GENETIC RELATEDNESS AMONG ENVIRONMENTAL, CLINICAL, AND DISEASED-EEL VIBRIO-VULNIFICUS ISOLATES FROM DIFFERENT GEOGRAPHIC REGIONS BY RIBOTYPING AND RANDOMLY AMPLIFIED POLYMORPHIC DNA PCR

Genetic relationships among 132 strains of Vibrio vulnificus (clinical , environmental, and diseased-eel isolates from different geographic o rigins, as well as seawater and shellfish isolates from the western Me diterranean coast, including reference strains) were analyzed by rando m amplified polymorphic DNA (RAPD) PCR. Results were validated by ribo typing. For ribotyping, DNAs were digested with KpnI and hybridized wi th an oligonucleotide probe complementary to a highly conserved sequen ce in the 23S rRNA gene. Random amplification of DNA was performed wit h M13 and T3 universal primers. The comparison between ribotyping and RAPD PCR revealed an overall agreement regarding the high level of hom ogeneity of diseased-eel isolates in contrast to the genetic heterogen eity of Mediterranean isolates, The latter suggests the existence of a utochthonous clones present in Mediterranean coastal waters. Both tech niques have revealed a genetic proximity among Spanish fish farm isola tes and a close relationship between four Spanish eel farm isolates an d some Mediterranean isolates, Whereas the differentiation within dise ased-eel isolates was only possible by ribotyping, RAPD PCR was able t o differentiate phenotypically atypical isolates of V. vulnificus. On the basis of our results, RAPD PCR is proposed as a better technique t han ribotyping for rapid typing in the routine analysis of new V. vuln ificus isolates.