Biological activities of two fungistatic antibiotics produced by Bacillus cereus UW85

Cultures and culture filtrates of Bacillus cereus UW85 suppress damping-off of alfalfa caused by Phytophthora medicaginis. We studied the role in disease suppression of two antibiotics from culture filtrates of UW85 that reversibly inhibited growth of P. medicaginis. We purified the two antibiotics by cation-exchange chromatography and high-voltage paper electrophoresis and showed that one of them, designated zwittermicin A, was an aminopolyol of 396 Da that was cationic at pH 7.0; the second, designated antibiotic B, appeared to be an aminoglycoside containing a disaccharide. Both antibiotics prevented disease of alfalfa seedlings caused by P. medicaginis. Purified zwittermicin A reversibly reduced elongation of germ tubes derived from cysts of P. medicaginis, and antibiotic B caused swelling of the germ tubes. Mutants generated with Tn917 or mitomycin C treatment were screened either for antibiotic accumulation in an agar plate diffusion assay or for the ability to suppress damping-off disease of alfalfa. Of 2,682 mutants screened for antibiotic accumulation, 5 mutants were substantially reduced in antibiotic accumulation and disease-suppressive activity. Of the 1,700 mutants screened for disease-suppressive activity, 3 mutants had reduced activity and they accumulated less of both antibiotics than did the parent strain. The amount of antibiotic accumulated by the mutants was significantly correlated with the level of disease suppression. Addition of either zwittermicin A or antibiotic B to alfalfa plants inoculated with a culture of a nonsuppressive mutant resulted in disease suppression. These results demonstrate that B. cereus UW85 produces two fungistatic antibiotics that contribute to suppression of damping-off disease of alfalfa