Characterization of a 4-methylbenzoate-degrading methanogenic consortium as determined by small-subunit rDNA sequence analysis.

A methanogenic consortium that degrades 4-methylbenzoate (MBA) as the sole carbon and energy source was successfully enriched in an upflow anaerobic sludge bed bioreactor and studied. Electron microscopic observation showed that long rods with a distinct collar feature resembling Desulfomonile tiedjei rods were the predominant population, and that these rods formed a close spatial orientation with Methanobrevibacter-like bacteria. In addition, thin filaments and bamboo-shaped filaments that highly resembled the acetoclastic Methanosaeta were also frequently observed. A 16S rDNA clone library was constructed for the domain Bacteria, and 20 sequence types or operational taxonomic units (OTUs) were found out of 139 clones screened. Phylogenetic analysis classified these 20 nearly full-length OTUs into the delta (50.3% of total clones) and gamma (4.3%) subdivisions of the division Proteobacteria, the green non-sulfur bacteria subdivision I (7.2%), Cytophagales (7.2%), Planctomycetes (5.7%), gram-positive low G + C group (8.6%), candidate divisions OP8, OP10 and OP11 (9.3%), and a novel candidate division MBA1 (7.2%) that had an interdivisional sequence similarity less than 75%. However, only 3 OTUs had a sequence similarity higher than 90% to known isolates or environmental 16S rDNA clones, suggesting that the microbial community was diversified and largely unidentified. In particular, those 8 OTUs found in the delta-Proteobacteria were either clustered into novel groups or showed a low sequence similarity to closely related bacteria. It is highly possible that the delta-Proteobacteria were the long rods with a distinct collar feature observed microscopically, and together with the methanogens were mainly responsible for the syntrophic degradation of MBA. The unique and novel microbial populations identified explained the requirement of a long start-up period of up to 426 d for the MBA-degrading consortium.