The indigenous microbial communities inhabiting anoxic and moderately thermophilic fracture water from two kilometers below land surface in the Witwatersrand Basin, South Africa, were shown to consist predominantly of sulfate reducing bacteria (SRB) closely related to the genus Desulfotomaculum and methanogenic Euryarchaeota. Temporal trends in the aqueous geochemistry correlated with a phylogenetic shift in the microbial community structure where the overriding determining factor in the community composition was found to be the free energy flux, which confirmed the dominance of the sulfate reducing and methanogenic metabolic pathways. Further examination of the microporous quartzite adjacent to the fracture revealed the presence of extremely small sulfate reducing bacteria inhabiting submicron pores. Through an adapted microautoradiographic technique metabolic rates of 3.6x10 ^-2 – 9.7 fmol cell^-1 day^-1 were determined. Such rates suggest that the SRB are in a state of quiescence termed 'maintenance metabolism' and the rates are slightly higher than those measured in marine environments, albeit at lower sulfate concentration and higher temperature. To verify whether such a rate is compatible with cellular maintenance and whether maintenance metabolism imparts a distinctive δ³⁴S isotopic signature, a model, thermophilic SRB of the genus Desulfotomaculum was cultured in a biomass recycling vessel or 'retentostat' under severe electron donor limitation. Cultivation through this method allowed for a state of maintenance metabolism to be achieved with an average metabolic rate of 0.2 fmol cell^-1 day^-1. The S isotopic fractionation measured was ϵ³⁴S = 20.9‰, which was at least double the fractionation observed in corresponding batch culture experiments (ϵ³⁴S = 9.7‰) and is consistent with the results reported from the fracture water. The results also suggest that the final reduction of sulfite to sulfide in Desulfotomaculum putei is a multi-step and potentially reversible process – possibly leading to the larger fractionations observed by maximizing the reverse flow of isotopically heavy sulfur species during sulfate reduction.