Phototrophic sulfur bacteria are generally able to use elemental sulfur as an electron donor for anoxygenic photosynthesis. Elemental sulfur is mainly a mixture of cyclo-octasulfur and polymeric sulfur. The purple sulfur bacterium Allochromatium vinosum strongly prefers the polymeric sulfur fraction showing that sulfur speciation has a strong influence on availability of elemental sulfur. X-ray absorption near edge structure (XANES) spectroscopy was used to investigate whether polymeric sulfur is also the preferred sulfur species in other purple sulfur bacteria belonging to the families Chromatiaceae and Ecothiorodospiraceae. The cultures were fed with 50 mM of elemental sulfur consisting of 68% polymeric sulfur and 30% cyclo-octasulfur. In all cultures, elemental sulfur was converted into intra- or extracellular sulfur globules, respectively, and further oxidized to sulfate. Sulfate concentrations were determined by HPLC and turbidometric assays, respectively. However, the added elemental sulfur was only partly used by the bacteria, one part of the 'elemental sulfur' remained in the cultures and was not taken up. XANES spectroscopy revealed that only the polymeric sulfur fraction was taken up by all cultures investigated. This strongly indicates that polymeric 'chain-like' sulfur is the form preferably used by phototrophic sulfur bacteria.