Real-time polymerase chain reaction and sequencing were used to characterize a 506-bp-long DNA fragment internal to the gyrB gene (gyrBint). The sequences obtained from 32 Enterobacteriaceae-type strains and those available in the Genbank nucleotide sequence database (n = 24) were used as a database to identify 240 clinical enterobacteria isolates. Sequence analysis of the gyrBint fragment of 240 strains showed that gyrBint constitutes a discriminative target sequence to differentiate between Enterobacteriaceae species. Comparison of these identifications with those obtained by phenotypic methods (Vitek 1 system and/or Rapid ID 32E; bioM?rieux, Marcy l'Etoile, France) revealed discrepancies essentially with genera Citrobacter and Enterobacter. Most of the strains identified as Enterobacter cloacae by phenotypic methods were identified as Enterobacter hormaechei strains by gyrBint sequencing. The direct sequencing of gyrBint would be useful as a complementary tool in the identification of clinical Enterobacteriaceae isolates.

Phylogenetic analysis of partial rpoB gene sequences of type and clinical strains belonging to different 16S rRNA gene-fingerprinting ribogroups within 11 species of enterobacteria of the genera Proteus, Morganella and Providencia was performed and allowed the definition of rpoB clades, supported by high bootstrap values and confirmed by ?2.5 % nucleotide divergence. None of the resulting clades included strains belonging to different species and the majority of the species were confirmed as discrete and homogeneous. However, more than one distinct rpoB clade could be defined among strains belonging to the species Proteus vulgaris (two clades), Providencia alcalifaciens (two clades) and Providencia rettgeri (three clades), suggesting that some strains represent novel species according to the genotypes outlined by rpoB gene sequence analysis. Percentage differences between the rpoB gene sequence of the type strain of Proteus myxofaciens and other members of the same genus (17.3-18.9 %) were similar to those calculated amongst strains of the genus Providencia (16.4-18.7 %), suggesting a genetic distance at the genus-level between Proteus myxofaciens and the rest of the Proteus-Providencia group. Proteus myxofaciens therefore represents a member of a new genus, for which the name Cosenzaea gen. nov., is proposed.

Proteus species are well-known opportunistic pathogens frequently associated with skin wound and urinary tract infections in humans and animals. O antigen diversity is important for bacteria to adapt to different hosts and environments, and has been used to identify serotypes of Proteus isolates. At present, 80 Proteus O-serotypes have been reported. Although the O antigen structures of most Proteus serotypes have been identified, the genetic features of these O antigens have not been well characterized. The O antigen gene clusters of Proteus species are located between the cpxA and secB genes. In this study, we identified 55 O antigen gene clusters of different Proteus serotypes. All clusters contain both the wzx and wzy genes and exhibit a high degree of heterogeneity. Potential functions of O antigen-related genes were proposed based on their similarity to genes in available databases. The O antigen gene clusters and structures were compared, and a number of glycosyltransferases were assigned to glycosidic linkages. In addition, an O serotype-specific suspension array was developed for detecting 31 Proteus serotypes frequently isolated from clinical specimens. To our knowledge, this is the first comprehensive report to describe the genetic features of Proteus O antigens and to develop a molecular technique to identify different Proteus serotypes.