Two Restriction and Modification Systems in Staphylococcus aureus NCTC8325

Iordănescu, S; Surdeanu, M

doi:10.1099/00221287-96-2-277

Cited by 184 publications

(77 citation statements)

References 10 publications

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“…S. aureus strains SA113 (ATCC 35556), a derivative of S. aureus NCTC8325 (15), and COL (34) were used in this study. All staphylococcal strains were cultured at 37°C in BM broth (1% soy peptone, 0.5% yeast extract, 0.5% NaCl, 0.1% K 2 HPO 4 , 0.1% glucose).…”

Section: Methodsmentioning

confidence: 99%

Characterization of the Oxygen-Responsive NreABC Regulon of Staphylococcus aureus

et al. 2008

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Here, we investigate the functionality of the oxygen-responsive nitrogen regulation system NreABC in the human pathogen Staphylococcus aureus and evaluate its role in anaerobic gene regulation and virulence factor expression. Deletion of nreABC resulted in severe impairment of dissimilatory nitrate and nitrite reduction and led to a small-colony phenotype in the presence of nitrate during anaerobic growth. For characterization of the NreABC regulon, comparative DNA microarray and proteomic analyses between the wild type and nreABC mutant were performed under anoxic conditions in the absence and presence of nitrate. A reduced expression of virulence factors was not observed in the mutant. However, both the transcription of genes involved in nitrate and nitrite reduction and the accumulation of corresponding proteins were highly decreased in the nreABC mutant, which was unable to utilize nitrate as a respiratory oxidant and, hence, was forced to use fermentative pathways. These data were corroborated by the quantification of the extracellular metabolites lactate and acetate. Using an Escherichia coli-compatible two-plasmid system, the activation of the promoters of the nitrate and nitrite reductase operons and of the putative nitrate/nitrite transporter gene narK by NreBC was confirmed. Overall, our data indicate that NreABC is very likely a specific regulation system that is essential for the transcriptional activation of genes involved in dissimilatory reduction and transport of nitrate and nitrite. The study underscores the importance of NreABC as a fitness factor for S. aureus in anoxic environments.

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Section: Methodsmentioning

confidence: 99%

Characterization of the Oxygen-Responsive NreABC Regulon of Staphylococcus aureus

et al. 2008

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“…S. aureus Newman (kindly provided by T. Foster, Dublin, Ireland) was used to generate the ⌬eap mutant. Recombinant plasmids cloned in Escherichia coli were passaged in a restriction-negative S. aureus strain, SA113 (16), before electroporation to S. aureus Newman. Staphylococcus carnosus TM300 (11) was used as an intermediate host in the construction of a complemented strain.…”

Section: Methodsmentioning

confidence: 99%

Insertional Inactivation of eap in Staphylococcus aureus Strain Newman Confers Reduced Staphylococcal Binding to Fibroblasts

et al. 2002

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In contrast, the mutant adhered to a significantly lesser extent to cultured fibroblasts (P < 0.001) than did the wild type, while adherence was restorable upon complementation. Furthermore, adherence to both epithelial cells (P < 0.05) and fibroblasts (not significant) could be blocked with antibodies against Eap, whereas preimmune serum was not active. In conclusion, Eap may contribute to pathogenicity by promoting adhesion of whole staphylococcal cells to complex eukaryotic substrates.Staphylococcus aureus continues to be a major cause of human disease, accounting for superficial skin infections as well as for serious invasive infections, such as endocarditis, osteomyelitis, and septic shock (22). Adherence of S. aureus to components of host tissues is an important first step in colonization and subsequent infection (10, 35) and is mediated by specific interactions between adhesins on the bacterial cell surface and host cell receptors (6). In addition to the wellcharacterized bacterial surface-located proteins (28) conferring adhesion to various extracellular matrix proteins (6) and invasion of eukaryotic cells (1, 31, 32), other adhesive S. aureus proteins are secreted. Three of these molecules, i.e., coagulase (29), the extracellular fibrinogen (Fg)-binding protein Efb (26), and the extracellular adherence protein Eap (24), have been shown to bind Fg. Eap of S. aureus Newman has been cloned and sequenced (14) and has previously been shown to bind to additional plasma proteins, including fibronectin (Fn) and prothrombin (Pt) (24). Eap can form oligomers, and by rebinding to the staphylococcal cell surface, it mediates bacterial agglutination. It also enhances binding to epithelial cells and fibroblasts by its dual affinity for eukaryotic components and the S. aureus surface (24). While these observations have been made with purified Eap, further precision in describing the role of Eap, as well as of the related molecule Map (for major histocompatibility complex class II analogous protein [19]), in intact S. aureus cells has been hampered by the lack of availability of defined Eap-negative mutants. Thus, we have constructed an eap-deficient mutant by allelic replacement, and here we report the genotypic and phenotypic characteristics of the mutant compared to the parent strain. MATERIALS AND METHODSBacterial strains and media. S. aureus Newman (kindly provided by T. Foster, Dublin, Ireland) was used to generate the ⌬eap mutant. Recombinant plasmids cloned in Escherichia coli were passaged in a restriction-negative S. aureus strain, SA113 (16), before electroporation to S. aureus Newman. Staphylococcus carnosus TM300 (11) was used as an intermediate host in the construction of a complemented strain. The following strains of E. coli were used as cloning hosts: E. coli DH5␣, E. coli TG1, and E. coli SCS 110 (Stratagene, La Jolla, Calif.).For cultivation of S. aureus, tryptic soy broth and agar (Difco, Detroit, Mich.), brain heart broth and agar (Merck, Darmstadt, Germany), Mueller-Hinton broth and agar (Mast, Me...

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“…As shown in Figure 4A, WT GBS COH1 made both glycoplipids (MGlcDAG and DGlcDAG), while the iagA mutant failed to synthesize DGlcDAG. A strain of Staphylococcus aureus, SA113 (18), that contains only DGlcDAG in detectable amounts is shown as control. This experiment confirmed the predicted activity of the GBS iagA gene product as a DGlcDAG synthase.…”

Section: Identification Of a Gbs Gene Associated With Hbmec Invasionmentioning

confidence: 99%

Blood-brain barrier invasion by group B Streptococcus depends upon proper cell-surface anchoring of lipoteichoic acid

Doran¹,

Engelson²,

Khosravi³

et al. 2005

J. Clin. Invest.

202

272

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Group B streptococci (GBSs) are the leading cause of neonatal meningitis. GBSs enter the CNS by penetrating the blood-brain barrier (BBB), which consists of specialized human brain microvascular endothelial cells (hBMECs). To identify GBS factors required for BBB penetration, we generated random mutant libraries of a virulent strain and screened for loss of hBMEC invasion in vitro. Two independent hypo-invasive mutants possessed disruptions in the same gene, invasion associated gene (iagA), which encodes a glycosyltransferase homolog. Allelic replacement of iagA in the GBS chromosome produced a 4-fold decrease in hBMEC invasiveness. Mice challenged with the GBS ∆iagA mutant developed bacteremia comparably to WT mice, yet mortality was significantly lower (20% vs. 90%), as was the incidence of meningitis. The glycolipid diglucosyldiacylglycerol, a cell membrane anchor for lipoteichoic acid (LTA) and predicted product of the IagA glycosyltransferase, was absent in the ∆iagA mutant, which consequently shed LTA into the media. Attenuation of virulence of the ∆iagA mutant was found to be independent of TLR2-mediated signaling, but bacterial supernatants from the ∆iagA mutant containing released LTA inhibited hBMEC invasion by WT GBS. Our data suggest that LTA expression on the GBS surface plays a role in bacterial interaction with BBB endothelium and the pathogenesis of neonatal meningitis.

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Two Restriction and Modification Systems in Staphylococcus aureus NCTC8325

Cited by 184 publications

References 10 publications

Characterization of the Oxygen-Responsive NreABC Regulon of Staphylococcus aureus

Characterization of the Oxygen-Responsive NreABC Regulon of Staphylococcus aureus

Insertional Inactivation of eap in Staphylococcus aureus Strain Newman Confers Reduced Staphylococcal Binding to Fibroblasts

Blood-brain barrier invasion by group B Streptococcus depends upon proper cell-surface anchoring of lipoteichoic acid

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