PubMed Health⌕ Search

Biomedical subjects

Tom P Brown

Publications and source records attributed to Tom P Brown.

4 recordsLinked to original sources
PubMed2004

Development of quantitative competitive-reverse transcriptase-polymerase chain reaction for detection and quantitation of avian leukosis virus subgroup J.

Infection with avian leukosis virus subgroup J (ALV-J) causes severe economic losses in the broiler industry by increasing mortality, producing tumors, and decreasing weight gain in chickens. The quantitation of ALV-J is difficult because of its failure to produce a cytopathic effect in cell culture systems and the nonspecificity of antigen-capture enzyme-linked immunosorbent assay (ELISA) tests. This study was performed to develop a quantitative competitive-reverse transcriptase-polymerase chain reaction (QC-RT-PCR) method based on coamplification of ALV-J genomic RNA and a known amount of a synthesized RNA competitor. The 369 bp RNA competitor was constructed by restriction enzyme treatment of an ALV-J specific 545 bp PCR product, ligation, transformation into Escherichia coli, and in vitro transcription. The competitor contained the same amplification primer annealing sites and sequence as the original viral RNA, except that it had a 176 bp internal deletion. Coamplified RT-PCR products were visualized by electrophoresis and ethidium bromide staining, and fluorescences were quantified using computer-assisted image analysis. The sensitivity of this new QC-RT-PCR method was 25 fg of viral RNA, and 10-fold dilutions were differentiable. This method allowed absolute and relative quantification of ALV-J RNA copy numbers and was simpler than previously published methods for ALV-J quantification.

Animals↗
PubMed2002

Comparison and verification of quantitative competitive reverse transcription polymerase chain reaction (QC-RT-PCR) and real time RT-PCR for avian leukosis virus subgroup J.

Avian leukosis virus subgroup J (ALV-J) infections cause significant economic losses because of increased mortality, tumor production, decreased production, and cost for eradication. Current quantification methods for ALV-J expressed by TCID(50) are difficult to determine because of the lack of cytopathic effect in cell cultures and non-specificity of currently available antigen-capture ELISA tests. In this study, a one-tube fluorescent probe based real time RT-PCR method was developed for quantification of ALV-J and compared with available quantification methods. Cell lysates with different TCID(50)s determined by cell culture and antigen capture ELISA (ag-ELISA) were used for one-tube real time RT-PCR using fluorogenic probe and quantitative competitive RT-PCR (QC-RT-PCR). The results of QC-RT-PCR and real time RT-PCR were highly correlated to the TCID(50)s determined by conventional culture methods. They were also very specific, sensitive, easy to perform, reproducible, and rapid compared with conventional methods. These RT-PCR based quantification methods of ALV-J viral RNA will be useful for virological and pathogenesis studies.

Animals↗
PubMedDate not supplied

Disseminated mycosis in layer cockerels and pullets.

Eight-wk-old layer cockerels and pullets were presented to the diagnostic lab with a history of increased mortality, ruffled feathers, lameness, and recent vaccination. At necropsy, the birds had large multifocal granulomas in multiple tissues. Only light bacterial growth was seen on culture. On histopathology, a mixed population of fungi was seen within the granulomas including zygomycetes and Aspergillus, with the zygomycetes being the predominant organism. Because of the coinfection with Aspergillus and Penicillium, obtaining the zygomycetes in pure culture was unsuccessful. The source of the zygomycete fungi remains unknown; however, zygomycetes are known to be ubiquitous. Serology was performed to evaluate the flock's immune status. There was no evidence of immunosuppression caused by chicken anemia virus or bursal disease infections. No flock treatment was initiated.

Air Sacs↗