Michael N. Oliver

Experimental reproduction of bovine fetal Neospora infection and death with a bovine Neospora isolate

Studies were conducted to determine the pathogenic potential of the recently isolated bovine Neospora protozoa (BPA-1) for the bovine fetus. Cows chosen for study had Neospora titers < 160 using an indirect immunofluorescent antibody (IFA) test. Four experimental groups were studied. In group 1, 2 fetuses were inoculated in utero at 118 days gestation with culture-derived Neospora tachyzoites. A pregnant control cow was housed in the same pen, observed daily and screened serologically for evidence of exposure to Neospora. In group 2, 2 cows were infected with Neospora tachyzoites at 138 or 161 days gestation, and 1 control cow was given uninfected cell culture suspension simultaneously at 154 days gestation. Groups 3 (85 days gestation) and 4 (120 days gestation) each consisted of 2 cows infected with Neospora tachyzoites and 1 control cow given uninfected material at the same stage of gestation. Dead fetuses were surgically removed from the infected cows in group 1 on postinfection day (PID) 17. The histopathology was compatible with protozoal fetal infection, and protozoa were identified by immunohistochemistry. Viable fetuses were removed surgically from cows in group 2 on PID 28-30. The histopathology was compatible with protozoal fetal infection, protozoa were identified by immunoperoxidase techniques, and Neospora tachyzoites were reisolated in vitro from tissues of the 2 infected fetuses. In groups 3 and 4, the control fetus and 1 infected fetus were removed surgically between PID 26 and PID 33. The remaining infected cows were observed until fetal death or abortion occurred. In group 3, the fetus that was surgically removed from 1 infected cow had no pathologic abnormalities, and parasites were not found (PID 26). The second fetus in group 3 died in utero, and expulsion of a mummified fetus was induced on PID 67. Brain histopathology was compatible with protozoal infection, and parasites were identified by immunoperoxidase techniques. The fetus that was surgically removed (PID 32) from 1 infected cow in group 4 had lesions compatible with protozoal infection, and Neospora tachyzoites were reisolated in vitro from fetal tissues. The second infected cow in group 4 produced a full-term live calf that had a precolostral Neospora titer of 20,480. Clinically, this calf had depressed conscious proprioception in all limbs. Very mild lesions were found in the central nervous system, but protozoa were not found in the tissues. The results demonstrate that the bovine Neospora protozoa can be transplacentally transmitted, resulting in fetal infection and death, and mimics the naturally occurring fetal disease.

Toxicity of storm‐water runoff after dormant spray application in a French prune orchard, Glenn County, California, USA: Temporal patterns and the effect of ground covers

Organophosphorous (OP) insecticides, especially diazinon, have been detected routinely in surface waters of the Sacramento and San Joaquin River watersheds, coincident with rainfall events following their application to dormant orchards during the winter months. Preventive best management practices (BMP) aim at reducing off-site movement of pesticides into surface waters. Two proposed BMPs are: The use of more hydrophobic pyrethroid insecticides believed to adsorb strongly to organic matter and soil and the use of various types of ground cover vegetation to increase the soils capacity for water infiltration. To measure the effectiveness of these BMPs, storm water runoff was collected in a California prune orchard (Glenn County, CA, USA) during several rainstorms in the winter of 2001, after the organophosphate diazinon and the pyrethroid esfenvalerate were applied to different orchard sections. We tested and compared acute toxicity of orchard runoff from diazinon- and esfenvalerate-sprayed sections to two species of fish (Pimephales promelas, Onchorhynchus mykiss) and three aquatic invertebrates (Ceriodaphnia dubia, Simocephalus vetelus, Chironomus riparius), and determined the mitigating effect of three ground cover crops on toxicity and insecticide loading in diazinon-sprayed orchard rows. Runoff from the esfenvalerate-sprayed orchard section was less toxic to waterflea than runoff from the diazinon-sprayed section. However, runoff from the orchard section sprayed with esfenvalerate was highly toxic to fish larvae. Samples collected from both sections one month later were not toxic to fish, but remained highly toxic to invertebrates. The ground cover crops reduced total pesticide loading in runoff by approximately 50%. No differences were found between the types of vegetation used as ground covers.

Experimental transmission of epizootic bovine abortion (foothill abortion)

Advances in defining the biology of epizootic bovine abortion (EBA), including identification of the etiologic agent, have been hampered by the inability to reproduce the disease with confidence. Experimental reproduction of EBA, by feeding the tick vector Ornithodoros coriaceus on susceptible pregnant heifers, is not reliable. The primary objectives of this study were to identify specific tissue(s) obtained from EBA-infected fetuses that could transmit the disease, and then utilize such an infectious challenge system to better define the pathogen, host immunity and geographic distribution of the agent. Described here is the ability to routinely reproduce EBA following inoculation of cryopreserved suspensions of homogenized thymus into susceptible pregnant heifers. This challenge system permitted experiments demonstrating the agent was non-filterable, inactivated upon sonication and susceptible to antibiotics. These findings suggest a prokaryotic microbe and represent a major advance in EBA research. Additional experiments demonstrated that inoculation of the cryopreserved EBA-infectious tissue into heifers, prior to breeding, conferred immunity. Furthermore, such immunized heifers were resistant to challenge with heterologous sources of infectious tissue, suggesting monovalent vaccine development might be feasible. Lastly, challenge studies employing animals from Central Nevada, an area considered free of EBA, demonstrated partial immunity, suggesting the pathogen, and possibly the disease, enjoy a broader distribution than previously thought.

Serologic Detection of Bluetongue Virus Infection of Black-tailed Deer: Comparison of Serum Neutralization, Agar Gel Immunodiffusion, and Competitive ELISA Assays

Three adult black-tailed deer (Odocoileus hemionus columbianus) and four fawns were inoculated with bluetongue virus (BTV) serotype 10 or 17, or epizootic hemorrhagic disease virus (EHDV) serotype 1. Animals were bled at irregular intervals thereafter and the presence of virus-specific antibodies in serum determined by agar gel immunodiffusion (AGID), serum neutralization (SN) and competitive enzyme-linked immunosorbent assay (C-ELISA) tests. Serum antibodies to BTV were detected in all three tests for 692 days after inoculation (DAI) of adult deer, but both the SN and AGID tests gave either erroneous or misleading results. Serum from one deer was negative by the AGID test at 409 DAI with BTV-10 but was positive at 248 and 692 DAI; also one adult and one fawn had antibodies by the SN test to serotypes of BTV with which they were not inoculated. The AGID test for EHDV had false positive results with some sera from animals inoculated only with BTV, and it consistently had false negative results with serum samples collected from an EHDV-inoculated deer at 140 DAI and thereafter. The C-ELISA was the most useful test for the detection of antibodies to BTV because it rapidly gave quantitative and accurate results.

Runoff from orchard floors—micro-plot field experiments and modeling

Hydrologic response patterns of three ground treatments (resident vegetation, bare soil and ripped resident vegetation) were investigated. An in situ, micro-scale, plot retention-tank system was employed and used in conjunction with a kinematic wave model [Seasonal changes in infiltration and soil water storage in conventional and alternative agricultural systems, MS Thesis, University of California, Davis (2000)] to calculate surface water runoff hydrographs via the orchard floor-specific infiltration function. The model optimized Green and Ampts infiltration function on the suction at the wetting front until the endpoint of the cumulative runoff hydrograph matched observed runoff volumes. Our approach aimed to minimize data collection in the field and was found useful in the evaluation of different ground treatments as well as in evaluating their ability to reduce runoff under dry and wetted conditions. Infiltration for ripped resident vegetation was approximately an order of magnitude greater than that for bare soil. Resident vegetation yielded intermediate results. However, under elevated soil water conditions (degree of saturation greater than 0.80), differences between response patterns of resident vegetation and bare soil markedly decreased.