Methicillin-sensitive S. Aureus Strains Research Articles

Distinguishing methicillin-resistant Staphylococcus aureus from methicillin-sensitive strains by combining Fe3O4 magnetic nanoparticle-based affinity mass spectrometry with a machine learning strategy

Pathogenic bacteria, including drug-resistant variants such as methicillin-resistant Staphylococcus aureus (MRSA), can cause severe infections in the human body. Early detection of MRSA is essential for clinical diagnosis and proper treatment, considering the distinct therapeutic strategies for methicillin-sensitive S. aureus (MSSA) and MRSA infections. However, the similarities between MRSA and MSSA properties present a challenge in promptly and accurately distinguishing between them. This work introduces an approach to differentiate MRSA from MSSA utilizing matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) in conjunction with a neural network-based classification model. Four distinct strains of S. aureus were utilized, comprising three MSSA strains and one MRSA strain. The classification accuracy of our model ranges from ~ 92 to ~ 97% for each strain. We used deep SHapley Additive exPlanations to reveal the unique feature peaks for each bacterial strain. Furthermore, Fe3O4 MNPs were used as affinity probes for sample enrichment to eliminate the overnight culture and reduce the time in sample preparation. The limit of detection of the MNP-based affinity approach toward S. aureus combined with our machine learning strategy was as low as ~ 8 × 103 CFU mL−1. The feasibility of using the current approach for the identification of S. aureus in juice samples was also demonstrated.Graphical

Prevalence of Panton-Valentine leucocidin (pvl) and exfoliative toxin A (eta) gene within methicillin resistant and sensitive Staphylococcus aureus in an urban tertiary referral hospital: A molecular epidemiology pilot study

Background: Staphylococcus aureus is well known to cause a multitude of clinical manifestations, from mild to severe bloodstream infections that could lead to death. Infections are common, either in community-acquired or hospital-acquired settings, and treatment remains a challenge due to methicillin-resistant Staphylococcus aureus (MRSA). The pathogenesis of S. aureus is mediated by several cell-surface and secreted virulence factors. The virulence factors discussed in this study are Panton-Valentine leucocidin (pvl) and exfoliative toxin A (eta). Our pilot study aimed to observe pvl and eta as virulence gene prevalence in a North Sumatera tertiary referral health center. Methods: Our study was a descriptive-analytical observational study with a cross-sectional design in which we collected isolates over a single time period. The frequency of genes is reported as a percentage comparison between MRSA and methicillin-sensitive S. aureus (MSSA). Qualitative gene prevalence analysis was carried out using the polymerase chain reaction (PCR). Results: Our results showed that from 38 MRSA sample isolates, six samples were found to be pvl-negative, or 15.7% of the total samples. From 40 MSSA sample isolates, one sample was found to be pvl-negative MSSA, or 0.025%. Regarding eta, from 38 MRSA sample isolates, 18.4% of the total sample did not have eta, while from 40 MSSA sample isolates, all samples were found to be positive for eta. We found that both pvl and eta were significantly more likely to be expressed in the MSSA strain. Conclusions: Our study shows that pvl and eta are more likely expressed in MSSA strains than in MRSA strains in Indonesia.

Anti-Biofilm Effects of Torilis japonica Ethanol Extracts against Staphylococcus aureus.

The spread of antibiotic-resistant strains of Staphylococcus aureus, a gram-positive opportunistic pathogen, has increased due to the frequent use of antibiotics. Inhibition of the quorum-sensing systems of biofilm-producing strains using plant extracts represents an efficient approach for controlling infections. Torilis japonica is a medicinal herb showing various bioactivities; however, no studies have reported the anti-biofilm effects of T. japonica extracts against drug-resistant S. aureus. In this study, we evaluated the inhibitory effects of T. japonica ethanol extract (TJE) on biofilm production in methicillin-sensitive S. aureus (MSSA) KCTC 1927, methicillin-resistant S. aureus (MRSA) KCCM 40510, and MRSA KCCM 40511. Biofilm assays showed that TJE could inhibit biofilm formation in all strains. Furthermore, the hemolysis of sheep blood was found to be reduced when the strains were treated with TJE. The mRNA expression of agrA, sarA, icaA, hla, and RNAIII was evaluated using reverse transcription-polymerase chain reaction to determine the effect of TJE on the regulation of genes encoding quorum sensing-related virulence factors in MSSA and MRSA. The expression of hla reduced in a concentration-dependent manner upon treatment with TJE. Moreover, the expression levels of other genes were significantly reduced compared to those in the control group. In conclusion, TJE can suppress biofilm formation and virulence factor-related gene expression in MSSA and MRSA strains. The extract may therefore be used to develop treatments for infections caused by antibiotic-resistant S. aureus.

MoWa: A Disinfectant for Hospital Surfaces Contaminated With Methicillin-Resistant Staphylococcus aureus (MRSA) and Other Nosocomial Pathogens.

Introduction Staphylococcus aureus strains, including methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA), are a main cause of nosocomial infection in the world. The majority of nosocomial S. aureus-infection are traced back to a source of contaminated surfaces including surgery tables. We assessed the efficacy of a mixture of levulinic acid (LA) and sodium dodecyl sulfate (SDS), hereafter called MoWa, to eradicate nosocomial pathogens from contaminated surfaces.Methods and ResultsA dose response study demonstrated that MoWa killed 24 h planktonic cultures of S. aureus strains starting at a concentration of (LA) 8.2/(SDS) 0.3 mM while 24 h preformed biofilms were eradicated with 32/1.3 mM. A time course study further showed that attached MRSA bacteria were eradicated within 4 h of incubation with 65/2 mM MoWa. Staphylococci were killed as confirmed by bacterial counts, and fluorescence micrographs that were stained with the live/dead bacterial assay. We then simulated contamination of hospital surfaces by inoculating bacteria on a surface prone to contamination. Once dried, contaminated surfaces were sprayed with MoWa or mock-treated, and treated contaminated surfaces were swabbed and bacteria counted. While bacteria in the mock-treated samples grew at a density of ~104 cfu/cm2, those treated for ~1 min with MoWa (1.0/0.04 M) had been eradicated below limit of detection. A similar eradication efficacy was obtained when surfaces were contaminated with other nosocomial pathogens, such as Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, or Staphylococcus epidermidis.ConclusionsMoWa kills planktonic and biofilms made by MRSA and MSSA strains and showed great efficacy to disinfect MRSA-, and MSSA-contaminated, surfaces and surfaces contaminated with other important nosocomial pathogens.

Molecular Characterization of Staphylococcus aureus Obtained from Blood Cultures of Paediatric Patients Treated in a Tertiary Care Hospital in Mexico.

PurposeStaphylococcus aureus is one of the main causative agents of hospital-acquired (HA) infections. In Mexico, information about the characteristics of clinical S. aureus isolates is limited. Our aim was to characterize S. aureus strains obtained from blood cultures of paediatric patients treated in a tertiary care hospital.Materials and MethodsWe analysed 249 S. aureus isolates over the period from 2006 to 2019, and their resistance profiles were determined. The isolates were classified into methicillin-resistant S. aureus (MRSA) or methicillin-sensitive S. aureus (MSSA). Staphylococcal cassettes chromosome mec (SCCmec) were detected. Virulence genes (cna, clfA, clfB, eta, etb, fnbA, fnbB, hla, pvl, sec, and tsst) were amplified, and their clonal relationships were established by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and clonal complex (CC) typing. We reviewed one hundred medical files to collect clinical information.ResultsThirty-eight percent of the isolates were MRSA and showed an expanded profile of resistance to other non-beta-lactam antibiotics, while MSSA strains presented a reduced resistance profile. SCCmec-II was the most frequent element (86.3%). Eight virulence factors were detected in MSSA and six in MRSA. The pvl gene was detected in four MRSA-SCCmec-IV isolates (P≤0.0001). MRSA isolates were distributed among 14 clones and were classified into 15 sequence types (ST); the most frequent was ST1011 (17%). The most common CC in MRSA was CC5 (69%, P≤0.0001), and in MSSA, it was CC30 (30%, P≤0.0001). Eighty-seven percent of MRSA isolates were HA-MRSA, and 13% were community-acquired MRSA (CA-MRSA). Of 21 HA-MRSA isolates, 17 had SCCmec-II, while two CA-MRSA isolates had SCCmec-IV. Of MSSA isolates, 77% were derived from HA infections and 23% from CA infections.ConclusionMSSA isolates had more virulence factors. MRSA isolates were resistant to more non-beta-lactam antibiotics, and those with SCCmec-IV expressed a greater variety of virulence factors. Most S. aureus isolates belonged to CC5.

The Role of Subinhibitory Concentrations of Daptomycin and Tigecycline in Modulating Virulence in Staphylococcus aureus.

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.

Control of Methicillin-Resistant Staphylococcus aureus Strains Associated With a Hospital Outbreak Involving Contamination From Anesthesia Equipment Using UV-C.

Methicillin-resistant Staphylococcus aureus (MRSA) is considered an opportunistic pathogen in humans and is mainly associated with healthcare-associated infections (HCAIs). This bacterium colonizes the skin and mucous membranes of healthy people and causes frequent hospital outbreaks. The aim of this study was to perform molecular typing of the staphylococcal cassette chromosome mec (SCCmec) and agr loci as wells as to establish the pulsotypes and clonal complexes (CCs) for MRSA and methicillin-sensitive S. aureus (MSSA) outbreaks associated with the operating room (OR) at a pediatric hospital. Twenty-five clinical strains of S. aureus (19 MRSA and 6 MSSA strains) were recovered from the outbreak (patients, anesthesia equipment, and nasopharyngeal exudates from external service anesthesia technicians). These clinical S. aureus strains were mainly resistant to benzylpenicillin (100%) and erythromycin (84%) and were susceptible to vancomycin and nitrofurantoin. The SCCmec type II was amplified in 84% of the S. aureus strains, and the most frequent type of the agr locus was agrII, which was amplified in 72% of the strains; however, the agrI and agrIII genes were mainly detected in MSSA strains. A pulsed-field gel electrophoresis (PFGE) analysis grouped the 25 strains into 16 pulsotypes (P), the most frequent of which was P1, including 10 MRSA strains related to the anesthesia equipment, external service anesthesia technicians, and hospitalized patients. Multilocus sequence typing (MLST) identified 15 sequence types (STs) distributed in nine CCs. The most prevalent ST was ST1011, belonging to CC5, which was associated with the SCCmec type II and agrII type. We postulate that the external service anesthesia technicians were MRSA carriers and that these strains were indirectly transmitted from the contaminated anesthesia equipment that was inappropriately disinfected. Finally, the MRSA outbreak was controlled when the anesthesia equipment disinfection was improved and hand hygiene was reinforced.

Rapid identification and discrimination of methicillin‐resistant Staphylococcus aureus strains via matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry

Methicillin-resistant Staphylococcus aureus (MRSA) is one of major clinical pathogens responsible for both hospital- and community-acquired infections worldwide. A delay in targeted antibiotic treatment contributes to longer hospitalization stay, higher costs, and increasing in-hospital mortality. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been integrated into the routine workflow for microbial identification over the past decade, and it has also shown promising functions in the detection of bacterial resistance. Therefore, we describe a rapid MALDI-TOF MS-based methodology for MRSA screening with machine-learning algorithms. A total of 452 clinical S. aureus isolates were included in this study, of which 194 were MRSA and 258 were methicillin-sensitive S. aureus (MSSA). The mass-to-charge ratio (m/z) features from MRSA and MSSA strains were binned and selected through Lasso regression. These features were then used to train a non-linear support vector machine (SVM) with radial basis function (RBF) kernels to evaluate the discrimination performance. The classifiers' accuracy, sensitivity, specificity, and the area under the receiver operating characteristic (ROC) curve (AUC) were evaluated and compared with those from the random forest (RF) model. A total of 2601 unique spectral peaks of all isolates were identified and 38 m/z features were selected for the classifying model. The AUCs of the non-linear RBF-SVM model and the RF model were 0.89 and 0.87, respectively, and the accuracy ranged between 0.86 (RBF-SVM) and 0.82 (RF). Our study demonstrates that MALDI-TOF MS coupled with machine-learning algorithms could be used to develop a rapid and easy-to-use method to discriminate MRSA from MSSA. Considering that this method is easy to implement in routine microbiology laboratories, it suggests a cost-effective and time-efficient alternative to conventional resistance detection in the future to improve clinical treatment.

<p>Efflux pumps and biofilm formation by both methicillin-resistant and methicillin-sensitive <em>Staphylococcus aureus</em> strains</p>

Staphylococcus aureus is the primary cause of nosocomial infections. It produces potent toxins and causes superficial lesions, systemic diseases, and several toxaemic syndromes. In this study, we determined the role of efflux pump in conferring resistance to fluoroquinolones in the biofilm of methicillin resistant Staphylococcus aureus (MRSA) and methicillin sensitive S. aureus (MSSA) strains. We selected five strains of S. aureus comprising three methicillin sensitive strains, including ATCC25923, and two strains of methicillin resistant S . aureus . Thioridazine, chlorpromazine, carbonyl cyanide 3-chloro-phenylhydrazone and reserpine showed activity against MSSA strains. Whereas, thioridazine and chlorpromazine were active against MRSAstrains too. Thioridazine and chlorpromazine showed a significant decrease in biofilm formation of MSSA strain 2493, and MRSA strain UC1079 compared to the control strain ATCC25923. Thioridazine showed a similar effect on MSSA strain N297214 and MRSA strain N307002. Four to eight-fold increase was found in the expression of efflux pump genes with ethidium bromide, ciprofloxacin, and moxifloxacin in these strains. However, a decrease in the expression of norB , norC , abcA and mepA was observed in MSSA, and MRSA strains with thioridazine, chlorpromazine and naringenin. This study, thus demonstrate that the major facilitator superfamily, multidrug and toxin compound extrusion, and ATP binding cassette family of efflux pumps play an essential role in developing antibiotic resistance and biofilm formation. It is the first report on existence of efflux pump genes in biofilm beside antibiotic resistance for fluoroquinolones in MRSA and MSSA strains, which need further characterization

Frequency of Vectors of Methicillin-resistant Staphylococcus Aureus Among Emergency Staff of Vali-e-Asr Hospital in Arak City, 2018

Background and Aim: Staphylococcus aureus is common pathogens of nosocomial infections. Nasal swabs in hospital staff is main sources of hospital infections are considered. The aim of this study was to determine the frequency of nasopharyngeal carriers of methicillin-resistant Staphylococcus aureus and microbial contamination of health care workers' cell phones in Emergency department of Vali-e-Asr Hospital in Arak City. Methods & Materials: In this descriptive study, nose swabs and cell phone levels were taken from 70 health care workers in the emergency ward of Vali-e-Asr Hospital. The Staphylococcus aureus clinical isolates were identified using standard microbiological methods (catalase, coagulase, mannitol fermentation and DNase). The susceptibility to oxacillin and cefoxitin was detected by the disk diffusion and the mecA genes in this clinically isolated strain of staphylococci was investigated by Polymerase Chain Reaction (PCR). Ethical Considerations: This study was approved by the Research Ethics committee of Arak University of Medical Sciences. (Code : IR.ARAKMU.REC.1396.282). Results: According the results, Staphylococcus aureus was isolated in 16 cases, which 5 cases were methicillin-sensitive S. aureus (MSSA), and 11 cases were Methicillin resistant S. aureus (MRSA). Also, 3 cell phones were infected with Staphylococcus aureus, which 1 case was MRSA and 2 cases were MSSA. Conclusion: The results of this study showed that frequency of MSSA strains is significant in emergency personnel of the hospital. Thus, regarding to the risk of epidemics due to nosocomial infections, periodic testing for the identification and treatment of carriers among employees for controlling and preventing hospital infections seems necessary.

Correlation Between Biofilm Formation and Antibiotic Resistance in MRSA and MSSA Isolated from Clinical Samples in Iran: A Systematic Review and Meta-Analysis.

Objectives: This study aimed at reviewing the correlation between biofilm formation and antibiotic resistance in methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) isolates. Materials and Methods: This review followed Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) protocols. The literature search was conducted in PubMed, Web of Science (ISI), and Scopus databases. Combinations of Mesh terms such as "biofilms" OR "biofilm formation," AND "Drug Resistance" OR "Antimicrobial Drug Resistance" OR "Antibiotic Resistance" AND "Staphylococcus aureus" OR "Methicillin-resistant Staphylococcus aureus" or "MRSA" AND "Methicillin-sensitive Staphylococcus aureus" OR "MSSA" AND "biofilm-related genes" AND "Prevalence" AND "Iran" were searched. Two reviewers independently searched the databases. Analyses were performed in Comprehensive Meta-Analysis software. The random-effects model was used to obtain the combined prevalence with a 95% confidence interval (CI). Results: The combined prevalence of MRSA retrieved from Iranian clinical samples was 48.3% (95% CI: 40.8-55.9). The pooled rate of biofilm formation in MRSA strains was reported as 80.9% (95% CI: 67.8-89.4). Overall, 52.9%, 45.3%, and 22.5% of MRSA isolates were strong, moderate, and weak biofilm producers, respectively. The highest frequency of biofilm-related genes was observed for icaD gene (67.7%) followed by clfA gene with a frequency of 64.7%. Among seven studies that addressed the relationship between biofilm formation and antibiotic resistance, six reported positive associations. Conclusions: Regarding the MRSA strains, they had a significantly higher ability of biofilm formation than MSSA strains; therefore, preventive measures against infections caused by them are required.

In Vitro Activity Of Ceftaroline And Comparators Against Staphylococcus aureus Isolates: Results From 6 Years Of The ATLAS Program (2012 To 2017).

BackgroundCeftaroline is effective against methicillin-resistant Staphylococcus aureus (MRSA), but the resistance patterns still need to be defined. This study aimed to investigate the susceptibility of S. aureus to ceftaroline and comparator antimicrobial agents in patients hospitalized due to infection and to observe the patterns among different regions and over the years.MethodsThe Antimicrobial Testing Leadership And Surveillance (ATLAS) program includes medical centers located in five geographical regions (Europe, Asia-Pacific, South America, Africa-West Asia, and the United States). The isolates were collected from different specimens from patients hospitalized between 2012 and 2017 due to documented complicated skin and soft tissue infection, complicated intra-abdominal infection, complicated urinary tract infection, lower respiratory tract infection, and bloodstream infection.ResultsDuring the study period, 61,045 isolates were tested, including 35,837 MRSA isolates (58.7%) and 25,208 methicillin-sensitive S. aureus (MSSA) isolates (41.3%). For MRSA, the minimal inhibitory concentration (MIC)50, MIC90, and MIC range of ceftaroline were 0.5, 2, and 0.015–64 μg/mL. The proportion of susceptible MRSA strains was 89.3%. The proportion of resistant MRSA strains was 0.7%. The susceptibility of all S. aureus, MRSA, and MSSA strains to ceftaroline remained relatively constant from 2012 to 2017. The susceptibility to ceftaroline of S. aureus, MRSA, and MSSA strains from the United States, Europe, South America, and Africa/West Asia was high, while the susceptibility of the strains from Asia-Pacific was lower, especially for MRSA.ConclusionThis study reveals the patterns of ceftaroline susceptibility of MRSA and MSSA around the world and over 6 years.

Staphylococcus aureus biofilm organization modulated by YycFG two-component regulatory pathway

BackgroundStaphylococcus aureus (S. aureus) infection accounts for more than 50% of the osteomyelitis cases. Currently, methicillin-resistant S. aureus (MRSA) strains present an urgent medical problem. The YycFG two-component regulatory system (TCS) can allow bacteria to rapidly adapt to physical, chemical, and biological stresses. To define the role of YycFG in modulation virulence of S. aureus in osteomyelitis, we isolated clinical MRSA strains and compared these with ATCC29213 methicillin-sensitive S. aureus (MSSA).MethodsIn the present study, 13 MRSA strains from chronic osteomyelitis tissues were isolated. The in-depth sequencing of 16S rRNA amplicons of the samples was conducted. Bacterial growth was monitored, and biofilm biomass was determined by crystal violet microtiter assay. Furthermore, quantitative RT-PCR analysis was adopted to identify the expression of yycF/G/H and icaA/D in MRSA and MSSA strains. Analysis of variance with one-way ANOVA was used for statistical analysis.ResultsThe in-depth sequencing of 16S rRNA amplicons of the clinical samples indicated a polymicrobial infection, with the phylum Firmicutes made up 13% of the microbial population. The MRSA strains showed an accelerated growth rate compared to the MSSA strains. Of note, MRSA biofilms showed an accumulation of an intercellular polysaccharides matrix and enhanced biomass upon microscopic examination. Furthermore, MRSA strains had a higher expression of the yycF/G/H and icaA/D genes and adhesion force.ConclusionsThese data suggested the roles of intercellular polysaccharide in S. aureus pathogenesis, indicating a possible association between YycFG pathways and MRSA strain virulence.

Impact of mass migrations on the clonal variation of clinical Staphylococcus aureus strains isolated from the Western region of Saudi Arabia

ObjectivesA rapid molecular typing system was used to determine the impact of mass migration on the clonal variation of Staphylococcus aureus isolates recovered from King Abdulaziz University Hospital (KAUH) Jeddah, in the western region of Saudi Arabia. This region experiences an annual influx of millions of pilgrims. MethodsSmaI-multiplex PCR typing (SMT) was used for the initial analysis of strains and the resulting data subsequently supported by Multi-Locus Sequence Typing (MLST). ResultsA total of 89 S. aureus isolates were SMT typed and revealed a high degree of genetic variation, with 40 SMT profiles detected among the isolates. Representatives of all forty SMT types were subsequently analysed by MLST, identifying 26 sequence types. A novel sequence type (ST), named ST3303, was identified in two methicillin-sensitive S. aureus (MSSA) isolates. MSSA strains exhibited more diversity than methicillin-resistant S. aureus (MRSA) strains, with community acquired MSSA and MRSA strains reaching alarmingly high levels. ConclusionThe relatively high degree of genetic diversity found among S. aureus isolates of single hospital was attributed to the fact that Jeddah is the principal gateway to Mecca, visited each year by millions of pilgrims from many countries. The observed diversity clearly reflects the impact of such mass migrations in the rapid dissemination of strains world-wide. Our findings suggest the importance of surveillance programmes in locations affected by mass migrations, both to monitor their impact on endemic strains and for the detection of pandemic strains. SMT provides a cost-effective and sensitive typing method for achieving this objective.

Association Between the Methicillin Resistance of Staphylococcus aureus Isolated from Slaughter Poultry, Their Toxin Gene Profiles and Prophage Patterns

In this work, 85 strains of Staphylococcus aureus were isolated from samples taken from slaughter poultry in Poland. Attempts were made to determine the prophage profile of the strains and to investigate the presence in their genome of genes responsible for the production of five classical enterotoxins (A–E), toxic shock syndrome toxin (TSST-1), exfoliative toxins (ETA and ETB) and staphylokinase (SAK). For this purpose, multiplex PCR was performed using primer-specific pairs for targeted genes. The presence of the mecA gene was found in 26 strains (30.6%). The genomes of one of the methicillin-resistant S. aureus (MRSA) strains and two methicillin-sensitive S. aureus (MSSA) strains contained the gene responsible for the production of enterotoxin A. Only one MRSA strain and two MSSA strains showed the presence of the toxic shock syndrome toxin (tst) gene. Only one of the MSSA strains had the gene (eta) responsible for the production of exfoliative toxins A. The presence of the staphylokinase gene (sak) was confirmed in 13 MRSA strains and in 5 MSSA strains. The study results indicated a high prevalence of prophages among the test isolates of Staphylococcus aureus. In all, 15 prophage patterns were observed among the isolates. The presence of 77-like prophages incorporated into bacterial genome was especially often demonstrated. Various authors emphasize the special role of these prophages in the spread of virulence factors (staphylokinase, enterotoxin A) not only within strains of the same species but also between species and even types of bacteria.

The Relationship Between Prevalence of Antibiotics Resistance and Virulence Factors Genes of MRSA and MSSA Strains Isolated from Clinical Samples, West Iran.

We sought to evaluate the relationship between the prevalence of antibiotics resistance and virulence factors genes in methicillin-resistant Staphylococcal aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) strains from clinical samples taken in west Iran. We performed a cross-sectional study using 100 MRSA and 100 MSSA samples isolated from clinical specimens. We used biochemical methods to identify the isolates, which were confirmed by the polymerase chain reaction (PCR) assay. Antibiotic susceptibility testing was performed using disk diffusion. PCR detected the presence of virulence factors, including enterotoxin genes, toxic shock syndrome toxin-1 (TSST-1), and exfoliative toxin. The majority of MRSA isolates exhibited a high level of resistance to common antibiotics and susceptible to vancomycin, while most MSSA isolates were also resistant to erythromycin and ciprofloxacin. The prevalence of Staphylococcal enterotoxins (SEs) were reported 147 (73.5%). Among 100 MRSA samples, 92 (92.0%) harbored SAg genes. The most frequent toxin gene was sea (45.0%) followed by sec (39.0%). Among 100 MSSA isolates, 89 (89.0%) harbored SAg genes and the most prevalent genes were sea (42.0%), sek (38.0%), sec (35.0%,), and TSST-1 (10.0%). The prevalence of TSST-1 and exfoliative toxin genes in MRSA samples were 12 (12.0%). The association of SAg genes with MRSA and MSSA isolates showed a high prevalence of enterotoxin seq, seg, and sei in MRSA than MSSA with a statistically significant difference (p < 0.050). The prevalence of MRSA and the association of pathogenic agents with antibiotics resistance genes can lead to the emergence of strains with higher pathogenicity and less susceptibility.

Antibiofilm activity of α-mangostin extracted from Garcinia mangostana L. against Staphylococcus aureus.

To isolate α-mangostin (AMG) from the peels of mangosteen (Garcinia mangostana L.), grown in Vietnam, and to investigate antibiofilm activity of this compound against three Staphylococcus aureus (S.aureus) strains, one of which was methicillin-resistant S.aureus (MRSA) and the other two strains were methicillin-sensitive S.aureus (MSSA). AMG in n-hexane fraction was isolated on a silica gel column and chemically analyzed by HPLC and NMR. The antibiofilm activity of this compound was investigated by using a 96-well plate model for the formation of biofilms. Biofilm biomass was quantified using crystal violet. The viability of cells was observed under confocal microscopy using LIVE/DEAD BacLight stains. Biofilm composition was determined using specific chemical and enzyme tests for polysaccharide, protein and DNA. Membrane-damaging activity was assayed by measuring the hemolysis of human red blood cells in presence of AMG. The results indicated that the isolated AMG, with a purity that exceeded 98%, had minimal inhibitory concentrations in the range of 4.6-9.2μmol/L for the three strains tested. Interestingly, the MSSA strains were more sensitive to AMG than the MRSA strain. Minimal bactericidal concentrations were 2-fold higher than the minimal inhibitory concentration values for the three strains, indicating that AMG was a bactericidal compound. AMG also prevented biofilm formation effectively, albeit that again the MRSA strain was the most resistant. Interestingly, biofilms of the MRSA strain contained protein as a main component of the extracellular matrix, whereas this was polysaccharide in the MSSA strains. This might relate to the resistance of the MRSA 252 strain to AMG. Assays using human red blood cells indicated that AMG caused significant membrane damage with 50% of cell lysis occurred at concentration of about 36μmol/L. Our results provide evidence that the isolated AMG has inhibitory activity against biofilm formation by S.aureus, including MRSA. Thus, isolated AMG proposes a high potential to develop a novel phytopharmaceutical for the treatment of MRSA.

Insights into chitosan antibiofilm activity against methicillin-resistant Staphylococcus aureus.

Chitosan is a natural compound that has been validated as a viable antimicrobial agent against Staphylococcus aureus. With this work we sought to evaluate the planktonic and sessile sensitivity of methicillin-resistant S. aureus to chitosan's activity and evaluate if methicillin-resistant S. aureus (MRSA) would be more or less sensitive to chitosan's activity than methicillin-sensitive S. aureus (MSSA). A group comprised of reference strains and clinical multiresistant isolates of MSSA and MRSA were used. Methicilin resistance effect upon chitosan activity was assessed in planktonic setting and in different phases of sessile colonization, namely adhesion, biofilm formation and mature biofilm through biomass and metabolism inhibition. The results obtained showed that S. aureus methicillin resistance mechanism did not impair chitosan's activity as the highest bacterial susceptibility was registered for MRSA. Chitosan was highly effective in inhibiting MSSA and MRSA strains in both planktonic and sessile settings with biofilm inhibition percentages reaching as high as 90% for MRSA. Staphylococcus aureus methicillin resistance did not impair chitosan's antimicrobial and antibiofilm activities and MRSA and MSSA were inhibited both in planktonic and sessile settings at low concentrations with great efficacy. Considering the obtained results chitosan shows potential as an alternative for the control of biofilm-related recalcitrant MRSA infections.

Investigation of biofilm formation properties of staphylococcus isolates

Biofilm production is an important virulence factor which allows staphylococci to adhere to medical devices. The principal component of biofilm is a "polysaccharide intercellular adhesin (PIA)" which is composed of a beta-1,6-N-acetylglucosamine polymer synthesized by an enzyme (N-acetylglucosamine transferase) encoded by the ica operon found on the bacterial chromosome. This operon is composed of four genes (A, B, C, and D), and a transposable element IS256. In this study, we aimed to determine the biofilm production characteristics of invasive/non-invasive staphylococcus isolates and different staphylococcus species. Biofilm production of 166 staphylococci was phenotypically investigated on Congo Red Agar (CRA); the presence of icaA, icaD and IS256 genes were investigated by polymerase chain reaction (PCR). 74 of the isolates (44.6%) were identified as methicillin resistant Staphylococcus aureus (MRSA), 25 (15.1%) as methicillin sensitive S.aureus (MSSA), 25 (37.3%) as Staphylococcus hominis, 20 (12%) as S.epidermidis, ten (15%) as Staphylococcus haemolyticus, nine (13.4%) as Staphylococcus capitis, two (3%) Staphylococcus saprophyticus and one (1.5%) as Staphylococcus warnerii. Of the MRSA strains, 52 were isolated from blood and 22 from nose; all MSSA strains were isolated from nose cultures. Coagulase-negative staphylococci (CoNS) strains were composed of invasive and non-invasive strains isolated from nose, catheter tip and blood cultures from patients with catheter. Production with CRA method was found to be statistically significant in invasive isolates (p< 0.001). It is concluded that; as the biofilm formation capacity of invasive isolates can cause refractory infections and the importance of carriage and hospital infections of these bacteria, it is important to prevent the spread of these isolates. A combination of phenotypic and genotypic tests is recommended for the investigation of biofilm formation in staphylococci. 40.3% of the CoNS isolates, and 85.8% of S.aureus isolates produced biofilm on CRA (p< 0.001) and with PCR method the ratio of carrying three genes was found to be statistically important in S.aureus when compared with CoNS. Carriage of three genes and biofilm formation capacity of invasive isolates can cause refractory infections and the importance of carriage and hospital infections of these bacteria, it is important to prevent the spread of these isolates. A combination of phenotypic and genotypic tests is recommended for the investigation of biofilm formation in staphylococci.

Antimicrobial Resistance and Molecular Characteristics of Nasal Staphylococcus aureus Isolates From Newly Admitted Inpatients.

Staphylococcus aureus, or methicillin-resistant S. aureus (MRSA), is a significant pathogen in both nosocomial and community infections. Community-associated MRSA (CA-MRSA) strains tend to be multi-drug resistant and to invade hospital settings. This study aimed to assess the antimicrobial resistance and molecular characteristicsof nasal S. aureus among newlyadmitted inpatients.In the present study, 66 S. aureus isolates, including 10 healthcare-associated MRSA (HA-MRSA), 8 CA-MRSA, and 48 methicillin-sensitive S. aureus (MSSA) strains, were found in the nasal cavities of 62 patients by screening 292 newlyadmitted patients. Antimicrobial resistance and molecular characteristics of these isolates, including spa-type, sequence type (ST) and SCCmec type, were investigated. All isolates were sensitive to linezolid, teicoplanin, and quinupristin/dalfopristin, but high levels of resistance to penicillin and erythromycin were detected. According to D-test and erm gene detection results, the cMLSB and iMLSB phenotypes were detected in 24 and 16 isolates, respectively. All 10 HA-MRSA strains displayed the cMLSB phenotypemediated by ermA or ermA/ermC, while the cMLSB CA-MRSA and MSSA strains carried the ermB gene. Molecular characterization revealedall 10 HA-MRSA strains were derived from the ST239-SCCmec III clone, and four out of eight CA-MRSA strains were t437-ST59-SCCmec V. The results suggest that patients play an indispensable role in transmitting epidemic CA-MRSA and HA-MRSA strains.