The effect of ovine peripheral blood mononuclear cells on Haemonchus contortus larval morbidity in vitro.

The objective of this study was to determine the effects of peripheral blood mononuclear cells (PBMC), derived from parasite-resistant St. Croix (STC) hair sheep and parasite-susceptible Suffolk (SUF) sheep, on Haemonchus contortus L3 stage larval death in vitro, with or without autologous serum. Larval morbidity was quantified by measuring larval ATP concentration following incubation with PBMC. Larvae exposed to either STC- or SUF-derived PBMC had lower ATP than live larvae (0.12 µmol/L ATP and 0.16 µmol/L ATP vs 0.27 µmol/L ATP, respectively) (P<.001) and greater ATP of dead larvae (0.03 µmol/L ATP) (P<.001). Breed differences were observed with addition of autologous serum. Larvae exposed to SUF-derived PBMC with autologous serum were not significantly different from live larval ATP. STC-derived serum did not significantly reduce larval ATP compared to PBMC alone (0.11 µmol/L ATP), but was significantly reduced compared to live larvae (0.22 µmol/L ATP) and SUF-derived PBMC with autologous serum (0.23 µmol/L ATP) (P<.001). These data indicate that a cellular response alone is capable of significantly reducing larval ATP in a breed-independent manner. However, addition of serum to SUF-PBMC failed to reduce larval ATP, indicating breed-dependent humoral response to H. contortus.

Serum-mediated Haemonchus contortus larval aggregation differs by larval stage and is enhanced by complement.

The objectives of this study were to measure Haemonchus contortus larval aggregation by complement/antibody complexes, determine effect of breed resistance and infection status and determine the effect of larval maturation on larval aggregation in vitro. Larval binding assays were performed on H. contortus L3, exsheathed L3 and L4 incubated with serum from either parasite naïve or H. contortus primed St. Croix (resistant) and Suffolk (susceptible) lambs. No differences in L3 aggregation were observed between serum from either breed or infection status. Exsheathed L3 (60%) and L4 (42%) aggregation by primed Suffolk serum was significantly reduced compared with L3 (80%, P<.001). Removal of either complement or antibody effectively eliminated L3 aggregation (P<.001). Combination of antibody-depleted and complement-inactivated serum restored L3 aggregation to levels consistent with unprocessed serum, supporting a role for antibody and complement in aggregation (P<.001). Use of fluorescence-labelled anti-sheep IgG antibody allowed documentation of IgG bound to serum complexes within L3 masses and was present only in larvae incubated with normal serum, and complement- and antibody-depleted serum combination. These data indicate that complement/antibody complexes inhibit larval motility through larval aggregation which may be critical in early larval clearance of H. contortus.

Early IL-4 gene expression in abomasum is associated with resistance to Haemonchus contortus in hair and wool sheep breeds.

Early immune events associated with reduced larval burden remain unclear in parasite-resistant breeds of sheep. Therefore, our objective was to determine breed differences in immune-related gene expression following infection with H. contortus. Gene expression in abomasal tissue and mucosa and in abomasal lymph nodes (ALN) was measured in 24 St. Croix (hair) lambs and 24 Dorset x (Finn-Rambouillet) (wool) lambs at 0 (uninfected), 3, 5 and 7 days after infection with 10 000 L3 H. contortus larvae. Expression of IL-4 in abomasal mucosa was detected on day 3 and increased to day 7 in hair lambs, but was not detectable in wool lambs. Genes that recruit neutrophils (CXCL1) and macrophages (MCP1) were upregulated in abomasal mucosa of hair lambs. Genes associated with alternative macrophage activation (ARG-1) and eosinophil activation (Gal-14) were also upregulated in the abomasal mucosa of hair lambs. Tissue remodeling genes (MMP13, PDGF) and tumour necrosis factor alpha (TNF-α) and MCP1 were upregulated in abomasal tissue of wool lambs; these lambs also had greater expression of forkhead box P3 in ALN. These data indicate a role for early IL-4 expression locally and demonstrate potential downregulation of immunity in wool sheep that could facilitate establishment of H. contortus.

Effects of peripheral blood mononuclear cells on Haemonchus contortus larval motility in vitro.

The objective of this experiment was to determine effects of peripheral blood mononuclear cells (PBMC), derived from parasite-resistant St. Croix (STC) and parasite-susceptible Suffolk (SF) sheep, on motility of Haemonchus contortus L3 stage larvae in vitro. Peripheral blood mononuclear cells were collected from 10 lambs of each breed, 5 naïve and 5 which had received a priming infection with H. contortus. Larval motility was quantified using a MIF Nikon Sweptfield microscope and NIS Elements AR software, and measurements included path length (PL) (µm), velocity (VEL) (µm/s) and acceleration (ACC) (µm/s2 ). After 18 h of incubation, PL and VEL were greatest in larvae cultured with SF-derived PMBC and were significantly different from all other groups (P < 0·01). No difference was observed in PL or VEL between larvae exposed to naïve or primed STC-derived PBMC and primed-SF PBMC. Differences in ACC were detected between larvae cultured with primed STC-derived PBMC (10·91 µm/s2 ) and naïve SF-derived PBMC (45·7 µm/s2 ) (P = 0·035). These data indicate an innate ability of STC-derived PBMC to severely inhibit larval motility.

Gene expression profiles of hair and wool sheep reveal importance of Th2 immune mechanisms for increased resistance to.

Management of gastrointestinal parasites is a critical issue for sheep producers worldwide. Increases in the prevalence of drug-resistant worms have complicated parasite control and increased economic losses. Therefore, other methods of parasite control need to be assessed, including the use of genetically resistant animals in breeding programs. Hair sheep breeds such as the St. Croix have greater parasite resistance than conventional wool breeds. However, the immune mechanisms that control parasite resistance in hair or wool breeds have not yet been fully determined, and information on cytokine expression profiles for both wool sheep selected for increased resistance and hair sheep is limited. Our objective was to investigate gene expression differences in 24 parasite-resistant hair and 24 susceptible wool sheep to identify immune effectors associated with resistance to . One-half of the lambs were infected and sacrificed at 3 or 27 d after infection. Remaining lambs were not infected. Breed differences in expression of genes associated with Th1 and Th2 immune responses in lymph nodes and abomasal tissue were determined. Th2-associated genes included IL-4, IL-13, IL-5, IgE, the α chain of the IL-4 receptor, and the α chain of the high-affinity IgE receptor (FcεRI). Th1-associated genes included interferon gamma (IFN-γ), the p35 subunit of IL-12 (IL-12 p35), and the ß1 and ß2 chains of the IL-12 receptor (IL-12 Rß1 and IL-12 Rß2, respectively). In both hair and wool sheep, infection with resulted in greater expression of IgE, IL-13, IL-5, and IL-12 p35 and somewhat reduced expression of IFNγ in lymph nodes. In abomasal tissue, parasite infection resulted in greater IgE, IL-13, FcεRI, and IL-12 p35 expression in infected lambs compared with control lambs. Between breeds, hair sheep had a stronger Th2 response after infection than wool sheep, with increased expression of IgE and IL-13 and decreased expression of IFNγ in lymph nodes and increased expression of IL-13 and decreased expression of IL-12 p35 in abomasal tissue. Expression of IL-4 in lymph nodes did not differ between hair and wool lambs, and IL-4, IL-5, IL-12 Rß1, and IL-12 Rß2 expression was too low to measure at the times sampled in abomasal tissue.