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- Active methanotrophs (1)
- Bacterial community (1)
- Cultivation (1)
- Methane oxidation (1)
- Oxic zone (1)
- PLFA (1)
- Paddy soil (1)
- Rice microcosms (1)
- Succession (1)
- pmoA gene (1)
Bacterial community succession corresponds to changes in the phylogenetic identity, growth-response time and rRNA operon (rrn) copy number of culturable populations. To test this hypothesis, we compared the bacterial fractions culturable from the oxic zone of flooded, unplanted paddy soil microcosms after 1-day (early succession) and 70-day (late succession) incubation periods. The proportion of bacteria that was cultivable on solid media corresponded for early and late succession to 3740% and 3135% of total DAPI cell counts, which were 7.40 (±0.36) × 108 and 5.54 (±0.28) × 108 cells per gram of dry soil, respectively. In colony-forming curve analysis, late successional bacteria showed a significant delay in their growth response compared with those from early succession. A total of 59 early successional isolates grouped into 16 species-level clusters (SLC) plus three Bacilli-like SLC, while 66 late successional isolates formed 25 SLC plus five Bacilli-like SLC. Except Bacilli-like spp., isolates from early succession always belonged to different SLC than those from late succession. Betaproteobacteria and Gammaproteobacteria were typical of the early stage, while Alphaproteobacteria and Actinobacteria prevailed in late succession. Considering all SLC except those assigned to Bacilli, growth-response time and rrn copy number were significantly correlated with successional stage. Isolates of most early successional SLC (14 of 16) formed visible colonies within 1 (11 SLC) or 2 days (three SLC) and contained = 4 rrn copies. In contrast, isolates of late successional SLC (23 of 25) formed visible colonies within 2 days (four SLC) or, in most cases, only within 315 days (19 SLC) and contained = 2 rrn copies. Regardless of whether isolated from early or late succession, Bacilli-like isolates always showed a colony-forming time of 2 days and had 911 rrn copies.
Methanotrophs in the rhizosphere of rice field ecosystems attenuate the emissions of CH4 into the atmosphere and thus play an important role for the global cycle of this greenhouse gas. Therefore, we measured the activity and composition of the methanotrophic community in the rhizosphere of rice microcosms. Methane oxidation was determined by measuring the CH4 flux in the presence and absence of difluoromethane as a specific inhibitor for methane oxidation. Methane oxidation started on day 24 and reached the maximum on day 32 after transplantation. The total methanotrophic community was analysed by terminal restriction fragment length polymorphism (T-RFLP) and cloning/sequencing of the pmoA gene, which encodes a subunit of particulate methane monooxygenase. The metabolically active methanotrophic community was analysed by stable isotope probing of microbial phospholipid fatty acids (PLFA-SIP) using 13C-labelled CH4 directly added to the rhizospheric region. Rhizospheric soil and root samples were collected after exposure to 13CH4 for 8 and 18 days. Both T-RFLP/cloning and PLFA-SIP approaches showed that type I and type II methanotrophic populations changed over time with respect to activity and population size in the rhizospheric soil and on the rice roots. However, type I methanotrophs were more active than type II methanotrophs at both time points indicating they were of particular importance in the rhizosphere. PLFA-SIP showed that the active methanotrophic populations exhibit a pronounced spatial and temporal variation in rice microcosms.