Genomic and Proteomic analysis of indigenous bacteria Under the stress of selected micropollutants

Abstract

Micropollutants are harmful for human health, therefore, this study was undertaken to address the concerns of micropollutant contamination of the environment. Inadequate biological information or molecular understanding of the cellular responses underpinning bacterial bioremediation hampers site-specific mineralization. It is necessary to generate information related to pollutant attenuating microorganisms and biotransforming genes as well as related phenomenon which could impact remediation. This work is an attempt to provide information necessary for a reliable evaluation of the state of the internal cellular environment as well as baseline information for crafting bioremediation strategies. Autocthonous bacteria were screened for their ability to resist pollutants and two bacteria, belonging to phylum Proteobacteria were studied at genome scale. Democratization of genome sequencing has made bacterial genomics a mature and easy approach for researchers from interdisciplinary fields like environment, evolution and scientists working in the biomedical disciplines. Whole genome sequencing of two isolates Alcaligenes faecalis strain MB207 and Serratia nematodiphila MB307 was attempted to provide a genomic foundation for micropollutant attenuation by these organisms. Genome sizes were roughly 4.16 (with 3749 protein coding genes) and 5.16 MB (with 4794 protein coding genes) for Alcaligenes faecalis MB207 and Serratia nematodiphila MB307, respectively. Both isolates yielded numerous gene sequences (such as monooxyenases, dioxygenases, azoreductases, peroxidases, metal resistance, porins etc.) with bioremediation potential. Pathways for biodegradation were searched in the KEGG database and 225 genes in Serratia nematodiphila MB307 while 201 genes in Alcaligenes faecalis MB207 were linked with microbial metabolism in varied environments. Whole genome analysis marked Alcaligenes faecalis strain MB207 and Serratia nematodiphila MB307 as prospective eco-friendly bacteria, provided their nutritional and physiological requirements are met for proper functioning of biotransformation pathways. Occurrence of metal resistance genes with antibiotic and biocide resistance DNA segments depicted a co-selection and common regulatory mechanism of these genes, which needs to be further explored. Availability of the sequence data in public genome repository of NCBI is also useful for future researchers to explore enzymes and apparatus for sustainable environmental clean-up. One isolate, identified as Serratia nematodiphila was found to have remarkable attenuation properties and further exploited for metabolite production and functional genomic/proteomic analysis under pollutant stress. Some novel metabolites were obtained which depict new or xiv evolved pathways in Serratia nematodiphila MB307 for biotransformation. A global view of the proteins was obtained which complemented genomic information. Differential regulation of several proteins was observed in the whole cell as well as in the enriched membrane fraction when subjected to 100 μg/ml pollutant for 24 hours. Numerous ribosomal proteins and chaperones were 2-fold differentially regulated during pollutant stress. This depicted either a change in protein production required to combat stress or utilization of energy for survival at the cost of cell growth/division. Although there were some common proteins differentially regulated under different type of stresses, a common pattern for pollution resistance could not be deciphered, depicting different mechanisms of coping with each class of pollutant (i.e. metals, azo dyes and pharmaceutical). Comparison with other related studies led to several interesting findings and made the study concurrently deeper as well as diffused, warranting more research in this domain. Indigenous bacteria are physiologically and genetically acclimatized to their surroundings and help detoxify the polluted environment in an efficient manner. This study sheds light on molecular mechanism of their catabolic traits as well as proteome rewiring behind their adaptation to stressed surroundings and biotransformation of environmental contaminants. Obtained results pave way for further systematic investigation (such as linking time resolved metabolome with degradation genes/pathways). Evolution of co-resistance and stress signatures of these biotopes after intermittent and prolonged pollutant exposure also needs to be studied.