Ostertagia ostertagi Mediates Early Host Immune Responses via Macrophage and Toll-Like Receptor Pathways

ABSTRACT Ostertagia ostertagi is an abomasal parasite with significant economic impact on the cattle industry. Early host immune responses are poorly understood. Here, we examined time course expression of Toll-like receptors (TLRs) in peripheral blood mononuclear cells (PBMC) during infection where PBMC macrophages (Mϕ) generated both pro- and anti-inflammatory responses when incubated with excretory/secretory products (ESP) from fourth-stage larvae (OoESP-L4) or adult worms (OoESP-Ad). First, changes in cell morphology clearly showed that both OoESP-L4 and OoESP-Ad activated PBMC-Mϕ in vitro, resulting in suppressed CD40 and increased CD80 expression. Expression of mRNAs for TLR1, -4, -5, and -7 peaked 7 days postinfection (dpi) (early L4), decreased by 19 dpi (postemergent L4 and adults) and then increased at 27 dpi (late adults). The proinflammatory cytokine tumor necrosis factor alpha (TNF-α) (transcript and protein) increased in the presence of OoESP-Ad, and the anti-inflammatory cytokine interleukin 10 (IL-10) (protein) decreased in the presence of OoESP-L4 or OoESP-Ad; however, IL-10 mRNA was upregulated, and IL-6 (protein) was downregulated by OoESP-L4. When PBMC-Mϕ were treated with ligands for TLR4 or TLR5 in combination with OoESP-Ad, the transcripts for TNF-α, IL-1, IL-6, and IL-10 were significantly downregulated relative to treatment with TLR4 and TLR5 ligands only. However, the effects of TLR2 ligand and OoESP-Ad were additive, but only at the lower concentration. We propose that O. ostertagi L4 and adult worms utilize competing strategies via TLRs and Mϕ to confuse the immune system, which allows the worm to evade the host innate responses.

Effect of Mixed Invasions of Hypoderma bovis and Ostertagia ostertagi in Cattle on Milk Yield and Contents in Polish Dairy Farms

Wide distribution of ecto- and endoparasites in cattle is a serious problem in the sustainability of a farm, due to the negative impact on animals’ health and productivity. The aim of this study was to determine the presence of antibodies against Ostertagia and Hypoderma in udder milk samples and the comparison of milk yield and content of the basic components of milk in ELISA-positive and ELISA-negative cows. Milk samples were collected from 148 lactating cows from 3 herds. Antibody detection was performed using specific ELISAs for Ostertagia ostertagi and Hypoderma bovis. Milk yield and content of protein, fat, and dry matter were studied in samples from each individual cow 11 times per year at 4 week intervals. The extensiveness of dual parasitic invasions in individual herds, estimated on the basis of udder milk testing with the ELISA test, varied and amounted to 3.22%, 11.36%, and 4.76% in the three studied herds, respectively. No antibodies were found in 61.2%, 22.7%, and 57.1% of the milk samples from the cows in each herd. The milk yield of ELISA-positive cows was significantly lower in comparison to the efficiency of ELISA-negative cows and amounted to 294 kg and even to 3672 kg of milk per year, per cow. No significant differences were found between the fat and protein contents of milk between ELISA-positive and -negative cows for both parasites.

Characterization of IL-10-producing neutrophils in cattle infected with Ostertagia ostertagi

IL-10 is a master regulator of immune responses, but its cellular source and function in cattle during the initial phase of immune priming have not been well established. Despite a massive B cell response in the abomasal draining lymph nodes in Ostertagia ostertagi (OO)-infected cattle, protective immunity is slow to develop, and partial protection requires years of repeated exposure. In addressing this problem, our initial hypothesis was that B cells produce IL-10 that downregulates the host protective immune response. However, our results showed that neutrophils made up the majority of IL-10-producing cells in circulation and in secondary lymphoid tissues, particularly the spleen (80%). Conversely, IL-10-producing B cells were rare. In addition, approximately 10% to 20% of the neutrophils in the blood and spleen expressed MHC II and were IL-10 negative, suggesting that neutrophils could also participate in antigen presentation. In vitro investigation of bovine neutrophils revealed that exposure thereof to OO extract increased IL-10 and MHC II expression in these cells in a dose-dependent manner, consistent with IL-10+/MHC II+ neutrophils detected in cattle shortly after experimental OO infection. Co-culture of untreated neutrophils with anti-CD3 antibody (Ab)-stimulated CD4+ T cells led to enhanced T cell activation; also, IL-10 depletion with neutralizing Ab enhanced the stimulatory function of neutrophils. OO extract depressed neutrophil stimulation of CD4+ T cells in the presence of IL-10-neutralizing Ab, suggesting that OO utilizes both IL-10-dependent and independent mechanisms to manipulate the bovine immune response. Finally, contact and viability were required for T cell-stimulatory neutrophil function. This report, to the best of our knowledge, is the first to demonstrate that neutrophil-derived IL-10 is directly involved in T cell regulation in cattle. Our data suggest that neutrophils and neutrophil-derived IL-10 are co-opted by nematode parasites and other pathogens to attenuate host immune responses and facilitate pathogen survival.

Electrochemical enzyme-linked immunosorbent assay (e-ELISA) for parasitic nematode Ostertagia ostertagi (brown stomach worm) infections in dairy cattle

A rapid and sensitive electrochemical immunoassay (e-ELISA) for the detection of the gastrointestinal parasitic nematode Ostertagia ostertagi (brown stomach worm) in serum samples is presented.