Flow-FISH analysis and isolation of clostridial strains in an anaerobic semi-solid bio-hydrogen producing system by hydrogenase gene target.
By using hydrogenase gene-targeted polymerase chain reaction (PCR) and reverse transcriptase PCR (RT-PCR), the predominant clostridial hydrogenase that may have contributed to biohydrogen production in an anaerobic semi-solid fermentation system has been monitored. The results revealed that a Clostridium pasteurianum-like hydrogenase gene sequence can be detected by both PCR and RT-PCR and suggested that the bacterial strain possessing this specific hydrogenase gene was dominant in hydrogenase activity and population. Whereas another Clostridium saccharobutylicum-like hydrogenase gene can be detected only by RT-PCR and suggest that the bacterial strain possessing this specific hydrogenase gene may be less dominant in population. In this study, hydrogenase gene-targeted fluorescence in situ hybridization (FISH) and flow cytometry analysis confirmed that only 6.6% of the total eubacterial cells in a hydrogen-producing culture were detected to express the C. saccharobutylicum-like hydrogenase, whereas the eubacteria that expressed the C. pasteurianum-like hydrogenase was 25.6%. A clostridial strain M1 possessing the identical nucleotide sequences of the C. saccharobutylicum-like hydrogenase gene was then isolated and identified as Clostridium butyricum based on 16S rRNA sequence. Comparing to the original inoculum with mixed microflora, either using C. butyricum M1 as the only inoculum or co-culturing with a Bacillus thermoamylovorans isolate will guarantee an effective and even better production of hydrogen from brewery yeast waste.
Jen
Chang Jui
CJ
Department of Life Sciences, National Chung Hsing University, Taichung 40227, Taiwan.
Chou
Chia-Hung
CH
Hsu
Ping-Chi
PC
Yu
Sian-Jhong
SJ
Chen
Wei-En
WE
Lay
Jiunn-Jyi
JJ
Huang
Chieh-Chen
CC
Wen
Fu-Shyan
FS
D000693:Anaerobiosis; D001483:Base Sequence; D003013:Clostridium / Q000145:classification / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification*; D005434:Flow Cytometry; D006859:Hydrogen / Q000378:metabolism*; D006864:Hydrogenase / Q000235:genetics*; D017404:In Situ Hybridization, Fluorescence; D008969:Molecular Sequence Data
D006859:Hydrogen; D006864:Hydrogenase