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Among other attributes, the Betaproteobacterial genus  Azoarcus  has biotechnological importance for plant growth-promotion and remediation of petroleum waste-polluted water and soils. It comprises at least two phylogenetically distinct groups. The “plant-associated” group includes strains that are isolated from the rhizosphere or root interior of the C4 plant Kallar Grass, but also strains from soil and/or water; all are considered to be obligate aerobes and all are diazotrophic. The other group (now partly incorporated into the new genus  Aromatoleum ) comprises a diverse range of species and strains that live in water or soil that is contaminated with petroleum and/or aromatic compounds; all are facultative or obligate anaerobes. Some are diazotrophs. A comparative genome analysis of 32 genomes from 30  Azoarcus-Aromatoleum  strains was performed in order to delineate generic boundaries more precisely than the single gene, 16S rRNA, that has been commonly used in bacterial taxonomy. The origin of diazotrophy in  Azoarcus-Aromatoleum  was also investigated by comparing full-length sequences of  nif  genes, and by physiological measurements of nitrogenase activity using the acetylene reduction assay. Based on average nucleotide identity (ANI) and whole genome analyses, three major groups could be discerned: (i)  Azoarcus  comprising  Az. communis ,  Az. indigens  and  Az. olearius , and two unnamed species complexes, (ii)  Aromatoleum  Group 1 comprising  Ar. anaerobium ,  Ar. aromaticum ,  Ar. bremense , and  Ar. buckelii , and (iii)  Aromatoleum  Group 2 comprising  Ar. diolicum ,  Ar. evansii ,  Ar. petrolei ,  Ar. toluclasticum ,  Ar. tolulyticum ,  Ar. toluolicum,  and  Ar. toluvorans . Single strain lineages such as  Azoarcus  sp. KH32C,  Az. pumilus,  and  Az. taiwanensis  were also revealed. Full length sequences of  nif -cluster genes revealed two groups of diazotrophs in  Azoarcus-Aromatoleum  with  nif  being derived from  Dechloromonas  in  Azoarcus sensu stricto  (and two  Thauera  strains) and from  Azospira  in  Aromatoleum  Group 2. Diazotrophy was confirmed in several strains, and for the first time in  Az. communis  LMG5514,  Azoarcus  sp. TTM-91 and  Ar. toluolicum  T T . In terms of ecology, with the exception of a few plant-associated strains in  Azoarcus  (s.s.), across the group, most strains/species are found in soil and water (often contaminated with petroleum or related aromatic compounds), sewage sludge, and seawater. The possession of  nar ,  nap ,  nir ,  nor,  and  nos  genes by most  Azoarcus-Aromatoleum  strains suggests that they have the potential to derive energy through anaerobic nitrate respiration, so this ability cannot be usefully used as a phenotypic marker to distinguish genera. However, the possession of  bzd  genes indicating the ability to degrade benzoate anaerobically plus the type of diazotrophy (aerobic vs. anaerobic) could, after confirmation of their functionality, be considered as distinguishing phenotypes in any new generic delineations. The taxonomy of the  Azoarcus-Aromatoleum  group should be revisited; retaining the generic name  Azoarcus  for its entirety, or creating additional genera are both possible outcomes.

genes

Comparative Genomics Provides Insights into the Taxonomy of Azoarcus and Reveals Separate Origins of Nif Genes in the Proposed Azoarcus and Aromatoleum Genera