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Taxonomic decisions within the order  Rhizobiales  have relied heavily on the interpretations of highly conserved 16S rRNA sequences and DNA–DNA hybridizations (DDH). Currently, bacterial species are defined as including strains that present 95–96% of average nucleotide identity (ANI) and 70% of digital DDH (dDDH). Thus, ANI values from 520 genome sequences of type strains from species of  Rhizobiales  order were computed. From the resulting 270,400 comparisons, a ≥95% cut-off was used to extract high identity genome clusters through enumerating maximal cliques. Coupling this graph-based approach with dDDH from clusters of interest, it was found that: (i) there are synonymy between  Aminobacter lissarensis  and  Aminobacter carboxidus ,  Aurantimonas manganoxydans  and  Aurantimonas coralicida , “ Bartonella mastomydis ,” and  Bartonella elizabethae ,  Chelativorans oligotrophicus , and  Chelativorans multitrophicus ,  Rhizobium azibense , and  Rhizobium gallicum ,  Rhizobium fabae , and  Rhizobium pisi , and  Rhodoplanes piscinae  and  Rhodoplanes serenus ; (ii)  Chelatobacter heintzii  is not a synonym of  Aminobacter aminovorans ; (iii) “ Bartonella vinsonii ” subsp.  arupensis  and “ B. vinsonii ” subsp.  berkhoffii  represent members of different species; (iv) the genome accessions GCF_003024615.1 (“ Mesorhizobium loti  LMG 6125 T ”), GCF_003024595.1 (“ Mesorhizobium plurifarium  LMG 11892 T ”), GCF_003096615.1 (“ Methylobacterium organophilum  DSM 760 T ”), and GCF_000373025.1 (“ R. gallicum  R-602 sp T ”) are not from the genuine type strains used for the respective species descriptions; and v) “ Xanthobacter autotrophicus ” Py2 and “ Aminobacter aminovorans ” KCTC 2477 T  represent cases of misuse of the term “type strain”.  Aminobacter heintzii  comb. nov. and the reclassification of  Aminobacter ciceronei  as  A. heintzii  is also proposed. To facilitate the downstream analysis of large ANI matrices, we introduce here ProKlust (“Prokaryotic Clusters”), an R package that uses a graph-based approach to obtain, filter, and visualize clusters on identity/similarity matrices, with settable cut-off points and the possibility of multiple matrices entries.

Genomic Metrics Applied to Rhizobiales (Hyphomicrobiales): Species Reclassification, Identification of Unauthentic Genomes and False Type Strains