Using genome comparisons of wild-type and resistant mutants of to help understand mechanisms of resistance to methane inhibitors.

Methane emissions from enteric fermentation in the ruminant digestive system generated by methanogenic archaea are a significant contributor to anthropogenic greenhouse gas emissions. Additionally, methane produced as an end-product of enteric fermentation is an energy loss from digested feed. To control the methane emissions from ruminants, extensive research in the last decades has been focused on developing viable enteric methane mitigation practices, particularly, using methanogen-specific inhibitors. We report here the utilization of two known inhibitors of methanogenic archaea, neomycin and chloroform, together with a recently identified inhibitor, echinomycin, to produce resistant mutants of S2 and S0001. Whole-genome sequencing at high coverage (> 100-fold) was performed subsequently to investigate the potential targets of these inhibitors at the genomic level. Upon analysis of the whole-genome sequencing data, we identified mutations in a number of genetic loci pointing to potential mechanisms of inhibitor action and their underlying mechanisms of resistance.