Identification and mapping of new chromosomal sites affecting response to β-lactams in Staphylococcus aureus

Vasilieva, E. A.; Labischinski, Harald; Berger‐Bächi, Brigitte

doi:10.1016/s0924-8579(96)00346-9

Cited by 7 publications

(5 citation statements)

References 32 publications

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“…The insertion ⍀2020::Tn551 was reported to increase susceptibility to ␤-lactams in S. aureus, but its nature remained unsolved (44). Sequencing of the insertion site revealed that the transposon had integrated into the intercistronic region of two divergently transcribed genes, tentatively termed msrA and msrR, which are separated by 136 nucleotides (Fig.…”

Section: Resultsmentioning

confidence: 99%

MsrR, a Putative Cell Envelope-Associated Element Involved in Staphylococcus aureus sarA Attenuation

Rossi

Bischoff

Wada

et al. 2003

Antimicrob Agents Chemother

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A novel membrane-associated protein, MsrR, was identified in Staphylococcus aureus which affects resistance to methicillin and teicoplanin, as well as the synthesis of virulence factors. MsrR belongs to the LytR-CpsA-Psr family of cell envelope-related transcriptional attenuators and was shown to be inducible by cell wall-active agents, such as ␤-lactams, glycopeptides, and lysostaphin. The expression of msrR peaked in the early exponential growth phase and decreased sharply thereafter. msrR mutants showed increased sarA transcription and an earlier and higher expression of RNAIII, resulting in altered expression of virulence factors such as alphatoxin and protein A. These observations suggest that MsrR is a new component involved in sarA attenuation and the regulatory network controlling virulence gene expression in S. aureus.Staphylococcus aureus, an inhabitant of the skin and mucous membranes in 10 to 30% of the healthy population, easily survives in the environment and is, moreover, the most common cause of community-acquired and nosocomial infections. The pathogenicity of S. aureus is based on an impressive repertoire of virulence factors. The diseases caused range from superficial infections of the skin to life-threatening infections such as septicemia, endocarditis, osteomyelitis, and toxic shock syndrome. Virulence factors comprise surface-associated and extracellular proteins such as toxins and enzymes. Cell surfaceassociated proteins, the MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), function in adhesion and colonization of the host and evasion of host defenses. Cell surface-associated factors are needed primarily during the initial stages of infection and are expressed mainly in the early exponential growth phase. The exoenzymes, which damage the host tissue and promote dissemination of the pathogen and most soluble exoproteins, have a role in a later stage of the infection and are produced in the postexponential phase. The coordinate and timely expression of those virulence factors during the growth cycle is governed by a complex network comprising two-component sensor transducers, global regulatory systems, and transcription factors, which are important for the pathogen to adapt to the changing host environment during different stages of infection (2,21,23,26). One of the two major global regulatory systems is the agr (accessory gene regulator) regulon. This locus consists of two divergent transcripts originating from the promoters P2 and P3, which produce RNAII and RNAIII, respectively. The RNAII transcript encodes four proteins, AgrA to AgrD. AgrA and AgrC form a two-component regulatory system whereby AgrC acts as the signal receptor and AgrA acts as sensor regulator. The signal is a small peptide processed from AgrD by AgrB. This self-encoded autoinducing peptide determines the specificity group of S. aureus (29). The autoinducer leads from the phosphorylation of AgrC to the phosphorylation of AgrA, which upregulates both P2 and P3 promoters of the agr regulon. T...

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Section: Resultsmentioning

confidence: 99%

MsrR, a Putative Cell Envelope-Associated Element Involved in Staphylococcus aureus sarA Attenuation

Rossi

Bischoff

Wada

et al. 2003

Antimicrob Agents Chemother

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“…Recently, Vasilieva et al (1997) isolated a ctaA mutant of S. aureus which shows an increased resistance to cefoxitin, a β-lactam. This mutant also forms a small colony variant similar to that seen in persistent and antibiotic-resistant S. aureus infections (Proctor et al, 1996).…”

Section: Discussionmentioning

confidence: 99%

Isolation and characterization of Staphylococcus aureus starvat ion4 nduced, stationary-phase mutants defective in survival or recovery

1998

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Ten Staphylococcus aureus mutants, defective in the starvation-induced stationary phase of growth were isolated from two independent Tn917-LTVI transposon insertion libraries and were designated suv as they had apparent -suryival defects. Seven of these mutants were defective under amino-acidlimiting conditions alone. Two mutants (suv-3 and suw-20) demonstrated lower plating efficiency when starved for glucose, phosphate or amino acids and one mutant (suv-11) had reduced plating efficiency after amino acid or glucose starvation. All of the mutants tested were as resistant to hydrogen peroxide assault as the parent, but six were more sensitive to low pH conditions. All the mutants were physically mapped on the 5. aureus chromosome using PFGE. Chromosomal DNA flanking the Tn917-LNI insertion sites was rescued by cloning into Escherichia coli. DNA sequence analysis resulted in the identification of a number of transposon-disrupted ORFs encoding putative components such as superoxide dismutase (suv-I), haem A synthase (suv-3)# a component of the 505 response (suv-9) and hypoxanthine-guanine phosphoribosyltransferase (suv-20). The Tn917-LTVI insertion created lac2 transcriptional fusions for some of the stationary-phase loci. Expression analysis indicated that suv-4 was induced at mid-exponential phase, whereas suv-3 and suv-II were induced at the onset of stationary phase. The possible roles of these suv components in stationary-phase survival or recovery is discussed. 2TN, UK

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“…Like in Bacillus subtilis, the S. aureus ctaA gene appears to be monocistronic, since potential rho-independent transcription terminators were located 9 bp downstream of the putative translation stop codon and 526 bp upstream of the predicted start codon, with ⌬G values of Ϫ36.4 and Ϫ3.6 kJ mol Ϫ1 respectively. Recently, Vasilieva et al (38) isolated a ctaA mutant of S. aureus during a screen for mutants with altered ␤-lactam (22) and B. firmus (28). Numbers represent positions in the amino acid sequence.…”

Section: Spw3 Has a Tn Insertion Located In A Homologue Of B Subtilimentioning

confidence: 99%

“…SPW3 has altered growth characteristics. For both B. subtilis and S. aureus, mutations within ctaA have been reported to result in a small-colony morphology on solid media (21,38). We clearly observed a small-colony morphology when S. aureus SPW3 was grown on either blood agar base (Difco) supplemented with sheep blood (5% [vol/vol]) or, after prolonged incubation, on CDM agar (37°C, 2 days), after which time the colonies were approximately 50% smaller than those of 8325-4 grown on the same plate (0.5 mm versus 1 mm in diameter).…”

Section: Spw3 Has a Tn Insertion Located In A Homologue Of B Subtilimentioning

confidence: 99%

CtaA ofStaphylococcus aureusIs Required for Starvation Survival, Recovery, and Cytochrome Biosynthesis

et al. 1999

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A Staphylococcus aureus mutant (SPW3) apparently unable to survive long-term starvation was shown to have a transposon insertion within a gene homologous to ctaA of Bacillus subtilis which encodes a heme A synthase. Analysis of the cytochrome profiles of SPW3 revealed the absence of heme A-containing cytochromes compared to the parental 8325-4 strain. SPW3 demonstrated a 100-fold reduction in the ability to survive starvation induced by glucose limitation, under aerated conditions, compared to 8325-4. Analysis of starved cultures revealed that greater than 90% of the cells which demonstrated metabolism (as shown by rhodamine 123 accumulation) were unable to recover and form colonies on agar. Analysis of the lag phase and initial growth kinetics of those cells which could recover also showed a defect. This recovery defect could be partially alleviated by the inclusion of catalase in the recovery medium, indicating the probable involvement of oxidative stress. SPW3 also exhibited reduced colony size similar to that of a small-colony variant, increased resistance to aminoglycoside antibiotics, and reduced hemolysin and toxic shock syndrome toxin 1 production, but no alteration in the ability to form lesions in a subcutaneous mouse infection model. MATERIALS AND METHODSCloning and sequencing of ctaA. Plasmid pSPW3, which contained chromosomal DNA flanking the lacZ proximal region of the Tn insertion in S. aureus SPW3, was generated in a previous study (42). The insert, which contains 750 bp of chromosomal DNA, was excised, digoxigenin (DIG) labeled according to the protocol of Boehringer GmbH (Mannheim, Germany), and used to probe a ZAP Express library of a partial Sau3A digest (2 to 10 kb) of S. aureus 8325-4 genomic DNA (10). A clone containing a 3.5-kb genomic DNA fragment, spanning the pSPW3 insert, was identified, and the stable phagemid pSPW30 was excised from ZAP Express in Escherichia coli XLOLR (Stratagene). A primerwalking-based approach was used to sequence a 1,100-bp region.Strains and growth conditions. Strains and plasmids used in this study are listed in Table 1. S. aureus 8325-4 (23) and SPW3 (42) were grown in either a chemically defined media (CDM) containing 0.1% glucose (14, 43) or brain heart infusion (BHI) broth (Oxoid). Agar plates were prepared by the addition of 1% (wt/vol) agar to the above. Catalase plates were prepared by overlaying 4 ml of CDM agar (1% [wt/vol]) containing bovine catalase (5 mg ml Ϫ1 ; Sigma) onto CDM agar. Sheep blood agar plates were made by the supplementation of blood agar base (Difco) with 5% (vol/vol) sheep blood (TCS Biologicals, Buckingham, United Kingdom). Starvation cultures were prepared by the inoculation of CDM to an optical density at 600 nm (OD 600 ) of 0.01, which was then incubated at 37°C with shaking (250 rpm). Stationary phase was reached after about 8 to 10 h. After 18 h of growth, cultures were either incubated statically or shaken further. Starved cells were recovered by the addition of 1/10 volume of glucose (1% [wt/vol]) and a mixture of 18 amino ...

show abstract

Identification and mapping of new chromosomal sites affecting response to β-lactams in Staphylococcus aureus

Cited by 7 publications

References 32 publications

MsrR, a Putative Cell Envelope-Associated Element Involved in Staphylococcus aureus sarA Attenuation

MsrR, a Putative Cell Envelope-Associated Element Involved in Staphylococcus aureus sarA Attenuation

Isolation and characterization of Staphylococcus aureus starvat ion4 nduced, stationary-phase mutants defective in survival or recovery

CtaA ofStaphylococcus aureusIs Required for Starvation Survival, Recovery, and Cytochrome Biosynthesis

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