Quantitative detection of culturable methanogenic archaea abundance in anaerobic treatment systems using the sequence-specific rRNA cleavage method

A method based on sequence-specific cleavage of rRNA with ribonuclease H was used to detect almost all known cultivable methanogens in anaerobic biological treatment systems. To do so, a total of 40 scissor probes in different phylogeny specificities were designed or modified from previous studies, optimized for their specificities under digestion conditions with 32 methanogenic reference strains, and then applied to detect methanogens in sludge samples taken from 6 different anaerobic treatment processes. Among these processes, known aceticlastic and hydrogenotrophic groups of methanogens from the families Methanosarcinaceae, Methanosaetaceae, Methanobacteriaceae, Methanothermaceae and Methanocaldococcaceae could be successfully detected and identified down to the genus level. Within the aceticlastic methanogens, the abundances of mesophilic Methanosaeta accounted for 5.7-48.5 of the total archaeal populations in mesophilic anaerobic processes, and those of Methanosarcina represented 41.7 of the total archaeal populations in thermophilic processes. For hydrogenotrophic methanogens, members of the Methanomicrobiales, Methanobrevibacter and Methanobacterium were detected in mesophilic processes (1.2-17.2), whereas those of Methanothermobacter, Methanothermaceae and Methanocaldococcaceae were detected in thermophilic process (2.0-4.8). Overall results suggested that those hierarchical scissor probes developed could be effective for rapid and possibly on-site monitoring of targeted methanogens in different microbial environments.