Mycobacterium smegmatis Grown in Static Culture

Mycobacterium smegmatis was grown on trace-metal-free medium in static culture. Throughout the growth phase, the concentration of mycobactin increased continuously, reaching a maximum of about 30 to 40 ,ug of mycobactin/mg of cell dry weight after 6 days; the concentration of salicylic acid remained approximately constant at 1 to 2 Mg of salicylic acid/gg of cell dry weight. Fe2+ (or Fe3+), Zn2+, Mn2+, and Mg2+ were all essential to a maximum formation of mycobactin. Optimum concentrations required were: Fe2+, about 1.8 sM; Mn2+ and Zn2+, about 0.5 Mm; and Mg2+, at least 0.17 mm. Higher levels of Fe2+ (9 to 90 gM) and Zn2+ (2 to 7 Mm) repressed mycobactin to about half the maximum value. No other cation or anion apparently is required for mycobactin biosynthesis. Salicylic acid concentration increased about fourfold when iron was omitted from the medium, but this is not as great as the increase reported previously for this strain of M. smegmatis. Mycobactin formation in another strain of M. smegmatis, NCIB 8548, showed similar dependencies on Fe2 , Zn2+, and Mn2 . Maximum accumulation of mycobactin with this strain was 85 ,g of mycobactin/mg of dry cell weight, under irondeficient (1.8 Mm Fe2+) conditions.