Quantitative real-time PCR monitoring dynamics of Thermotoga neapolitana in synthetic co-culture for biohydrogen production

Abstract This study demonstrates the potential for biohydrogen production in a co-culture of two ecologically distant species, Thermatoga neapolitana and Caldicellulosiruptor saccharolyticus , and the development of a quantitative real-time PCR (qPCR) method for quantifying the hyperthermophilic bacterium of the genus Thermotoga. Substrate utilization and H 2 production performance was compared to those of their individual cultures. The highest H 2 yields obtained were 2.7 ± 0.05, 2.5 ± 0.07 and 2.8 ± 0.09 mol H 2 /mol glucose for C. saccharolyticus , T. neapolitana, and their co-culture respectively. Statistical analysis comparing the H 2 production rate of the co-culture to either C. saccahrolyticus or T. neapolitana pure cultures indicated a significant difference in the H 2 production rate (p  t -test), with the highest rate of H 2 production (36.02 mL L −1  h −1 ) observed from the co-culture fermentations. In order to monitor the presence of T. neapolitana in the bioprocess, we developed a qPCR method using 16S rRNA gene and hydrogenase (hydA) gene targets. The qPCR data using hydA primers specific to T. neapolitana showed an increase in hydA gene copies from 3.32 × 10 7 to 4.4 × 10 8 hydA gene copies per mL confirming the influence of T. neapolitana in the synthetic consortium.