Molecular detection and identification of enteroviruses using enzymatic amplification and nucleic acid hybridization.
Analysis of enteroviral genomes has revealed that the 5' nontranslated region is highly conserved, providing consensus sequences for the design of oligonucleotides which should anneal to most, if not all, human enteroviral RNAs. We designed and used a pair of such generic primers to enzymatically amplify cDNA from coxsackievirus group B types 1 through 6, poliovirus types 1 through 3, 4 coxsackievirus A types, and 29 echoviruses. The polymerase chain reaction (PCR) products generated with these enteroviral primers were analyzed by agarose gel electrophoresis, Southern blotting, or slot blot hybridization. A genotype-specific PCR was used to detect coxsackievirus B3, to the exclusion of other enteroviruses, by using a coxsackievirus B3 genome-specific primer pair that was derived from sequences coding for part of a capsid protein. A technique is demonstrated by which individual genotypes, for which no sequence information is known, can be identified by high-criterion hybridization analysis following amplification with generic enterovirus PCR primers.
Chapman
N M
NM
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha 68105.
Tracy
S
S
Gauntt
C J
CJ
Fortmueller
U
U
D001483:Base Sequence; D004247:DNA / Q000235:genetics; D015342:DNA Probes; D004770:Enterovirus / Q000235:genetics* / Q000302:isolation & purification; D004769:Enterovirus Infections / Q000175:diagnosis; D005784:Gene Amplification; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction
D015342:DNA Probes; D004247:DNA