Document Type

Thesis - University Access Only

Award Date

2001

Degree Name

Master of Science (MS)

Department / School

Biology and Microbiology

Abstract

Bovine viral diarrhea virus (BVDV) is responsible for a complex respiratory and reproductive disease in cattle. BVDV may lead to abortions, persistent infections and Mucosa! Disease (MD). BVDV may be present as a persistent infection (PI), affecting the calf for the remainder of its life. These animals are a great concern to producers as reservoirs for successive BVDV outbreaks. The need to identify and remove infectious animals is imperative to the well-managed herd. BVDV is a member of the Pestivirus family and is related to border disease virus (BDV) in sheep and classical swine fever virus (CSFV) in pigs. Strains of BVDV are characterized as cytopathic (CP) or non-cytopathic (NCP) by their growth characteristics. BVDV strains are further characterized as BVDV Type 1 or Type 2 by antigenic or nucleic acid testing. BVDV has a high mutation rate, allowing it to change quickly and making control difficult. Current testing methods for the detection of BVDV are based on detection of infectious virus, antigen, antibody or nucleic acid. This research project assessed the current methods to detect BVDV from a variety of cattle samples. The detection methods chosen for this research were: virus isolation (VI), microtiter virus isolation (MVI), antigen capture ELISA (AG-ELISA), flow cytometry and reverse transcription polymerase chain reaction (RT-PCR). Serum and whole blood (buffy coat) samples were obtained weekly following vaccination with either a modified-live (ML) or inactivated (IN) multivalient vaccine containing BVDV and three other bovine respiratory viruses. The animals were challenged with a non-cytopathic type 2 BVDV (strain 890) at 14 weeks post-vaccination. Serum and whole blood samples were collected daily for 14 days post-challenge to detect BVD virus. Following vaccination, MVI detected BVDV from a ML animal (#26, week 2). This was the only isolation obtained for post-vaccination phase. VI, MVI, nested RTPCR and flow cytometry detected BVDV acutely infected animals during the post: challenge phase. The AG-ELISA was not specific and the data could not be used. Comparisons of the clinical data, white blood cell (WBC) counts, detection methods and the flow cytometry data indicate: 1) viremia is present in BVDV infected animals 2-4 days preceding clinical symptoms and was correlated with leukopenia, 2) detection of BVDV infected PBMC does not indicate viral shedding and 3) the methods evaluated demonstrated comparable results for BVDV detection. A second experiment was devised to test pooled samples with the nested RT-PCR as a herd screening method for PI animals. Five pools were created by adding 100μ1 of the following samples: 1) five negative samples, 2) four negative and 1 positive sample, 3) nine negative and 1 positive sample, 4) 19 negative and 1 positive sample, 5) 49 negative and 1 positive sample. An aliquot of 300μ1 was removed from each of the pools and was processed for RNA extraction and nested BVDV RT-PCR. The result of these experiments demonstrated the ability to detect one positive sample from a pool of 50 samples. This is another alternative for herd screening and is a reasonable approach to an annual BVDV monitoring program.

Library of Congress Subject Headings

Bovine viral diarrhea -- Diagnosis -- Evaluation Cattle -- Virus diseases -- Diagnosis -- Evaluation

Format

application/pdf

Number of Pages

104

Publisher

South Dakota State University

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