A Putatively New Family of Alphaproteobacterial Chloromethane Degraders from a Deciduous Forest Soil Revealed by Stable Isotope Probing and Metagenomics

Background: Chloromethane (CH3 Cl) is the most abundant halogenated organic compound in the atmosphere and substantially responsible for the destruction of the stratospheric ozone layer. Since anthropogenic CH 3 Cl sources have become negligible with the application of the Montreal Protocol (1987), natural sources, such as vegetation and soils, have increased proportionally in the global budget. CH3 Cl-degrading methylotrophs occurring in soils might be an important and overlooked sink.Results & Conclusions: The objective of our study was to link the biotic CH3 Cl sink with the identity of active microorganisms and their biochemical pathways for CH3 Cl degradation in a deciduous forest soil. When tested in laboratory microcosms, biological CH3 Cl consumption occurred in leaf litter, senescent leaves, and organic and mineral soil horizons. Highest consumption rates, around 2 mmol CH3 Cl g -1 dry weight h -1 , were measured in organic soil and senescent leaves, suggesting that top soil layers are active (micro-)biological CH 3 Cl degradation compartments of forest ecosystems. The DNA of these [13C]-CH3 Cl-degrading microbial communities was labelled using stable isotope probing (SIP), and the corresponding taxa and their metabolic pathways studied using high-throughput metagenomics sequencing analysis. [ 13C]-labelled Metagenome-Assembled Genome closely related to the family Beijerinckiaceae may represent a new methylotroph family of Alphaproteobacteria, which is found in metagenome databases of forest soils samples worldwide. Gene markers of the only known pathway for aerobic CH3 Cl degradation, via the methyltransferase system encoded by the CH3 Cl utilisation genes (cmu), were undetected in the DNA-SIP metagenome data, suggesting that biological CH3 Cl sink in this deciduous forest soil operates by a cmu-independent metabolism.