Dataset Information

Presence of Bacterial Virulence Gene Homologues in the dibenzo-p-dioxins degrading bacterium Sphingomonas wittichii.

ABSTRACT: Sphingomonas wittichii, a close relative of the human pathogen Sphingomonas paucimobilis, is a microorganism of great interest to the bioremediation community for its ability of biodegradation to a large number of toxic polychlorinated dioxins. In the present study we investigated the presence of different virulence factors and genes in S. wittichii. We utilized phylogenetic, comparative genomics and bioinformatics analysis to investigate the potentiality of S. wittichii as a potential virulent pathogen. The 16SrDNA phylogenetic tree showed that the closest bacterial taxon to S. wittichii is Brucella followed by Helicobacter, Campylobacter, Pseudomonas then Legionella. Despite their close phylogenetic relationship, S. wittichii did not share any virulence factors with Helicobacter or Campylobacter. On the contrary, in spite of the phylogenetic divergence between S. wittichii and Pseudomonas spp., they shared many major virulence factors, such as, adherence, antiphagocytosis, Iron uptake, proteases and quorum sensing. S. wittichii contains several major virulence factors resembling Pseudomonas sp., Legionella sp., Brucella sp. and Bordetella sp. virulence factors. Similarity of virulence factors did not match phylogenetic relationships. These findings suggest horizontal gene transfer of virulence factors rather than sharing a common pathogenic ancestor. S. wittichii is a potential virulent bacterium. Another possibility is that reductive evolution process attenuated S. wittichii pathogenic capabilities. Thus plenty of care must be taken when using this bacterium in soil remediation purposes.

SUBMITTER: Saeb AT

PROVIDER: S-EPMC5290665 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Presence of Bacterial Virulence Gene Homologues in the dibenzo-p-dioxins degrading bacterium Sphingomonas wittichii.

Saeb Amr T M AT

Bioinformation 20160726 4

Sphingomonas wittichii, a close relative of the human pathogen Sphingomonas paucimobilis, is a microorganism of great interest to the bioremediation community for its ability of biodegradation to a large number of toxic polychlorinated dioxins. In the present study we investigated the presence of different virulence factors and genes in S. wittichii. We utilized phylogenetic, comparative genomics and bioinformatics analysis to investigate the potentiality of S. wittichii as a potential virulent ...[more]

PMID: 28197061

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Project description:Sphingomonas wittichii strain RW1 (RW1) is one of the few strains that can grow on dibenzo-p-dioxin (DD). We conducted a transcriptomic study of RW1 using RNA-Seq to outline transcriptional responses to DD, dibenzofuran (DF), and the smectite clay mineral saponite with succinate as carbon source. The ability to grow on DD is rare compared to growth on the chemically similar DF even though the same initial dioxygenase may be involved in oxidation of both substrates. Therefore, we hypothesized the reason for this lies beyond catabolic pathways and may concern genes involved in processes for cell-substrate interactions such as substrate recognition, transport, and detoxification. Compared to succinate (SUC) as control carbon source, DF caused over 240 protein-coding genes to be differentially expressed, whereas more than 300 were differentially expressed with DD. Stress response genes were up-regulated in response to both DD and DF. This effect was stronger with DD than DF, suggesting a higher toxicity of DD compared to DF. Both DD and DF caused changes in expression of genes involved in active cross-membrane transport such as TonB-dependent receptor proteins, but the patterns of change differed between the two substrates. Multiple transcription factor genes also displayed expression patterns distinct to DD and DF growth. DD and DF induced the catechol ortho- and the salicylate/gentisate pathways, respectively. Both DD and DF induced the shared down-stream aliphatic intermediate compound pathway. Clay caused category-wide down-regulation of genes for cell motility and chemotaxis, particularly those involved in the synthesis, assembly and functioning of flagella. This is an environmentally important finding because clay is a major component of soil microbes' microenvironment influencing local chemistry and may serve as a geosorbent for toxic pollutants. Similar to clay, DD and DF also affected expression of genes involved in motility and chemotaxis.

Project description:Sphingomonas wittichii strain RW1 (RW1) is one of few strains that can grow on dibenzo-p-dioxin (DD). We conducted a transcriptomic study of RW1 using RNA-Seq to outline transcriptional responses to DD, dibenzofuran (DF), and the smectite clay mineral saponite (SAP), an important geosorbent of toxicants in soil, with succinate as carbon source. The ability to grow on DD is rare compared to growth on the chemically similar DF even though the same initial dioxygenase may be involved in oxidation of both substrates. Therefore we hypothesized the reason for this lies beyond catabolic pathways and may concern genes involved in processes for cell-substrate interactions such as substrate recognition and transport, and detoxification. Compared to succinate (SUC) as control carbon source, DF caused over 240 protein-coding genes to be differentially expressed, whereas more than 300 were differentially expressed with DD. Stress response genes were up-regulated by both DD and DF. This effect was stronger with DD than DF, suggesting a higher toxicity of DD compared to DF. Both DD and DF caused changes in expression of genes involved in active cross-membrane transport such as TonB dependent receptor proteins, but the patterns of changes differed between the two substrates. Multiple transcription factor genes also displayed expression patterns distinct to DD and DF growth. DD and DF induced the catechol ortho- and the salicylate/gentisate pathways, respectively. Both DD and DF induced the shared down-stream aliphatic intermediate compound pathway. Clay (SAP) caused category-wide down-regulation of genes for cell motility and chemotaxis, particularly those involved in the synthesis, assembly and functioning of flagella. This is environmentally important finding because clay is a major component of soil microbes’ microenvironment influencing local chemistry and may serve as geosorbent for toxic pollutants. Similar to clay, DD and DF also affect motility and chemotaxis.

Project description:Chlorinated congeners of dibenzo-p-dioxin and dibenzofuran are widely dispersed pollutants that can be treated using microorganisms, such as the Sphingomonas wittichii RW1 bacterium, able to transform some of them into non-toxic substances. The enzymes of the upper pathway for dibenzo-p-dioxin degradation in S. wittichii RW1 have been biochemically and genetically characterized, but its genome sequence has indicated the existence of a tremendous potential for aromatic compound transformation, with 56 ring-hydroxylating dioxygenase subunits, 34 extradiol dioxygenases, and 40 hydrolases. To further characterize this enzymatic arsenal, new methodological approaches should be employed. Here, a large shotgun proteomic survey has been performed on cells grown on dibenzofuran, dibenzo-p-dioxin, and 2-chlorodibenzo-p-dioxin, and compared to growth on acetate. Changes in the proteome were monitored over time. Peak lists were generated with the Mascot Daemon software (version 2.3.2; Matrix Science, London, UK) using the extract_msn.exe data import filter (Thermo Fisher Scientific) from the Xcalibur FT package (version 2.0.7; Thermo Fisher Scientific). Data import filter options were set to 400 (minimum mass), 5000 (maximum mass), 0 (grouping tolerance), 0 (intermediate scans) and 1000 (threshold). The mgf files from technical triplicates were merged, and MS/MS spectra were assigned using the Mascot Daemon 2.3.2 (Matrix Science) and a database containing the nonredundant RefSeq protein entries for S. wittichii RW1 (NCBI Taxonomy ID: 392499), comprising 5345 protein sequences totalling 1 800 684 amino acids. The search was performed using the following criteria: tryptic peptides with a maximum of two miscleavages, mass tolerances of 5 ppm on the parent ion and 0.5 Da on the MS/MS, fixed modification for carbamidomethylated cysteine, and variable modification for methionine oxidation. Mascot results were parsed using the IRMa 1.28.0 software. Peptides were identified with a P-value threshold below 0.05. Proteins were considered validated when at least two distinct peptides were detected. Using a selection of 11 result files and the appropriate decoy database, the false discovery rate for protein identification was estimated to be 0.33% with these parameters.

Project description:Sphingomonas wittichii RW1 is one of a few strains known to grow on the related compounds dibenzofuran (DBF) and dibenzo-p-dioxin (DXN) as the sole source of carbon. Previous work by others (B. Happe, L. D. Eltis, H. Poth, R. Hedderich, and K. N. Timmis, J Bacteriol 175:7313-7320, 1993, https://doi.org/10.1128/jb.175.22.7313-7320.1993) showed that purified DbfB had significant ring cleavage activity against the DBF metabolite trihydroxybiphenyl but little activity against the DXN metabolite trihydroxybiphenylether. We took a physiological approach to positively identify ring cleavage enzymes involved in the DBF and DXN pathways. Knockout of dbfB on the RW1 megaplasmid pSWIT02 results in a strain that grows slowly on DBF but normally on DXN, confirming that DbfB is not involved in DXN degradation. Knockout of SWIT3046 on the RW1 chromosome results in a strain that grows normally on DBF but that does not grow on DXN, demonstrating that SWIT3046 is required for DXN degradation. A double-knockout strain does not grow on either DBF or DXN, demonstrating that these are the only ring cleavage enzymes involved in RW1 DBF and DXN degradation. The replacement of dbfB by SWIT3046 results in a strain that grows normally (equal to the wild type) on both DBF and DXN, showing that promoter strength is important for SWIT3046 to take the place of DbfB in DBF degradation. Thus, both dbfB- and SWIT3046-encoded enzymes are involved in DBF degradation, but only the SWIT3046-encoded enzyme is involved in DXN degradation.IMPORTANCE S. wittichii RW1 has been the subject of numerous investigations, because it is one of only a few strains known to grow on DXN as the sole carbon and energy source. However, while the genome has been sequenced and several DBF pathway enzymes have been purified, there has been very little research using physiological techniques to precisely identify the genes and enzymes involved in the RW1 DBF and DXN catabolic pathways. Using knockout and gene replacement mutagenesis, our work identifies separate upper pathway ring cleavage enzymes involved in the related catabolic pathways for DBF and DXN degradation. The identification of a new enzyme involved in DXN biodegradation explains why the pathway of DBF degradation on the RW1 megaplasmid pSWIT02 is inefficient for DXN degradation. In addition, our work demonstrates that both plasmid- and chromosomally encoded enzymes are necessary for DXN degradation, suggesting that the DXN pathway has only recently evolved.

Dataset Information

Presence of Bacterial Virulence Gene Homologues in the dibenzo-p-dioxins degrading bacterium Sphingomonas wittichii.

Publications

Presence of Bacterial Virulence Gene Homologues in the dibenzo-p-dioxins degrading bacterium Sphingomonas wittichii.

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