Differentially expressed genes in rough and smooth colony isolates of Clostridioides difficile
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA664385 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR12671773_1.fastq.gz | Fastqsanger.gz | |||
| SRR12671773_2.fastq.gz | Fastqsanger.gz | |||
| SRR12671774_1.fastq.gz | Fastqsanger.gz | |||
| SRR12671774_2.fastq.gz | Fastqsanger.gz | |||
| SRR12671775_1.fastq.gz | Fastqsanger.gz |
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Project description:Purpose: to determine the differentially expressed genes in the phase-variable rough and smooth colony isolates of C. difficile Methods: C. difficile R20291 was grown on BHIS agar to obtain distinct colonies. Individual rough and smooth colonies were chosen for propagation on BHIS-agar for 24 hours as described in Garrett, et al., PLoS Biology, 2019. Growth was collected from n = 2 biological replicates. RNA was purified using TriSure and chloroform, beadbeating, and isopropanol/ethanol precipitation. Quality was verified with Bioanalyzer Assay. Samples were submitted to Genewiz for depletion of rRNA using the TruSeq RiboZero Gold Kit, library preparation, and single-end sequencing on the Illumina HiSeq 2500 platform. RNA sequencing analysis was done using CLC Genomic Workbench v20. Reads were mapped to C. difficile R20291 genome using the software's default menalties for mismatch, deletion, and insertion differences from the reference genome. Transcript reads were normalized as RPKM. Results: 88 genes were differentially expressed between bacteria from rough versus smooth colonies, with equal to or greater than a 2-fold change and p < 0.05 with FDR correction.
2020-09-27 | GSE158225 | GEO
Project description:In this study, we were interested to get deeper insights into the molecular mechanisms that govern the formation and selection of the different colony morphologies in Mycobacterium abscessus strains, including the potential reversibility of the rough (R) phenotype into a smooth (S) phenotype. We used next generation sequencing (NGS) and micro-array / RNAseq approaches to determine the genome sequences and transcriptomic profiles of three isogenic S/R strain couples of M. abscessus. One clinical isolate strain named CF and two collection strains referred as 19977-AT and 19977-IP. To perform the transcriptomic comparison of the rough variant versus the smooth variant for each M. abscessus strain, a customized micro-array has been manufactured by Agilent (8 x 15k format). The design of oligonucleotides covering all protein coding sequences was done using OligoArray version 2.1 on the basis of the 4920 predicted coding sequences composing the entire M. abscessus genome. The experimental data for each of the 3 strains consisted of 6 hybridizations (3 biological replicates with dye-swap).
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Project description:Salmonella enterica serovar Typhimurium (S. Typhimurium) is a major cause of salmonellosis, with various multidrug-resistant pathovariants emerging. Here, we investigated a distinct rough colony variant with potentially enhanced environmental adaptation in China. Whole-genome sequencing of 2,212 Chinese isolates and 1,739 publicly available isolates revealed the population structure and evolutionary history of the rough colony variants. These variants showed macro, red, dry, and rough (mrdar) colonies with better biofilm-forming abilities at 28 °C and 37 °C compared to typical rdar colonies. The mrdar variants displayed expansive multidrug resistance, mainly attributed to an IncHI2 plasmid carrying 19 antimicrobial resistance genes. Phylogenomic analysis divided global collections into six lineages, with the majority of mrdar variants in sublineage L6.5, which originated from Chinese smooth colony strains and possibly emerged circa 1977. Upregulation of the csgDEFG operons was observed among mrdar variants, probably due to a distinct point mutation (-44G>T) in the csgD gene promoter. Pangenome and genome-wide association analyses identified 87 specific accessory genes and 72 distinct single nucleotide polymorphisms associated with the mrdar morphotype. This study presents the first large-scale, genome-based study of a distinct rdar colony morphology variant of S. Typhimurium in China, highlighting its global epidemic potential. Ongoing surveillance and genomic epidemiological studies are required to control epidemics, monitor bacterial population changes, and assess associated risks to global public health.
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Project description:High-throughput sequencing of human macrophages infected with Mycobacterium abscessus Smooth and Rough variants
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Project description:The protein inventory of the anaerobic pathogen Clostridioides difficile grown in colony or aggregate biofilms was investigated in a comprehensive LC-MS/MS approach. C. difficile has a versatile metabolism and is perfectly equipped to survive and persist inside the mammalian intestine. However, knowledge on how C. difficile exactly resides inside the intestine is still scare. Aggregate or biofilms attached to the epithelium are discussed as possible growth forms and have become a focus in research. However, recent data suggest that the choice of the model strain, timepoint of analysis and the biofilm model significantly impact the outcome of biofilm experiments leading to contradictory results. To address some of these issues, a comprehensive proteomics approach was applied to investigate the metabolism of C. difficile strain 630erm grown in colony or aggregate biofilms. Besides verification of recent findings, a comprehensive overview on the differential expression profile of C. difficile’s cell surface proteins as well as its energy and stress metabolism during the two different forms of biofilm growth could be provided. Additionally, regulatory circuits that were activated in the two different biofilm models were analysed whereby RpoN was determined to be an important regulator of aggregate biofilm formation in C. difficile.
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Project description:Mycobacterium abscessus is an emerging pathogen causing pulmonary infections in those with inflammatory lung disorders, such as Cystic Fibrosis (CF), and is associated with the highest fatality rate among rapidly growing mycobacteria (RGM). Phenotypically, MAB manifests as either a Smooth (MAB-S) or a Rough (MAB-R) morphotype, which differ in their levels of cell wall glycopeptidolipids (GPLs) and in their pathogenicity in vivo. As one of the primary immune cells encountered by MAB, we sought to examine the early transcriptional events within macrophages, following infection with both MAB-S or MAB-R. We sampled the small RNA (sRNA) transcriptome of THP-1-derived macrophages infected with both MAB-R and MAB-S at 1, 4 and 24 hours post-infection (hpi) using RNA-seq. MAB-S elicited a more robust transcriptional response at the miRNA level, reflecting higher cytokine levels in culture supernatants. However, and a direct comparison identified no differentially expressed miRNAs between MAB-R- and MAB-S-infected cells. Most of the induced miRNAs have previously been associated with mycobacterial infection and overall miRNA expression patterns were similarly highly correlated between the morphotypes.
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Project description:Members of the Mycobacterium (M.) abscessus complex (MABC) are rapidly growing mycobacteria showing smooth and/or rough colony morphotype. While not as virulent as M. tuberculosis, they can cause soft tissue infection and fatal pulmonary disease, especially in patients with cystic fibrosis. Diagnosing MABC pulmonary disease is challenging since the isolation of M. abscessus from respiratory samples is in itself not diagnostic and the clinical features are often non-specific. Immunologic assays, which could aid in the understanding and diagnosis of the disease, are not available. In this study eight rough and six smooth colony morphotype isolates were collected from seven clinical MABC strains and the M. abscessus reference strain ATCC19977, as six strains showed both morphotypes simultaneously and two strains only showed a rough morphotype. Clinical isolates were submitted to whole genome sequencing. Quantitative proteomic analysis was performed on bacterial lysates and the culture supernatant of all 14 isolates. Supernatant proteins present in all isolates were compared in a BLAST search against other clinically significant mycobacterial species to determine species-specific proteins of MABC. In silico B- and T-cell epitope prediction was performed for species-specific proteins. All clinical strains were found to be M. abscessus ssp. abscessus. Six of seven rough colony clinical isolates contained genetic changes in the MAB_4099c gene, which is a likely genetic basis for the rough morphotype. Proteomic analysis detected 3 137 different proteins in total of which 79 proteins were found in the culture supernatants of all isolates. BLAST analyses of these 79 proteins identified 12 of those exclusively encoded by all members of MABC plus M. immunogenum. In silico prediction of epitopes predicted B- and T-cell epitopes in all these 12 species-specific proteins, rendering them promising candidates for future studies on immune pathogenesis and immune diagnostic tools for MABC disease.
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