De novo transcriptome analysis of halotolerant bacterium Staphylococcus sp. strain P-TSB-70 isolated from East coast of India: In search of salt stress tolerant genes

Abstract

In the present study, we identified salt stress tolerant genes from the marine bacterium Staphylococcus sp. strain P-TSB-70 through transcriptome sequencing. In favour of whole-genome transcriptome profiling of Staphylococcus sp. strain P-TSB-70 (GenBank Accn. No. KP117091) which tolerated upto 20% NaCl stress, the strain was cultured in the laboratory condition with 20% NaCl stress. Transcriptome analyses were performed by SOLiD4.0 sequencing technology from which 10280 and 9612 transcripts for control and treated, respectively, were obtained. The coverage per base (CPB) statistics were analyzed for both the samples. Gene ontology (GO) analysis has been categorized at varied graph levels based on three primary ontology studies viz. cellular components, biological processes, and molecular functions. The KEGG analysis of the assembled transcripts using KAAS showed presumed components of metabolic pathways which perhaps implicated in diverse metabolic pathways responsible for salt tolerance viz. glycolysis/gluconeogenesis, oxidative phosphorylation, glutathione metabolism, etc. further involving in salt tolerance. Overall, 90 salt stress tolerant genes were identified as of 186 salt-related transcripts. Several genes have been found executing normally in the TCA cycle pathway, integral membrane proteins, generation of the osmoprotectants, enzymatic pathway associated with salt tolerance. Recognized genes fit diverse groups of salt stress genes viz. abc transporter, betaine, sodium antiporter, sodium symporter, trehalose, ectoine, and choline, that belong to different families of genes involved in the pathway of salt stress. The control sample of the bacterium showed elevated high proportion of transcript contigs (29%) while upto 20% salt stress treated sample of the bacterium showed a higher percentage of transcript contigs (31.28%). A total of 1,288 and 1,133 transcript contigs were measured entirely as novel transcript contigs in both control and treated samples, respectively. The structure and function of 10 significant salt stress tolerant genes of Staphylococcus sp. have been analyzed in this study. The information acquired in the present study possibly used to recognize and clone the salt stress tolerant genes and support in developing the salt stress-tolerant plant varieties to expand the agricultural productivity in the saline system.

Fig 1. Schematic workflow of the methodologies of transcriptome analysis of both control and treated bacterial samples.

Fig 2

Bioanalyzer profiles of total RNA of A) P-TSB-70_1.5% control sample and B) P-TSB-70_20% Treated sample on Pico 6000 chip.

Fig 3

GO Level distribution of both control (A, C, E) and treated (B, D, F) transcripts in three domains.

Fig 4. GO Level 2distribution of both control and treated transcripts in three domains.

Fig 5

Pie diagram of (A) control transcripts involved in biological processes in GO study (B) treated transcripts involved in biological processes in GO study (C) control transcripts involved in molecular functions in GO study, (D) treated transcripts involved in molecular functions in GO study, (E) control transcripts involved in cellular components in GO study, (F) treated transcripts involved in cellular components in GO study.

Fig 6. Heat map (cluster) showing the top 100 differentially expressed genes.

Fig 7

3D structural information of salt tolerance functional genes showing N and C-Terminal, where (A) designated with treatment_transcript_1371-betaine carnitine choline family, (B) treatment_transcript_1069-bacterial abc transporter protein, (C) treatment_transcript_6679-proline betaine transporter, (D) treatment_transcript_4687-betaine aldehydede hydrogenase and (E) treatment_transcript_1432-dass family.

Fig 8

3D structural information of salt tolerance related functional genes showing N and C-Terminal, where (A) treatment_transcript_194-succinate dehydrogenase iron-sulfur subunit, (B) treatment_transcript_5134-atp synthase gamma subunit, (C) treatment_transcript_6954-succinate dehydrogenase flavoprotein subunit, (D) treatment_transcript_7764-succinate semialdehyde dehydrogenase and (E) treatment_transcript_10031- fumarate class ii.