Molecular characterization of Drosophila C virus isolates

Reverse transcription coupled with polymerase chain reaction and restrictio n enzyme analysis was used to characterize 12 Drosophila C virus isolates f rom geographically different regions. A 1.2-kb fragment was amplified from cDNA and profiles from digestion with 20 restriction enzymes were generated . Analysis of the restriction fragment data gave estimates of nucleotide di vergence of 0-10% between isolates. The isolates were grouped on the basis of genetic distance estimates derived from the restriction data. For the is olates from which a single genotype could be purified, a geographical patte rn in the distribution of viral genotypes was identified. The 4 Moroccan is olates were very closely related to each other, differing in only 1 restric tion profile. The 2 Australian isolates were each other's closest relatives , as were the 2 isolates first recovered in France. The PCR-RFLP technique used in this study has provided us with a simple procedure which can be use d to characterize DCV isolates. A single enzyme, Tag I, generated 5 distinc t and diagnostic restriction fragment patterns, which allowed easy assignme nt of isolates to one of the five viral genotypes identified in this study. (C) 1999 Academic Press.