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Journal of Clinical Microbiology, March 2000, p. 1170-1174, Vol. 38, No. 3
0095-1137/00/$04.00+0

Comparison of Classic and Molecular Approaches for the Identification of Untypeable Enterovirusesdagger

M. Steven Oberste,* Kaija Maher, Mary R. Flemister, George Marchetti,Dagger David R. Kilpatrick, and Mark A. Pallansch

Respiratory and Enteric Viruses Branch, Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30333

Received 20 October 1999/Returned for modification 23 November 1999/Accepted 9 December 1999

Members of the family Picornaviridae are the most common viruses infecting humans, and species in several genera also infect a wide variety of other mammals. Picornaviruses have traditionally been classified by antigenic type, based on a serum neutralization assay. However, this method is time-consuming and labor-intensive, is sensitive to virus aggregation and antigenic variation, and requires a large number of antisera to identify all serotypes, even when antiserum pools are used. We developed generic reverse transcription (RT)-PCR primers that will amplify all human enterovirus serotypes, as well as many rhinoviruses and other picornaviruses, and used RT-PCR amplification of the VP1 gene and amplicon sequencing to identify enteroviruses that were refractory to typing by neutralization with pooled antisera. Enterovirus serotypes determined by sequencing were confirmed by neutralization with monospecific antisera. Of 55 isolates tested, 49 were of known enterovirus serotypes, two were rhinoviruses, and four were clearly picornaviruses but did not match any known picornavirus sequence. All four untyped picornaviruses were closely related to one another in sequence, suggesting that they are of the same serotype. RT-PCR, coupled with amplicon sequencing, is a simple and rapid method for the typing and classification of picornaviruses and may lead to the identification of many new picornavirus serotypes.

* Corresponding author. Mailing address: Centers for Disease Control and Prevention, 1600 Clifton Rd. NE, Mailstop G-17, Atlanta, GA 30333. Phone: (404) 639-2751. Fax: (404) 639-4011. E-mail: mbo2{at}cdc.gov.

dagger This paper is dedicated to the memory of our friend and colleague George Marchetti.

Dagger Deceased.

Journal of Clinical Microbiology, March 2000, p. 1170-1174, Vol. 38, No. 3
0095-1137/00/$04.00+0


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