Evaluation of enzyme-linked immunosorbent assays performed on milk and serum samples for detection of neosporosis and leukosis in lactating dairy cows.

Serum and milk samples from 1229 cows on 22 Ontario dairy farms were individually tested for antibodies specific for bovine leukosis virus (BLV) and Neospora caninum by enzyme-linked immunosorbent assay (ELISA). Antibodies against BLV were present in 361 serum samples (29.4%) and 369 milk samples (30.0%). Comparing the 2 tests, agreement was almost perfect (k = 0.86; 95% CI = 0.83 to 0.90) and the proportions of samples positive were not significantly different (P = 0.56). Both tests identified the same 3 herds free of bovine leukosis virus. Antibodies against N. caninum were detected in 138 serum samples (11.2%), and 111 milk samples (9.0%). Agreement between the 2 tests was moderate (k = 0.52; 95% CI = 0.43 to 0.59). Four herds were free of neosporosis by the serum test, while 10 herds were negative by the milk test. The ELISA on milk samples facilitates sample collection to classify herds free of BLV; the milk N. caninum ELISA was less reliable in predicting herd-level infection.

Évaluation des tests ELISA réalisés sur des échantillons de lait et de sérum pour la détection de la néosporose et de la leucose chez les vaches laitières en lactation. Des échantillons de sérum et de lait provenant de 1229 vaches dans 22 fermes laitières de l'Ontario ont été testés individuellement pour déceler des anticorps particuliers au virus de la leucose bovine (VLB) et de Neospora caninum à l'aide d'un test ELISA. Les anticorps contre le VLB étaient présents dans 361 échantillons de sérum (29,4 %) et 369 échantillons de lait (30,0 %). En comparant les 2 tests, la concordance était quasiment parfaite (k = 0,86; IC de 95 % = de 0,83 à 0,90) et les proportions d'échantillons positifs n'étaient pas significativement différentes (P = 0,56). Les deux tests ont identifié les même 3 troupeaux comme étant libres du virus de la leucose bovine. Des anticorps contre N. caninum ont été détectés dans 138 échantillons de sérum (11,2 %) et 111 échantillons de lait (9,0 %). La concordance entre les 2 tests était modérée (k = 0,52; IC de 95 % = de 0,43 à 0,59). Quatre troupeaux étaient libres de néosporose lors du test pour le sérum, tandis que 10 troupeaux étaient négatifs lors du test pour le lait. Le test ELISA sur les échantillons de lait facilite le prélèvement d'échantillons pour déclarer les troupeaux comme étant libre du VLB; le test ELISA du lait pour N. caninum était moins fiable pour prédire l'infection au niveau du troupeau.(Traduit par Isabelle Vallières).

Characteristics of a very virulent infectious bursal disease virus from California.

An outbreak of infectious bursal disease (IBD) in two California layer flocks resulted in the isolation of two infectious bursal disease viruses designated rA and rB. Increased mortality plus gross and histopathology in the layer flocks suggested rA and rB could be very virulent infectious bursal disease virus (vvIBDV). Preliminary studies indicated that high mortality resulted when bursa homogenates from the layer farms were used to inoculate specific-pathogen-free (SPF) chicks. In addition, rA and rB contained VP2 amino acid sequences typically seen in vvIBDV. Molecular and in vivo studies were conducted to more thoroughly identify and characterize the rA and rB viruses. Nucleotide sequence analysis demonstrated that rA and rB had identical sequences across the hypervariable VP2 (hvVP2) and segment B regions examined, and their amino acid sequences in the hvVP2 region were identical to the vvwIBDV type strains UK 661, OKYM, and Harbin. Furthermore, the genome segment B nucleotide sequences examined for rA and rB were a 98.1% match with vvIBDV and only an 88.0% match with classic IBDV strains. Phylogenetic analysis placed the rA and rB viruses with other vvIBDV and confirmed these viruses were close genetic descendants of vvIBDV seen around the world. Pathogenicity studies in 4-wk-old SPF chicks demonstrated that at a high dose (105.5 50% egg infective dose [EID50]) and a low dose (102.0 EID50) of rA and rB, mortality ranged from 91% to 100%. A pathogenic classic virus, standard challenge (STC), at similar doses did not cause mortality in the SPF chicks. In addition, mortality occurred in three out of four SPF birds exposed by direct contact to rA and rB inoculated chicks. Serum from convalescent birds inoculated with rA had high titers to IBDV but were negative for antibodies to infectious bronchitis virus, avian influenza virus, chicken anemia virus, Newcastle disease virus, Mycoplasma gallisepticum, and Mycoplasma synoviae. Virus isolation attempts on the rA and rB bursa homogenate inocula also indicated that no contaminating microorganisms contributed to the high mortality and pathology observed in the SPF chicks. In one experiment, broilers with maternal immunity to IBDV were protected from infection and disease when they were challenged with 10(2) EID50 and 10(5) EID50 of the STC virus. When challenged with 10(2) EID50 of the rA virus, the maternally immune broilers were protected from disease but not infection as evidenced by a positive reverse transcription-polymerase chain reaction (RT-PCR) assay for the virus. When the broilers were challenged with 10(5) EID50 of the rA virus, they had typical gross and histopathologic signs of IBD but no mortality by 7 days postinoculation. It was concluded that the rA and rB viruses meet the genotypic and phenotypic characteristics of a vvIBDV.

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.

Bovine viral diarrhea viruses (BVDV) cause both acute and persistent infections. While diagnostic tests have been designed to detect animals persistently infected (PI) with BVDV, the reliability of these tests in detecting acute BVDV infections is not known. It is also possible that acute BVDV infections may be confused with persistent infections in surveys for PI animals. In this study, 2 tests presently in use in diagnostic laboratories to test for PI animals, polymerase chain reaction amplification followed by probe hybridization (RT-PCR/probe) of serum samples and immunohistochemical detection of viral antigen in skin biopsies (IHC), were evaluated for their ability to detect acute BVDV infections. Sixteen colostrum-deprived, BVDV-free, and BVDV-antibody-free calves were infected with 6 different BVDV strains. Clinical signs, seroconversion, and virus isolation indicated that inoculated animals did replicate virus. Virus could be detected in 19% (3/16) of acutely infected animals by the RT-PCR/probe technique. No acutely infected animals were positive by IHC.