Xanthobacter tagetidis grew as a chemolithotrophic autotroph on thiosulfate and other inorganic sulfur compounds, as a heterotroph on thiophene-2-carboxylic acid, acetic acid and alpha-ketoglutaric acid, and as a mixotroph on thiosulfate in combination with thiophene-2-carboxylic acid and/or acetic acid. Autotrophic growth on one-carbon organosulfur compounds, and intermediates in their oxidation are also reported. Thiosulfate enhanced the growth yields in mixotrophic cultures, presumably by acting as a supplementary energy source, since ribulose bisphosphate carboxylase was only active in thiosulfate-grown cells and was not detected in mixotrophic cultures using thiosulfate with thiophene-2-carboxylic acid. Bacteria grown on thiophene-2-carboxylic acid also oxidized sulfide, thiosulfate and tetrathionate, indicating these as possible sulfur intermediates in thiophene-2-carboxylic acid degradation. Thiosulfate and tetrathionate were oxidized completely to sulfate and, consequently, did not accumulate as products of thiophene-2-carboxylic acid oxidation in growing cultures. Km and Vmax values for the oxidation of thiosulfate, tetrathionate or sulfide were 13 &mgr;M and 83 nmol O2 min-1 (mg dry wt.)-1, respectively; thiosulfate and tetrathionate became autoinhibitory at concentrations above 100 &mgr;M. The true growth yield (Ymax) on thiophene-2-carboxylic acid was estimated from chemostat cultures (at dilution rates of 0.034-0.094 h-1) to be 112.2 g mol-1, with a maintenance coefficient (m) of 0.3 mmol thiophene-2-carboxylic acid (g dry wt.)-1 h-1, and the maximum specific growth rate (&mgr;max) was 0.116 h-1. Growth in chemostat culture at a dilution rate of 0. 041 h-1 indicated growth yields [g dry wt. (mol substrate)-1] of 8.1 g (mol thiosulfate)-1, 60.9 g (mol thiophene-2-carboxylic acid)-1, and 17.5 g (mol acetic acid)-1, with additive yields for growth on mixtures of these substrates. At a dilution rate of 0.034 h-1, yields of 57.8 g (mol alpha-ketoglutaric acid)-1 and 60.7 g (mol thiophene-2-carboxylic acid)-1 indicated some additional energy conservation from oxidation of the thiophene-sulfur. SDS-PAGE of cell-free preparations indicated a polypeptide (Mr, 21.0 kDa) specific to growth on thiophene-2-carboxylic acid for which no function can yet be ascribed: no metabolism of thiophene-2-carboxylic acid by cell-free extracts was detected. It was shown that X. tagetidis exhibits a remarkable degree of metabolic versatility and is representative of facultatively methylotrophic and chemolithotrophic autotrophs that contribute significantly to the turnover of simple inorganic and organic sulfur compounds (including substituted thiophenes) in the natural environment.