Immune responses during pregnancy in heifers naturally infected with Neospora caninum with and without immunization.
ABSTRACT This study was designed to identify changes in parasite-specific immune responses that occur during vertical transmission of Neospora caninum and can be used as indicators of parasite reactivation in naturally infected heifers. Ten heifers were unimmunized and 11 immunized with killed tachyzoites. One unimmunized heifer, which aborted at week 19 of gestation, had an increase in parasite-specific antibodies, mainly IgG(2), from week 15 to week 19 and a concomitant decline in parasite-specific cell-mediated immune (CMI) responses. Eight unimmunized heifers, which had live full-term congenitally infected calves, had an increase in antibodies, mainly IgG(2), from week 21 onwards. All immunized heifers delivered live full-term congenitally infected calves, and had a bimodal increase in antibodies; primarily IgG(1) following immunization and predominantly IgG(2) from week 17 onwards. Immunized heifers had significantly greater overall mean humoral and CMI responses than unimmunized heifers. Nine uninfected control heifers and their calves were seronegative. These results indicate that reactivation of a latent infection occurred in the naturally infected heifers, regardless of their immunization status, and was associated with increased parasite-specific antibodies, mainly IgG(2).
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ABSTRACT: The aim of this study was to evaluate whether Neospora caninum tachyzoites (Nc-1) inoculated by the conjunctival route in pregnant cows were able to generate infection in their fetuses. Group 1 contained 2 naturally infected cows; group 2 contained two cows inoculated intravenously with 2.5×10(8) tachyzoites, group 3 contained two cows inoculated with 2.5×10(8) tachyzoites by the conjunctival route, and group 4 contained two uninfected control cows. The four inoculated cows from groups 2 and 3 were challenged at 23 weeks of gestation. An indirect fluorescent antibody test (IFAT), recombinant NcGRA7-based ELISA, ELISA for IgG subisotypes and Western blot analysis were assessed to characterize the humoral immune response in dams. Sera from their fetuses were tested also using Western blot analysis. Routine microscopic evaluation of H&E stained fetal tissues was made and any fetal tissues and placentas with lesions compatible with Neospora-infection were processed by immunohistochemistry (IHC). DNA extraction from fresh and formalin-fixed, paraffin-embedded fetal tissues were tested by nested PCR. All dams from groups 1, 2 and 3 were seropositive by IFAT, rNcGRA7-based-ELISA and Western blot. IgG1/IgG2 ratios were ≤1 at weeks 27 and 29 of gestation. Only fetuses from groups 1 and 2 developed N. caninum specific antibodies by Western blot. Histopathological lesions compatible with those caused by N. caninum were observed in fetuses from groups 1 and 2. N. caninum cysts and tachyzoites were observed by IHC on fetal tissues from groups 1 and 2. Only fetal samples from group 2 were positive by PCR. Further work is needed not only to characterize the cellular immune response but also to clarify the consequences on the dam after conjunctival inoculation of N. caninum tachyzoites. This study shows that N. caninum tachyzoites inoculated by the conjunctival route were not vertically transmitted in pregnant cows.Veterinary Parasitology 10/2013; 199(1-2). DOI:10.1016/j.vetpar.2013.10.006 · 2.55 Impact Factor
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ABSTRACT: The aim of the present study was to compare the immunogenicity and protective efficacy of live tachyzoites and native antigen extract obtained from the NC-6 Argentina strain against vertical transmission of Neospora caninum, following experimental challenge in pregnant heifers with the NC-1 strain. Sixteen pregnant heifers were divided in 4 groups of 4 animals, each receiving different inoculation before mating: group A animals were intravenously (iv) inoculated with 6.25×10(7) live tachyzoites of the NC-6 strain, group B heifers were inoculated twice subcutaneously (sc) with N. caninum native antigen extract formulated with ISCOMs, group C heifers were sc injected with sterile phosphate-buffered saline (PBS) and group D heifers received sc ISCOM-matrix (ISCOMs without antigen). All groups were iv challenged with the NC-1 strain at 70 days of gestation. Serum and heparinized blood samples were collected eight times on weeks 0, 2, 3, 5, 9, 13, 16 and 17 post-inoculation. Dams were slaughtered at the 17th week of experiment (104 days of pregnancy) and placental and fetal tissue samples were collected. Specific antibody responses in heifers were tested by indirect enzyme linked immunosorbent assay (iELISA). The cellular immune response in dams was assessed by quantifying IFN-γ production and the percentages of T-cells (CD4(+), CD8(+) and γδ(+)) and monocytes in peripheral blood mononuclear cells (PBMC). Fetal fluids and tissue samples were tested using the indirect fluorescence antibody test, western blot, histopathology, immunohistochemistry and nested-PCR. A significant increase in N. caninum antibody response was detected in heifers of groups A and B from week 3 after inoculation (P<0.001). IFN-γ production was similar in groups A and B at week 13 (P>0.05). All fetuses were viable at necropsy. Specific IgG against N. caninum was detected in 1/4 fetal fluids recovered from groups A, C and D heifers and 3/4 fetal fluids from group B. Transplacental transmission could be determined in one fetus from group A and three fetuses from group B by nPCR. All fetuses from groups C and D were positive by nPCR. It is noteworthy that dams with higher CD4(+)/CD8(+) ratios in PBMC, regardless of the experimental group, had lower pathology scores. The results of this study confirm that inoculation with live parasites pre-mating may provide at least partial protection against vertical transmission of N. caninum following challenge in heifers at early gestation.Veterinary Parasitology 07/2013; 197(3-4). DOI:10.1016/j.vetpar.2013.07.027 · 2.55 Impact Factor
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ABSTRACT: Neospora caninum is a protozoan which can cause abortions in caprines. However, information regarding the humoral immune response and the occurrence of reproductive disorders is scarce. This is the first study in which the kinetics of antibodies is studied in pregnant goats naturally infected by N. caninum, as well as their respective conceptuses. The subclasses of IgG (IgG1 and IgG2) were also evaluated in pregnant goats. Reproductive problems related to neosporosis (abortion and stillbirth) occurred in 15.38% of the goats. There was a statistically significant association between the increased titres of maternal IgG in the second half of the gestational period with the occurrence of endogenous transplacental transmission. The rate of congenital transmission was 77%. During the gestational period of the seropositive goats, there was mainly a predominance of the subclass IgG2, although mixed patterns of IgG2-IgG1 and the IgG1 pattern were also observed. These results indicate that N. caninum is responsible for the occurrence of important alterations in the humoral immune response of naturally infected goats, and is also a potential causative agent for reproductive disorders in goats. The high proportion of infected conceptuses reinforces the suggestion that congenital infection is one of the main routes of parasite transmission in goats.Veterinary Parasitology 03/2013; 196(3-4). DOI:10.1016/j.vetpar.2013.03.002 · 2.55 Impact Factor