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Thermococcus  may be an important alternative source of H 2  in the hot subseafloor in otherwise low H 2  environments such as some hydrothermal vents and oil reservoirs. It may also be useful in industry for rapid agricultural waste treatment and concomitant H 2  production.  Thermococcus paralvinellae  grown at 82°C without sulfur produced up to 5 mmol of H 2  L −1  at rates of 5–36 fmol H 2  cell −1  h −1  on 0.5% (wt vol −1 ) maltose, 0.5% (wt vol −1 ) tryptone, and 0.5% maltose + 0.05% tryptone media. Two potentially inhibiting conditions, the presence of 10 mM acetate and low pH (pH 5) in maltose-only medium, did not significantly affect growth or H 2  production. Growth rates, H 2  production rates, and cell yields based on H 2  production were the same as those for  Pyrococcus furiosus  grown at 95°C on the same media for comparison. Acetate, butyrate, succinate, isovalerate, and formate were also detected as end products. After 100 h,  T. paralvinellae  produced up to 5 mmol of H 2  L −1  of medium when grown on up to 70% (vol vol −1 ) waste milk from cows undergoing treatment for mastitis with the bacterial antibiotic Ceftiofur and from untreated cows. The amount of H 2  produced by  T. paralvinellae  increased with increasing waste concentrations, but decreased in  P. furiosus  cultures supplemented with waste milk above 1% concentration. All mesophilic bacteria from the waste milk that grew on Luria Bertani, Sheep's Blood (selective for  Staphylococcus , the typical cause of mastitis), and MacConkey (selective for Gram-negative enteric bacteria) agar plates were killed by heat during incubation at 82°C. Ceftiofur, which is heat labile, was below the detection limit following incubation at 82°C.  T. paralvinellae  also produced up to 6 mmol of H 2  L −1  of medium when grown on 0.1–10% (wt vol −1 ) spent brewery grain while  P. furiosus  produced < 1 mmol of H 2  L −1 . Twelve of 13 enzyme activities in  T. paralvinellae  showed significant ( p  < 0.05) differences across six different growth conditions; however, methyl viologen-dependent membrane hydrogenase activity remained constant across all media types. The results demonstrate the potential of at least some  Thermococcus  species to produce H 2  if protein and α-glucosides are present as substrates.

Hydrogen Production and Enzyme Activities in the Hyperthermophile Thermococcus paralvinellae Grown on Maltose, Tryptone, and Agricultural Waste