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Genome wide computational identification of potential therapeutic targets of Clostridium botulinum ATCC 3502 using comparative genomics

Student Name: Ms. Tulika Bhardwaj
Guide: Dr Pallavi Somvanshi
Year of completion: 2019

Abstract:

Food borne botulism is a paralytic illness caused by ingestion of preformed toxins of rod shaped anaerobic bacterium, Clostridium botulinum. Its spores are commonly available in soil, water but germinate only in specific environments (low acidity, high water content). Global distribution of sudden hospital outbreaks and diseases surveillance reflects its genetic diversity and ability of evading host immune response from a range of virulence factors and host pathogen interactions. Due to this biological diversity, comparative analysis of Clostridium botulinum strains reveals different strategies and evolutionary pattern of successive lineages. This represents the clear picture of bacterial lifestyle, its adaptation into wide range of ecological niches. Computation of pangenome, core genome and singletons assists in identifying conserved genome content and specific phenotypic traits of new genome evolved. A high genomic similarity among the genomes was observed by phylogenomic analysis and thus constitutes highly conserved core genome. Pan genome trend indicates an open pan genome i.e. successive addition of genes (accessory genes and singletons) with every genome introduction. Core genome assists in identifying genetic fingerprints of Clostridial family which serve as biological markers in pathogenicity identification.

Clostridium botulinum ATCC 3502 was targeted to identify potential therapeutic targets using subtractive proteome approach. This computational approach speeds up drug discovery process by eliminating protein sequence showing homology to host proteome dataset. Comparative assessment with potential drug targets of C.difficile strain 630 (screened out by following same computational pipeline as for Clostridium botulinum ATCC 3502) was performed to identify common drug targets which will provide a platform for the novel chemical scaffold development to be used as an effective wide-spectrum drug inhibiting their catalytic activities against multidrug resistant Clostridium in the near future. This study renders identification of 5 common; present in both pathogens (Uridylate kinase, D-alanine--D-alanine ligase, Diaminopimelate (DAP) epimerase, 1-deoxy-D-xylulose 5-phosphate reductoisomerase and 3-methyl-2-oxobutanoate hydroxymethyltransferase, 34 in C.botulinum ATCC 3502 and 42 in C.difficile str. 630 drug targets. In addition, to explore all possible pathways of host-pathogen interaction responsible for pathogenesis, a computational pipeline was developed to identify homologous sequences of C. botulinum ATCC 3502 and the host and identifies of 20 potential mimics between the pathogen (C. botulinum ATCC 3502) and the host (Homo sapiens).

Further, KEGG pathway analysis reveals participation of identified five common drug targets in pathogen specific pathways of C.botulinum ATCC 3502. Metabolic pathway analysis enables representation of reconstructed metabolic pathway in the form of stoichiometric matrix enabling the computation of flux distributed in the objective solution space. Further, simulation of mathematical model generated by imposing constraints of thermodynamic stability and non-decomposability constraints scrutinize elementary flux nodes. These elementary flux modes are considered essential for bacterial survival and maintenance, therefore considered as potential drug targets. The study validates essentiality of identified five potential targets for pathogen cell survival and consideration for future drug discovery.

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