Campylobacter spp. are small, asaccharolytic bacteria exhibiting unique nutritional and environmental requirements. Campylobacter spp. exist as commensal organisms in some animal species, yet are estimated to be the most common causative agents of foodborne illness in humans. C. jejuni is most often associated with poultry, while C. coli are more frequently associated with swine. Temperature has been suggested to trigger potential colonization or virulence factors in C. jejuni, and recent studies have demonstrated temperature-dependent genes are important to colonization. It is possible that temperature-dependent colonization factors are in part responsible for the species-specific colonization characteristics of C. coli also. We determined utilization of 190 different sole carbon substrates by C. coli ATCC 49941 at 37 and 42A degrees C using phenotype microarray (PM) technology. Temperature did affect amino acid utilization. l-asparagine and l-serine allowed significantly (P = 0.004) more respiration by C. coli ATCC 49941 at the lower temperature of 37A degrees C as compared to 42A degrees C. Conversely, l-glutamine was utilized to a significantly greater extent (P = 0.015) at the higher temperature of 42A degrees C. Other organic substrates exhibited temperature-dependent utilization including succinate, d,l-malate, and propionate which all supported active respiration by C. coli to a significantly greater extent at 42A degrees C. Further investigation is needed to determine the basis for the temperature-dependent utilization of substrates by Campylobacter spp. and their possible role in species-specific colonization.