Phylogenetic relationships within the genus Pseudomonas were examined by comparing partial (about 1000 nucleotides) rpoB gene sequences. A total of 186 strains belonging to 75 species of Pseudomonas sensu stricto and related species were studied. The phylogenetic resolution of the rpoB tree was approximately three times higher than that of the rrs tree. Ribogroups published earlier correlated well with rpoB sequence clusters. The rpoB sequence database generated by this study was used for identification. A total of 89 isolates (79.5%) were identified to a named species, while 16 isolates (14.3%) corresponded to unnamed species, and 7 isolates (6.2%) had uncertain affiliation. rpoB sequencing is now being used for routine identification of Pseudomonas isolates in our laboratory.

A significant proportion of protein-encoding gene phylogenies in bacteria is inconsistent with the species phylogeny. It was usually argued that such inconsistencies resulted from lateral transfers. Here, by further studying the phylogeny of the oprF gene encoding the major surface protein in the bacterial Pseudomonas genus, we found that the incongruent tree topology observed results from a long-branch attraction (LBA) artifact and not from lateral transfers. LBA in the oprF phylogeny could be explained by the faster evolution in a lineage adapted to the rhizosphere, highlighting an unexpected adaptive radiation. We argue that analysis of such artifacts in other inconsistent bacterial phylogenies could be a valuable tool in molecular ecology to highlight cryptic adaptive radiations in microorganisms.

During the microbiological research performed within the scope of activities of Czech expeditions based at the Johann Gregor Mendel Station at James Ross Island, Antarctica, two psychrotrophic gram-stain negative non-fluorescent strains CCM 8506T and CCM 8507 from soil were extensively characterized using genotypic and phenotypic methods. Initial characterization using ribotyping with HindIII restriction endonuclease and phenotyping implies that both isolates belong to a single Pseudomonas species. Sequencing of rrs, rpoB, rpoD and glnA genes of strain CCM 8506(T) confirmed affiliation of investigated strains within the genus Pseudomonas. Further investigation using automated ribotyping with EcoRI (RiboPrinter(?) Microbial Characterisation System), whole-cell protein profiling using the Agilent 2100 Bioanalyzer system, extensive biochemical testing and DNA-DNA hybridization experiments confirmed that both investigated strains are members of a single taxon which is clearly separated from all hitherto described Pseudomonas spp. Based on all findings, we describe a novel species Pseudomonas gregormendelii sp. nov. with the type strain CCM 8506(T) (=LMG 28632T).

Pseudomonas spp. are common inhabitants of aquatic environments, including drinking water. Multi-antibiotic resistance in clinical isolates of P. aeruginosa is widely reported and deeply characterized. However, the information regarding other species and environmental isolates of this genus is scant. This study was designed based on the hypothesis that members of the genus Pseudomonas given their high prevalence, wide distribution in waters and genetic plasticity can be important reservoirs of antibiotic resistance in drinking water. With this aim, the diversity and antibiotic resistance phenotypes of Pseudomonas isolated from different drinking water sources were evaluated. The genotypic diversity analyses were based on six housekeeping genes (16S rRNA, rpoD, rpoB, gyrB, recA and ITS) and on pulsed field gel electrophoresis. Susceptibility to 21 antibiotics of eight classes was tested using the ATB PSE EU (08) and disk diffusion methods. Pseudomonas spp. were isolated from 14 of the 32 sampled sites. A total of 55 non-repetitive isolates were affiliated to twenty species. Although the same species were isolated from different sampling sites, identical genotypes were never observed in distinct types of water (water treatment plant/distribution system, tap water, cup fillers, biofilm, and mineral water). In general, the prevalence of antibiotic resistance was low and often the resistance patterns were related with the species and/or the strain genotype. Resistance to ticarcillin, ticarcillin with clavulanic acid, fosfomycin and cotrimoxazol were the most prevalent (69-84%). No resistance to piperacillin, levofloxacin, ciprofloxacin, tetracycline, gentamicin, tobramycin, amikacin, imipenem or meropenem was observed. This study demonstrates that Pseudomonas spp. are not so widespread in drinking water as commonly assumed. Nevertheless, it suggests that water Pseudomonas can spread acquired antibiotic resistance, preferentially via vertical transmission.

A multiplex PCR based on oprI and oprL, coding for the outer membrane lipoprotein I and the peptidoglycan-associated lipoprotein OprL, respectively, was developed for the detection of Pseudomonas strains from a bacterial collection isolated from a small river. To study the diversity of these Pseudomonas isolates, an oprI-oprL gene sequence database of 94 Pseudomonas type strains was constructed. Phylogenetic analysis of the concatenated oprI and oprL gene sequences of the Pseudomonas type strains showed that they were largely congruent with the classification based on the MLSA approach based on 16S rRNA, gyrB, rpoB and rpoD gene sequences of Mulet et al. in 2010. Identification of the isolates demonstrated a high diversity of Pseudomonas isolates at the source of the river located in a forest of which most isolates belonged to the Pseudomonas fluorescens lineage. On the other hand, the Pseudomonas population isolated at an anthropized site at the mouth of the river, receiving waste water from both households and industry, was very different and contained many Pseudomonas aeruginosa isolates.