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Abstract

Phylogenetically higher termites emit higher amounts of methane than lower termites, but the reason for this phenomenon has not been clear. Our comparative study based on 16S rRNA gene sequencing and qPCR analysis of archaeal communities in the guts of higher termites revealed that unlike the lower termites, which host mainly members of Methanobacteriales in their guts, higher termites host a diverse assemblage of methanogenic euryarchaeota comprising representatives of four major orders: Methanobacteriales, Methanosarcinales, Methanomicrobiales and the recently discovered Methanoplasmatales. 16S rRNA-based diversity of archaea was highest in soil-feeding taxa, where nearly all major archaeal groups were represented. Besides the euryarchaeotal lineages, the gut contained also lineages closely related to ammonia-oxidizing Thaumarchaeota and a deep-branching termite specific group of uncultured archaea loosely affiliated to Crenarchaeota. Archaeal diversity in the fungus-cultivating Macrotermes species, in the grass-feeding Trinervitermes sp., and in the wood-feeding Microcerotermes sp., which show low methane emission rates, was much lower. These results show the high methane emission rates in higher termites is reflected in the high diversity, density and complex community structure of archaea in the termite hindguts. Higher termites host gut-specific archaeal communities different from those of lower termites and from other environments and these communities seem to co-evolve with the host termite probably with shift in feeding behavior. Higher termites harbor archaeal lineages which are specific to their gut environment and are different from communities from lower termites and from other environments. Methanogenic archaea are heterogeneously distributed in the highly compartmented gut. Density and diversity of archaea was highest in posterior gut compartments, which also harbored most of the methanogenic activities. The highly alkaline anterior gut compartments were preferentially colonized by Methanosarcinales. Archaeal community structure differed strongly among gut compartments, with communities in the more methanogenic posterior gut sections being distinct from those of the anterior sections, a phenomenon that is reflected in the different micro-environmental conditions in the compartments. Experimental stimulation of methanogenesis in isolated gut sections of soil-feeding termites revealed significant activities of hydrogenotrophic methanogens that are obligately dependent on methanol and formate. Our results suggest that community structure in the different microhabitats is shaped by exogenous factors, such as pH, oxygen status and the availability of methanogenic substrates. The recently discovered Methanoplasmatales are the seventh order of methanogenic euryarchaeota comprising methylotrophic lineages which colonizes higher termite guts and various other environments, and helps to explain the high methane emission rates in higher termites. The methylotrophic nature of termite-derived lineage demonstrates that substrates other than hydrogen drive methanogenesis in higher termites.