DEGRADATION OF N-ACYLHOMOSERINE LACTONE QUOURM SENSING SIGNAL BY BACILLUS THURINGIENSIS AHL LACTONASE

Abstract

Bacterial species within the community correspond with each other by producing, secreting, and detecting small diffusible molecules in a process known as quorum sensing (QS). Once these molecules reach the threshold concentration, they facilitate gene expression and cause phenotypic changes, including pigment production and others. N-acyl-homoserine lactone (AHL) is the most QS common molecule secreted by Gram-negative bacteria. On the other hand, some bacteria produce quorum sensing inhibitory (QSI) enzymes such as AHL lactonase degrade AHL molecular structure and disrupt the function of QS. This study focuses on the degradation of AHL molecules by AHL lactonase in Bacillus thuringiensis (Bt) strains isolated from Middle Tennessee. B. thuringiensis strains were grown overnight and diluted to OD600 of 2.0 as well as OD600 of 1.0. Chromobacterium violaceum (CV026) and Escherichia coli were used in the study as a biosensor and negative control, respectively. N- hexanoyl-homoserine lactone (C6-HSL), as well as N-3-oxo-hexanoyl homoserine lactone (3-oxo-C6-HSL), were applied to CV026 to stimulate the expression of pigment production. The outcome results showed that out of 62 Bt strains, 58 strains possess the aiiA gene: the gene responsible for AHL lactonase expression. Among the 58 strains carrying the aiiA gene, 48 strains grown overnight degrade 10.0 μM of C6- HSL molecules and inhibit pigment production. When the OD600 of 1.0 was tested on 5.0 μM C6-HSL, 23 strains inhibited pigment production. Moreover, when Bt strains were grown at OD600 of 1.0, and the concentration of C6-HSL was 10.0 μM, only 13 strains were able to degrade C6-HSL completely. Verification was done by incubating non-functional AHL at low pH, and whether CV026 produced the pigment. AHL lactonase in Bt strains only degrade C6-HSL but not 3-oxo-C6-HSL molecules. It was concluded that there were variations of degrading AHL molecules in Bt strains and some strains showed a potent activity on breaking down AHL molecules.