Rational optimization of generic primers used for Norwalk-like virus detection by reverse transcriptase polymerase chain reaction

Abstract

Norwalk-like caliciviruses (NLV) are increasingly recognized as causes of gastro-enteritis world-wide. Because these viruses can not be grown in tissue culture, molecular detection by reverse-transcriptase PCR (RT-PCR) has been used since the genome sequence of the prototype virus named Norwalk virus was published (Jiang et al., 1993). With the recognition that NLV are a quite diverse genus of viruses, several protocols have been published for generic detection methods that differ substantially in sensitivity and specificity (Vinje et al., submitted). A key issue is that due to the great diversity of NLV, it is difficult to do a complete assay validation, using a broad range of genetically distinct viruses. To address this, we have recently completed the evaluation of a sequence-directed modification of a primer set for NLV RT-PCR detection. 2. Approach