Phylogeny and Molecular Identification of Vibrios on the Basis of Multilocus Sequence Analysis

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Abstract
We analyzed the usefulness ofrpoA,recA, andpyrHgene sequences for the identification of vibrios. We sequenced fragments of these loci from a collection of 208 representative strains, including 192 well-documentedVibrionaceaestrains and 16 presumptiveVibrioisolates associated with coral bleaching. In order to determine the intraspecies variation among the three loci, we included several representative strains per species. The phylogenetic trees constructed with the different genetic loci were roughly in agreement with former polyphasic taxonomic studies, including the 16S rRNA-based phylogeny of vibrios. The familiesVibrionaceae,Photobacteriaceae,Enterovibrionaceae, andSalinivibrionaceaewere all differentiated on the basis of each genetic locus. Each species clearly formed separated clusters with at least 98, 94, and 94%rpoA,recA, andpyrHgene sequence similarity, respectively. The genusVibriowas heterogeneous and polyphyletic, withVibrio fischeri,V. logei, andV. wodanisgrouping closer to thePhotobacteriumgenus.V. halioticoli-,V. harveyi-,V. splendidus-, andV. tubiashii-related species formed groups within the genusVibrio. Overall, the three genetic loci were more discriminatory among species than were 16S rRNA sequences. In some cases, e.g., within theV. splendidusandV. tubiashiigroup,rpoAgene sequences were slightly less discriminatory thanrecAandpyrHsequences. In these cases, the combination of several loci will yield the most robust identification. We can conclude that strains of the same species will have at least 98, 94, and 94%rpoA,recA, andpyrHgene sequence similarity, respectively.

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