A Six-well Plate Method: Less Laborious and Effective Method for Cultivation of Obligate Anaerobic Microorganisms

• Published in: Microbes and Environments Vol. 26; no. 4; pp. 301 - 306
• Main Authors: Nakamura, Kohei, Tamaki, Hideyuki, Kang, Myung Suk, Mochimaru, Hanako, Lee, Sung-Taik, Nakamura, Kazunori, Kamagata, Yoichi
• Format: Journal Article
• Language: English
• Published: Japan Japanese Society of Microbial Ecology / Japanese Society of Soil Microbiology / Taiwan Society of Microbial Ecology / Japanese Society of Plant Microbe Interactions / Japanese Society for Extremophiles 2011
• Subjects: Anaerobiosis; Archaea; Archaea - growth & development; Archaea - isolation & purification; Bacteria - growth & development; Bacteria - isolation & purification; Colony Count, Microbial; gellan gum; methanogen; Microbiological Techniques - methods; sulfate-reducing bacterium (SRB); uncultured anaerobes
• ISSN: 1342-6311; 1347-4405
• DOI: 10.1264/jsme2.ME11120

We developed a simple, less laborious method to cultivate and isolate obligate anaerobic microorganisms using a six-well plate together with the AnaeroPack System, designated as the six-well plate method. The cultivation efficiency of this method, based on colony-forming units, colony formation time, and colony size, was evaluated with four authentic obligate anaerobes (two methanogenic archaea and two sulfate-reducing bacteria). The method was found to be comparable to or even better than the roll tube method, a technique that is commonly used at present for the cultivation of obligate anaerobes. Further experiments using 21 representative obligate anaerobes demonstrated that all examined anaerobes (11 methanogens, 5 sulfate- or thiosulfate-reducing bacteria, and 5 syntrophs) could form visible colonies on the six-well plate and that these colonies could be successfully subcultured in fresh liquid media. Using this method, an unidentified sulfate-reducing bacterium was successfully isolated from an environmental sample.