Abstract
BackgroundThe genus Vibrio is a diverse group of Gram-negative bacteria comprised of 74 species. Furthermore, the genus has and is expected to continue expanding with the addition of several new species annually. Consequently, it is of paramount importance to have a method which is able to reliably and efficiently differentiate the numerous Vibrio species.ResultsIn this study, a novel and rapid polymerase chain reaction (PCR)-based intergenic spacer (IGS)-typing system for vibrios was developed that is based on the well-known IGS regions located between the 16S and 23S rRNA genes on the bacterial chromosome. The system was optimized to resolve heteroduplex formation as well as to take advantage of capillary gel electrophoresis technology such that reproducible analyses could be achieved in a rapid manner. System validation was achieved through testing of 69 archetypal Vibrio strains, representing 48 Vibrio species, from which an 'IGS-type' profile database was generated. These data, presented here in several cluster analyses, demonstrated successful differentiation of the 69 type strains showing that this PCR-based fingerprinting method easily discriminates bacterial strains at the species level among Vibrio. Furthermore, testing 36 strains each of V. parahaemolyticus and V. vulnificus, important food borne pathogens, isolated from a variety of geographical locations with the IGS-typing method demonstrated distinct IGS-typing patterns indicative of subspecies divergence in both populations making this technique equally useful for intraspecies differentiation, as well.ConclusionThis rapid, reliable and efficient IGS-typing system, especially in combination with 16S rRNA gene sequencing, has the capacity to not only discern and identify vibrios at the species level but, in some cases, at the sub-species level, as well. This procedure is particularly well-suited for preliminary species identification and, lends itself nicely to epidemiological investigations providing information more quickly than other time-honoured methods traditionally used in these types of analyses.
Highlights
The genus Vibrio is a diverse group of Gram-negative bacteria comprised of 74 species
The study began by confirming that the 69 Vibrio type strains obtained from American Type Culture Collection (ATCC) and the Belgian Co-Ordinated Collection of Micro Organisms (BCCM) used in this study were correctly identified
Analysis using BioNumerics yielded an unweight pair group method with arithmetic mean (UPGMA) dendrogram that demonstrated that the patterns generated were sufficiently different from one another so that all species could be separated by virtue of their own unique “species-specific” intergenic spacer (IGS)-type patterns (Figure 2)
Summary
The genus Vibrio is a diverse group of Gram-negative bacteria comprised of 74 species. The United States Centers for Disease Control and Prevention (CDC) estimates that 8,028 Vibrio infections and 57 deaths occur annually in the United States. Of these infections, 5,218 are foodborne in origin [1]. Within the genus Vibrio, V. cholerae, V. parahaemolyticus and V. vulnificus have long been established as important human pathogens in various parts of the world. These organisms are contracted after the patient has consumed raw or undercooked seafood, such as oysters, shrimp, and fish [2]. Identification and subtyping of Vibrio isolates are of significant importance to public health and the safety of the human food supply
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