Optimization of methods for the isolation of Marek's disease viruses in primary chicken cell cultures

Tan, J, Cooke, J, Clarke, N and Tannock, G 2008, 'Optimization of methods for the isolation of Marek's disease viruses in primary chicken cell cultures', Journal of Virological Methods, vol. 147, no. 2, pp. 312-318.


Document type: Journal Article
Collection: Journal Articles

Title Optimization of methods for the isolation of Marek's disease viruses in primary chicken cell cultures
Author(s) Tan, J
Cooke, J
Clarke, N
Tannock, G
Year 2008
Journal name Journal of Virological Methods
Volume number 147
Issue number 2
Start page 312
End page 318
Total pages 7
Publisher Elsevier
Abstract A real-time PCR was used to measure increases in viral DNA in Marek's disease virus (MDV)-infected primary chicken cell cultures in order to optimize methods for viral isolation. Serotype-1 and -3 vaccine and serotype-1 challenge strains exhibited similar growth characteristics, with increases in viral DNA being proportional to inoculum size. Studies of viral growth revealed a linear relationship between increase in MDV copy number and infectious titre, although the rate of increase for copy number was greater. Using real-time PCR, viral DNA yields of the virulent Woodlands strain in infected chicken kidney cultures were shown to be slightly, but not significantly, higher than in chicken embryo kidney cultures and significantly higher than in chicken embryo fibroblast cultures. Viral DNA levels in freshly trypsinised cells suspended in growth medium and infected with the Woodlands strain were higher than levels obtained following the inoculation of monolayer cultures. For cells infected in suspension, no significant enhancement of yield was observed following a medium change after 2-3 days. Peak yields were obtained at days 6-8 after inoculation of all cultures. Findings obtained from the optimization of viral DNA levels were applied to a program for the isolation of Australian strains of serotype-1 viruses from problem flocks over 3 years. Significant improvements were obtained in the isolation rate of strains capable of growing to high titre (>104 plaque-forming units/mL) for use in challenge studies.
Subject Veterinary Virology
Keyword(s) Real-time PCR
Marek's
copy number
isolation
cell cultures
DOI - identifier 10.1016/j.jviromet.2007.09.011
Copyright notice © 2007 Elsevier B.V. All rights reserved.
ISSN 0166-0934
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