Development and evaluation of one-step TaqMan real-time reverse transcription-PCR assays targeting nucleoprotein, matrix, and hemagglutinin genes of equine influenza virus.

The objective of this study was to develop and evaluate new TaqMan real-time reverse transcription-PCR (rRT-PCR) assays by the use of the minor groove binding probe to detect a wide range of equine influenza virus (EIV) strains comprising both subtypes of the virus (H3N8 and H7N7). A total of eight rRT-PCR assays were developed, targeting the nucleoprotein (NP), matrix (M), and hemagglutinin (HA) genes of the two EIV subtypes. None of the eight assays cross-reacted with any of the other known equine respiratory viruses. Three rRT-PCR assays (EqFlu NP, M, and HA3) which can detect strains of the H3N8 subtype were evaluated using nasal swabs received for routine diagnosis and swabs collected from experimentally inoculated horses. All three rRT-PCR assays have greater specificity and sensitivity than virus isolation by egg inoculation (93%, 89%, and 87% sensitivity for EqFlu NP, EqFlu M, and EqFlu HA3 assays, respectively). These assays had analytical sensitivities of >or=10 EIV RNA molecules. Comparison of the sensitivities of rRT-PCR assays targeting the NP and M genes of both subtypes with egg inoculation and the Directigen Flu A test clearly shows that molecular assays provide the highest sensitivity. The EqFlu HA7 assay targeting the H7 HA gene is highly specific for the H7N7 subtype of EIV. It should enable highly reliable surveillance for the H7N7 subtype, which is thought to be extinct or possibly still circulating at a very low level in nature. The assays that we developed provide a fast and reliable means of EIV diagnosis and subtype identification of EIV subtypes.

Serologic and reproductive findings after a herpesvirus-1 abortion storm in goats.

CASE DESCRIPTION: An abortion storm occurred in a goat herd, resulting in 75 aborted kids and 1 neonatal death from December 2004 to February 2005. CLINICAL FINDINGS: Aborted fetuses ranged from being premature to past term. Laboratory findings in 4 of 5 aborted fetuses were consistent with herpesvirus abortion. A virus that yielded positive results with a fluorescent antibody test for bovine herpesvirus-1 was isolated and identified as caprine herpesvirus-1 (CpHV-1) via DNA sequence analysis. TREATMENT AND OUTCOME: Many does that aborted were rebred for kidding in late summer. Most of the young wethers born in 2005 were sold; however, all of the young does were kept for breeding in fall. In November 2005, all 241 goats in the herd were tested for antibodies against CpHV-1 to identify goats that had seroconverted during the outbreak. No complications attributable to CpHV-1 were identified during kidding in 2006. CLINICAL RELEVANCE: On the basis of serologic findings, infection with CpHV-1 was not associated with reduced reproductive success in the subsequent breeding.

Evaluation of a human group a rotavirus assay for on-site detection of bovine rotavirus.

Neonatal diarrhea induced by bovine group A rotavirus causes significant economic loss in the dairy and beef industry due to increased morbidity and mortality, treatment costs, and reduced growth rates. The objective of this study was to evaluate a human group A rotavirus assay (ImmunoCardSTAT Rotavirus [ICS-RV]) as an on-site diagnostic test for bovine rotavirus. When used with a collection of bovine diarrhea samples submitted to the Virology Section of the Diagnostic Center for Population and Animal Health at Michigan State University and compared to a bovine group A rotavirus-specific reverse transcription-PCR (RT-PCR), the ICS-RV assay had a sensitivity and specificity of 87.0 and 93.6%, respectively. A commercially available group A rotavirus enzyme-linked immunosorbent assay (ELISA) (Pathfinder; Sanofi Diagnostics, Redmond, Wash.), when used with the same fecal sample collection and compared to the same RT-PCR, had a sensitivity and specificity of 78.3 and 67.7%, respectively. Subsequently, the ICS-RV assay, RT-PCR, and a different commercially available group A rotavirus ELISA (Rotaclone; Meridian Diagnostics, Cincinnati, Ohio) were used to evaluate fecal samples collected from neonatal calves experimentally infected with bovine rotavirus. When diarrheic fecal samples that were positive for bovine rotavirus by RT-PCR were evaluated, the ICS-RV assay and the Rotaclone assay detected bovine rotavirus 85 and 95% of the time, respectively. Based on these studies, the ICS-RV assay appears to be an excellent test for detecting group A bovine rotaviruses. This assay may be useful as an on-site diagnostic test for veterinarians as an aid in the management of bovine neonatal diarrhea.