Restriction endonucleases whose sites are predictable from the amino acid sequence offer an improved strategy for typing bovine rotaviruses

Variation in the third base of a codon hampers genotypic characterization, particularly of RNA viruses. Some restriction endonucleases, however, have a recognition site with a variable base at the third position and will always cleave when a certain amino acid pair occurs (such as glycine-proline forSau96I and glutamic or aspartic acid followed by serine usually forHinfI). We developed a restriction fragment length polymorphism (RFLP) procedure based on these enzymes for P-typing bovine group A rotaviruses (BRV). Employing this procedure 20 BRV local strains, isolated in tissue culture as well as the original faecal sample, could be typed in one of three patterns. More variability was observed when restriction endonucleases were employed whose cleavage sites cannot be predicted from the amino acid sequence (TaqI andTsp509I). These RFLP results agreed with the PCR-VP4 typing assay, neutralization tests, and nucleotide sequence analysis. RFLP withSau96I andHinfI provided quick and objective P-typing of strains and could detect multiple genotypes in the same sample.