Composition and Dynamics of Methanogenic Archaeal Community in Japanese Paddy Field Soils.
Takeshi Watanabe, Makoto Kimura, and Susumu Asakawa. Graduate School of Bioagricultural Sciences, Nagoya Univ, Furocho, Chikusa-ku, Nagoya, Japan
Methane is one of greenhouse gases and of final degradation products of organic materials in anoxic environments. Methanogenic archaea monopolize biological methane production in those environments. Paddy field is one of major sources for methane emission and released methane contributes to global warming. Therefore, it is important to understand the ecology of methanogenic archaea in paddy fields. In this study, the composition and the dynamics of methanogenic archaea in paddy field soils under double cropping (rice [Oryza sativa L.] and wheat [Triticum aestivum L.]) were studied by denaturing gradient gel electrophoresis (DGGE) method and real-time RT-PCR, targeting 16S rDNA and 16S rRNA. Soil samples were collected from two paddy fields (Anjo [Anthraquic Yellow Soil; Oxyaquic Dystrudept] and Chikugo [Gray Lowland Soil; Endoquepts] in central and south Japan, respectively) under flooded and upland conditions throughout a year in 2003. Total DNAs and RNAs were extracted by bead-beating method separately, and two primer sets (0357F-GC/0691R and 1106F-GC/1378R) were used for DGGE analysis. real-time RT-PCR was carried out with the primers 1106F/1378R. DGGE band patterns of the methanogenic archaeal community obtained from 16S rDNA and 16S rRNA were stable throughout the year, which included cultivation periods of rice under flooded conditions and of wheat under upland conditions, in the both sampling sites. Almost all the bands detected on DGGE gels of 16S rDNA were also observed on the gels of 16S rRNA. Cluster analysis of DGGE band patterns showed that the soil type (sampling site; Anjo and Chikugo) mainly influenced the community structures of methanogenic archaea in paddy field soil. Totally 51 sequences were retrieved from the DGGE gels of 16S rDNA with the two primer sets. Most of the sequences were closely related to Methanomicrobiales, Methanosarcinaceae, Methanosaetaceae and uncultured Rice cluster I, suggesting that these species of methanogenic archaea were predominated in the paddy fields. The copy number of methanogenic archaeal 16S rRNAs estimated by real-time RT-PCR did not fluctuate between flooded and upland conditions. In addition, methanogenic archaeal 16S rRNAs were detected in air-dry soils collected from the paddy fields in 1990 (Chikugo) and 2005 (Anjo). These findings suggested that strictly anaerobic methanogenic archaea survive in oxic soils and/or the activity of methanogenic archaea is maintained under upland conditions.