Polyphasic systematics of marine bacteria and their alpha-glucosidase inhibitor activity

Abstract

Microbes are the most diverse group of organisms present on the earth. Microbial diversity is the variety and variability of an organism’s and the ecosystem in which they occur. Microbes have been adapted to global environmental changes as and when it happened on the earth.The enormous microbial diversity also gives rise to a largely untapped amount of genetic information, bioactive compounds and biomaterials which could deliver important beneficial traits and applications of societal interest, for example, to improve the medical treatments,fisheries and aquaculture applications, supply of energy for the development of industrial products and processes. It has been estimated that, only one to five per cent of microorganisms have been studied extensively. Identification of a variety of microorganisms is utmost important as it describes the positioning of an organism into the phylogenetic group and various biotechnological applications are dependent on known microorganisms. Microbial taxonomy is the branch of science that deals with the study of identification, classification and nomenclature of an organism. It classifies and arranges the microorganisms into taxonomical order. With the advancement of molecular, chromatographic and microscopy techniques, knowledge and the number of well characterized microorganisms in taxonomy has been increased in the past two decades. Polyphasic systematics is the recent approach used in the identification of bacteria. It comprises phylogenetic, phenotypic, genotypic and chemotaxonomic methods. This thesis deals with the study of the identification of microorganisms isolated from the sediment sample of Chorao Island, Goa and investigation of these bacteria for their α-glucosidase inhibition activities. Chapter 1: Introduction This chapter covers the literature on scope of bacterial systematics, polyphasic approach for the identification of bacteria, potentials of marine bacteria, types of marine sediments, types of alpha-glucosidase inhibitors and alpha-glucosidase inhibitors from marine isolates. This chapter concludes with the objectives and scope of the thesis. Chapter 2: Polyphasic systematic approach This chapter focused on the isolation and identification of bacteria from the sediment sample of Chorao Island, Goa. Sixty four unique isolates have been isolated and further characterized with the help of the polyphasic approach. The bacteria were characterized using 16S rRNA gene sequencing, morphological, physiological, biochemical, DNA G+C content, DNA-DNA hybridization, AP-PCR and chemotaxonomical. In this present study, we have been succesfully identified and published ten novel species from the sediment sample using polyphasic taxonomy approach. Chapter 3: Screening, isolation and identifiaction of alpha-glucosidase inhibitors from marine novel bacteria This chapter focused on screening the marine bacteria for their alpha-glucosidase inhibition activity. Alpha-glucosidase hydrolyzes the breakdown of carbohydrates with α,1-4 linkage into glucose units. Inhibition of these enzymes delays the release of glucose into the blood stream and forms hypoglycemia condition after the meal, thus used in the treatment of diabetes mellitus. In this study, isolated strains were screened for the presence of sedoheptulose 7-phosphate cyclase gene which corelates for α-glucosidase inhibition activity from their crude supernatant and detection of acarbose like molecules and their related products using thin layer chromatography. From overall study, we found that the two potential strains Arthrobacter enclensis NIO-1008T and Deinococcus enclensis NIO-1023T showed a highest inhibition towards α-glucosidase with 75.42 ±1.3 and 73.26 ±1.8 % inhibition, respectively compared with standard acarbose (Sigma) which showed 91.11±1.1 % inhibition. Further, these two strains were subjected to purification and characterization of α-glucosidase inhibitors. The purified compound from both the strains was characterized using HPLC, FTIR and tandem mass spectrometry (MS/MS). The purified compound was annotated and identified by a Web tool CFM-ID (Competitive Fragmentation Modeling For Metabolite Identification). From the above analysis, it was confirmed that both the strains produces acarbose like molecule. These compounds showed an inhibition at IC50 value of 500 μg/ml and 530 μg/ml for A. enclensis NIO-1008T and D. enclensis NIO-1023T respectively. Further, anti-SMASH analysis of whole genome of A. enclensis NIO-1008T confirms the presence of biosynthetic gene clusters of acarviostatin and acarbose, with 11% and 7% homology respectively. Chapter 4: Summary and future prospectives This chapter covers the overall summary and conclusion of polyphasic systematic approach for the identification of bacteria from the sediment sample collected from Chorao island, Goa and the alpha-glucosidase inhibition activity of these isolated strains. These marine isolates could be a source of bioactive molecules. Further, work needs to be anticipated for the identification of enzymes and gene clusters for the inhibitor.