We have developed a simple and efficient procedure for adding an epitope-encoding tail to one or more genes of interest in the bacterial chromosome. The is modification gene replacement method Datsenko Wanner [Datsenko, K. A. & Wanner, B. L. (2000) Proc. Natl. Acad. Sci. USA 97, 6640–6645]. A DNA module that begins with sequence includes selectable marker amplified by PCR primers carry extensions (as short as 36 nt) homologous last portion targeted region downstream from it....

Gene transfer between separate lineages of a bacterial pathogen can promote recombinational divergence and the emergence new pathogenic variants. Temperate bacteriophages, by virtue their ability to carry foreign DNA, are potential key players in this process. Our previous work has shown that representative strains Salmonella typhimurium (LT2, ATCC14028 SL1344) lysogenic for two temperate bacteriophages: Gifsy‐ 1 2. Several lines evidence suggested both elements genes contribute virulence....

We show that Salmonella typhimurium harbours two fully functional prophages, Gifsy-1 and Gifsy-2, can be induced by standard treatments or, more effectively, exposing bacteria to hydrogen peroxide. Curing for the Gifsy-2 prophage significantly reduces Salmonella's ability establish a systemic infection in mice. Cured strains recover their virulence properties upon relysogenization. Phage carries sodC gene periplasmic [Cu,Zn]-superoxide dismutase previously implicated bacterial defences...

A relevant, yet little recognized feature of antisense regulation is the possibility switching roles between regulatory and regulated RNAs. Here we show that induction a Salmonella gene relies on conversion small RNA from effector to target. The chiP (formerly ybfM ), identified characterized in present study, encodes conserved enterobacterial chitoporin required for uptake chitin-derived oligosaccharides. In absence inducer, kept silent by action constitutively made RNA, ChiX SroB, RybC),...

Cocultures of Salmonella strains carrying or lacking specific prophages undergo swift composition changes as a result phage-mediated killing sensitive bacteria and lysogenic conversion survivors. Thus, spontaneous prophage induction in few cells enhances the competitive fitness lysogen population whole, setting selection regime that forces maintenance spread viral DNA. This is likely to account for profusion sequences bacterial genomes may contribute evolutionary success certain phylogenetic...

A model is proposed that accounts for regulation of the histidine operon by a mechanism involving alternative configuration mRNA secondary structure (the stem model). New evidence includes sequence data on three regulatory mutations. The first (hisO1242) mutation deletes sequences needed to form attenuator and causes constitutive expression. second (hisO9654) His- ochre (UAA) in leader peptide gene; existence this constitutes gene translated. third (hisO9663) remarkable. It neither generates...

Gene regulation by bacterial trans -encoded small RNAs (sRNAs) is generally regarded as a post-transcriptional process bearing exclusively on the translation and/or stability of target messenger RNA (mRNA). The work presented here revealed existence transcriptional component in bicistronic operon—the chiPQ locus—by ChiX sRNA Salmonella . By studying mechanism which ChiX, upon pairing near 5′ end transcript, represses distal gene operon, we discovered that action induces Rho-dependent...

In all living cells, genomic DNA is compacted through interactions with dedicated proteins and/or the formation of plectonemic coils. bacteria, compaction achieved dynamically, coordinated dense and constantly changing transcriptional activity. H-NS, a major bacterial nucleoid structuring protein, special interest due to its interplay RNA polymerase. H-NS:DNA nucleoprotein filaments inhibit transcription initiation by However, discovery that genes silenced H-NS can be activated originating...

Summary Ubiquitous RNA‐binding protein Hfq mediates the regulatory activity of many small RNAs (sRNAs) in bacteria. To identify potential targets for Hfq‐mediated regulation Salmonella , we searched lacZ translational fusions whose varied presence or absence Hfq. Fusions downregulated by were more common than showing opposite response. Surprisingly, a subset isolates from major class, higher was due to transcriptional activation alternative sigma factor RpoE (σ E ). Activation σ regulon...

Many species of bacteria harbor multiple prophages in their genomes. Prophages often carry genes that confer a selective advantage to the bacterium, typically during host colonization. can convert infectious viruses through process known as induction, which is relevant spread bacterial virulence genes. The paradigm prophage set by phage Lambda model, sees initiated RecA-stimulated self-proteolysis repressor. Here we show large family lambdoid found Salmonella genomes employs an alternative...

RNA-binding protein CsrA is a key regulator of variety cellular processes in bacteria, including carbon and stationary phase metabolism, biofilm formation, quorum sensing, virulence gene expression pathogens. binds to bipartite sequence elements at or near the ribosome loading site messenger RNA (mRNA), most often inhibiting translation initiation. Here we describe an alternative novel mechanism through which achieves negative regulation. We show that binding upstream portion 5' untranslated...

Summary Prokaryotic regulatory small RNAs act by a conserved mechanism and yet display stunning structural variability. In the present study, we used mutational analysis to dissect functional anatomy of RybB, σ E ‐dependent sRNA that regulates synthesis major porins in Escherichia coli Salmonella . Mutations chromosomal rybB locus altered expression an ompC – lac fusion were identified. Some mutations cluster within seven‐nucleotide segment at 5′ end affect its ability pair with sequence 40...

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Numerous intracellular bacterial pathogens interfere with macrophage function, including polarization, to establish a niche and persist. However, the spatiotemporal dynamics of polarization during infection within host remain be investigated. Here, we implement model persistent Salmonella Typhimurium in zebrafish, which allows visualization polarized macrophages bacteria real time at high resolution. While polarize toward M1-like phenotype control early infection, later stages, persists...

ABSTRACT The Gifsy-2 temperate bacteriophage of Salmonella enterica serovar Typhimurium contributes significantly to the pathogenicity strains that carry it as a prophage. Previous studies have shown encodes SodCI, periplasmic Cu/Zn superoxide dismutase, and at least one additional virulence factor. island 2 type III secreted effector protein. Sequence analysis genome also identifies several open reading frames with homology those known genes. However, we found null mutations in these genes...

Summary In Escherichia coli and Salmonella enterica , activation of σ E ‐dependent envelope stress response leads to the abrupt decline in synthesis all major outer membrane proteins (OMPs). Recent studies found that two ‐controlled small RNAs (sRNAs), MicA RybB, downregulate a number OMPs. While RybB targets several different mRNAs, including ompC ompD was up date thought act solely on ompA . Here we present evidence showing downregulates second OMP: LamB maltoporine. strains overexpressing...

RNA-binding protein Hfq is a key component of the adaptive responses many proteobacterial species including Escherichia coli, Salmonella enterica and Vibrio cholera. In these organisms, importance largely stems from its participation to regulatory mechanisms involving small non-coding RNAs. contrast, function in Gram-positive bacteria has remained elusive somewhat controversial. present study, we have further addressed this point by comparing growth phenotypes transcription profiles between...

Significance Mg 2+ is an important signal for the regulation of virulence and thermotolerance in Salmonella enterica . Transcription mgtA gene, which encodes a transporter , highly induced by limitation. The 5′ leader mRNA mgtL 17-codon, proline-rich open reading frame, whose translation controls transcription : efficient results termination upstream protein-coding region, whereas slow or incomplete antagonizes this termination. We show that proline codons present impediment to at low but...

We present evidence for the existence of two large (≈50 kb) excisable segments in chromosome Salmonella typhimurium . The elements — designated Gifsy ‐1 and ‐2 cover, respectively, 57 units 24 genetic map where they contribute indicative rare restriction sites. are closely interrelated both contain a region sequence similarity to recE locus Rac prophage Escherichia coli. Mutations within this Gifsy‐ 1 yield classical ‘Sbc’ phenotype: suppress recombination defect recB mutants, apparently by...