Antibacterial Activity of Compounds Derived from Endophytic Bacteria of Juniperus sabina: Enterobacter hormaechei subsp. Xiangfangensis and Bacillus tequilensis

BACKGROUND AND OBJECTIVESEndophytes are microorganisms that live inside plants and have been found to produce biologically active compounds. The study aimed to explore the antimicrobial properties of compounds synthesized by two endophytic bacteria, B. tequilensis and E. hormaechei subsp. xiangfangensis, isolated from the Juniperus sabina plant.MATERIALS AND METHODSThe isolation of endophytes involved a process of surface sterilization (۲.۵% sodium hypochlorite and ۷۰% ethanol) followed by cultivation and purification on NA medium. Identification was performed via DNA extraction, amplification of the ۱۶S rRNA region via PCR and sequencing by the Sanger method. To investigate the antimicrobial activity of the isolated bacteria, the supernatant of ۷ days culture in minimum media was clarified by centrifugation at ۱۲۰۰۰ rpm, ۴ ºC for ۲۰ minutes and dried using a rotary evaporator at ۴۰ ºC. Desalting and extraction was performed by methanol (۱:۱۰, W/V) and extracted materials were dried and kept at ۴ ºC. Thereafter, well diffusion method was followed by the determination of the Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) according to CLSI protocol with some modifications, against four bacterial standard strains.RESULTS AND DISCUSSIONChromaspro software was utilized to edit the sequenced genes. Sequences were blasted on the NCBI and EzBioCloud sites to identify the bacteria. Finally, genes of the two identified bacteria were submitted to the NCBI site . According to the results of well diffusion method supernatant of B. tequilensis and E. hormaechei subsp. xiangfangensis, cultures could inhibit Bacillus subtilis ATCC ۶۰۵۱ and Escherichia coli ATCC ۱۱۷۷۵ (۲۰ and ۱۵ mm for ۱۰ mm diameter wells, respectively). While the methanolic extract of B. tequilensis showed MIC values of ۱۲.۵ and ۲۵ mg/ml against S. aureus and B. subtilis, sample of E. hormaechei subsp. xiangfangensis demonstrated a MIC value of ۲۵ mg/ml against S. aureus. Salmonella typhi PTCC ۱۶۰۹ could not be inhibited by evaluated samples.CONCLUSIONThe study showed a mild antibacterial activity of two endophytic bacteria isolated from the J. sabina plant, and determination of effective antibacterial compound (s) will be performed in ongoing analysis with RP-HPLC in parallel with antibacterial assessment of purified compounds.

کلیدواژه ها:

Juniperus sabina ، Endophyte ، ۱۶S rRNA ، PCR ، MIC ، MBC