MOLECULAR CHARACTERIZATION OF HUMAN ROTAVIRUS VP4 GENES BY POLYMERASECHAIN-REACTION AND RESTRICTION-FRAGMENT-LENGTH-POLYMORPHISM ASSAY

A restriction fragment length polymorphism (RFLP) assay was developed to examine the genetic variability and similarity of the VP4 genes of human rotaviruses. The VP4 genes of 14 human rotavirus strains, includ ing VP4 serotype PIA strains (Wa, P, VA70), serotype P1B strain (DS-1) , serotype P2 strains (M37, 1076, McN, ST3) and serotype P3 strains (A U-1, AU228, K8, PA151, PCP5, MZ58), and those of 2 feline strains (FRV -1 and Cat2) were reverse-transcribed and amplified by the polymerase chain reaction (PCR). The amplified VP4 cDNAs were then digested with a panel of restriction endonucleases (HindIII, NruI, HaeIII, and EcoRI ), resulting in the identification of at least one enzyme with which d igestion produced an RFLP profile specific for a particular P serotype . Of interest was the presence of two distinct RFLP patterns within th e serotype P3 VP4 genes: one corresponding to the VP4 gene carried by the members of the AU-1 genogroup and the other corresponding to the V P4 genes carried by naturally-occurring reassortants between members o f the AU-1 and other genogroups.