Design and validation of a multiplex PCR method for the simultaneous quantification of Clostridium acetobutylicum, Clostridium carboxidivorans and Clostridium cellulovorans.

Co-cultures of clostridia with distinct physiological properties have emerged as an alternative to increase the production of butanol and other added-value compounds from biomass. The optimal performance of mixed tandem cultures may depend on the stability and fitness of each species in the consortium, making the development of specific quantification methods to separate their members crucial. In this study, we developed and tested a multiplex qPCR method targeting the 16S rRNA gene for the simultaneous quantification of Clostridium acetobutylicum, Clostridium carboxidivorans and Clostridium cellulovorans in co-cultures. Designed primer pairs and probes could specifically quantify the three Clostridium species with no cross-reactions thus allowing significant changes in their growth kinetics in the consortia to be detected and correlated with productivity. The method was used to test a suitable medium composition for simultaneous growth of the three species. We show that higher alcohol productions were obtained when combining C. carboxidivorans and C. acetobutylicum compared to individual cultures, and further improved (> 90%) in the triplet consortium. Altogether, the methodology could be applied to fermentation processes targeting butanol productions from lignocellulosic feedstocks with a higher substrate conversion efficiency.

Bacteria coated cathodes as an in-situ hydrogen evolving platform for microbial electrosynthesis.

Hydrogen is a key intermediate element in microbial electrosynthesis as a mediator of the reduction of carbon dioxide (CO2) into added value compounds. In the present work we aimed at studying the biological production of hydrogen in biocathodes operated at - 1.0 V vs. Ag/AgCl, using a highly comparable technology and CO2 as carbon feedstock. Ten bacterial strains were chosen from genera Rhodobacter, Rhodopseudomonas, Rhodocyclus, Desulfovibrio and Sporomusa, all described as hydrogen producing candidates. Monospecific biofilms were formed on carbon cloth cathodes and hydrogen evolution was constantly monitored using a microsensor. Eight over ten bacteria strains showed electroactivity and H2 production rates increased significantly (two to eightfold) compared to abiotic conditions for two of them (Desulfovibrio paquesii and Desulfovibrio desulfuricans). D. paquesii DSM 16681 exhibited the highest production rate (45.6 ± 18.8 µM min-1) compared to abiotic conditions (5.5 ± 0.6 µM min-1), although specific production rates (per 16S rRNA copy) were similar to those obtained for other strains. This study demonstrated that many microorganisms are suspected to participate in net hydrogen production but inherent differences among strains do occur, which are relevant for future developments of resilient biofilm coated cathodes as a stable hydrogen production platform in microbial electrosynthesis.