Abstract Anaerobic methanotrophic (ANME) archaea conserve energy from the breakdown of methane, an important driver of global warming, yet the extrachromosomal genetic elements that impact the activities of ANME archaea are little understood. Here we describe large plasmids associated with ANME archaea of the Methanoperedens genus. These have been maintained in two bioreactors that contain enrichment cultures dominated by different Methanoperedens species and co-occur with Methanoperedens species in other anoxic environments. By manual curation we show that two of the plasmids are large (155,607 bp and 191,912 bp), circular, and replicate bidirectionally. The group of Methanoperedens species that carry these plasmids is related to “ Ca . Methanoperedens nitroreducens”, “ Ca . Methanoperedens ferrireducens”, “ Ca . Methanoperedens manganicus" and the plasmids occur in the same copy number as the main chromosome. The larger plasmid encodes transporters that potentially enhance access to Ni, which is required for the methyl-CoM reductase (Mcr), Co required for the cobalamin cofactor needed for methyltransferases, and amino acid uptake. We show that many plasmid genes are actively transcribed, including genes involved in plasmid chromosome maintenance and segregation, a Co 2+ /Ni 2+ transporter and cell protective proteins. Notably, one plasmid carries three tRNAs and two colocalized genes encoding ribosomal protein uL16 and elongation factor eEF2. These are not encoded in the host Methanoperedens genome and uL16 and eEF2 were highly expressed, indicating an obligate interdependence between this plasmid and its host. The finding of plasmids of Methanoperedens opens the way for the development of genetic vectors that could be used to probe little understood aspects of Methanoperedens physiology. Ultimately, this may provide a route to introduce or alter genes that may enhance growth and overall metabolism to accelerate methane oxidation rates.

A widespread group of large plasmids in methanotrophic Methanoperedens archaea