Strains of Clostidum thermoaceticum were tested for H2-and CO-dependent growth in a defined medium containing metals, minerals, vitamins, cysteine-sulfide, C02-bicarbonate, and H2 or CO. Ten of the thirteen strains tested grew at the expense of H2 and CO, and C. thermoaceticum ATCC 39073 was chosen for further study. The doubling times for H2-and CO-dependent growth under chemolithotrophic conditiods (the defined medium with nicotinic acid as sole essential vitamin and sulfide as sole reducer) were 25 and 10 h, respectively.Product stoichiometries for chemolithotrophic cultures approximated: 4.1H2 + 2.4CO2-+CH3COOH + 0.1 cell C + 0.3 unrecovered C and 6.8CO-*CH3COOH + 3.5CO2 + 0.4 cell C + 0.9 unrecovered C. H2-dependent growth produced signifiantly higher acetate concentrations per unit of biomass synthesized than did CO-or glucose-dependent growth. In dated the minimal nutritional requirements of this acetogen (37) so that a definitive assessment could be made of its heterotrophic and chemolithotrophic potentials. In the study presented here, numerous strains of C. thermoaceticum were obtained from various sources and evaluated. In addition, Acetogenium kivui (33, 34), a thermophilic nonclostridial acetogen which is capable of H2-dependent chemolithotrophic growth, was also included in this evaluation and used for comparative purposes. In this report, we demonstrate for the first time that certain strains of C. thermoaceticum grow chemolithotrophically (requiring only trace levels of nicotinic acid as the sole vitamin) at the expense of H2 or CO. Besides the overall metabolic properties exhibited by C. thermoaceticum and A. kivui under chemolithotrophic conditions, evidence is also presented which suggests that the type of energy source used during growth (e.g., H2 versus glucose) influences the expression or activity of hydrogenase and CO dehydrogenase in both of these acetogens. MATERIALS AND METHODSBacterial strains and cultivation. C. thermoaceticum (see Table 1 for strains used in this study) and A.

The nickel transport system of Clostridium thermoaceticum was investigated with 63NiC12 and an anaerobic microfiltration transport assay. Transport was optimal at pH 7 to pH 7.5 and 65°C and decreased in the presence of metabolic uncouplers and inhibitors. Exogenous nickel was concentrated 3,000-fold over the apparent nickel concentration gradient during typical transport assays. Stored Nickel is a biologically active trace metal, its biological roles ranging from the induction of carcinomas to the catalysis of essential met?abolic processes. In general, the biological activities of nickel are dependent upon cellular internalization of the metal (8). Two principal types of energy-dependent nickel transport system have been described (8, 10, 16). One is characterized by a high-affinity magnesium transporter which translocates other divalent cations (e.g., nickel) with decreased affinity. The second type is a high-affinity nickel transport system which is less affected by other divalent cations, in particular, magnesium.As an essential element of carbon monoxide (CO) dehydrogenase (acetyl coenzyme A synthetase), nickel plays a vital role in the Wood pathway of acetogenesis (6,9,20,33). In this study, we report that nickel translocation by Clostridium thermoaceticum (i) is energy dependent, (ii) is not inhibited by other divalent cations, and (iii) may be powered by cellular energy reserves.MATERIALS AND METHODS Cultivation. C. thermoaceticum ATCC 39073 was cultivated at 55°C in a defined medium containing low phosphates (3.7 mM) and 1 ,uM NiCl2 (as described previously, with the exclusion of yeast extract [5] (25). Protein was estimated by the Bradford method (2). Acetate was quantitated by highperformance liquid chromatography as previously described (3). Cellular extracts for polyacrylamide gel electrophoretic analysis were prepared by lysozyme digestion (23), and electrophoresis and in situ gel staining for CO dehydrogenase were done as previously described (7). 63NiCl2 was purchased from New England Nuclear Corp., Boston, Mass. RESULTS AND DISCUSSIONOptimal conditions for and kinetics of nickel transport. Cells from the mid-log phase of growth displayed the highest rates of nickel transport (data not shown), and subsequent studies were performed with cells obtained from this period of growth. Nickel transport was optimal at pH 7 to pH 7.5 and 65°C (data not shown). However, to more closely approximate the standard growth conditions of the organism, we conducted the standard nickel transport assay at pH