Combined Omics Approach Reveals Key Differences between Aerobic and Microaerobic Xylene-Degrading Enrichment Bacterial Communities: ─A Hitherto Unknown Player Emerges from the Microbial Dark Matter.

Among monoaromatic hydrocarbons, xylenes, especially the and isomers, are the least biodegradable compounds in oxygen-limited subsurface environments. Although much knowledge has been gained regarding the anaerobic degradation of xylene isomers in the past 2 decades, the diversity of those bacteria which are able to degrade them under microaerobic conditions is still unknown. To overcome this limitation, aerobic and microaerobic xylene-degrading enrichment cultures were established using groundwater taken from a xylene-contaminated site, and the associated bacterial communities were investigated using a polyphasic approach. Our results show that the xylene-degrading bacterial communities were distinctly different between aerobic and microaerobic enrichment conditions. Although members of the genus were the most dominant in both types of enrichments, the and lineages were only abundant under microaerobic conditions, while entirely replaced them under aerobic conditions. Analysis of a metagenome-assembled genome of a -related bacterium revealed aromatic hydrocarbon-degrading ability by identifying two catechol 2,3-dioxygenases in the genome. Moreover, phylogenetic analysis indicated that both enzymes belonged to a newly defined subfamily of type I.2 extradiol dioxygenases (EDOs). Aerobic and microaerobic xylene-degradation experiments were conducted on strains sp. AS12 and sp. MAP12, isolated from the aerobic and microaerobic enrichments, respectively. The obtained results, together with the whole-genome sequence data of the strains, confirmed the observation that members of the genus are excellent aromatic hydrocarbon degraders but effective only under clear aerobic conditions. Overall, it was concluded that the observed differences between the bacterial communities of aerobic and microaerobic xylene-degrading enrichments were driven primarily by (i) the method of aromatic ring activation (monooxygenation vs dioxygenation), (ii) the type of EDO enzymes, and (iii) the ability of degraders to respire utilizing nitrate.