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Journal of Clinical Microbiology, April 2002, p. 1368-1375, Vol. 40, No. 4
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.4.1368-1375.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Detection of Mammalian Reovirus RNA by Using Reverse Transcription-PCR: Sequence Diversity within the 3-Encoding L1 Gene

Thomas P. Leary,^1* James C. Erker,¹ Michelle L. Chalmers,¹ Andrea T. Cruz,^2,3 J. Denise Wetzel,^2,3 Suresh M. Desai,¹ Isa K. Mushahwar,¹ and Terence S. Dermody^2,3,4

Virus Discovery Group, Abbott Diagnostics Division, Abbott Laboratories, North Chicago, Illinois 60064-6269 ,¹ Departments of Pediatrics,² Microbiology and Immunology,⁴ Elizabeth B. Lamb Center for Pediatric Research, Vanderbilt University School of Medicine, Nashville, Tennessee 37232³

Received 31 July 2001/ Returned for modification 20 October 2001/ Accepted 19 January 2002

Reoviruses infect virtually all mammalian species, and infection of humans is associated with mild gastrointestinal or upper respiratory illnesses. To improve reovirus detection strategies, we developed a reverse transcription-PCR technique to amplify a fragment of the reovirus L1 gene segment. This assay was capable of detecting 44 of 44 reovirus field isolate strains and was sufficiently sensitive to detect nearly a single viral particle (1.16 ± 0.13) per PCR of prototype strain type 3 Dearing. Pairwise comparisons of the 44 partial L1 gene sequences revealed that nucleotide variability ranged from 0 to 24.7%, with most of the nucleotide polymorphism occurring at synonymous positions. Phylogenetic trees generated from amplified L1 gene sequences suggest that multiple alleles of the L1 gene cocirculate in nature and that genetic diversity of the L1 gene is largely independent of the host species, geographic locale, or date of isolation. Phylogenetic trees constructed from the L1 gene sequences are distinct from those constructed from the four reovirus S-class gene segments, which supports the hypothesis that reovirus gene segments reassort in nature. This study establishes a new sensitive and specific technique for the identification of mammalian reoviruses and enhances our understanding of reovirus evolution.

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* Corresponding author. Mailing address: Virus Discovery Group, Experimental Biology Research, Department 90D, Building L3, Abbott Laboratories, 1401 Sheridan Rd., North Chicago, IL 60064-6269. Phone: (847) 937-3512. Fax: (847) 938-6042. E-mail: Thomas.Leary{at}abbott.com.

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Journal of Clinical Microbiology, April 2002, p. 1368-1375, Vol. 40, No. 4
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.4.1368-1375.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

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