Effects of avian triggering receptor expressed on myeloid cells (TREM-A1) activation on heterophil functional activities.

A class of innate receptors called the triggering receptors expressed on myeloid cells (TREM) has been discovered and shown to be involved in innate inflammatory responses. The TREM family has been found in the chicken genome and consists of one activating gene (TREM-A1) and two inhibitory genes (TREM-B1 and TREM-B2). However, to date, there have been no reports on the effects of activating the TREM molecules on the functional activity of the primary avian polymorphonuclear cell, the heterophil. To characterize the activation of avian heterophils, we evaluated the effect of receptor ligation on heterophil effector functions. A specific agonistic antibody (Ab) was generated against the peptide sequence of chicken TREM-A1 38-51aa (YNPRQQRWREKSWC). To study TREM-A1 mediated activation, purified peripheral blood heterophils were incubated with various concentrations of the anti-TREM-A1 Ab or control Ab against an irrelevant antigen. Activation via TREM-A1 induces a significant increase in phagocytosis of Salmonella enteritidis, a rapid degranulation, and a dramatic up-regulation in gene expression of the pro-inflammatory cytokine, IL-6, and the inflammatory chemokine, CXCLi2. However, we found no direct TREM-A1 stimulation of the heterophil oxidative burst. Like mammalian TREM, avian TREM-A1 ligation synergizes with the activation of Toll-like receptor-4 (TLR4) ligand, LPS. In addition, the synergistic activity of LPS and TREM-A1 resulted in a significantly (p⩽0.05) increased production of an oxidative burst. Taken together, these results suggest, unlike in mammalian neutrophils, TREM-A1 engagement activates a differential functional activation of avian heterophils, but like mammalian neutrophils, acts in synergy with TLR agonists. These results provide evidence of the function of TREM-A1 in heterophil biology and avian innate immunity.

Evaluation of the effect of gallium maltolate on fecal Salmonella shedding in cattle.

Strategies aimed at reducing fecal shedding of Salmonella and other foodborne pathogens may be effective for limiting transmission of pathogens from food animals to humans. The objective of this study was to determine the effectiveness of gallium maltolate (GaM) against Salmonella in vitro and to determine whether oral administration of GaM would reduce fecal shedding of Salmonella in cattle. Gallium is a semimetal exhibiting antimicrobial properties against some pathogenic bacteria, including Salmonella, by exploiting their need for iron to survive and replicate. In vitro growth studies were performed in pure cultures of Salmonella and in mixed cultures from ruminal fluid. Inclusion of GaM in culture medium or in mixed cultures of ruminal fluid resulted in a significant reduction in growth of Salmonella, suggesting that GaM may be effective for limiting growth and survival in vivo. Therefore, we subsequently administered two doses of GaM to Holstein steers, experimentally infected them with Salmonella, and quantitatively and qualitatively monitored fecal shedding at 12-h intervals. Sixty hours after beginning treatment, cattle were euthanized, and luminal contents and tissue were aseptically harvested from the rumen, jejunum, spiral colon, cecum, and rectum. The luminal contents were processed for quantitative and qualitative analysis of the challenge strains of Salmonella, and tissue samples were enriched and plated for qualitative analysis. We found no significant differences between control and treated animals in quantitative levels of Salmonella in the feces or the luminal contents. Likewise, we observed no pattern between control and treated animals in the frequency of positive or negative results from enriched feces, luminal contents, or tissue samples. These results suggest that GaM was not effective for reducing Salmonella in cattle.

Expression profile of toll-like receptors within the gastrointestinal tract of 2-day-old Salmonella enteriditis-infected broiler chickens.

Salmonella enterica serovar Enteriditis (SE) causes a majority of foodborne illness in the U.S. A more productive avian innate immune response could reduce bacterial colonization and the incidence of infection in humans. However, quantification and comparison of the toll-like receptors (TLR), a component of the innate immune system that recognize bacterial pathogens, and their response to SE colonization across the avian gastrointestinal (GI) tract has not been reported. Therefore, we assessed these changes using real-time qRT-PCR to measure expression of TLR 1LA, 2A, 2B, 3, 4, 5, 7, 15, and 21 in the duodenum, jejunum, ileum, cecal tonsil, ceca, and large intestine of uninfected and SE-infected 2-day-old broiler chickens. Samples were collected soon after hatch to approximate natural SE exposure and to measure initial changes in the immune response to infection. All TLRs had measurable expression within the duodenum, jejunum, ileum, cecal tonsil, ceca, and large intestine. The general expression pattern, with the exception of TLR 21, showed distal GI segments had higher TLR mRNA expression than proximal segments. Infected chickens had increased expression of TLR 1LA, 2A, 4, and 15 in distal GI segments and upregulation of TLR 2B, 3, and 15 in proximal segments, including the duodenum. Interestingly, SE-infection caused downregulation of TLR 5, with no change in TLR 7 or 21. Overall, we provide a comprehensive report of mRNA expression profiles for the TLR family of innate immune receptors in the GI tract of 2-day-old broilers and their differential response to SE colonization.

In ovo treatment with CpG oligodeoxynucleotides decreases colonization of Salmonella enteriditis in broiler chickens.

Induction of the innate immune response in newly hatched chickens is important for limiting infections with bacteria, such as Salmonella enterica serovar Enteriditis (SE). CpG oligodeoxynucleotides (CpG-ODN) can stimulate the innate immune response of young chickens. Therefore, we examined the effectiveness of CpG-ODN administered in ovo on intestinal colonization by SE and the ability to modulate the function of heterophils in young chickens. Heterophils were isolated from 2-day-old chickens and were stimulated with heat-killed SE (HK-SE) or PMA for oxidative burst and HK-SE or live SE for degranulation assays. CpG-ODN treatment had no effect on heterophil oxidative burst when stimulated with HK-SE or PMA. However, HK-SE and live SE increased degranulation (P<0.01) in heterophils from CpG-ODN-treated birds compared to PBS-treated controls. In a second experiment, chickens were orally infected with SE on day 10 post-hatch and cecal contents were collected 6 days later for assessment of SE intestinal colonization. CpG-ODN treatment reduced SE colonization by greater than 10-fold (P<0.001) compared to PBS-injected control birds. Overall, we show for the first time that CpG-ODN given in ovo stimulates innate immune responsiveness of chicken heterophils and increases resistance of young chickens to SE colonization.