A thermophilic bacterium producing xylose isomerase was isolated from Yunnan Tengchong Hot Spring water samples. The strain was finally identified as Anoxybacillus flavithermus WL based on morphological and 16S rDNA gene sequence analysis. Its optimal fermentation conditions for xylose isomerase were determined as follows:cultivation temperature 65℃,culture initial pH 8.0,inoculum density 3% and 15 g/L xylose for 24h. Under these conditions,enzyme activity reached 13.6 U/mL. Xylose isomerase from this strain performed the highest activity at 90℃.