Influence of Nisin-Biogel at Subinhibitory Concentrations on Virulence Expression in Staphylococcus aureus Isolates from Diabetic Foot Infections.
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SUBMITTER: Jesus C
PROVIDER: S-EPMC8698857 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundDiabetic foot infections (DFIs) are a frequent complication of Diabetes mellitus and a major cause of nontraumatic limb amputations. The Gram-positive bacterium Staphylococcus aureus, known for its resilient biofilms and antibiotic resistant profile, is the most frequent DFI pathogen. It is urgent to develop innovative treatments for these infections, being the antimicrobial peptide (AMP) nisin a potential candidate. We have previously proposed the use of a guar gum biogel as a delivery system for nisin. Here, we evaluated the potential of the nisin-biogel to enhance the efficacy of conventional antibiotics and antiseptics against DFIs S. aureus clinical isolates.MethodsA collection of 23 S. aureus strains isolated from DFI patients, including multidrug- and methicillin-resistant strains, was used. The antimicrobial activity of the nisin-biogel was tested alone and in different combinations with the antiseptic chlorhexidine and the antibiotics clindamycin, gentamicin and vancomycin. Isolates' in vitro susceptibility to the different protocols was assessed using broth microdilution methods in order to determine their ability to inhibit and/or eradicate established S. aureus biofilms. Antimicrobials were added to the 96-well plates every 8 h to simulate a typical DFI treatment protocol. Statistical analysis was conducted using RCBD ANOVA in SPSS.ResultsThe nisin-biogel showed a high antibacterial activity against biofilms formed by DFI S. aureus. The combined protocol using nisin-biogel and chlorhexidine presented the highest efficacy in biofilm formation inhibition, significantly higher (p<0.05) than the ones presented by the antibiotics-based protocols tested. Regarding biofilm eradication, there were no significant differences (p>0.05) between the activity of the combination nisin-biogel plus chlorhexidine and the conventional antibiotic-based protocols.ConclusionsResults provide a valuable contribution for the development of complementary strategies to conventional antibiotics protocols. A combined protocol including chlorhexidine and nisin-biogel could be potentially applied in medical centres, contributing for the reduction of antibiotic administration, selection pressure on DFI pathogens and resistance strains dissemination.
| S-EPMC6655664 | biostudies-literature
Project description:AIM:l-benzyl-3-cetyl-2-methylimidazolium iodide (NH125) can inhibit Staphylococcus aureus growth. We investigated the effects of sub-MIC concentrations of NH125 on S. aureus biofilm and virulence. Methodology & results: Three strains of S. aureus were tested. Sub-lethal concentrations of NH125 repressed biofilm formation. At partial sub-MICs, NH125 downregulated the expression of most virulence, while strain-dependent effects were found in the production of ?-hemolysin, ?-hemolysin, coagulase and nuclease. In Galleria mellonella model, methicillin-resistant S. aureus pre-exposed to NH125 demonstrated significantly lower killing (p = 0.032 for 1/16 and 1/8 MICs; 0.008 for 1/4 MIC; and 0.001 for 1/2 MIC). CONCLUSION:Sub-MIC concentrations of NH125 inhibited biofilm formation and virulence of S. aureus. These findings provide further support for evaluating the clinical efficacy of NH125 in staphylococcal infection.
| S-EPMC6088273 | biostudies-literature
Project description:The most prevalent microorganism in diabetic foot infections (DFI) is Staphylococcus aureus, an important multidrug-resistant pathogen. The antimicrobial peptide nisin is a promising compound for DFI treatment, being effective against S. aureus. However, to avoid the selection of resistant mutants, correct drug therapeutic doses must be established, being also important to understand if nisin subinhibitory concentrations (subMIC) can potentiate resistant genes transfer between clinical isolates or mutations in genes associated with nisin resistance. The mutant selection window (MSW) of nisin was determined for 23 DFI S. aureus isolates; a protocol aiming to prompt vanA horizontal transfer between enterococci to clinical S. aureus was performed; and nisin subMIC effect on resistance evolution was assessed through whole-genome sequencing (WGS) applied to isolates subjected to a MEGA-plate assay. MSW ranged from 5-360 μg/mL for two isolates, from 5-540 μg/mL for three isolates, and from 5-720 μg/mL for one isolate. In the presence of nisin subMIC values, no transconjugants were obtained, indicating that nisin does not seem to promote vanA transfer. Finally, WGS analysis showed that incubation in the presence of nisin subMIC did not promote the occurrence of significant mutations in genes related to nisin resistance, supporting nisin application to DFI treatment.
| S-EPMC9311964 | biostudies-literature
Project description:Staphylococcus aureus (S. aureus) infections are notoriously complicated by the ability of the organism to grow in biofilms and are difficult to eradicate with antimicrobial therapy. The purpose of the current study was to clarify the influence of sub-inhibitory concentrations (sub-MICs) of daptomycin and tigecycline antibiotics on biofilm adhesion factors and exoproteins expressions by S. aureus clinical isolates. Six clinical isolates representing positive biofilm S. aureus clones (3 methicillin-sensitive S. aureus (MSSA) and 3 methicillin-resistant S. aureus (MRSA)) were grown with sub-MICs (0.5 MIC) of two antibiotics (daptomycin and tigecycline) for 12 h of incubation. RNA extracted from culture pellets was used via relative quantitative real-time-PCR (qRT-PCR) to determine expression of specific adhesion (fnbA, fnbB, clfA, clfB, fib, ebps, cna, eno) and biofilm (icaADBC) genes. To examine the effect of sub-MIC of these antibiotics on the expression of extracellular proteins, samples from the culture supernatants of six isolates were collected after 12 h of treatment with or without tigecycline in order to profile protein production via 2D gel sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2D gel-SDS-PAGE). Sub-MIC treatment of all clinical MRSA and MSSA strains with daptomycin or tigecycline dramatically induced or suppressed fnbA, fnbB, clfA, clfB, fib, ebps, cna, eno, and icaADBC gene expression. Furthermore, sub-MIC use of tigecycline significantly reduced the total number of separated protein spots across all the isolates, as well as decreasing production of certain individual proteins. Collectively, this study showed very different responses in terms of both gene expression and protein secretion across the various isolates. In addition, our results suggest that sub-MIC usage of daptomycin and tigecycline could signal virulence induction by S. aureus via the regulation of biofilm adhesion factor genes and exoproteins. If translating findings to the clinical treatment of S. aureus, the therapeutic regimen should be adapted depending on antibiotic, the virulence factor and strain type.
| S-EPMC7823975 | biostudies-literature
Project description:BackgroundThe pathogenicity of staphylococcus aureus is dependent largely upon its ability to secrete a number of virulence factors, therefore, anti-virulence strategy to combat S. aureus-mediated infections is now gaining great interest. It is widely recognized that some plant essential oils could affect the production of staphylococcal exotoxins when used at subinhibitory concentrations. Perilla [Perilla frutescens (L.) Britton], a natural medicine found in eastern Asia, is primarily used as both a medicinal and culinary herb. Its essential oil (perilla oil) has been previously demonstrated to be active against S. aureus. However, there are no data on the influence of perilla oil on the production of S. aureus exotoxins.Methodology/principal findingsA broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) of perilla oil against S. aureus strains. Hemolysis, tumour necrosis factor (TNF) release, Western blot, and real-time RT-PCR assays were performed to evaluate the effects of subinhibitory concentrations of perilla oil on exotoxins production in S. aureus. The data presented here show that perilla oil dose-dependently decreased the production of α-toxin, enterotoxins A and B (the major staphylococcal enterotoxins), and toxic shock syndrome toxin 1 (TSST-1) in both methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA).Conclusions/significanceThe production of α-toxin, SEA, SEB, and TSST-1 in S. aureus was decreased by perilla oil. These data suggest that perilla oil may be useful for the treatment of S. aureus infections when used in combination with β-lactam antibiotics, which can increase exotoxins production by S. aureus at subinhibitory concentrations. Furthermore, perilla oil could be rationally applied in food systems as a novel food preservative both to inhibit the growth of S. aureus and to repress the production of exotoxins, particularly staphylococcal enterotoxins.
| S-EPMC3023776 | biostudies-literature
Project description:Staphylococcus aureus biofilms are extremely difficult to treat. They provide a protected niche for the bacteria, rendering them highly recalcitrant toward host defenses as well as antibiotic treatment. Bacteria within a biofilm are shielded from the immune system by the formation of an extracellular polymeric matrix, composed of polysaccharides, extracellular DNA (eDNA), and proteins. Many antibiotics do not readily penetrate biofilms, resulting in the presence of subinhibitory concentrations of antibiotics. Here, we show that subinhibitory concentrations of clindamycin triggered a transcriptional stress response in S. aureus via the alternative sigma factor B (?(B)) and upregulated the expression of the major biofilm-associated genes atlA, lrgA, agrA, the psm genes, fnbA, and fnbB Our data suggest that subinhibitory concentrations of clindamycin alter the ability of S. aureus to form biofilms and shift the composition of the biofilm matrix toward higher eDNA content. An understanding of the molecular mechanisms underlying biofilm assembly and dispersal in response to subinhibitory concentrations of clinically relevant antibiotics such as clindamycin is critical to further optimize antibiotic treatment strategies of biofilm-associated S. aureus infections.
| S-EPMC5038281 | biostudies-literature
Project description:Glycopeptides are known to select for heterogeneous vancomycin-intermediate Staphylococcus aureus (h-VISA) from susceptible strains. In certain clinical situations, h-VISA strains have been isolated from patients without previous exposure to glycopeptides, such as cystic fibrosis patients, who frequently receive repeated treatments with beta-lactam antibiotics. Our objective was to determine whether prolonged exposure to beta-lactam antibiotics can induce h-VISA. We exposed 3 clinical vancomycin-susceptible methicillin-resistant Staphylococcus aureus (MRSA) strains to ceftazidime, ceftriaxone, imipenem, and vancomycin (as a control) at subinhibitory concentrations for 18 days in vitro. Population analyses showed progressive increases in vancomycin resistance; seven of the 12 derived strains obtained after induction were classified as h-VISA according to the following criteria: area under the curve (AUC) on day 18/AUC of Mu3 of ?90% and/or growth on brain heart infusion (BHI) agar with 4 mg/liter vancomycin. The derived isolates had thickened cell walls proportional to the level of glycopeptide resistance. Genes known to be associated with glycopeptide resistance (vraSR, yvqF, SA1703, graRS, walKR, and rpoB) were PCR sequenced; no de novo mutations were observed upon beta-lactam exposure. To determine whether trfA, a gene encoding a glycopeptide resistance factor, was essential in the selection of h-VISA upon beta-lactam pressure, a trfA-knockout strain was generated by allelic replacement. Indeed, beta-lactam exposure of this mutated strain showed no capacity to induce vancomycin resistance. In conclusion, these results showed that beta-lactam antibiotics at subinhibitory concentrations can induce intermediate vancomycin resistance in vitro. This induction required an intact trfA locus. Our results suggest that prior use of beta-lactam antibiotics can compromise vancomycin efficacy in the treatment of MRSA infections.
| S-EPMC4135816 | biostudies-other
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.
| S-EPMC7038260 | biostudies-literature
Project description:ObjectiveThe purpose of this study was to assess the virulence potential of Staphylococcus aureus strains isolated from diabetic foot ulcers and to discriminate noninfected from infected ulcers.Research design and methodsDiabetic patients hospitalized in a diabetic foot department with a foot ulcer were prospectively enrolled if they had been free of antibiotic treatment over the previous 6 months. At admission, ulcers were classified as infected or noninfected on the basis of clinical examination, according to the International Working Group on the Diabetic Foot system. Only patients carrying S. aureus as the sole pathogen were included. In individuals with a grade 1 ulcer, a second bacterial specimen was obtained 1 month later. Using virulence genotyping markers, clonality tools, and an in vivo Caenorhabditis elegans model, we correlated the virulence of 132 S. aureus strains with grade, time of collection, and ulcer outcome.ResultsAmong virulence genes, the most relevant combination derived from the logistic regression was the association of cap8, sea, sei, lukE, and hlgv (area under the curve 0.958). These markers were useful to distinguish noninfected (grade 1) from infected (grades 2-4) ulcers and to predict wound status at the follow-up. With use of the nematode model, S. aureus strains isolated from grade 1 ulcers were found to be significantly less virulent than strains from ulcers at or above grade 2 (P < 0.001).ConclusionsThis study highlights the coexistence of two S. aureus populations on diabetic foot ulcers. A combination of five genes that may help distinguish colonized grade 1 from infected grade >or=2 wounds, predict ulcer outcome, and contribute to more appropriate use of antibiotics was discovered.
| S-EPMC2584189 | biostudies-literature
Project description:Nisin A is a bacteriocin produced by Lactococcus lactis and is widely used as a food preservative. Staphylococcus aureus has the BraRS-VraDE system that provides resistance against low concentrations of nisin A. BraRS is a two-component system that induces the expression of the ABC transporter VraDE. Previously, we isolated a highly nisin A-resistant strain with increased VraDE expression due to a mutation in braRS In this study, we isolated S. aureus MW2 mutants with BraRS-VraDE-independent nisin A resistance. These mutants, designated SAN2 ( S.aureusnisin resistant) and SAN469, had a mutation in pmtR, which encodes a transcriptional regulator responsible for the expression of the pmtABCD operon. As a result, these mutants exhibited increased expression of PmtABCD, a transporter responsible for the export of phenol-soluble modulin (PSM). Characterization of the mutants revealed that they have decreased susceptibility to human β-defensin-3 (hBD3) and LL37, which are innate immune factors. Additionally, these mutants showed higher hemolytic activity than the original MW2 strain. Furthermore, in a mouse bacteremia model, the SAN2 strain exhibited a lower survival rate than the original MW2 strain. These results indicate that the increased expression of pmtABCD due to a pmtR mutation is an alternative nisin A resistance mechanism that also affects virulence in S. aureusIMPORTANCE Recently, the emergence of antibiotic-resistant bacteria has resulted in serious problems for chemotherapy. In addition, many antibacterial agents, such as disinfectants and food additives, are widely used. Therefore, there is a possibility that bacteria are becoming resistant to some antibacterial agents. In this study, we investigated whether Staphylococcus aureus can become resistant to nisin A, one of the bacteriocins applied as a food additive. We isolated a highly nisin A-resistant strain designated SAN2 that displayed increased expression of Pmt proteins, which are involved in the secretion of virulence factors called phenol-soluble modulins (PSMs). This strain also showed decreased susceptibility to human antimicrobial peptides and increased hemolytic activity. In addition, SAN2 showed increased lethal activity in a mouse bacteremia model. Our study provides new insights into the possibility that the acquisition of resistance against food preservatives may modulate virulence in S. aureus, suggesting that we need to pay more attention to the use of food preservatives together with antibiotics.
| S-EPMC7117933 | biostudies-literature