Yunnan Baiyao reduces hospital-acquired pressure ulcers via suppressing virulence gene expression and biofilm formation of Staphylococcus aureus.
ABSTRACT: Yunnan Baiyao (YB) as a kind of famous Chinese herbal medicine, possessed hemostatic, invigorating the circulation of blood, and anti-inflammatory effects. Identifying strategies to protect patients at risk for hospital-acquired pressure ulcers (HAPU) is essential. Herein, our results showed that YB treatment can effectively reduce the acne wound area and improve efficacy in a comparative study of 60 cases HAPU patients with S. aureus positive of acne wound pathogens. Furthermore, YB inhibited HIa expression and suppressed accessory gene regulator (agr) system controlled by regulatory RNA II and RNA III molecule using pALC1740, pALC1742 and pALC1743 S. aureus strain linked to gfpuvr reporter gene. Moreover, YB downregulated cao mRNA expression and inhibited coagulase activity by RT-PCR, slide and tube coagulase test. Additionally, YB downregulated seb, sec, sed, and tsst-1 mRNA expression to suppress enterotoxin and tsst-1 secretion and adhesion function related genes sarA, icaA, and cidA mRNA expression. Taken together, the data suggest that YB may reduce HAPU via suppressing virulence gene expression and biofilm formation of S. aureus.
Project description:Previous studies have shown that the administration of antibiotics at subinhibitory concentrations stimulates biofilm formation by the majority of multidrug-resistant Staphylococcus aureus (MRSA) strains. Here, we investigated the effect of subinhibitory concentrations of mupirocin on biofilm formation by the community-associated (CA) mupirocin-sensitive MRSA strain USA300 and the highly mupirocin-resistant clinical S. aureus SA01 to SA05 isolates. We found that mupirocin increased the ability of MRSA cells to attach to surfaces and form biofilms. Confocal laser scanning microscopy (CLSM) demonstrated that mupirocin treatment promoted thicker biofilm formation, which also correlated with the production of extracellular DNA (eDNA). Furthermore, quantitative real-time PCR (RT-qPCR) results revealed that this effect was largely due to the involvement of holin-like and antiholin-like proteins (encoded by the cidA gene), which are responsible for modulating cell death and lysis during biofilm development. We found that cidA expression levels significantly increased by 6.05- to 35.52-fold (P?<?0.01) after mupirocin administration. We generated a cidA-deficient mutant of the USA300 S. aureus strain. Exposure of the ?cidA mutant to mupirocin did not result in thicker biofilm formation than that in the parent strain. We therefore hypothesize that the mupirocin-induced stimulation of S. aureus biofilm formation may involve the upregulation of cidA.
Project description:BACKGROUND:Neonatal Staphylococcus aureus (S. aureus) bacteremia is an important cause of morbidity and mortality. In this study, we examined whether methicillin-susceptible S. aureus (MSSA) transmission and genetic makeup contribute to the occurrence of neonatal S. aureus bacteremia. METHODS:A retrospective, single-centre study was performed. All patients were included who suffered from S. aureus bacteremia in the neonatal intensive care unit (NICU), Erasmus MC-Sophia, Rotterdam, the Netherlands, between January 2011 and November 2017. Whole-genome sequencing (WGS) was used to characterize the S. aureus isolates, as was also done in comparison to reference genomes. Transmission was considered likely in case of genetically indistinguishable S. aureus isolates. RESULTS:Excluding coagulase-negative staphylococci (CoNS), S. aureus was the most common cause of neonatal bacteremia. Twelve percent (n?=?112) of all 926 positive blood cultures from neonates grew S. aureus. Based on core genome multilocus sequence typing (cgMLST), 12 clusters of genetically indistinguishable MSSA isolates were found, containing 33 isolates in total (2-4 isolates per cluster). In seven of these clusters, at least two of the identified MSSA isolates were collected within a time period of one month. Six virulence genes were present in 98-100% of all MSSA isolates. In comparison to S. aureus reference genomes, toxin genes encoding staphylococcal enterotoxin A (sea) and toxic shock syndrome toxin 1 (tsst-1) were present more often in the genomes of bacteremia isolates. CONCLUSION:Transmission of MSSA is a contributing factor to the occurrence of S. aureus bacteremia in neonates. Sea and tsst-1 might play a role in neonatal S. aureus bacteremia.
Project description:Staphylococcus aureus is an important pathogen in hospital and community infections. Fusidic acid is particularly effective in treating skin and wound infections caused by staphylococci. The purpose of our study was to clarify the effect of fusidic acid on the biofilm formation and ?-toxin expression of S. aureus at subinhibitory concentrations [1/64, 1/32, and 1/16 × minimum inhibitory concentration (MIC)]. A total of 504 genes greater than a twofold or less than twofold change in expression of S. aureus effected by subinhibitory concentrations of fusidic acid were found, including 232 up-regulated genes and 272 down-regulated genes, which were determined by transcriptome sequencing. Our results showed subinhibitory concentrations of fusidic acid significantly inhibited the expression of hla, spa, icaA, and cidA at the mRNA level in clinical S. aureus strains tested. And subinhibitory concentrations of fusidic acid can significantly reduce the hemolysis activity and ?-toxin production of S. aureus. In addition, the subinhibitory concentrations of fusidic acid significantly inhibited biofilm formation, autolysis, cell aggregation, and polysaccharide intercellular adhesin (PIA) production of S. aureus. Moreover, fusidic acid effectively reduces the damage of mouse skin lesion area. Furthermore, fusidic acid reduced the expression of the two-component regulatory system saeRS and staphylococcal accessory gene regulator (sarA). In conclusion, our results suggested that the subinhibitory concentrations of fusidic acid may reduce the virulence of S. aureus by down-regulating sarA and saeRS to reduce biofilm formation and ?-toxin expression, which will provide a theoretical basis for the clinical treatment of S. aureus infection. This is the first report that fusidic acid has an inhibitory effect on the virulence of S. aureus, and this broadens the clinical application of fusidic acid.
Project description:Recent studies have shown that expression of the Staphylococcus aureus lrgAB operon inhibits murein hydrolase activity and decreases sensitivity to penicillin-induced killing. It was proposed that the lrgAB gene products function in a manner analogous to an antiholin, inhibiting a putative holin from transporting murein hydrolases out of the cell. In the present study the cidAB operon was identified and characterized based on the similarity of the cidA and cidB gene products to the products of the lrgAB operon. Zymographic and quantitative analyses of murein hydrolase activity revealed that mutation of the cidA gene results in decreased extracellular murein hydrolase activity compared to that of S. aureus RN6390, the parental strain. Complementation of cidA expression restored the wild-type phenotype, indicating that expression of the cidAB operon has a positive influence on extracellular murein hydrolase activity. The cidA mutant also displayed a significant decrease in sensitivity to the killing effects of penicillin. However, complementation of the cidA defect did not restore penicillin sensitivity to wild-type levels. Reverse transcriptase PCR also revealed that cidAB is maximally expressed during early exponential growth, opposite of what was previously observed for lrgAB expression. Based on these results, we propose that the cidAB operon encodes the holin-like counterpart of the lrgAB operon and acts in a manner opposite from that of lrgAB by increasing extracellular murein hydrolase activity and increasing sensitivity to penicillin-induced killing.
Project description:The cell surface-associated extracellular adherence protein (Eap) mediates adherence of Staphylococcus aureus to host extracellular matrix components and inhibits inflammation, wound healing, and angiogenesis. A well-characterized collection of S. aureus and non-S. aureus staphylococcal isolates (n = 813) was tested for the presence of the Eap-encoding gene (eap) by PCR to investigate the use of the eap gene as a specific diagnostic tool for identification of S. aureus. Whereas all 597 S. aureus isolates were eap positive, this gene was not detectable in 216 non-S. aureus staphylococcal isolates comprising 47 different species and subspecies of coagulase-negative staphylococci and non-S. aureus coagulase-positive or coagulase-variable staphylococci. Furthermore, non-S. aureus isolates did not express Eap homologs, as verified on the transcriptional and protein levels. Based on these data, the sensitivity and specificity of the newly developed PCR targeting the eap gene were both 100%. Thus, the unique occurrence of Eap in S. aureus offers a promising tool particularly suitable for molecular diagnostics of this pathogen.
Project description:Sodium houttuyfonate (SH), an addition compound of sodium bisulfite and houttuynin, showed in vitro antibacterial activity against 21 Staphylococcus aureus (S. aureus) strains grown in planktonic cultures. Microarray results showed decreased levels of autolysin atl, sle1, cidA and lytN transcripts in the SH-treated strain as compared to the control strain, consistent with the induction of the autolytic repressors lrgAB and sarA and with the downregulation of the positive regulators agrA and RNAIII. Triton X-100-induced autolysis was significantly decreased by SH in S. aureus ATCC 25923, and quantitative bacteriolytic assays and zymographic analysis demonstrated SH-mediated reduction of extracellular murein hydrolase activity in these cells. Anti-biofilm assay showed that SH is poorly active against S. aureus grown in biofilm cultures, whereas SH diminished the amounts of extracellular DNA (eDNA) of S. aureus in a dose-dependent manner, which suggested that SH may impede biofilm formation by reducing the expression of cidA to inhibit autolysis and eDNA release in the early phase. Some of the microarray results were confirmed by real-time RT-PCR.
Project description:Asymptomatic carriers of toxigenic Staphylococcus aureus are potential source of diseases, including food poisoning. Toxigenic potential and genetic traits of colonizing S. aureus were investigated for 563 healthy food handlers in Myanmar. Carriage of S. aureus was found in 110 individuals (19.5%), and a total of 144 S. aureus isolates were recovered from nasal cavities (110 isolates) and hands (34 isolates). Panton-Valentine leucocidin genes (pvl) were detected in 18 isolates (12.5%), among which 11 isolates were classified into coa-VIa, agr type III, and ST1930 (CC96) that had been also detected in pvl-positive clinical isolates in Myanmar. A pvl-positive, ST2250 nasal isolate was identified as S. argenteus, a novel coagulase-positive staphylococcus species. Toxic shock syndrome toxin-1 (TSST-1) gene was detected in five pvl-negative isolates. All of the 144 isolates harbored at least one of the 21 enterotoxin(-like) gene(s). The most prevalent enterotoxin(-like) gene was selw (98%), followed by selx (97%), sei (28%), sely (28%), sem (26%), sel (24%), and sea and sec (22% each). Considerable genetic diversity with five groups was detected for selw. The present study revealed the relatively high rate of pvl, as well as the wide distribution of enterotoxin(-like) genes among colonizing S. aureus in Myanmar.
Project description:Staphylococcus lugdunensis has gained recognition as an atypically virulent pathogen with a unique microbiological and clinical profile. S. lugdunensis is coagulase negative due to the lack of production of secreted coagulase, but a membrane-bound form of the enzyme present in some isolates can result in misidentification of the organism as Staphylococcus aureus in the clinical microbiology laboratory. S. lugdunensis is a skin commensal and an infrequent pathogen compared to S. aureus and S. epidermidis, but clinically, infections caused by this organism resemble those caused by S. aureus rather than those caused by other coagulase-negative staphylococci. S. lugdunensis can cause acute and highly destructive cases of native valve endocarditis that often require surgical treatment in addition to antimicrobial therapy. Other types of S. lugdunensis infections include abscess and wound infection, urinary tract infection, and infection of intravascular catheters and other implanted medical devices. S. lugdunensis is generally susceptible to antimicrobial agents and shares CLSI antimicrobial susceptibility breakpoints with S. aureus. Virulence factors contributing to this organism's heightened pathogenicity remain largely unknown. Those characterized to date suggest that the organism has the ability to bind to and interact with host cells and to form biofilms on host tissues or prosthetic surfaces.
Project description:Although coagulase-positive staphylococci are considered to be the main factor responsible for food poisoning, an increasing role for the coagulase-negative staphylococci in the production of enterotoxins has been observed in recent years. This study was conducted to assess the occurrence of genes responsible for the production of staphylococcal enterotoxins (SE), enterotoxin-like toxins (SEI) and toxic shock syndrome toxin-1 (TSST-1) in coagulase-negative staphylococci (CoNS) isolated from ready-to-eat food from bars and restaurants. One hundred and eighteen CoNS strains were tested using polymerase chain reaction (PCR) to five superantigenic toxin genes, including five different types of classical enterotoxins (sea, seb, sec, sed and see) and the toxic shock syndrome toxin-1 (tsst-1) as well as to supertoxin-like genes. PCR-positive isolates were then tested using immunoenzymatic methods (SET-RPLA, Vidas SET 2) for toxin expression. Out of 118 CoNS strains, the presence of staphylococcal enterotoxins was confirmed in 72% of them. The most frequently found enterotoxin-like genotype was ser, selu. Two of the tested strains had up to ten different enterotoxin genes in the genome at the same time. Although no production of enterotoxins was detected in the CoNS, which means that their possible role in the epidemiology of food-borne diseases is minimal, the data demonstrated that the toxigenic capacity of the CoNS should not be ignored, and that this group of microorganisms should be continuously monitored in food.
Project description:PURPOSE:Extracellular protein toxins contribute to the pathogenesis of Staphylococcus aureus infections. The present study compared the effects of iclaprim and trimethoprim - two folic acid synthesis inhibitors - with nafcillin and vancomycin on production of Panton-Valentine leukocidin (PVL), alpha haemolysin (AH) and toxic-shock syndrome toxin I (TSST-1) in methicillin-resistant and vancomycin-intermediate S. aureus (MRSA and VISA, respectively). METHODOLOGY:Northern blotting and RT-PCR were used to assess gene transcription; toxin-specific bioassays were used to measure protein toxin production. RESULTS:As shown previously, sub-inhibitory concentrations (sub-MIC) of nafcillin increased and prolonged MRSA toxin gene transcription and enhanced PVL, TSST-1 and AH production. Sub-inhibitory doses of iclaprim and trimethoprim delayed maximal AH gene (hla) transcription and suppressed AH production; both drugs delayed, but neither reduced, maximal TSST-1 production. Trimethoprim significantly increased lukF-PV expression and PVL production compared to both untreated and iclaprim-treated cultures. Higher concentrations of iclaprim and trimethoprim markedly suppressed MRSA growth, mRNA synthesis and toxin production. In VISA, iclaprim, vancomycin and nafcillin variably increased tst and hla expression, but only nafcillin increased toxin production. Despite its ability to increase hla expression, iclaprim was the most potent inhibitor of AH production. CONCLUSIONS:We conclude that, due to its ability to suppress toxin production, iclaprim should be effective against severe staphylococcal infections caused by toxin-producing MRSA and VISA strains, especially given its ability to concentrate at sites of infection such as skin and skin structures and the lung.