Isolation, molecular characterization and investigation of antimicrobial potential of bacteria from intestines of some aquatic insect species

The gut flora of beetles plays an important role, especially in their adaptation to the environment. Recently, these flora members have become important resources for biotechnology. Insects belonging to the Hydrophilidae and Dytiscidae families are seen as a rich source of microorganisms because they are highly adapted to aquatic life. This thesis aimed to isolate bacteria from the gut of some aquatic beetles and screen antibacterial peptide-producing species to discover new antibacterial peptide-producing species. For this purpose, five different species belonging to Hydrophilidae and ten different species belonging to Dytiscidae were obtained in field studies conducted between July- September 2020. To determine the intestinal flora, dissections of beetles were performed under aseptic conditions, and gut samples were homogenized and spread on TSA medium. Molecular characterizations of bacteria were performed according to the 16SrDNA region. Aeromonas veronii, Bacillus pumilus, Carnobacterium divergens, Hafnia paralvei, Kurthia gibsonii, Lactococcus garvieae, Proteus hauseri, Providencia rettgeri, Pseudomonas otitidis, Serratia fonticola and S. liquefaciens from members of Hydrophilidae; Acinetobacter radioresistens, Aeromonas sp., A.allosaccharophila, A.veronii, B.pumilus, Carnobacterium divergens, Exiguobacterium mexicanum, Hafnia paralvei, Kurthia gibsonii, Lactococcus garvieae, Paenibacillus amylolyticus, Providencia rettgeri and P.otitidis species were obtained from Dytiscidae members. Culture filtrates were tested against Staphylococcus aureus and P.aeruginosa by well diffusion test. As a result of pre-selection, none of the culture filtrates caused zone formation against P.aeruginosa; 5 filtrates gave zones against S.aureus. Firstly, bacteria were screened by Trypsin/Proteinase K digestion method to confirm the activity. Secondly, the filtrates of the two selected bacteria were run on Native-Page gel to determine the protein band responsible for the activity, and a gel overlay test was performed. As a result, it was determined that the peptide of P.otitidis M29 with approximately 12 kDa showed antibacterial activity.