Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1211591 | ENA |
REPOSITORIES: ENA
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Project description:Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant challenge to public health due to its evolving resistance to antibiotics. In addressing this concern, we employed a multi-omics approach, integrating proteomics and Transposon Directed Insertion-site Sequencing (TraDIS), to comprehensively investigate adaptive response of MRSA to β-lactam antibiotics and identify key genetic determinants influencing susceptibility. Proteomic analysis revealed significant shifts in the protein spectrum under sub-inhibitory concentrations of oxacillin and cefazolin, characterized by a stringent response. This included the upregulation of amino acid metabolism and oligopeptide transport pathways, along with the downregulation of arginine biosynthesis. Concurrent TraDIS screening identified 50 genes whose inactivation conferred a fitness advantage in the presence of both β-lactams, implicating processes such as nucleotide second messenger signaling, DNA repair, and histidine transport. Validation of selected mutants (relA::Tn, mutY::Tn, nsaR::Tn, aroC::Tn, SAUSA300_0432::Tn, SAUSA300_0846::Tn, SAUSA300_0595::Tn) confirmed these findings, demonstrating improved fitness in the presence of β-lactams. Integrative analysis of proteomic and TraDIS datasets highlighted the implication of the stringent response in mediating MRSA adaptation to β-lactams. These findings establish the 'susceptome' concept, a collection of genes that maintain or potentiate MRSA's vulnerability to β-lactams, providing new targets for therapeutic intervention and strategies to counteract resistance evolution.
2025-12-12 | PXD062516 | Pride
Project description:Transposon-directed insertion-site sequencing (TraDIS) of a 1 million mutant library of Salmonella Typhi
| PRJEB2004 | ENA
Project description:Generation of a Tn5 transposon library in Haemophilus parasuis and analysis by Transposon-Directed Insertion-Site Sequencing (TraDIS)
| PRJEB4161 | ENA
Project description:A whole genome screen was used to assay every gene of Escherichia coli strain BW25113 to identify genes involved in susceptibility to the monobactam (beta-lactam) antibiotic aztreonam. The methodology has been called TraDIS-Xpress, and is a version of TraDIS or Tn-seq. A transposon mutant library consisting of several hundred thousand mutants was constructed using a Tn5-derived transposon incorporating an inducible outward transcribing promoter. All the mutants were grown in LB broth cultures supplemented with aztreonam at 2 x, 1 x, 0.5 x and 0.25 x MIC with induction of the transposon promoter using 0.2 mM IPTG or 1 mM IPTG or without induction. Following growth, mutants with increased susceptibility show reduced numbers and those with reduced susceptibility show increased numbers. Each condition was performed in duplicate. The methodology enable genes to be assayed by insertional inactivation or by changes in expression. Expression changes result from altered transcription from upstream transposon insertions transcribing into the gene, or downstream insertions transcribing into the gene in the reverse direction leading to RNA interference through the generation of reverse and complementary RNA. Thus, essential genes into which transposon insertions are not tolerated may be assayed also by changes in numbers of upstream or downstream insertion mutants. Changes to high throughput sequencing protocols permit the generation of nucleotide sequence reads from the known transposon sequences into the surrounding insertion site for all the mutants in the mixture simultaneously. Matching the sequence of the reads to the genome nucleotide sequence of E. coli BW25113 then allows the precise locations of all the transposon insertion sites of all the mutants to be mapped simultaneously. The relative changes in mutants between control (without) and selective condition (with aztreonam) then indicates which genes are involved in susceptibility. The numbers of sequence reads that match is reflected by the number of mutants, and so the degree of susceptibility can also be estimated.
2024-07-06 | E-MTAB-11576 | biostudies-arrayexpress
Project description:Genome-wide identification of essential genes in Verrucomicrobium spinosum using transposon-directed insertion-site sequencing (TraDIS)
| PRJNA1267754 | ENA
Project description:Whole genome sequence and transposon-directed insertion site sequencing (TraDIS) reads of a uropathogenic Escherichia coli strain
| PRJNA1162300 | ENA
Project description:Transposon insertion site sequencing (TIS) is a powerful method for associating genotype to phenotype. However, all TIS methods described to date use short nucleotide sequence reads which cannot uniquely determine the locations of transposon insertions within repeating genomic sequences where the repeat units are longer than the sequence read length. To overcome this limitation, we have developed a TIS method using Oxford Nanopore sequencing technology that generates and uses long nucleotide sequence reads; we have called this method LoRTIS (Long Read Transposon Insertion-site Sequencing). This experiment data contains sequence files generated using Nanopore and Illumina platforms. Biotin1308.fastq.gz and Biotin2508.fastq.gz are fastq files generated from nanopore technology. Rep1-Tn.fastq.gz and Rep1-Tn.fastq.gz are fastq files generated using Illumina platform. In this study, we have compared the efficiency of two methods in identification of transposon insertion sites.
2022-05-30 | E-MTAB-11351 | biostudies-arrayexpress