Molecular Characterization of Endophytic Bacteria from Some Medicinal Plants and Their Potential Application in Agriculture and Industry

Abstract

Medicinal plants are known to harbor diverse species of endophytic bacteria which are known for their secretion of beneficial secondary metabolites like enzymes and antimicrobial compounds. Medicinal plants of Artemisia annua, Ocimum lamiifolium, and Moringa oleifera are widely used in Ethiopia for medicinal, culinary, and nutritional applications. The current study aimed to isolate the endophytic bacteria of these medicinal plants and identify the strains using biochemical tests and 16S rRNA molecular sequencing and evaluate the ability of the isolates for important plant growth promotion features as well as their ability to produce industrially important exo-enzymes. In this study, 45 endophytic bacteria were isolated from the three medicinal plants, 15 from each medicinal plant’s leaf, stem, and root parts. Preliminary tests of Gram’s staining and Endospore staining were done. Biochemical tests including catalase test, citrate utilization test, H2S production, motility, methyl-red, voges-proskauer, indole, and urease tests were conducted for each of the isolates. Enzymatic activities of the endophytic bacteria such as the amylase, cellulase, pectinase, and protease producing ability were checked for each isolate. The plant growth-promoting abilities of the isolates like phosphate solubilization, zinc solubilization, and ACC deaminase production were checked. The secondary metabolites of isolates were extracted using ethyl acetate and the antibacterial activities of the extracts were evaluated. In this study, MALDI-TOF MS analysis of the isolated endophytic bacteria revealed the species diversity of the endophytic bacteria isolated from the medicinal plants. These endophytic bacterial isolates were found to be Bacillus cereus, Bacillus subtilis, Citrobacter freundii, Enterobacter asburiae, Enterobacter cloacae, Enterobacter kobei, Enterobacter ludwigii, Enterococcus faecium, and Pseudomonas monteilli. Bacillus and Enterobacter species were isolated in the recent study at the highest frequency. The most frequently isolated bacterial species were sent for 16S rRNA gene sequencing and found to be Enterobacter species with 99.47% sequence similarity to Enterobacter cloacae. This result was the same as the result obtained from MALDI-TOF MS analysis. Since this isolate was obtained from root part of Artemisia annua, its name is designated as Enterobacter sp. RPAAI8. This isolate was found to be capable of using certain cheap and cost-effective agro-wastes for its growth. The isolates were found to be excellent pectinase and protease producers which suggested the possibility of the isolates to be used in industrial applications. The isolates were also able to exhibit plant growth promoting attributes by solubilizing insoluble phosphate and zinc as well as ACC deaminase production. The ethyl acetate extract of the isolates showed inhibitory activity against test organisms.