Isolation of Pectinase Producing Bacteria from the Rhizosphere of <i>Andrographis paniculata</i> Nees and 16S rRNA Gene Sequence Comparison of Some Potential Strains

Abstract

Pectinases, the enzymes which break down pectic substances, have a wide range of applications in food, agriculture and environmental sectors. In the present study, attempts were made to isolate highly efficient pectinase producer from the rhizosphere of a medicinal plant, Andrographis paniculata Nees, known as the “King of bitters”. The total heterotrophic bacterial count of the rhizosphere soil of A. paniculata Nees ranged from 1.53 × 109 to 2.52 × 109 cfu/g. A total of 65 bacterial colonies were randomly selected from the nutrient agar plates, purified and assessed for pectinase activity. Out of the 65 isolates, 62 (95.38%) showed varying degree of pectinase activity in plate assay using pectin as a sole source of carbon. Among the pectinase producing strains, JBST36 showed best pectinase activity which is followed by the JBST22 and JBST27. Morphological characterization, biochemical tests and 16S rRNA gene sequencing were performed to identify the three most potential strains. Based on the morphological, biochemical and molecular data, JBST22 was identified as Bacillus flexus and the other two were identified as Bacillus subtilis. Furthermore, nucleotide sequences of the 16S rRNA gene of these 3 strains were compared and a phylogenetic tree was constructed. The study reveals that there are at least 66 base differences in the 16S rRNA gene sequences of B. flexus JBST22 and the B. subtilis JBST36.