Many microorganisms live in anaerobic environments. Most of these microorganisms have not yet been cultivated. Here, we present, from a metagenomic analysis of an anaerobic digester of a municipal wastewater treatment plant, a reconstruction of the complete genome of a bacterium belonging to the WWE1 candidate division. In silico proteome analysis indicated that this bacterium might derive most of its carbon and energy from the fermentation of amino acids, and hence, it was provisionally classified as "Candidatus Cloacamonas acidaminovorans." "Candidatus Cloacamonas acidaminovorans" is probably a syntrophic bacterium that is present in many anaerobic digesters. This report highlights how environmental sequence data might provide genomic and functional information about a new bacterial clade whose members are involved in anaerobic digestion.The use of molecular techniques over the past few decades has shown the extent of microbial diversity and that the majority of archaeal and bacterial phyla still lack a cultivable representative (18,24). The recent advent of whole-community genome sequencing, or metagenomics, is rapidly changing this view and will revolutionize our understanding of the functional diversity of complex environments. For instance, a recent study has surprisingly revealed that ammonia-oxidizing Crenarchaeota are very abundant in soils (20). However, apart from the pioneering work in the Sargasso Sea (31) and the recent addition of millions of sequences from the Global Ocean Sampling expedition that revealed the extent of the ocean microbial diversity (25), most of the community sequencing programs have focused on relatively simple ecosystems. However, most microbial ecosystems are complex. Anaerobic digestion is a complex biological process that involves several metabolic pathways for the decomposition of organic matter into methane and carbon dioxide. The overall reactions-depolymerization, primary and secondary fermentation, acidogenesis, acetogenesis, and methanogenesis-are performed by a complex microbial community. Despite the industrial, technological, economic, and ecological importance of this community, little is known about the roles and activities of the microorganisms that inhabit anaerobic niches. The anaerobic digestion of organic matter involved in wastewater processing represents a good example of a complex and active microflora. During exploration of the bacterial diversity of an anaerobic mesophilic digester, a new bacterial candidate division called WWE1 was discovered (8). It was found that WWE1 bacteria could represent up to 10% of the bacterial microflora and thus could be a subdominant group. Using metagenomic sequence data and a specific genome assembly procedure, we were able to reconstruct the genome of a representative bacterium of the WWE1 division. We have for the first time obtained the complete genome sequence from a complex environment and from a bacterial candidate division with no cultivated representative. Because the metabolic pathways of anaerobic bacteria are ...

Cloacibacillus evryensis gen. nov., sp. nov., a novel asaccharolytic, mesophilic, amino-acid-degrading bacterium within the phylum 'Synergistetes', isolated from an anaerobic sludge digester T were non-motile, rodshaped (2.0-3.0 ¾ 0.8-1.0 mm) and stained Gram-negative. Optimal growth occurred at 37 6C and pH 7.0 in an anaerobic basal medium containing 1 % Casamino acids. Strain 158 T fermented arginine, histidine, lysine and serine and showed growth on yeast extract, brain-heart infusion (BHI) medium and tryptone, but not on carbohydrates, organic acids or alcohols. The end products of degradation were: acetate, butyrate, H 2 and CO 2 from arginine; acetate, propionate, butyrate, H 2 and CO 2 from lysine; and acetate, propionate, butyrate, valerate, H 2 and CO 2 from histidine, serine, BHI medium, Casamino acids and tryptone. The DNA G+C content was 55.8 mol%. The 16S rRNA gene sequence of strain 158 T showed only 92.6 % sequence similarity with that of Synergistes jonesii, the only described species of the 'Synergistes' group. The major cellular fatty acids were iso-C 15 : 0 (16.63 %), iso-C 15 : 0 3-OH (12.41 %) and C 17 : 1 v6c (9.46 %) and the polar fatty acids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and phosphatidylmonomethylamine; these fatty acid profiles did not resemble those of any recognized bacterial species. Due to the considerable differences in genotypic, phenotypic and phylogenetic characteristics between strain 158 T and those of its nearest relative, it is proposed that strain 158 T represents a novel species in a new genus, Cloacibacillus evryensis gen. nov., sp. nov., in the phylum 'Synergistetes'. The type strain is 158Proteins, as well as carbohydrates and lipids, comprise a major part of the organic load in anaerobic sludges and wastewaters. For example, the protein component of dairy wastewater and domestic sewage is reportedly greater than 40 % (Ramsay & Pullammanappallil, 2001). Protein hydrolysis in anaerobic environments has not been well studied except in the animal gut, e.g. the rumen (Debroas & Blanchart, 1993), and has been restricted to molecular inventories in protein-containing wastewaters (Tang et al., 2005). The major difference between protein degradation in the rumen and in anaerobic digesters is that, in the rumen, the ruminal bacteria (both saccharolytic and asaccharolytic) grow on amino acids and peptides as majorAbbreviations: CFA, cellular fatty acid; DHP, 3,4-dihydroxypyridine; SGOs, Synergistes group of organisms; VFA, volatile fatty acid; WWTP, wastewater treatment plant.The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of Cloacibacillus evryensis strain 158 T is CU463952.