To evaluate herd-level risk factors for seropositive status of cattle to 1 or more bluetongue viruses. 110 herds of cattle in Nebraska, North Dakota, and South Dakota. Blood samples were collected before and after the vector season. Samples were tested for antibodies against bluetongue virus by use of a commercially available competitive ELISA. Factors evaluated included descriptors of geographic location and management practices. Trapping of insect vectors was conducted to evaluate vector status on a subset of 57 operations. A multivariable logistic regression model was constructed to evaluate associations. For the full data set, altitude and latitude were associated with risk of having seropositive cattle (an increase in altitude was associated with an increase in risk, and a more northerly location was associated with a decrease in risk of a premise having seropositive cattle). Import of cattle from selected states was associated with an increase in risk of having seropositive cattle. From the subset of herds with data on vector trapping, altitude and latitude were associated with risk of having seropositive cattle, similar to that for the full model. However, commingling with cattle from other herds was associated with a decrease in risk of seropositivity. Findings reported here may be useful in generating additional hypotheses regarding the ecologic characteristics of bluetongue viruses and other vector-borne diseases of livestock. Sentinel surveillance programs are useful for documenting regionalization zones for diseases, which can be beneficial when securing international markets for animals and animal products.

A regional prospective study of the epidemiology of bluetongue virus (BTV) serotypes covering 11 countries in Central America and the Caribbean took place between 1987 and 1992. Active surveillance revealed BTV infection to be endemic in the absence of confirmed indigenous cases of bluetongue. During the 6-year span of the study, over 300 BTV isolations were obtained from cattle and sheep. Results of the earlier years of the study were summarized, and surveillance activities in the concluding months of the study from November 1990 to February 1992 were evaluated. Forty-five BTV isolations were made during this time, 44 from sentinel cattle and 1 from a ram with clinical signs compatible with contagious ecthyma. Virus isolation from potential vectors also was attempted, yielding a further 9 BTV isolates from parous Culicoides insignis and C pusillus, 2 BTV isolates from blood-engorged C filarifer, and 1 epizootic hemorrhagic disease virus type-2 isolate from parous C pusillus. Our extensive network of sentinel herds in the region detected BTV-1 as the predominant serotype in Central America in 1991, after an apparent absence of 1 year in the sentinel animals. Other serotypes in Central America at that time included BTV-3 and BTV-6. In Puerto Rico and the Dominican Republic, BTV-4 became the predominant serotype, without detection of BTV-8 and BTV-17, which were common in recent years of the study. The serotypes found in the Caribbean Basin continued to have marked differences from those in North America. The importance of viewing bluetongue as an infection, the distribution of which is determined principally by ecologic factors, is emphasized.

The consequences of inoculation of bluetongue virus (BTV) serotype 11 into 16 susceptible cows either at the time of breeding or at specified stages of pregnancy were studied. The cows were free of BTV or epizootic hemorrhagic disease virus, and none had antibodies to BTV before virus inoculation. A group of 4 cows was mated naturally to a bull reported to shed BTV (CO75B300 strain) in the semen. The bull was suspected of infecting cows at mating with BTV-11, which subsequently transplacentally infected the developing fetuses and induced persistently infected and congenitally malformed progeny. Two groups of 4 pregnant cows were inoculated with an insect-derived strain of BTV-11 (CO75B300), one group by direct deposit into the uterus at estrus, the other, by intradermal and sc administrations. A 90-day fetus was inoculated in utero with virus from the same pool. Four pregnant cows were inoculated with sheep blood-passaged virus of the same BTV-11 strain (CO75B300) by intradermal and sc routes. Three cows were inoculated with BTV-free suspending fluids and ovine erythrocytes by the intrauterine and intradermal-sc routes and were used as in-contact controls.Infection with insect-derived BTV-11 was confirmed in 3 cows of 1 group by virus isolation and by detection of serum antibodies. The 4 cows inoculated with sheep blood suspension of BTV-11 developed viremia and produced antibodies to the virus. None of the cattle had clinical signs of bluetongue, other than 2 cows that had a slight rectal temperature increase on postinoculation day 4.All cows and fetuses that ranged in gestational age from 69 to 217 days appeared grossly normal at necropsy. Antibodies were not detected in fetal blood. Viral antigen was not detected in fetal tissues by inoculation into sheep or by immunofluoerscence, and viral RNA was not detected by use of the polymerase chain reaction. Developmental deformities were not seen in any fetus. The BTV-11 was not transmitted via the bull semen after natural mating. The BTV-11 strain CO75B300, isolated from this bull and passaged either as insect-derived or ovine erythrocyte suspensions, infected 8 cows. However, the virus was not transplacentally transmitted to their fetuses. It was concluded that there was no evidence for congenital BTV-11 infection in this study.

An antigen-capture ELISA was used to detect bluetongue virus (BTV) from blood of infected sheep. A rabbit-origin capture antibody and a mouse-origin detection antibody combined with biotin-avidin amplification were used for the assay. The antigen-capture ELISA could not detect virus directly from the blood of infected sheep because of low virus titer. To enhance detection, virus from infected blood was amplified in cell culture. Virus could then be detected from cell culture supernatant fluids, using the ELISA. This amplification step increased the sensitivity of the assay comparable to that of assays performed in cell culture measuring cytopathic effects. The ELISA procedure was specific for BTV and did not mistakenly identify the antigenically related epizootic hemorrhagic disease virus. The antigen-capture ELISA permitted indirect quantitation and identification of BTV from the blood of infected sheep.

Sheep had viremias that were first detected on day 3 (+/- 1) after infection with several strains of bluetongue virus (BTV) representing United States serotypes 10, 11, 13, and 17. Diphasic peaks of infectivity were attained on days 6 and 10 (+/- 2). Interferon (IFN) was first detected in serum samples on day 5 (+/- 1), and reached greatest concentrations on day 6 (+/- 2), which coincided with the first viremic peak; IFN concentrations then decreased toward zero by day 10 (+/- 2). Interferon peak concentrations induced approximately a 90% decrease in virus titer. The decrease in IFN concentrations by day 9 (+/- 2) corresponded with the second viremic peak on day 10 (+/- 2). Onset of the decrease in detectable concentrations of virus after the second peak of viremia corresponded to the initial detection of serum antibody to BTV by day 10 (+/- 2). Virus titer decreased and antibody production increased until approximately days 21 to 28, when the titers plateaued and virus was not detected. Febrile responses peaked on day 7 (+/- 1) during the peak viremic period. The WBC count was depressed at the time the virus titer increased, but returned to normal values while the sheep were still viremic. Diphasic viremias in BTV-infected sheep were attributed to induction of high concentrations of IFN concurrent with the first virus titer peak, followed by production of antibody to specific BTV strains and a subsequent reduction in viremia at the second virus titer peak.

Inoculation of 53 ewes after 35, 45, 60, or 80 days of gestation with bluetongue virus serotypes 10, 11, 13, or 17, or with epizootic hemorrhagic disease virus serotypes 1 or 2, resulted in overt clinical disease in the 47 ewes inoculated with bluetongue virus but not in the 6 ewes inoculated with epizootic hemorrhagic disease virus. None of the lambs produced by these ewes had developmental defects or any evidence of persistence of viremia.

Each of 5 US-origin serotypes of bluetongue virus (BTV) was inoculated into a separate pair of sheep. The duration of each animal's ensuing viremia was monitored, using a BTV serogroup-specific nested polymerase chain reaction (PCR) method and an embryonating chicken egg (ECE) inoculation procedure. Mean duration of viremia was 100 and 38 days for the PCR and ECE methods, respectively. This difference was significant (P < 0.001) and documents a more prolonged viremia in virus-exposed sheep than has been reported. A dual internal oligonucleotide solution hybridization procedure was developed for the rapid (2 hours) colorimetric detection and identification of BTV-specific PCR products. This enzyme-linked oligonucleotide sorbent assay (ELOSA) relied on annealing of separate biotinylated and fluoresceinated probes to the amplified BTV nucleic acid; these complexes were captured on streptavidin-coated microtitration wells and were detected, using a horseradish peroxidase-labeled antifluorescein antibody conjugate. End-point dilution analyses of PCR products indicated that the ELOSA was more sensitive than gel electrophoretic or comparable colorimetric slot-blot hybridization techniques. The BTV PCR-ELOSA system represents a more sensitive and expeditious means of diagnosing BTV-induced viremia than does the ECE procedure currently used. The combination of ELOSA with PCR should facilitate practical application of nucleic acid technology to diagnostic veterinary medicine.