Bacterial community of the colon.We pooled the colon samples of six representative cockroaches (see Fig. S1 in the supplemenatal material) and constructed a clone library of 16S rRNA genes. A total of 265 randomly selected clones were sequenced; 14 were putative chimeras and were excluded from further analysis. The remaining clones were assigned to 132 different phylotypes (>97% sequence similarity). Phylogenetic analysis revealed that the clones represented 11 bacterial phyla. Almost half of the clones belonged to Bacteroidetes (49 phylotypes), followed by Firmicutes (58 phylotypes, mostly Clostridia), and diverse Proteobacteria (12 phylotypes). Three phylotypes clustered among the Planctomycetes, two each among Deferribacteres, Elusimicrobia, and Actinobacteria, and only a single phylotype each among Fusobacteria, Chloroflexi, Synergistetes, and the candidate division TM7. Members of the Spirochaetes and Fibrobacteres, which are consistently found in the gut of termites, were absent from the clone library. Rarefaction analysis indicated that the number of clones analyzed was not sufficient to describe bacterial diversity within the cockroach gut at either genus (95% sequence similarity) or species level (97% sequence similarity) (Fig. S2); however, reasonable coverage was obtained at a sequence similarity threshold of 90%, indicating that the most abundant groups within the six individuals sampled were adequately covered.
Clones assigned to Bacteroidetes.The 120 clones assigned to Bacteroidetes fell mostly within the order Bacteroidales (Fig. 5). The most abundant phylotype was SL41 (5.2% of the library), which clustered together with SL35 (3.2%) and other phylotypes among a group consisting exclusively of termite and Cryptocercus clones in termite group IV of Bacteroidales. They represented a total of 28 clones in the library and were distantly related to bacteria in the genus Parabacteroides (92 to 94% sequence similarity to Parabacteroides distasonis from the mammalian gut). Another abundant group, represented by SL14 (4.0%), was loosely affiliated with clones from other intestinal sources, including Bacteroides cellulosilyticus (90 to 94% sequence similarity). Several phylotypes, with SL20 being the most abundant, formed a large cluster with clones in cluster V of Bacteroidales (4.4% of clones), which consists exclusively of clones from termites and Cryptocercus punctulatus, many of which represent symbionts of gut flagellates from Candidatus Azobacteroides pseudotrichonymphae (33). Phylotypes SL48 to SL51 clustered with sequences originating from the guts of higher termites within Bacteroidales cluster I (50), distantly related (91 to 93% sequence identity) to bacteria in the genus Alistipes.