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Cultivation of Important Methanotrophs From Indian Rice Fields.

ABSTRACT: Methanotrophs are aerobic to micro-aerophilic bacteria, which oxidize and utilize methane, the second most important greenhouse gas. The community structure of the methanotrophs in rice fields worldwide has been studied mainly using culture-independent methods. Very few studies have focused on culturing methanotrophs from rice fields. We developed a unique method for the cultivation of methanotrophs from rice field samples. Here, we used a modified dilute nitrate mineral salts (dNMS) medium, with two cycles of dilution till extinction series cultivation with prolonged incubation time, and used agarose in the solid medium. The cultivation approach resulted in the isolation of methanotrophs from seven genera from the three major groups: Type Ia (Methylomonas, Methylomicrobium, and Methylocucumis), Type Ib (Methylocaldum and Methylomagnum), and Type II (Methylocystis and Methylosinus). Growth was obtained till 10^-6-10^-8 dilutions in the first dilution series, indicating the culturing of dominant methanotrophs. Our study was supported by 16S rRNA gene-based next-generation sequencing (NGS) of three of the rice samples. Our analyses and comparison with the global scenario suggested that the cultured members represented the major detected taxa. Strain RS1, representing a putative novel species of Methylomicrobium, was cultured; and the draft genome sequence was obtained. Genome analysis indicated that RS1 represented a new putative Methylomicrobium species. Methylomicrobium has been detected globally in rice fields as a dominant genus, although no Methylomicrobium strains have been isolated from rice fields worldwide. Ours is one of the first extensive studies on cultured methanotrophs from Indian rice fields focusing on the tropical region, and a unique method was developed. A total of 29 strains were obtained, which could be used as models for studying methane mitigation from rice fields and for environmental and biotechnological applications.

SUBMITTER: Rahalkar MC

PROVIDER: S-EPMC8447245 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Rice-frog cultivation is a traditional farming system in China and has been reintroduced as an agricultural practice in China in recent years. The microbial community in paddy rhizospheric soils has attracted much attention because many microorganisms participate in functional processes in soils. In this study, Illumina MiSeq sequencing-based techniques were used to investigate soil microbial communities and functional gene patterns across samples obtained by conventional rice cultivation (CR) and rice-frog cultivation (RF). The results showed that RF significantly affected the microbial community composition and richness, which indicated that the rhizospheric microorganisms responded to the introduction of tiger frogs into the paddy fields. Operational taxonomic units (OTUs) from Sandaracinaceae, Anaerolineaceae, Candidatus Nitrososphaera, Candidatus Nitrosotalea, Candidatus Nitrosoarchaeum and some unclassified OTUs from Euryarchaeota and Agaricomycetes were significantly enriched by RF. The abiotic parameters soil organic carbon (SOC), nitrate nitrogen (NO3 --N), and available phosphorus (AP) changed under RF treatment and played essential roles in establishing the soil bacterial, archaeal, and fungal compositions. Correlations between environmental factors and microbial communities were described using network analysis. SOC was strongly correlated with Anaerolineaceae, Methanosaeta, and Scutellinia. NO3 --N showed strong positive correlations with Opitutus, Geobacter, and Methanosaeta. NH4 ++-N was strongly positively associated with Sideroxydans, and TN was strongly positively correlated with Candidatus Nitrotoga. Compared to conventional CR, RF greatly enriched specific microbial taxa. These taxa may be involved in the decomposition of complex organic matter and the transformation of soil nutrients, thus promoting plant growth by improving nutrient cycling. The unique patterns of microbial taxonomic and functional composition in soil profiles suggested functional redundancy in these paddy soils. RF could significantly affect the bacterial, archaeal, and fungal communities though changing SOC and AP levels.

Project description:The methanotrophs in rice field soil are crucial in regulating the emission of methane. Drainage substantially reduces methane emission from rice fields. However, it is poorly understood how drainage affects microbial methane oxidation. Therefore, we analyzed the dynamics of methane oxidation rates, composition (using terminal restriction fragment length polymorphism [T-RFLP]), and abundance (using quantitative PCR [qPCR]) of methanotroph pmoA genes (encoding a subunit of particulate methane monooxygenase) and their transcripts over the season and in response to alternate dry/wet cycles in planted paddy field microcosms. In situ methane oxidation accounted for less than 15% of total methane production but was enhanced by intermittent drainage. The dry/wet alternations resulted in distinct effects on the methanotrophic communities in different soil compartments (bulk soil, rhizosphere soil, surface soil). The methanotrophic communities of the different soil compartments also showed distinct seasonal dynamics. In bulk soil, potential methanotrophic activity and transcription of pmoA were relatively low but were significantly stimulated by drainage. In contrast, however, in the rhizosphere and surface soils, potential methanotrophic activity and pmoA transcription were relatively high but decreased after drainage events and resumed after reflooding. While type II methanotrophs dominated the communities in the bulk soil and rhizosphere soil compartments (and to a lesser extent also in the surface soil), it was the pmoA of type I methanotrophs that was mainly transcribed under flooded conditions. Drainage affected the composition of the methanotrophic community only minimally but strongly affected metabolically active methanotrophs. Our study revealed dramatic dynamics in the abundance, composition, and activity of the various type I and type II methanotrophs on both a seasonal and a spatial scale and showed strong effects of dry/wet alternation cycles, which enhanced the attenuation of methane flux into the atmosphere.

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Cultivation of Important Methanotrophs From Indian Rice Fields.

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