Vaccination against Eimeria tenella infection using a fraction of E. tenella sporozoites selected by the capacity to activate T cells.

At 8 days after a primary Eimeria tenella infection, a subset of T cells, of which the protective role is as yet unclear, circulates in the peripheral blood. In order to investigate this, the in vitro cellular responsiveness of these peripheral blood lymphocytes has been used as selection criterion to identify potentially protective E. tenella sporozoite antigens. The hydrophilic protein phase of purified E. tenella sporozoite homogenates obtained by Triton X-114 extraction was fractionated using preparative gel electrophoresis. Nine fractions, separated according to different molecular weight, were tested for their ability to stimulate T-cell responses. Both the proliferation of peripheral blood lymphocytes and the macrophage activating activity released in the culture supernatants were measured. On the basis of this responsiveness, four fractions were selected and used to vaccinate chickens. All vaccine preparations induced strong T-cell responses. One fraction immunised chickens against subsequent challenge infection, in that the caecal lesion scores were significantly lower as compared with that of the unvaccinated controls. This fraction contained hydrophilic polypeptides with a molecular mass that ranged from 26 to 30 kDa.

Vaccination of broilers with HVT expressing an Eimeria acervulina antigen improves performance after challenge with Eimeria.

The sequence encoding part of the 100 kDa refractile body protein (Ea1A) from Eimeria acervulina was cloned into the US10 locus of herpesvirus of turkeys (HVT) downstream of LTR promoter. Expression of the fusion protein was shown in vitro. Recombinant HVT showed a delayed and slightly reduced level of viremia compared to the parent strain in SPF chickens as well as in broilers. Effect on the performance of broilers vaccinated with recombinant or parent HVT was measured by challenge at day 24 with a high dose of E. acervulina and E. maxima oocysts. A significant improvement in weight of animals vaccinated with the recombinant HVT was detected at the end of the challenge period.

Peripheral blood lymphocytes from Eimeria tenella infected chickens produce gamma-interferon after stimulation in vitro.

Protective immunity to infection by Eimeria parasites has been demonstrated to be dependent on T-cell mediated immune responses and may be associated with the release of cytokines. We have previously shown that the proportion of CD8-expressing T-cells in the peripheral blood of chicken increases transiently at 8 days after a primary infection with Eimeria tenella oocysts. The increase in the CD8+ population coincided with an increased proliferative lymphocyte response upon stimulation with E. tenella sporozoite antigen in vitro. In this study, we further investigated the functional activity of these peripheral blood leucocytes (PBL) by determining both the potential to proliferative and to produce IFN upon stimulation with E. tenella sporozoite antigens and mitogens. Enhanced proliferative responses to parasite antigen were accompanied by reduced responses to T-cell mitogens around 1 week of infection. The IFN activity in the supernatants of the stimulated PBL was measured by the ability to inhibit Semliki Forest Virus (SFV) replication in chicken embryo fibroblasts (CEF) and to activate macrophages, as measured by nitric oxide production. At eight days after infection the highest levels of virus inhibition and NO-production were detected upon stimulation with both E. tenella sporozoite antigen and mitogen. A strong correlation between the individual data of the two methods was found at this timepoint indicating that the produced cytokine was indeed IFN-gamma. These results suggest that around eight days after a primary E. tenella infection a parasite specific T-cell subset with the capacity of produce IFN(-gamma) is circulating which would be involved in the induction of protective immunity against Eimeria tenella.

Immunity to eimeria tenella in chickens: phenotypical and functional changes in peripheral blood T-cell subsets.

The changes in peripheral blood leukocyte (PBL) T-cell subsets following Eimeria tenella infection in outbred white leghorn chickens were studied, using a panel of murine monoclonal antibodies specific for the chicken homologues of the mammalian CD3, CD8, and CD4 markers on day-to-day samples of PBLs. Both flow cytometric analysis (FCA) and immunofluorescence microscopy with fixed cells on slides were used as read-out systems. The changes in the composition of the T-cell subsets measured with both techniques were similar. At 8 days post primary infection, a sharp transitory increase in the proportion of CD8-expressing cells was found. With FCA, CD8-expressing cells could be discriminated in CD8(Dim+) and CD8(Bright+) populations, which have not been described before. The proportion of CD4-expressing cells was decreased at days 9-10 after primary infection, which coincided with a less marked decrease in CD3-expressing cells. Such effects were not seen after secondary infection. When PBLs collected at day 8 post primary infection were stimulated in vitro with E. tenella sporozoite antigen, the response was higher than that in uninfected control chickens. The effects we observed coincide with the onset of recovery from primary infection. We speculate that the increase in CD8-expressing PBLs is the result of stimulation and expansion of a specific subset involved in the induction of protective immunity against Eimeria tenella.