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Applied and Environmental Microbiology, September 1998, p. 3403-3410, Vol. 64, No. 9
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Genetic Relatedness among Environmental, Clinical, and Diseased-Eel Vibrio vulnificus Isolates from Different Geographic Regions by Ribotyping and Randomly Amplified Polymorphic DNA PCR

Covadonga R. Arias,1 María Jesús Pujalte,1 Esperanza Garay,1,* and Rosa Aznar1,2

Departamento de Microbiología, Universitat de València,1 and Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas,2 Burjassot, E-46100 Valencia, Spain

Received 9 February 1998/Accepted 23 June 1998

Genetic relationships among 132 strains of Vibrio vulnificus (clinical, environmental, and diseased-eel isolates from different geographic origins, as well as seawater and shellfish isolates from the western Mediterranean coast, including reference strains) were analyzed by random amplified polymorphic DNA (RAPD) PCR. Results were validated by ribotyping. For ribotyping, DNAs were digested with KpnI and hybridized with an oligonucleotide probe complementary to a highly conserved sequence in the 23S rRNA gene. Random amplification of DNA was performed with M13 and T3 universal primers. The comparison between ribotyping and RAPD PCR revealed an overall agreement regarding the high level of homogeneity of diseased-eel isolates in contrast to the genetic heterogeneity of Mediterranean isolates. The latter suggests the existence of autochthonous clones present in Mediterranean coastal waters. Both techniques have revealed a genetic proximity among Spanish fish farm isolates and a close relationship between four Spanish eel farm isolates and some Mediterranean isolates. Whereas the differentiation within diseased-eel isolates was only possible by ribotyping, RAPD PCR was able to differentiate phenotypically atypical isolates of V. vulnificus. On the basis of our results, RAPD PCR is proposed as a better technique than ribotyping for rapid typing in the routine analysis of new V. vulnificus isolates.

* Corresponding author. Mailing address: Departamento de Microbiología y Ecología, Universitat de València, Burjassot, E-46100 Valencia, Spain. Phone: 34-6-398 31 43. Fax: 34-6-386 43 72. E-mail: Esperanza.Garay{at}uv.es.

Applied and Environmental Microbiology, September 1998, p. 3403-3410, Vol. 64, No. 9
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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