Abstract
Three notable members of the Harveyi clade, Vibrio harveyi, Vibrio alginolyticus and Vibrio parahaemolyticus, are best known as marine pathogens of commercial and medical import. In spite of this fact, the discrimination of Harveyi clade members remains difficult due to genetic and phenotypic similarities, and this has led to misidentifications and inaccurate estimations of a species' involvement in certain environments. To begin to understand the underlying genetics that complicate species level discrimination, we compared the genomes of Harveyi clade members isolated from different environments (seawater, shrimp, corals, oysters, finfish, humans) using microarray-based comparative genomic hybridization (CGH) and multilocus sequence analyses (MLSA). Surprisingly, we found that the only two V. harveyi strains that have had their genomes sequenced (strains BAA-1116 and HY01) have themselves been misidentified. Instead of belonging to the species harveyi, they are actually members of the species campbellii. In total, 28% of the strains tested were found to be misidentified and 42% of these appear to comprise a novel species. Taken together, our findings correct a number of species misidentifications while validating the ability of both CGH and MLSA to distinguish closely related members of the Harveyi clade.
Highlights
The eight Vibrio species currently recognized as members of the Harveyi clade (V. harveyi, V. campbellii, V. alginolyticus, V. rotiferianus, V. parahaemolyticus, V. natrigens, V. mytili and V. azureus) (Sawabe et al, 2007; Yoshizawa et al, 2009) are a subset of the Vibrio core group (Reichelt et al, 1976; Dorsch et al, 1992)
A total of 43 previously characterized isolates (29 V. harveyi, seven V. campbellii, two V. parahaemolyticus, three V. rotiferianus and two V. alginolyticus), from a wide temporal, geographical and environmental distribution were selected for this study (Table 1)
A subset of 38 isolates were analysed via comparative genomic hybridization (CGH) using a custom-designed Affymetrix DNA microarray (Vharveyi520694F) that targets 4831 total coding sequence (CDS) from the fully assembled and annotated V. harveyi BAA-1116 genome (Naval Research Laboratory sequencing effort GenBank CP001223-5) and 965 CDS unique to the unfinished V. harveyi HY01 genome sequence (GenBank AAWP00000000)
Summary
The eight Vibrio species currently recognized as members of the Harveyi clade (V. harveyi, V. campbellii, V. alginolyticus, V. rotiferianus, V. parahaemolyticus, V. natrigens, V. mytili and V. azureus) (Sawabe et al, 2007; Yoshizawa et al, 2009) are a subset of the Vibrio core group (Reichelt et al, 1976; Dorsch et al, 1992). V. harveyi, V. campbellii and V. rotiferianus, which form the most recent subclade of speciation within the Harveyi clade (Pascual et al, 2009), have nearly indistinguishable phenotypes (Bryant et al, 1986; Gomez-Gil et al, 2004). We employed CGH using a custom-designed Affymetrix V. harveyi BAA-1116/HY01 DNA microarray to delineate 38 geographically, environmentally and temporally distributed members of the Harveyi clade and confirmed the resulting cluster assignments using two MLSAs
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