Sharon K. Hietala

NEOSPORA CANINUM ANTIBODIES IN COWS DURING PREGNANCY AS A PREDICTOR OF CONGENITAL INFECTION AND ABORTION

A cohort study was undertaken on a dairy experiencing endemic Neospora caninum abortions, to characterize dam serologic variations during pregnancy, and to determine if dam N. caninum antibody levels during gestation predicted congenital infection or abortion. Blood samples were collected monthly during pregnancies of 254 cows and precolostrally from 87 of their calves. Antibody levels, as measured by an enzyme-linked immunosorbent assay, indicated 60.6% of cows were seropositive at some time during pregnancy and 87.4% of seropositive cows were seropositive throughout pregnancy. The rate of seroconversion was 8.5/100 cows/yr. The risk of abortion for seropositive cows at the time of pregnancy diagnosis and during gestation was twice that for seronegative cows (P = 0.025, P = 0.006). Calves born to seropositive cows were more likely to be seropositive at birth if the dam had high antibody levels at 240 days of gestation (P = 0.04) and an increase in antibody levels between 90 and 240 days (P = 0.08) than if the respective values of the dam were low or decreasing. Seropositive cows with high antibody levels at 180 and 210 days of gestation were less likely to abort than cows with low antibody levels at those times (P = 0.05, P = 0.03). Results support a causal effect between exposure to N. caninum and abortion, indicate that acquisition of infection during pregnancy is not necessary for congenital infection or abortion to occur, and suggest that maternal immune response influences congenitial infection and abortion.

An Enzyme-Linked Immunosorbent Assay (ELISA) for Serological Diagnosis of Neospora Sp. Infection in Cattle

A kinetic enzyme-linked immunosorbent assay (ELISA) was developed and optimized for detection of antibodies to Neospora sp. in cattle. Sonicated tachyzoites of Neospora sp. isolated from an aborted bovine fetus were used as antigen. Variability in immunoblot patterns among positive sera, and the fact that all life stages of the parasites are unknown, justified use of a multiple-antigen ELISA to allow for maximum sensitivity. Immunoblot analysis revealed negligible cross-reactions between Toxoplasma gondii antigen and Neospora sp. antisera and between Neospora sp. antigen and antisera from various apicomplexan parasites. The maximum positive-to-negative Vmax (average maximum slope of the optical density over time) ratio was obtained using 200 ng/well of sonicated tachyzoite antigen and a 1:200 serum dilution. Using logistic regression to determine the optimal cutoff point between known infected and noninfected cattle, a sample-to-positive control Vmax ratio of 0.45 was found to maximize the percent correct classification, with an estimated sensitivity of 88.6% and specificity of 96.5%. Use of Neospora caninum antigen following the same protocol demonstrated no difference in ELISA interpretation. Comparison with an existing indirect immunofluorescent antibody (IFA) test showed the ELISA to be the more sensitive and specific test for serodiagnosis of Neospora infection in cattle.

Interpretation of an Indirect Fluorescent Antibody Test for Diagnosis of Neospora sp. Infection in Cattle

Neospora is a recently discovered protozoan that may cause bovine protozoal abortion (BPA) in cows and encephalomyelitis in congenitally infected calves. An indirect fluorescent antibody (IFA) test has been described for diagnosing Neospora sp. infection in cattle. We report here results using the IFA test on peritoneal and pleural fluids from aborted fetuses, precolostral calf sera, and selected adult cattle. The IFA test employed was the same as described elsewhere, with the following modifications. Teflon-coated slides were precleaned in absolute ethanol and fixed in an-

Evidence suggesting a point source exposure in an outbreak of bovine abortion due to neosporosis

A Holstein dairy farm suffered an abortion outbreak due to neosporosis. Abortion losses were > 18%. Cows with the highest Neospora antibody titers were at the greatest risk of aborting. Mummified fetuses were found after the 43rd day of the outbreak. The epidemic curve was suggestive of a point source exposure, which is consistent with the hypothesis that Neospora can be spread by a definitive host.

Interaction of Rhodococcus equi with phagocytic cells from R. equi-exposed and non-exposed foals

The interaction of Rhodococcus equi with alveolar macrophages from adult horses, foals experimentally exposed to R. equi (sensitized foals) and non-exposed foals was studied using in vitro bactericidal assays, cytochemical staining and transmission electron microscopy. It was demonstrated that R. equi is a facultative intracellular parasite, able to survive and multiply within the alveolar macrophages of the host by interfering with phagosome-lysosome fusion. Opsonization of R. equi with antibody against capsular components was associated with increased phagosome-lysosome fusion and significantly enhanced (P less than 0.05) killing of the organism by alveolar macrophages from non-exposed foals. Macrophages from non-exposed foals were able to ingest the non-opsonized organism, but unable to kill greater than 65% of the infective dose by 6 h post-exposure. Alveolar macrophages from sensitized foals behaved as adult macrophages, able to kill greater than 95% of the infective dose by 6 h. Lymphocyte factors, derived by in vitro incubation of sensitized peripheral blood lymphocytes with R. equi surface antigens, enhanced macrophage bactericidal activity. Macrophages from non-exposed foals incubated in the presence of the lymphocyte factors had a 50% increase in killing of R. equi, while sensitized macrophages incubated with lymphocyte factors had a greater than 100% increase in killing capacity.

Pooled-Sample Testing as a Herd-Screening Tool for Detection of Bovine Viral Diarrhea Virus Persistently Infected Cattle

The study was conducted to develop methodology for least-cost strategies for using polymerase chain reaction (PCR)/probe testing of pooled blood samples to identify animals in a herd persistently infected with bovine viral diarrhea virus (BVDV). Cost was estimated for 5 protocols using Monte Carlo simulations for herd prevalences of BVDV persistent infection (BVDV-PI) ranging from 0.5% to 3%, assuming a cost for a PCR/probe test of

Seroepidemiologic study of natural transmission of Mycoplasma hyopneumoniae in a swine herd

20. The protocol associated with the least cost per cow involved an initial testing of pools followed by repooling and testing of positive pools. For a herd prevalence of 1%, the least cost per cow was

Evaluation of the reverse transcription-polymerase chain reaction/probe test of serum samples and immunohistochemistry of skin sections for detection of acute bovine viral diarrhea infections.

2.64 (95% prediction interval =

Diagnosis and seroprevalence of leptospirosis in california sea lions from coastal California

1.72,

Neutrophil phagocytic and serum opsonic response of the foal to Corynebacterium equi

3.68), where pool sizes for the initial and repooled testing were 20 and 5 blood samples per pool, respectively. Optimization of the least cost for pooled-sample testing depended on how well a presumed prevalence of BVDV-PI approximated the true prevalence of BVDV infection in the herd. As prevalence increased beyond 3%, the least cost increased, thereby diminishing the competitive benefit of pooled testing. The protocols presented for sample pooling have general application to screening or surveillance using a sensitive diagnostic test to detect very low prevalence diseases or pathogens in flocks or herds.