Diversity of Formyltetrahydrofolate Synthetases in the Guts of the Wood-Feeding Cockroach Cryptocercus punctulatus and the Omnivorous Cockroach Periplaneta americana

We examined the diversity of a marker gene for homoacetogens in two cockroach gut microbial communities. Formyltetrahydrofolate synthetase (FTHFS or fhs) libraries prepared from a wood-feeding cockroach, Cryptocercus punctulatus, were dominated by sequences that affiliated with termite gut treponemes. No spirochete-like sequences were recovered from the omnivorous roach Periplaneta americana, which was dominated by Firmicutes-like sequences.The guts of wood-feeding termites and Cryptocercus punctulatus cockroaches share an unusual pattern of electron flow, as high rates of CO₂-reductive acetogenesis typically supplant methanogenesis as the terminal electron sink (2, 3). Past studies have shown that from 10 to 30% of gut acetate produced in environments of termites and wood-feeding cockroaches is microbially generated from CO₂ (3, 28), ultimately powering 18 to 26% of the host insect''s own respiratory energy metabolism (25). Nevertheless, most termites emit methane (2), and termite emissions constitute approximately 4% of the global methane budget (27). Cockroaches have been proposed to represent an additional source of note (9). Interestingly, methanogenic termites and cockroaches exhibit increased acetogenesis following addition of exogenous H₂ (3, 29). This suggests that these insects are host to a robust population of bacteria that are capable of homoacetogenesis but may be primarily using alternative electron donors (and other substrates and pathways) in vivo.Acetogenic bacteria belonging to two bacterial phyla, Firmicutes and Spirochaetes, have been isolated from the guts of termites (1, 4, 11, 12, 14). Several surveys have targeted and used the gene for formyltetrahydrofolate synthetase (FTHFS), a key gene in the Wood-Ljungdahl pathway of acetogenesis (16), as a potential marker for the pathway (15, 18). For the wood-feeding termites that have been examined, the studies have revealed an abundance of FTHFS sequences that form a coherent phylogenetic cluster, together with genes from homoacetogenic termite gut spirochetes belonging to the genus Treponema (24, 26, 30). This suggests that treponemes may be among the more abundant of the homoacetogens active in these environments.Little is known about the population structure and biology of CO₂-reducing, acetogenic bacteria in the guts of either omnivorous or wood-feeding cockroaches. The wood-feeding cockroach Cryptocercus hosts an abundance of flagellate protozoa closely related to those believed to dominate polysaccharide fermentation in the guts of termites (5, 6, 22), suggesting that at least one key environmental niche is filled by similar microbes in both termites and Cryptocercidae. Additionally, Cryptocercidae cockroaches, like termites, house diverse spirochetes and are the site of intense CO₂ reduction into acetate (3, 7). Possibly, spirochetes capable of CO₂ reduction into acetate are present in the hindguts of cockroaches. However, no evidence has yet been presented for the existence of homoacetogenic treponemes in environments other than the guts of termites, and FTHFS surveys of human (21) or horse (15) fecal matter and bovine rumen samples (20) revealed only Firmicutes-like and other FTHFS alleles that are distinct from those comprising the termite treponeme cluster.Here, by examining FTHFS gene diversity in Cryptocercus punctulatus and Periplaneta americana guts, we endeavored to learn more about the distribution and origins of homoacetogenic treponemes (and their genes) that are found in wood-feeding termites. In particular, we wished to ascertain whether FTHFS genes present in either of the two cockroaches are termite treponeme-like and, if so, whether analysis reveals any obvious signal indicating recent or ancient lateral community transfer events between insect lineages.