  A Bacillus subtilis derivative was obtained from strain ATCC 6633 by
replacement of the native promoter of the mycosubtilin operon by a constitutive
promoter originating from the replication gene repU of the Staphylococcus
aureus plasmid pUB110. The recombinant strain, designated BBG100, produced up
to 15-fold more mycosubtilin than the wild type produced. The overproducing
phenotype was related to enhancement of the antagonistic activities against
several yeasts and pathogenic fungi. Hemolytic activities were also clearly
increased in the modified strain. Mass spectrometry analyses of enriched
mycosubtilin extracts showed similar patterns of lipopeptides for BBG100 and
the wild type. Interestingly, these analyses also revealed a new form of
mycosubtilin which was more easily detected in the BBG100 sample. When tested
for its biocontrol potential, wild-type strain ATCC 6633 was almost ineffective
for reducing a Pythium infection of tomato seedlings. However, treatment of
seeds with the BBG100 overproducing strain resulted in a marked increase in the
germination rate of seeds. This protective effect afforded by mycosubtilin
overproduction was also visualized by the significantly greater fresh weight of
emerging seedlings treated with BBG100 compared to controls or seedlings
inoculated with the wild-type strain.
