in suppressing autoimmune and inflammatory responses in the gut. Following dietary supplementation, β-glucans have been shown to reduce the T(h)17 signature molecule IL-17a in the porcine colon. To expand this observation we examined the effects of supplementing feeds with β-glucans derived from seaweeds Laminaria hyperborea and Laminaria digitata and the yeast Saccharomyces cerevisiae on gene expression of a range of cytokines, receptors, and signal transducing molecules relevant to the T(h)17 and T(REG) pathways in the porcine colon. All sources of β-glucans significantly decreased the expression of T(h)17-related cytokines (IL-17a, IL-17F, and IL-22), receptor IL23R, and IL-6. There was no alteration to the T(REG)-related target, Foxp3, or to TGF-β, although a significant reduction in IL-10 was observed in the L. digitata supplementation group.

Immunogenomics of gastrointestinal nematode infection in ruminants - Breeding for resistance to produce food sustainably and safely.

Gastrointestinal nematode (GIN) infection of ruminants represents a major health and welfare challenge for livestock producers worldwide. The emergence of anthelmintic resistance in important GIN species and the associated animal welfare concerns have stimulated interest in the development of alternative and more sustainable strategies aimed at the effective management of the impact of GINs. These integrative strategies include selective breeding using genetic/genomic tools, grazing management, biological control, nutritional supplementation, vaccination and targeted selective treatment. In this review, the logic of selecting for “resistance” to GIN infection as opposed to “resilience” or “tolerance” is discussed. This is followed by a review of the potential application of immunogenomics to genetic selection for animals that have the capacity to withstand the impact of GIN infection. Advances in relevant genomic technologies are highlighted together with how these tools can be advanced to support the integration of immunogenomic information into ruminant breeding programmes.

Effects of dietary β-glucans supplementation on cytokine expression in porcine liver.

As dietary supplementation with β-glucans can stimulate the innate immune response in the porcine gastrointestinal system (GIT), the aim of this study was to determine if the effects of β-glucan supplementation extend beyond the GIT to systemic levels. Hence, the effects of dietary supplementation of β-glucans derived from Laminara digitata, Laminara hyperborea, and Sacharomyces cerevisiae on cytokine expression in the porcine liver with or without ex vivo lipopolysaccharide (LPS) challenge were examined. No significant differences in gene expression were observed in the unchallenged liver tissue, but differences were observed in all supplementation groups in the LPS challenged tissue. Relative to the basal diet, IL-6 (P < 0.05) was less expressed in the S. cerevisiae supplementation group, IL-6 (P < 0.05) and TLR-4 (P < 0.05) were less expressed in the L. digitata supplementation group, and IL-10 (P = 0.06) and IL-1α (P = 0.02) were more expressed in the L. hyperborea supplementation group. There was a ≈ 3-fold increase in both IL-10 and IL-1α in the liver samples of L. hyperborea relative to the L. digitata supplementation groups (P < 0.01). The results indicate that supplementation with β-glucans from both yeast and seaweed sources have systemic effects evidenced by changes in cytokine expression in the liver in response to LPS challenge; however, the cytokines affected varied according to the source of the β-glucan.