The hindgut of soil-feeding termites is highly compartmentalized and characterized by pronounced axial dynamics of the intestinal pH and microbial processes such as hydrogen production, methanogenesis, and reductive acetogenesis. Nothing is known about the bacterial diversity and the abundance or axial distribution of the major phylogenetic groups in the different gut compartments. In this study, we showed that the variety of physicochemical conditions is reflected in the diversity of the microbial communities in the different gut compartments of two Cubitermes species (Termitidae: Termitinae). 16S rRNA gene clones from the highly alkaline first proctodeal segment (

Methanogenesis represents an important electron sink reaction in the hindgut of soil-feeding termites. This is the first comprehensive analysis of the archaeal community structure within the highly compartmentalized intestinal tract of a humivorous insect, combining clonal analysis and terminal restriction fragment (T-RF) length polymorphism (T-RFLP) fingerprinting of the archaeal communities in the different gut compartments of Cubitermes orthognathus. We found that the morphological and physicochemical heterogeneity of the gut is reflected in a large phylogenetic diversity and pronounced axial differences in the composition of the archaeal gut microbiota, notably among those clones or ribotypes that could be assigned to methanogenic taxa. Comparative analysis of the relative frequencies of different archaeal lineages among the small-subunit rRNA gene (SSU rDNA) clones and their corresponding T-RF indicated that the archaeal community in the anterior, extremely alkaline hindgut compartment (P1) consists mainly of members of the Methanosarcinaceae, whereas Methanobacteriaceae and Methanomicrobiales predominate in the subsequent, more posterior compartments (P3/4a and P4b). The relative abundance of Thermoplasmales increased towards the rectum (P5). SSU rDNA sequences representing Crenarchaeota, which have not yet been reported to occur in the intestinal tracts of arthropods, were detected in all gut sections. We discuss how the spatial distribution of methanogenic populations may be linked to axial heterogeneity in the physicochemical gut conditions and to functional adaptations to their respective ecological niches.Termites are considered an important source of the climaterelevant greenhouse gas methane (for a review, see references 5 and 40), which is formed by methanogenic Archaea located in the enlarged hindguts of these insects. Methane emission rates differ strongly between termite species; these differences are closely correlated with the feeding habits of the respective taxa.In the hindgut of most wood-feeding species, homoacetogenesis is the major hydrogen sink reaction (8, 9). The methane emission rates of the extremely abundant and globally important soil-feeding species, however, exceed those of woodfeeding termites considerably. In the humivorous Termitinae, the rates of reductive acetogenesis measured in gut homogenates were about 10-fold lower than the rates of methanogenesis (8). Methanogenesis represents a major electron sink in the hindgut metabolism of these termites; in Cubitermes orthognathus, methane production amounts to almost 10% of the respiratory activity of the insect (46).Initially, the factors influencing the outcome of the competition of these CO 2 -reducing processes for hydrogen were quite enigmatic (9). However, several studies using microsensors and radiotracer analysis of intestinal CO 2 -reduction rates have provided evidence that the radial and axial distribution of the microbial populations involved in the production and consumption of hydrogen may play a key role in contro...

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Differences in microenvironment and interactions of microorganisms within and across habitat boundaries should influence structure and diversity of the microbial communities within an ecosystem. We tested this hypothesis using the well characterized gut tract of the European subterranean termite Reticulitermes santonensis as a model. By cloning and sequencing analysis and molecular fingerprinting (terminal restriction fragment length polymorphism), we characterized the bacterial microbiota in the major intestinal habitats - the midgut, the wall of the hindgut paunch, the hindgut fluid and the intestinal protozoa. The bacterial community was very diverse (> 200 ribotypes) and comprised representatives of several phyla, including Firmicutes (mainly clostridia, streptococci and Mycoplasmatales-related clones), Bacteroidetes, Spirochaetes and a number of Proteobacteria, all of which were unevenly distributed among the four habitats. The largest group of clones fell into the so-called Termite group 1 (TG-1) phylum, which has no cultivated representatives. The majority of the TG-1 clones were associated with the protozoa and formed two phylogenetically distinct clusters, which consisted exclusively of clones previously retrieved from the gut of this and other Reticulitermes species. Also the other clones represented lineages of microorganisms that were exclusively recovered from the intestinal tract of termites. The termite specificity of these lineages was underscored by the finding that the closest relatives of the bacterial clones obtained from R. santonensis were usually derived also from the most closely related termites. Overall, differences in diversity between the different gut habitats and the uneven distribution of individual phylotypes support conclusively that niche heterogeneity is a strong determinant of the structure and spatial organization of the microbial community in the termite gut.

The highly compartmentalized gut of soil-feeding termites is characterized by pronounced axial dynamics in physicochemical conditions and microbial processes. The hindgut of soil-feeding termites is highly compartmentalized and characterized by pronounced axial dynamics of oxygen, intestinal pH, and redox potential (1,3,4,14) and of microbial processes such as hydrogen production, methanogenesis, and reductive acetogenesis (26,29). Recent studies have provided strong evidence for an important role of the special physicochemical gut conditions and the pronounced intestinal microbiota in the chemical and microbial transformation of organic matter and microbial biomass during gut passage (11,12,13).A study of the phylogenetic diversity and axial distribution of microorganisms in the intestinal tract of Cubitermes orthognathus revealed that this variety of physicochemical conditions is reflected in the diversity of microbial communities in the different gut compartments and provided first evidence for the presence of a specific intestinal bacterial community (27).Although sequencing and phylogenetic analysis of cloned 16S rRNA genes provides information on the different phylotypes in a given community, the effort and costs involved in this approach limit investigations of changes in the structure of complex microbial communities over space and time. Molecular fingerprinting methods such as denaturing gradient gel electrophoresis (8,21,22) and terminal restriction fragment length polymorphism (T-RFLP) (9,18,19) analyses avoid these problems and allow the comparison of microbial communities in a larger number of samples, which has made these methods well-established tools in microbial ecology.A recent study combining clone analysis and T-RFLP to investigate archaeal community structure in the gut of the soil-feeding termite C. orthognathus provided evidence for pronounced differences among the microbiota not only between the gut and the ingested soil, but also among the different compartments of the intestinal tract (9). Based on the information on the phylogenetic diversity of the bacterial microbiota in the major gut compartments of C. orthognathus described in the companion paper (27), the present study employed T-RFLP analysis to follow changes in the bacterial community structure of ingested soil during gut passage. Comparing the terminal restriction fragment patterns of the homologous gut segments of three different species of Cubitermes, the study also addressed questions regarding the specificity of the gut microbiota for the compartments and the temporal stability of gut microbial communities during maintenance of termites under laboratory conditions. MATERIALS AND METHODSTermites and DNA extraction. Cubitermes orthognathus Emerson was collected in grassland near Busia, and Cubitermes ugandensis Fuller was collected in a glade of the Kakamega rainforest; both sites are located in the highlands of Western Kenya. Species were identified by Julius Muli, National Museums of Kenya. Cubitermes niokoloensis was coll...