Dataset Information

The CRISPR/Cas immune system is an operon regulated by LeuO, H-NS, and leucine-responsive regulatory protein in Salmonella enterica serovar Typhi.

ABSTRACT: Prokaryotes have developed multiple strategies to survive phage attack and invasive DNA. Recently, a novel genetic program denominated the CRISPR/Cas system was demonstrated to have a role in these biological processes providing genetic immunity. This defense mechanism is widespread in the Archaea and Bacteria, suggesting an ancient origin. In the last few years, progress has been made regarding the functionality of the CRISPR/Cas system; however, many basic aspects of the system remain unknown. For instance, there are few studies about the conditions and regulators involved in its transcriptional control. In this work, we analyzed the transcriptional organization of the CRISPR/Cas system as well as the positive and negative regulators involved in its genetic expression in Salmonella enterica serovar Typhi. The results obtained show that in S. Typhi the CRISPR/Cas system is a LeuO-dependent operon silenced by the global regulator LRP, in addition to the previously known nucleoid-associated protein H-NS; both LRP and H-NS bind upstream and downstream of the transcriptional start site of casA. In this study, relevant nucleotides of the casA regulatory region that mediate its LeuO transcriptional activation were identified. Interestingly, specific growth conditions (N-minimal medium) were found for the LeuO-independent expression of the CRISPR/Cas system in S. Typhi. Thus, our work provides evidence that there are multiple modulators involved in the genetic expression of this immune system in S. Typhi IMSS-1.

SUBMITTER: Medina-Aparicio L

PROVIDER: S-EPMC3133143 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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The CRISPR/Cas immune system is an operon regulated by LeuO, H-NS, and leucine-responsive regulatory protein in Salmonella enterica serovar Typhi.

Medina-Aparicio L L Rebollar-Flores J E JE Gallego-Hernández A L AL Vázquez A A Olvera L L Gutiérrez-Ríos R M RM Calva E E Hernández-Lucas I I

Journal of bacteriology 20110311 10

Prokaryotes have developed multiple strategies to survive phage attack and invasive DNA. Recently, a novel genetic program denominated the CRISPR/Cas system was demonstrated to have a role in these biological processes providing genetic immunity. This defense mechanism is widespread in the Archaea and Bacteria, suggesting an ancient origin. In the last few years, progress has been made regarding the functionality of the CRISPR/Cas system; however, many basic aspects of the system remain unknown. ...[more]

PMID: 21398529

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Project description:Typhoid fever, caused by Salmonella enterica serovar Typhi (S. Typhi), is a global health concern and its treatment is problematic due to the rise in antimicrobial resistance (AMR). Rapid detection of patients infected with AMR positive S. Typhi is, therefore, crucial to prevent further spreading. Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated genes (CRISPR-Cas), is an adaptive immune system that initially was used for typing purposes. Later, it was discovered to play a role in defense against phages and plasmids, including ones that carry AMR genes, and, at present, it is being explored for its usage in diagnostics. Despite the availability of whole-genome sequences (WGS), very few studied the CRISPR-Cas system of S. Typhi, let alone in typing purposes or relation to AMR. In the present study, we analyzed the CRISPR-Cas system of S. Typhi using WGS data of 1059 isolates obtained from Bangladesh, India, Nepal, and Pakistan in combination with demographic data and AMR status. Our results reveal that the S. Typhi CRISPR loci can be classified into two groups: A (evidence level >2) and B (evidence level ≤2), in which we identified a total of 47 unique spacers and 15 unique direct repeats. Further analysis of the identified spacers and repeats demonstrated specific patterns that harbored significant associations with genotype, demographic characteristics, and AMR status, thus raising the possibility of their usage as biomarkers. Potential spacer targets were identified and, interestingly, the phage-targeting spacers belonged to the group-A and plasmid-targeting spacers to the group-B CRISPR loci. Further analyses of the spacer targets led to the identification of an S. Typhi protospacer adjacent motif (PAM) sequence, TTTCA/T. New cas-genes known as DinG, DEDDh, and WYL were also discovered in the S. Typhi genome. However, a specific variant of the WYL gene was only identified in the extensively drug-resistant (XDR) lineage from Pakistan and ciprofloxacin-resistant lineage from Bangladesh. From this work, we conclude that there are strong correlations between variations identified in the S. Typhi CRISPR-Cas system and endemic AMR positive S. Typhi isolates.

Project description:BackgroundTyphoid fever is an acute systemic infection of humans caused by Salmonella enterica subspecies enterica serovar Typhi (S. Typhi). In chronic carriers, the bacteria survive the harsh environment of the gallbladder by producing biofilm. The phenotype of S. Typhi biofilm cells is significantly different from the free-swimming planktonic cells, and studies have shown that they are associated with antibiotic resistance, immune system evasion, and bacterial persistence. However, the mechanism of this transition and the events leading to biofilm formation are unknown. High throughput sequencing was performed to identify the genes involved in biofilm formation and to postulate the mechanism of action.ResultsPlanktonic S. Typhi cells were cultured using standard nutrient broth whereas biofilm cells were cultured in a stressful environment using high shearing-force and bile to mimic the gallbladder. Sequencing libraries were prepared from S. Typhi planktonic cells and mature biofilm cells using the Illumina HiSeq 2500 platform, and the transcriptome data obtained were processed using Cufflinks bioinformatics suite of programs to investigate differential gene expression between the two phenotypes. A total of 35 up-regulated and 29 down-regulated genes were identified. The identities of the differentially expressed genes were confirmed using NCBI BLAST and their functions were analyzed. The results showed that the genes associated with metabolic processes and biofilm regulations were down-regulated while those associated with the membrane matrix and antibiotic resistance were highly up-regulated.ConclusionsIt is proposed that the biofilm phenotype of S. Typhi allows the bacteria to increase production of the membrane matrix in order to serve as a physical shield and to adhere to surfaces, and enter an energy conservation state in response to the stressful environment. Conversely, the planktonic phenotype allows the bacteria to produce flagella and increase metabolic activity to enable the bacteria to migrate and form new colonies of infection. This data provide a basis for further studies to uncover the mechanism of biofilm formation in S. Typhi and to discover novel genes or pathways associated with the development of the typhoid carrier state.

Dataset Information

The CRISPR/Cas immune system is an operon regulated by LeuO, H-NS, and leucine-responsive regulatory protein in Salmonella enterica serovar Typhi.

Publications

The CRISPR/Cas immune system is an operon regulated by LeuO, H-NS, and leucine-responsive regulatory protein in Salmonella enterica serovar Typhi.

Similar Datasets

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