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The possible role of chitinase in in vitro growth inhibition of the wheat pathogens Fusarium graminearum and Bipolaris sorokiniana by Bacillus pumilus SG2 was investigated. B. pumilus SG2, a chitinolytic bacterium producing two different chitinases, was previously isolated from the saline deserts of Iran. When grown in Spizizen salts medium with colloidal chitin, B. pumilus SG2 secreted two chitinases into the medium, resulting in growth inhibition of F. graminearum and abortion of hyphal elongation of B. sorokiniana. In contrast, when glucose was used as the carbon source, the chitinases were not expressed and antifungal activity of the B. pumilus SG2 was completely abolished. These results confirmed that expression of the B. pumilus SG2 chitinases is under the control of two types of regulation, special regulation by chitin and global regulation by glucose. We demonstrated that chitinases are the main components that caused hyphal inhibition activity of B. pumilus SG2. Hyphal inhibition of F. graminearum and B. sorokiniana was stable in agar for a minimum of 14 days.

Bacillus pumilus SG2 is a chitinolytic bacterium that produces two chitinases, namely ChiS and ChiL. The chiS and chiL genes are consecutively expressed under a common promoter. Regulation of the chiS and chiL genes is under the control of carbon catabolite repression (CCR) in B. pumilus. This study aimed to investigate the cis-acting elements of the chitinase promoter. For this purpose, we transferred the chiS gene along with its specific promoter to Bacillus subtilis as a host. Primer extension analysis revealed two transcription start sites located 287 and 65 bp upstream of the chiS start codon. The distal promoter was highly compatible with the consensus sequence of the σ(A)-type promoters in B. subtilis, whereas the proximal promoter sequence showed less similarity to the σ(A)-type consensus sequence. A catabolite responsive element (cre), which is required for CCR in Bacillus species, was found to be 136 to 123 bp upstream of the chiS start codon. Interestingly, this cre site was located upstream of the -35 of the proximal promoter and downstream of the distal promoter. Deletion of this cre site sequence rendered the chiS expression constitutive.