• Title/Summary/Keyword: Methanosarcina mazei

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Isolation and characterization of new Methanosarcina mazei strains KOR-3, -4, -5, and -6 from an anaerobic digester using pig slurry

  • Battumur, Urantulkhuur;Yoon, Youngman;Bae, Gui Sek;Kim, Chang-Hyun
    • Asian-Australasian Journal of Animal Sciences
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    • v.30 no.8
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    • pp.1198-1205
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    • 2017
  • Objective: An experiment was conducted to isolate and identify new methanogens in Korea from an anaerobic digester that uses pig slurry. Methods: An anaerobic digestate sample was collected from an anaerobic digester using pig slurry. Pre-reduced media were used for the growth and isolation of methanogens. Growth temperature range, pH range, NaCl concentration range, substrate utilization, and antibiotic tolerance were investigated to determine the physiological characteristics of isolated methanogens. The isolates were also examined microscopically for their morphology and Gram-stained. Polymerase chain reaction of 16S rRNA and mcrA gene-based amplicons was used for identification purpose. Results: Four strains, designated KOR-3, -4, -5, and -6, were isolated and were non-motile, irregular coccoid, and 0.5 to $1.5{\mu}m$ in diameter. Moreover, the cell walls of isolated strains were Gram-negative. KOR-3 and KOR-4 strains used acetate for methane production but did not use $H_2+CO_2$, formate, or methanol as a growth substrate KOR-5 and KOR-6 strains utilized acetate, methanol, and trimethylamine for methanogenesis but did not use $H_2+CO_2$ or formate as a growth substrate. The optimum temperature and pH for growth of four strains were $39^{\circ}C$ and 6.8 to 7.2, respectively. The optimum concentration of NaCl for growth of KOR-3, KOR-5, and KOR-6 were 1.0% (w/v). The optimum NaCl concentration for KOR-4 was 0.5% (w/v). All of the strains tolerated ampicillin, penicillin G, kanamycin, streptomycin, and tetracycline; however, chloramphenicol inhibited cell growth. Phylogenetic analysis of 16S rRNA and mcrA genes demonstrated that strains KOR-3, -4, -5, and -6 are related to Methanosarcina mazei (M. mazei, 99% sequence similarity). Conclusion: On the basis of physiological and phylogenetic characteristics, strains KOR-3, -4, -5, and -6 are proposed to be new strains within the genus Methanosarcina, named M. mazei KOR-3, -4, -5, and -6.

Study on Low Temperature Tolerant Methane-Producing Bacteria for the Treatment of Agricultural and Livestock Wastes;III. Isolation of Low Temperature Tolerant Methanogens (농축산(農畜産) 폐기물(廢棄物) 처리(處理)를 위(爲)한 저온내성(低溫耐性) 메탄 생성균(生成菌)의 특성(特性)에 관(關)한 연구(硏究);III. 저온내성(低溫耐性) Methanogens의 분리(分離))

  • Kim, Kwang-Yong;Kim, Jai-Joung;Daniels, Lacy
    • Korean Journal of Environmental Agriculture
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    • v.15 no.3
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    • pp.362-371
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    • 1996
  • This study was conducted to investigate the biochemical properties of isolated bacteria, low temperature tolerant methanogens which were selected for use as inoculum for anaerobic fermentation of agricultural and livestock wasted at low temperature. The results, obtained were summarized as follows: Low temperature tolerant methanogens were isolated from the samples which showed the high methanogenesis rate by enrichment culture at low temperature in methanol medium. These methanogens, Methanobacterium M-251 and Methanobacterium M-253 were isolated from swampy sediment at latitude $56.9^{\circ}$, Methanosarcina mazei M-372 from lake sediment IV at latitude $55.0^{\circ}$ N, and Methanobacterium formicicum M-375 from tidal land soil at latitude $37.0^{\circ}N$, respectively. The isolated anaerobic bacteria could not use sugars as carbon sources. The optimum pH value for the growth of M-251 and M-375 was 6.8, but those for M-253 and M-372 6.5 and 7.0, respectively. The minimum growth temperature of isolated, M-251 and M-253 were $8^{\circ}C$ and the optimum temperature $30^{\circ}C$, while the minimum of M-392 and M-395 were $13^{\circ}C$ and the optimum $37^{\circ}C$. The growth rate of isolates at $17.5^{\circ}C$ were lower by 32-50% than that of $30^{\circ}C$. The isolated Methanobacterium strains such as M-251, M-253, and M-375 have lower cell yield, 0.38-1.21g/1M $CH_4$ than 1.14-1.51g/1M $CH_4$ of Methanosarcina mazei M-372.

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