Comparison of specific and random priming in the reverse transcriptase polymerase chain reaction for genotyping group A rotaviruses

This study describes an approach to the molecular typing of rotaviruses whi ch requires only a single RNA extraction and reverse transcription (RT) rea ction using random primers. Random-primed RT provides complementary DNA (cD NA) which can be used not only for G- and P-typing polymerase chain reactio ns (PCR), but also for the detection of other RNA viruses which may act as enteric pathogens. It is a sensitive and specific method that can detect 10 virus particles/ml of 10% faecal suspension provided the cDNA is amplified in a nested typing-PCR. Of 121 specimens positive for rotavirus by EM and analysed using this method, only 8% could pot be G- or P-genotyped. The unt yped samples were tested again performing the RT reaction with G- and P-spe cific primers, achieving a 5% increase in sensitivity. Comparing G-genotypi ng against G-serotyping, 92% were genotyped through random priming RT-PCR w hereas only 64% were serotyped using G-serotype specific monoclonal antibod ies (MAbs). (C) 1999 Elsevier Science B.V. All rights reserved.