Distribution of 13 virulence genes among clinical and environmental Aeromonas spp. in Western Australia.

We evaluated the pathogenic potential of 98 clinical and 31 environmental Aeromonas isolates by detecting the presence of 13 virulence genes using a polymerase chain reaction (PCR)-based method. The majority (96 %) of the strains contained at least one of the virulence genes. The overall distribution was aerA/haem (77 %), alt (53 %), lafA (51 %), ast (39 %), flaA (32 %), aspA (29 %), vasH (26 %), ascV (16 %) and aexT (13 %). No amplification products were detected for the genes encoding a bundle-forming pilus (BfpA and BfpG) or a Shiga-like toxin (stx-1 and stx-2). Five or more virulence genes were detected in 42 % of environmental and 24 % of clinical isolates. Among the major species, 48 % of A. hydrophila and 42 % of A. dhakensis isolates harboured five or more virulence genes compared with 19 % in A. veronii bv. sobria and none in A. caviae isolates. Our results suggest that, in Western Australia, strains of A. dhakensis and A. hydrophila are potentially more virulent than those of A. veronii bv. sobria and A. caviae, although the pathogenic potential of Aeromonas spp. is probably strain- rather than species-dependent.

Canibacter oris gen. nov., sp. nov., isolated from an infected human wound.

A facultatively anaerobic, Gram-reaction-positive, catalase- and oxidase-negative, rod-shaped bacterium isolated from an infected human wound caused by a dog bite was characterized by phenotypic and molecular genetic methods. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain IMMIB Q2029717T was a member of the order Micrococcales of the class Actinobacteria, displaying 91.6% to 96% sequence similarity with members of the family Microbacteriaceae. Phylogentic trees generated by different algorithms indicated that the strain forms an independent phylogenetic line of descent that consistently clustered proximal to the base of the genus Leucobacter. Chemical studies revealed the presence of a cell-wall murein based on L-lysine (type B1α), major menaquinone (MK-10) and a DNA G+C content of 56.9 mol%. The distinct phylogenetic position, ribotyping and matrix-assisted laser desorption/ionization time-of-flight MS profiles and the significant phenotypic differences clearly separate strain IMMIB Q2029717T from its nearest phylogenetic neighbour and support its classification as a representative of a novel genus and species, with the suggested name Canibacter oris gen. nov., sp. nov. The type strain is IMMIB Q2029717T (=DSM 27064T=CCUG 64069T).

Cruoricaptor ignavus gen. nov., sp. nov., a novel bacterium of the family Flavobacteriaceae isolated from blood culture of a man with bacteraemia.

A Gram-reaction-negative bacterium, strain IMMIB L-12475(T), was isolated from blood cultures of a human with septicaemia. The yellowish orange pigmented strain contained flexirubin pigment. Phylogenetic analysis based on 16S rRNA gene sequence revealed that strain IMMIB L-12475(T) belonged to the family Flavobacteriaceae, forming a distinct phyletic line that is distantly related (79.1-89.4% sequence similarity) to described genera of this family. Membership to the family was confirmed by a fatty acid profile consisting of branched-chain and 3-hydroxy fatty acids with major amounts of iso-C(17:0) 3-OH and iso-C(15:0), by the presence of menaquinone MK-6 as the only respiratory quinone and a polyamine pattern that contained sym-homospermidine as major component. The phospholipids consisted of phosphatidylethanolamine and an unknown phospholipid. The genomic DNA mol% G+C content was 45.6%. The distant phylogenetic position as compared to other representative of the family and the significant phenotypic properties such as pigment composition, morphology, and physiology support the proposal of a novel genus and species Cruoricaptor ignavus gen. nov., sp. nov. The type strain is IMMIB L-12475(T) (=DSM 25479(T)=CCUG 62025(T)).

Corynebacterium aquatimens sp. nov., a lipophilic Corynebacterium isolated from blood cultures of a patient with bacteremia.

An unknown lipophilic coryneform bacterium isolated from the blood cultures of a patient with bacteremia was characterized by phenotypic and molecular genetic methods. Chemical analysis revealed the presence of short chain mycolic acids consistent with the genus Corynebacterium. The DNA G+C content was 60.8 mol%. Comparative 16S rRNA gene sequence analysis demonstrated that the isolate represents a new subline within the genus Corynebacterium. The closely phylogenetic relative of the unknown bacterium was found to be C. tuscaniense (97.8% sequence similarity). Partial rpoB gene sequence revealed that strain IMMIB L-2475(T) exhibited 13.5% sequence divergence with C. tuscaniense. The unknown bacterium was distinguished from C. tuscaniense by, DNA-DNA hybridization, cellular fatty acid profiles, MALDI-TOF analyses of cell extracts and biochemical tests. Based on the phylogenetic and phenotypic criteria, it is proposed that this bacterium be classified as new species, Corynebacterium aquatimens sp. nov., and is represented by strain IMMIB L-2475(T) (=DSM 45632(T)=CCUG 61574(T)).

Corynebacterium pilbarense sp. nov., a non-lipophilic corynebacterium isolated from a human ankle aspirate.

A non-lipophilic coryneform bacterium isolated from an anaerobic Bactec bottle inoculated with an ankle aspirate from a male patient was characterized by phenotypic and molecular taxonomic methods. Chemotaxonomic investigations revealed the presence of short-chain mycolic acids in the cell wall of the bacterium, a feature consistent with members of the genus Corynebacterium. Comparative 16S rRNA gene sequence analysis demonstrated that the isolate displayed 92.0-99.0 % gene sequence similarity with members of the genus Corynebacterium, with Corynebacterium ureicelerivorans as the most closely related phylogenetic species (99.0 % gene sequence similarity). However, the isolate could be genomically separated from C. ureicelerivorans on the basis of DNA-DNA hybridization studies (39.5 % relatedness). Furthermore, the isolate could also be differentiated from C. ureicelerivorans and other species of the genus Corynebacterium on the basis of biochemical properties. Based on both phenotypic and phylogenetic evidence, it is proposed that this isolate be classified as representing a novel species, Corynebacterium pilbarense sp. nov. (type strain IMMIB WACC 658(T)=DSM 45350(T)=CCUG 57942(T)).

Evaluation of the RapID CB plus system for identification of coryneform bacteria and Listeria spp.

In a stress test, the recently introduced RapID CB Plus system (Remel Inc. [formerly Innovative Diagnostic Systems], Norcross, Ga.) was challenged with a diverse set of gram-positive rods comprising 345 strains of coryneform bacteria and 33 strains of Listeria spp. representing a total of 49 different taxa. Overall, within 4 h, the system correctly identified 80.9% of the strains on the species level and 12.2% of the strains on the genus level. Only 3.7% strains were misidentified, and for 3.2% of the strains no identification was provided. Difficulties with the system were mainly due to occasional uncertainties in reading reactions for acid production from carbohydrates and, to a lesser extent, aminopeptidase reactions. It is concluded that the system may also perform well under the conditions of a routine clinical laboratory.

Polymerase chain reaction for the detection of toxigenic Corynebacterium diphtheriae.

Several conventional methods have been described for the detection of Corynebacterium diphtheriae toxin, including Elek immunodiffusion, tissue culture using VERO cells and guinea pig inoculation. All these methods have the disadvantage of being either slow to complete or technically demanding, particularly when performed infrequently. We examined 64 strains of C. diphtheriae by PCR and Elek immunodiffusion, and strains showing a positive result in either assay were inoculated into guinea pigs. Seven isolates were positive in both Elek and PCR assays and subsequently positive in guinea pig inoculation assay. One isolate was negative in Elek testing but positive in PCR assay and guinea pig inoculation. All other isolates were negative in both Elek and PCR assays. The PCR assay is rapid with cycling and detection complete within 3-4 hrs of receipt of strains. PCR has now become the routine method for detection of C. diphtheriae toxin in our laboratory.

Identification of Staphylococcus epidermidis and Staphylococcus hominis from blood cultures by testing susceptibility to desferrioxamine.

Testing susceptibility to desferrioxamine has recently been described as a method for the identification of Staphylococcus epidermidis. This method was compared to a commercial test and the tube coagulase test for the identification of staphylococci from blood cultures and other fluid specimens. A total of 216 isolates was tested over a 13-month period. Sensitivity of the desferrioxamine test in identifying isolates of Staphylococcus epidermidis and Staphylococcus hominis was 97.3%, while specificity was 91.8%. When isolates displaying discrepant desferrioxamine results were characterized using recently described interpretive criteria, sensitivity and specificity of the desferrioxamine test improved to 100%. The desferrioxamine test was reliable, inexpensive and simple to perform, and should prove useful in the diagnostic laboratory.