The relationship between butirosin biosynthesis and certain biochemical characteristics related to sporulation in a strain of BaciUus circulans NRRL B-3313 was examined. The cellular content of dipicolinic acid increased while the amount of poly-13-hydroxybutyrate decreased with changes in antibiotic productivity.Oligosporogenous mutants failed to synthesize the antibiotic and to degrade poly-4-hydroxybutyrate. These observations suggest that spore formation may be related to antibiotic production in this strain of B. circulans.A close relationship between sporulation and the production of secondary metabolites in microorganisms has been demonstrated by biochemical and genetic analysis of some organisms. In bacilli, the polypeptide antibiotics produced have been found to affect the spore formation directly or indirectly (2,6,9,10,16,(18)(19)(20)23). It has also been observed that antibiotic production is closely related to the production of serine proteases during sporulation (14,23) and that some polypeptide antibiotics produced are specific inhibitors of RNA synthesis (10,18,19). The relationship between antibiotic production and morphological differentiation in Cephalosporium acremonium, a cephalosporin C producer (11,13,17), in Streptomyces lactamdurans, a cephamycin C producer (3), and in Penicillium urticae, a patulin and griseofulvin producer (21), has also been reported.The production of butirosin and another related compound, Bu-1975 (4'-deoxybutirosin), by a strain of Bacillus circulans ( Fig. 1) (4, 7) represents the only reported example of production by an organism other than the filamentous Actinomycetes. In this unique aminoglycoside-producing strain of bacillus, the relationship between the production of butirosin and several biochemical characteristics related to sporulation was studied.
MATERIALS AND METHODSMicroorganisms and cultivation. A strain of B. circulans NRRL B-3313 was used in this study. Four strains, B-3313-SM1 to B-3313-SM4, were independently isolated as spontaneous oligosporogenous mutants from B. circulans NRRL B-3313.The inoculum was prepared in 100 ml of medium (pH 7.5) containing 3.0% soybean meal, 0.4% ammonium chloride, and 0.5% calcium carbonate in a 500-ml Erlenmeyer flask and incubated at 30°C for 48 h on a rotary shaker at 300 rpm.The same medium supplemented with 4.0%o glycerol was seeded with 5% 48-h inoculum and fermented for 5 days under the same conditions used for the inoculum (4).Analytical procedures. To investigate the relationship between antibiotic production and sporulation, the production of butirosin and several other biochemical products related to the spore formation was measured after centrifugation of 120-h culture broth. The antibacterial activity of butirosin in the supematant was estimated by an agar diffusion method as reported earlier (12). Dipicolinic acid found in the spore * Corresponding author.coat was assayed by the method of Janssen et al. (5), and poly-p-hydroxybutyrate in the cell mass was determined by the procedure described by Law and ...