Reliable Identification of Environmental Pseudomonas Isolates Using the rpoD Gene.

The taxonomic affiliation ofPseudomonasisolates is currently assessed by using the 16S rRNA gene, MultiLocus Sequence Analysis (MLSA), or whole genome sequencing. Therefore, microbiologists are facing an arduous choice, either using the universal marker, knowing that these affiliations could be inaccurate, or engaging in more laborious and costly approaches. TherpoDgene, like the 16S rRNA gene, is included in most MLSA procedures and has already been suggested for the rapid identification of certain groups ofPseudomonas.However, a comprehensive overview of therpoD-based phylogenetic relationships within thePseudomonasgenus is lacking. In this study, we present therpoD-based phylogeny of 217 type strains ofPseudomonasand defined a cutoff value of 98% nucleotide identity to differentiate strains at the species level. To validate this approach, we sequenced therpoDof 145 environmental isolates and complemented this analysis with whole genome sequencing. TherpoDsequence allowed us to accurately assignPseudomonasisolates to 20 known species and represents an excellent first diagnostic tool to identify newPseudomonasspecies. Finally,rpoDamplicon sequencing appears as a reliable and low-cost alternative, particularly in the case of large environmental studies with hundreds or thousands of isolates.