Project description:Relentless mining operations have destroyed our environment significantly. Soil inhabiting microbes play a significant role in ecological restoration of these areas. Microbial weathering processes like chemical dissolution of rocks significantly promotes the soil properties and enhances the rock to soil ratio respectively. Earlier studies have reported that bacteria exhibit efficient rock-dissolution abilities by releasing organic acids and other chemical elements from the silicate rocks. However, rock-dissolving mechanisms of the bacterium remain to be unclear till date. Thus, we have performed rock-dissolution experiments followed by genome and transcriptome sequencing of novel Pseudomonas sp.NLX-4 strain to explore the efficiency of microbe-mediated habitat restoration and its molecular mechanisms underlying this biological process. Results obtained from initial rock dissolution experiments revealed that Pseudomonas sp. NLX-4 strain efficiently accelerates the dissolution of silicate rocks by secreting amino acids, exopolysaccharides, and organic acids with elevated concentrations of potassium, silicon and aluminium elements. The rock dissolution experiments of NLX-4 strain exhibited an initial increase in particle diameter variation values between 0-15 days and decline after 15 days-time respectively. The 6,771,445-base pair NLX-4 genome exhibited 63.21 GC percentage respectively with a total of 6041 protein coding genes. Genome wide annotations of NLX-4 strain exhibits 5045-COG, 3996-GO, 5342-InterPro, 4386-KEGG proteins respectively Transcriptome analysis of NLX-4 cultured with/without silicate rocks resulted in 539 (288-up and 251-down) differentially expressed genes (DEGs). Fifteen DEGs encoding for siderophore transport, EPS and amino acids synthesis, organic acids metabolism, and bacterial resistance to adverse environmental conditions were highly up-regulated by cultured with silicate rocks. This study has not only provided a new strategy for the ecological restoration of rock mining areas, but also enriched the applicable bacterial and genetic resources.
Project description:To know whether the microarray technique could be used to detect bacterial gene expression in soil, large quantity of RNA was extracted from soil cultures of Pseudomonas putida KT2440 containing a chloroaromatic degrading plasmid at the presence or absence of the growth substrate, 3-chlorobenzoate (3CB). The quality and quantity of the extracted RNA were proper for a typical microarray analysis. Gene expression patterns of soil cultures were analyzed by DNA microarray using the extracted RNA. Among 5346 genes on the array, 5% and 4.5% of genes showed up- or down-regulation. Analysis done at the DAVID Bioinformatics Resources server suggested that the benzoate degradation via hydroxylation pathway had the most significant changes after treatment with 3CB. Expression of the 3CB degradation genes located in the genome was confirmed by real-time RT-PCR. In addition, real time RT-PCR analysis revealed that the fluorescent signals from plasmid genes on the microarray were saturated so that the induction ratio of the genes located in the plasmid was underestimated in microarray analysis. To our best knowledge, this report represents the first trial to use microarray technique to detect genome-wide bacterial gene expression in soil.
Project description:Here we presented the detailed transcriptomic analysis for Pseudomonas sp. AP3_22, an effective sodium dodecyl sulfate degrader isolated from the soil sample from wastewater treatment plant, cultured in the presence of SDS to get the first insight in the global bacterial response toward Sthis anionic detergent. Our results suggest showed that although SDS could be used as a carbon source, in the first place it acts influence on integrity of the cell envelopes and causes global stress response together combined with cell wall modification and repair induction. These results suggest that the modulation of the membrane content composition is first adaptation step in a typical response to detergent exposure. As the second response to the sodium dodecyl sulfate the AP3_22 strain metabolism was shifted from the lipid biosynthesis to the lipid catabolism and the SDS degradation started.
Project description:To know whether the microarray technique could be used to detect bacterial gene expression in soil, large quantity of RNA was extracted from soil cultures of Pseudomonas putida KT2440 containing a chloroaromatic degrading plasmid at the presence or absence of the growth substrate, 3-chlorobenzoate (3CB). The quality and quantity of the extracted RNA were proper for a typical microarray analysis. Gene expression patterns of soil cultures were analyzed by DNA microarray using the extracted RNA. Among 5346 genes on the array, 5% and 4.5% of genes showed up- or down-regulation. Analysis done at the DAVID Bioinformatics Resources server suggested that the benzoate degradation via hydroxylation pathway had the most significant changes after treatment with 3CB. Expression of the 3CB degradation genes located in the genome was confirmed by real-time RT-PCR. In addition, real time RT-PCR analysis revealed that the fluorescent signals from plasmid genes on the microarray were saturated so that the induction ratio of the genes located in the plasmid was underestimated in microarray analysis. To our best knowledge, this report represents the first trial to use microarray technique to detect genome-wide bacterial gene expression in soil. A study using total RNA extracted from soil cultures of Pseudomonas putida KT2440/pSL1. Each chip measures the expression level of 5,341 genes from Pseudomonas putida KT2440 genome and 5 genes from an introduced plasmid pSL1 with fourteen 60-mer probes per gene which have five-fold technical redundancy.
Project description:The identification of processes activated by specific microbes during microbiota colonization of plant roots has been hampered by technical constraints in metatranscriptomics. These include lack of reference genomes, high representation of host or microbial rRNA sequences in datasets, or difficulty to experimentally validate gene functions. Here, we recolonized germ-free Arabidopsis thaliana with a synthetic, yet representative root microbiota comprising 106 genome-sequenced bacterial and fungal isolates. We used multi-kingdom rRNA depletion, deep RNA-sequencing and read mapping against reference microbial genomes to analyse the in-planta metatranscriptome of abundant colonizers. We identified over 3,000 microbial genes that were differentially regulated at the soil-root interface. Translation and energy production processes were consistently activated in planta, and their induction correlated with bacterial strains’ abundance in roots. Finally, we used targeted mutagenesis to show that several genes consistently induced by multiple bacteria are required for root colonization in one of the abundant bacterial strains (a genetically tractable Rhodanobacter). Our results indicate that microbiota members activate strain-specific processes but also common gene sets to colonize plant roots.
Project description:Untargeted metabolomics analysis of in vitro headspace volatiles from 81 Pseudomonas aeruginosa bacterial isolates from individuals with cystic fibrosis. Headspace volatiles were collected using solid-phase microextraction (SPME) (in triplicate) and comprehensive two-dimensional gas chromatography and time-of-flight mass spectrometry (GCxGC-TOFMS). 15 replicates of un-inoculated media were prepared and analyzed in parallel, for a total of 258 samples.