| Summary: | The genus <i>Pseudomonas</i> hosts an extensive genetic diversity and is one of the largest genera among Gram-negative bacteria. Type strains of <i>Pseudomonas</i> are well known to represent only a small fraction of this diversity and the number of available <i>Pseudomonas</i> genome sequences is increasing rapidly. Consequently, new <i>Pseudomonas</i> species are regularly reported and the number of species within the genus is constantly evolving. In this study, whole genome sequencing enabled us to define 43 new <i>Pseudomonas</i> species and provide an update of the <i>Pseudomonas</i> evolutionary and taxonomic relationships. Phylogenies based on the <i>rpoD</i> gene and whole genome sequences, including, respectively, 316 and 313 type strains of <i>Pseudomonas</i>, revealed sixteen groups of <i>Pseudomonas</i> and, together with the distribution of cyclic lipopeptide biosynthesis gene clusters, enabled the partitioning of the <i>P. putida</i> group into fifteen subgroups. Pairwise average nucleotide identities were calculated between type strains and a selection of 60 genomes of non-type strains of <i>Pseudomonas</i>. Forty-one strains were incorrectly assigned at the species level and among these, 19 strains were shown to represent an additional 13 new <i>Pseudomonas</i> species that remain to be formally classified. This work pinpoints the importance of correct taxonomic assignment and phylogenetic classification in order to perform integrative studies linking genetic diversity, lifestyle, and metabolic potential of <i>Pseudomonas</i> spp.
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