Bitespiramycin (BT), a multi-component antibiotic consisted mainly of 4"-isovalerylspiramycin I, II and III, is produced by Streptomyces spiramyceticus WSJ-1, a recombinant spiramycin-production strain that harbored the 4"-O-acyltransferase gene (ist) from Streptomyces mycarofaciens 1748, which could isovalerylate the 4"-OH of spiramycin. To eliminate the production of components 4"-isovalerylspiramycin II and III, therefore reducing the component complexity of BT, inactivation of the sspA gene, which encodes the 3-O-acyltransferase responsible for the acylation of spiramycin I to spiramycin II and III, was performed in Streptomyces spiramyceticus WSJ-1, by in-frame partial deletion. The resulting strain, Streptomyces spiramyceticus WSJ-2, is a 4"-isovalerylspiramycin-I-producing strain as expected.