報告書番号

U02034

タイトル（和文）

電気による微生物の制御（その7） --難培養性微生物単離のためのマイクロアレイ電気培養法の開発--

タイトル（英文）

Electrochemical control of microorganisms (part 7). -Development of microarray electrochemical cultivation for separation of uncultureable environmental bacteria.-

概要 （図表や脚注は「報告書全文」に掲載しております）

自然界に生息する、難培養性微生物単離のためのマイクロアレイ電気培養法の開発を行った。アレイ培養では、炭素板に設けた穴を微小な培養器（セル）として使用し、イオン交換膜を隔てた白金電極との間に通電する事で個々のセルを独立的に電気培養槽とした。本装置による高濃度培養を確認するため、植菌する微生物の希釈倍率を変化させ、嫌気的環境下で2週間の通電培養を実施した。その結果、すべての濃度領域において非通電時の10倍超の濃度の微生物が得られた。環境由来の混合微生物を希釈植菌し、0.85Vの通電の下マイクロアレイ電気培養に供したところ、1か月の培養により18個所中13個所で微生物の増殖が確認された。原子間力顕微鏡による微生物観察の結果、3種類以上の微生物が混在していた初期状態に対し、アレイ電気培養後には、単一の微生物が増殖している様子が確認され、従来不可能だった難培養性微生物の有効利用の足掛かりができた。

概要 （英文）

It is well known that there are many naturally occurring bacteria with useful properties. For example, bacteria have been identified capable degradation of toxic organic compounds, and reduction of harmful metal ions into their oxidized forms. Unfortunately, the difficulty of culturing naturally occurring bacteria has prevented investigating of all but a few species. In a recent study, we have investigated an electrochemical cultivation under anaerobic condition with Fe(III) ions and have been able to culture several previously uncultured species. In our previous work, those bacteria were grown as a mixture, but separation and isolation will be required to utilize them. We report here on a microarray electrochemical cultivation system developed for isolation of electrochemically cultureable environmental bacteria. The microarray electrochemical cultivation system consisted of eighteen holes made in a carbon plate. Each well works as small electrochemical cell with a volume of 250 microliters. A cathode bath with a Pt electrode was separated from the carbon plate anode by an ion exchange membrane and a membrane filter. This microarray system allows cell isolation by limiting dilution followed by electrochemical culturing. In order to evaluate a performance of the microarray system, an electrochemical cultivation of iron reducing bacterium Thiobacillus ferrooxidans (JCM7811), was undertaken. After 2 weeks cultivation under anaerobic conditions, 10-fold more cells were found in wells energized by +1.2 volts on the anode compared to unenergized wells. Cell isolation using the microarray electrochemical system was performed on a sample of bacterial cells from the environment. After 1 month of anaerobic cultivation under +0.85 V of electrochemical excitation, cell growths were observed on 13 wells. Visual inspection revealed only a single type of bacteria in 9 of the 13 wells, indicating that the microarray electrochemical cultivator is useful for isolating previously uncultureable bacteria from environment.

報告者

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