GROUP-SPECIFIC IDENTIFICATION OF POLIOVIRUSES BY PCR USING PRIMERS CONTAINING MIXED-BASE OR DEOXYINOSINE RESIDUES AT POSITIONS OF CODON DEGENERACY

We have developed a method for differentiating polioviruses from nonpo lio enteroviruses using PCR. A pair of panpoliovirus PCR primers were designed to match intervals encoding amino acid sequences within VP1 t hat are strongly conserved among polioviruses. The initiating primer h ybridizes with codons of a 7-amino-acid sequence that has been found o nly in polioviruses; the second primer matches codons of a domain thou ght to interact with the cell receptor. The panpoliovirus PCR primers contain mixed-base and deoxyinosine residues to compensate for the hig h degeneracy of the targeted codons. All RNAs from 48 vaccine-related and 110 wild poliovirus isolates of all three serotypes served as effi cient templates for amplification of the 79-bp product. None of the ge nomic sequences of 49 nonpolio enterovirus reference strains were ampl ified under equivalent reaction conditions. Sensitivities of polioviru s detection were as low as 100 fg (equivalent to similar to 25,000 gen omic copies or 25 to 250 PFU) when the amplified products were visuali zed by ethidium bromide fluorescence. These degenerate PCR primers sho uld aid in the detection of all polioviruses, including those wild pol iovirus isolates for which genotype-specific reagents are unavailable.