A previous report described the presence of a self-splicing group I intron in a flagellin gene from a thermophilic Bacillus species. Here, we present evidence that the splicing reaction of the flagellin introns is dependent on temperature. Furthermore, a complementation analysis using a Bacillus subtilis flagellin-deficient mutant indicated that the intron-containing flagellin gene significantly restored the motility of the mutant at higher temperatures.