The gut microflora of Reticulitermes flavipes, its relation to oxygen, and evidence for oxygen-dependent acetogenesis by the most abundant Enterococcus sp

We previously demonstrated a potentially deep penetration of oxygen into the hindgut lumen of wood-feeding termites, high oxygen consumption rates of the symbiotic gut microbiota, and an indispensable role of oxygen in the mineralization of aromatic compounds. Here we characterize the gut microflora of the wood-feeding termite Reticulitermes flavipes, focusing on metabolic capacities and relation to oxygen of the most prevalent physiotypes. The carbohydrate-utilizing isolates, enumerated on liquid and solid growth media (3.1x10hoch5 cells per gut; gut liquid volume ca.0.3 Mikroliter), consisted of aerotolerant lactic acid bacteria (58%) and surprisingly large numbers of facultatively aerobic (20%) and even strictly aerobic bacteria (22%). The second largest group of cultivable gut bacteria were hydrogen-oxidizing methanogens (3.0x10hoch5 cells per gut), which were cultivated only on liquid media. Together, these two groups represented 10% of the total microscopic counts obtained by DAPI staining. The large number of lactic acid bacteria agreed with the metabolic product profiles in anoxic serial dilutions of gut homogenates in which lactate was the main product from glucose in the highest dilutions. Other physiological groups, e.g. propionate and butyrate producers, were much less abundant. Lactate was not, however, a major metabolite in the hindgut fluid, a phenomenon hitherto explained by a rapid turnover of lactate by other gut bacteria. In view of the relatively small number of lactate-oxidizing and lactate-fermenting bacteria, this explanation is not sufficient. We show that the isolate Enterococcus strain RfL6, representing the most abundant physiotype among the carbohydrate-utilizing gut bacteria, was not purely fermentative, but consumed oxygen during growth on glucose, accompanied by a complete shift in the product spectrum from lactate to acetate, and was able to oxidize lactate to acetate when oxygen was present. This agrees with the observation that acetate is the major product of the hindgut metabolism of R. flavipes (ca. 70 mM in the hindgut fluid). Together with previous findings, these results underline the obvious but so far neglected importance of oxygen and contribute to the emerging concept that the termite hindguts are far from simple, anoxic fermenters, but axially and radially structured, heterogeneous systems characterized by steep gradients of metabolites.