Methicillin-resistant Strains Of Staphylococcus Aureus Research Articles

Mucus Mimic Hydrogel Coating for Lubricous, Antibiofouling, and Anti-Inflammatory Urinary Catheters.

Silicone is a common elastomer used in indwelling urinary catheters, and catheters are widely used in various medical applications due to their exceptional biocompatibility, hypoallergenic properties, and flexibility. However, silicones exhibit hydrophobic characteristics, lack inherent biolubrication, and are susceptible to nonspecific biosubstance adsorption, resulting in complications including but not limited to tissue trauma, postoperative pain, and urinary tract infections (UTIs). The development of effective surface designs for biomedical catheters to mitigate invasive damage and UITs has been a longstanding challenge. Herein, we present a novel approach to prepare a mucus mimic hydrogel coating. A thin layer of hydrogel containing xylitol is fabricated via photopolymerization. The surface modification technique and the interface-initiated hydrogel polymerization method ensure robust interfacial coherence. The resultant coating exhibits a low friction coefficient (CoF ≈ 0.1) for urinary catheter applications. Benefiting from the hydration layer and the antifouling of the xylitol unit, the xylitol hydrogel-coated surfaces (pAAAMXA) demonstrate outstanding antibiofouling properties against proteins (98.9% reduction relative to pristine polydimethylsiloxane (PDMS)). Furthermore, the pAAAMXA shows general adhesion resistance against bacteria primarily responsible for UITs (Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), Methicillin-resistant strains of Staphylococcus aureus (MRSA), and Staphylococcus epidermidis (S. epidermidis)) without compromising biotoxicity (cell viability 98%). In vivo, catheters coated with the mucus mimic hydrogel displayed excellent biocompatibility, resistance to adhesion of bio substance, and anti-inflammatory characteristics. This work describes a promising alternative to conventional silicone catheters, offering potential for clinical interventional procedures with minimized complications.

Investigating the Antimicrobial Activity and Pigmentation of the Extract of Rhazya stricta Plant Against Human Bacteria

Background: Due to the increasing resistance of pathogenic bacteria to common antibiotics, researchers are looking for antimicrobial agents of plant origin as alternative drugs. Objectives: This study aimed to investigate the antimicrobial activity and pigmentation of the extract of the Rhazya stricta plant against human bacteria. Methods: The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) in six concentrations were evaluated by the dilution method in a liquid medium on the determined bacteria. The pink color of the plant extract was assessed using ELISA. Results: The results of the diameter of the inhibition zone showed that the maximum diameter of the inhibition zone for ethyl acetate (2 mm), aqueous (7 mm), ethanolic (8 mm), methanolic (9 mm), and hydroalcoholic (10 mm) extracts was against Staphylococcus aureus. The smallest diameter of the inhibition zone was related to the ethyl acetate extract of R.stricta against methicillin-resistant strains of Staphylococcus aureus (MRSA) (2 mm), while the largest diameter of the inhibition zone was related to the methanolic extract of R.stricta (8 mm). Conclusions: The results of the study showed that different medicinal plant solvents inhibit the growth of pathogenic bacteria at different concentrations, which can be used in the treatment of infections caused by these bacteria. Additionally, this medicinal plant can be used in the dyeing industry due to its pigmentation properties.

Elevated acetate kinase (ackA) gene expression, activity, and biofilm formation observed in methicillin-resistant strains of Staphylococcus aureus (MRSA)

BackgroundStaphylococcus aureus spreads its infections through biofilms. This usually happens in the stationary phase of S. aureus growth where it utilizes accumulated acetate as a carbon source via the phosphotrans-acetylase-acetate kinase (Pta-Ack) pathway. In which acetate kinase (ackA) catalyzes the substrate-level phosphorylation, a vital secondary energy-yielding pathway that promotes biofilms formation aids bacterium survival in hostile environments. In this study, we describe the cloning, sequencing, and expression of S. aureus ackA gene. The expression analysis of ackA gene in methicillin-resistant strains of S. aureus (MRSA) correlates with ackA activity and biofilm units. The uniqueness of ackA was analyzed by using in silico methods. ResultsElevated ackA gene expression was observed in MRSA strains, which correlates with increased ackA activity and biofilm units, explaining ackA role in MRSA growth and pathogenicity. The pure recombinant acetate kinase showed a molecular weight of 44 kDa, with enzyme activity of 3.35 ± 0.05 μM/ml/min. The presence of ACKA-1, ACKA-2 sites, one ATP, and five serine/threonine-protein kinase sites in the ackA gene (KC954623.1) indicated that acetyl phosphate production is strongly controlled. The comparative structural analysis of S. aureus ackA with ackA structures of Mycobacterium avium (3P4I) and Salmonella typhimurium (3SLC) exhibited variations as indicated by the RMSD values 1.877 Å and 2.141 Å respectively, explaining why ackA functions are differently placed in bacteria, concurring its involvement in S. aureus pathogenesis. ConclusionsOverall findings of this study highlight the correlation of ackA expression profoundly increases survival capacity through biofilm formation, which is a pathogenic factor in MRSA and plays a pivotal role in infection spreading.

A comparison of passive and active dust sampling methods for measuring airborne methicillin-resistant Staphylococcus aureus in pig farms.

Methicillin-resistant strains of Staphylococcus aureus (MRSA) are resistant to most β-lactam antibiotics. Pigs are an important reservoir of livestock-associated MRSA (LA-MRSA), which is genetically distinct from both hospital and community-acquired MRSA. Occupational exposure to pigs on farms can lead to LA-MRSA carriage by workers. There is a growing body of research on MRSA found in the farm environment, the airborne route of transmission, and its implication on human health. This study aims to directly compare two sampling methods used to measure airborne MRSA in the farm environment; passive dust sampling with electrostatic dust fall collectors (EDCs), and active inhalable dust sampling using stationary air pumps with Gesamtstaubprobenahme (GSP) sampling heads containing Teflon filters. Paired dust samples using EDCs and GSP samplers, totaling 87 samples, were taken from 7 Dutch pig farms, in multiple compartments housing pigs of varying ages. Total nucleic acids of both types of dust samples were extracted and targets indicating MRSA (femA, nuc, mecA) and total bacterial count (16S rRNA) were quantified using quantitative real-time PCRs. MRSA could be measured from all GSP samples and in 94% of the EDCs, additionally MRSA was present on every farm sampled. There was a strong positive relationship between the paired MRSA levels found in EDCs and those measured on filters (Normalized by 16S rRNA; Pearson's correlation coefficient r = 0.94, Not Normalized; Pearson's correlation coefficient r = 0.84). This study suggests that EDCs can be used as an affordable and easily standardized method for quantifying airborne MRSA levels in the pig farm setting.

Comparative In Vitro Activity of New Lipoglycopeptides and Vancomycin Against Ocular Staphylococci and Their Toxicity on the Human Corneal Epithelium.

The purpose of this study was to assess the potential of new lipoglycopeptides as novel topical therapies for improved treatment of recalcitrant ocular infections. We evaluated the in vitro antimicrobial activity of oritavancin, dalbavancin, and telavancin compared with vancomycin (VAN) against a large collection of ocular staphylococcal isolates and their cytotoxicity on human corneal epithelial cells (HCECs). Antimicrobial susceptibility testing was performed by broth microdilution against 223 Staphylococcus spp. clinical isolates. Time-kill kinetics were determined for methicillin-resistant strains of Staphylococcus aureus (MRSA) (n = 2) and Staphylococcus epidermidis (MRSE) (n = 1). In vitro cytotoxicity assays were performed with AlamarBlue and live/dead staining on HCECs. All new lipoglycopeptides showed strong in vitro potency against ocular staphylococci, including multidrug-resistant MRSA strains, with dalbavancin showing a slightly higher potency overall [minimum inhibitory concentration (MIC) 90 0.06 μg/mL] compared with telavancin and oritavancin (MIC 90 0.12 μg/mL), whereas VAN had the lowest potency (MIC 90 2 μg/mL). Oritavancin exerted rapid bactericidal activity within 1 h for MRSA and 2 h for MRSE. All other drugs were bactericidal within 24 h. At a concentration commonly used for topical preparations (25 mg/mL), cytotoxicity was observed for VAN after 5 min of incubation, whereas reduction in HCEC viability was not seen for telavancin and was less affected by oritavancin and dalbavancin. Cytotoxicity at 25 mg/mL was seen for all drugs at 30 and 60 min but was significantly reduced or undetected for lower concentrations. Our study demonstrates that new lipoglycopeptides have substantially better in vitro antimicrobial activity against ocular staphylococcal isolates compared with VAN, with a similar or improved toxicity profile on HCECs.

The Therapeutic Potential of 4-Methoxy-1-methyl-2-oxopyridine-3-carbamide (MMOXC) Derived from Ricinine on Macrophage Cell Lines Infected with Methicillin-Resistant Strains of Staphylococcus aureus.

The incidences of methicillin-resistant strains of Staphylococcus aureus (MRSA) and their survival inside the macrophages are the major attributes of the relapsed infections after antimicrobial therapy, and it is a global problem. In this context, we have previously demonstrated 4-methoxy-1-methyl-2-oxopyridine-3-carbamide (MMOXC), a Ricinine derivative exhibiting anti-S. aureus and anti-biofilm characteristics by competitively inhibiting uridine monophosphate kinase (UMPK), UDP-N-acetyl muramyl pentapeptide ligase (Mur-F), and peptidyl deformylase, (PDF). In the present study, the stability of this competitive inhibitor MMOXC was evaluated by showing its ability to remain bound to the active sites of UMPK, Mur-F, and PDF even after increasing the incubation time, temperature, pH, and substrate concentration. On growing MRSA in fewer concentrations of MMOXC, these strains could not attain resistance to MMOXC and at the same time distinct reductions in the expression of UMPK, Mur-F, and PDF genes were noted. In vitro, infective models were generated by infecting MRSA to RAW 264.7 and human monocyte-derived macrophage (hMDM) cell lines. In these infected cell lines, in spite of increased nitric oxide synthase (NOS), NADPH-P450 reductase, superoxide dismutase, catalase, and peroxidase activities, the MRSA survived. At 640µM/ml, the concentration of MMOXC penetrated into these infected cells and obliterated MRSA. While treating uninfected macrophage cell lines with MMOXC, no appreciable effect was observed indicating that MMOXC is the most suitable drug for the treatment of infections caused by MRSA.

A new flavonol derivative and other compounds from the leaves of Bauhinia thonningii Schum with activity against multidrug-resistant bacteria

Investigation of the leaves of Bauhinia thonningii Schum led to the isolation and identification of a new flavonol derivative, 6-C-methylquercetin-3,4'-dimethyl ether (1) together with eleven known compounds (2–13), with two of them (10 and 11) obtained as a mixture. Their structures were established by extensive spectroscopic analyses. Antibacterial activity of compound 1 as well as the reference antibiotic, ciprofloxacin was tested on Gram-negative multidrug-resistant bacteria overexpressing active efflux pumps, and against methicillin-resistant strains of Staphylococcus aureus (MRSA). Samples were tested alone and in combination with an efflux pump inhibitor (EPI), phenylalanine-arginine-β-naphthylamide (PAßN). Results show that when compound 1 was tested alone, its inhibitory effects were obtained on 7/10 tested bacteria with the highest MIC value of 128 µg/mL whilst in the presence of EPI, this activity significantly increase in all the 10 bacteria. An interesting antibacterial activity was obtained with compound 1 against Klebsiella pneumoniae ATCC11296 (MIC of 4 µg/mL), KP55 and Staphylococcus aureus MRSA6 in the presence of the PaßN.

Radiology imaging equipment and accessories as possible fomites of nosocomial pathogens.

Radiology is a technical service that provides medical imaging for all sectors of healthcare. Hospital-acquired infections (HAIs) is a major challenge in radiology and this is exacerbated in contexts where the healthcare system is unable to provide adequate funding and attention to effective infection control measures. The objectives of this study were to audit current cleaning procedures through the observation of practices in a radiology department, and to determine the types and numbers of nosocomial pathogens present on selected radiology imaging equipment and accessories before and after decontamination. In phase one we observed seven radiographers to audit cleaning procedures and practices. In phase two we collected swab samples from selected radiology imaging equipment and accessories and then cultured them for identification of microbes. It was observed that radiographers partially practiced infection control measures. This was due to the absence of documented protocol for infection control procedures. Our results indicated that all the selected equipment and accessories were contaminated with microorganisms pre- and post-cleaning. The identified microbes were Staphylococcus aureus, Coagulase negative Staphylococci (CoNS), Bacillus species (spp.), Shigella spp., Shigella sonnei., Klebsiella spp., Salmonella paratyphi A (S. paratyphi A), Salmonella typhi (S. typhi), Providencia rettgeri, Enterobacter spp. and Citrobacter spp. and Methicillin resistant strains of Staphylococcus aureus (MRSA). The research concluded that the recommended cleaning agents did not effectively reduce the number of microorganisms making the selected equipment and accessories fomites for nosocomial pathogens.

Methicillin resistance in Staphylococcus aureus modulates the transcriptome and disease pathology in a murine model of endophthalmitis

The ever-increasing incidence of methicillin-resistant strains of Staphylococcus aureus (MRSA) endophthalmitis is of particular concern as they are associated with poor outcomes. To compare the histology and whole transcriptome of Methicillin resistant (MRSA) and Methicillin-susceptible (MSSA) Staphylococcus aureus endophthalmitis in an experimental murine model. MRSA and MSSA endophthalmitis was induced in C57BL/6 mice and disease progression was scored clinically and histologically at 24 h p.i. Retinal changes were monitored by H&E, CD45, MPO and GFAP staining followed by retinal cell death evaluation. Whole Transcriptome was analysed using the SuperPrint G3 Mouse Gene Expression v2 chip. Differential gene expression analysis (Limma package, R) was done followed by enrichment of pathways (KEGG database). Increased corneal haze, diminished vitreous clarity and red reflex was observed in MRSA infected mice eye compared to MSSA (p = 0.04). Histological assessment also corroborated with increased disease severity in MRSA (p = 0.02). Although MRSA infected eye displayed higher CD45+ cells and greater GFAP intensity, the difference was not statistically significant. However, higher retinal cell death was found to be associated with the MRSA infection (p = 0.007). Our study also revealed that MRSA infection induces changes in host transcriptome (FC = 1.5, p = 0.05), revealing the involvement of several interleukins (IL-11,15,10,1ra), chemokines (CCL-11, CXCL-1), Interferon receptors, GM-CSF, M-CSF, MMPs, Neruopilin2 (NRP-2), Ubiquitin associated peptidase and apoptotic ligands. ErbB signalling, JAK-STAT, adipocytokine and Ras signalling were the top divergently enriched pathways. Our study confirms the differential host immune response triggered by MRSA infection in the eye. Our study may help to elucidate the mechanisms of pathogenesis and to identify additional candidate drug targets for the treatment of MRSA endophthalmitis.

Design, synthesis, antimicrobial evaluations and in silico studies of novel pyrazol-5(4H)-one and 1H-pyrazol-5-ol derivatives

A new series of 1,4-disubstituted 3-methylpyrazol-5(4 H )-one derivatives were synthesized by reacting various substituted aromatic aldehydes with 3-methylpyrazol-5(4 H )-one derivatives through Knoevenagel condensation by conventional as well as by exposure to microwave irradiations. After that newly synthesized compounds of 1,4-disubstituted 3-methyl-1 H -pyrazol-5-ol were prepared from these derivatives by reduction reaction of sodium borohydride at 0–5 °C. Sixty-four heterocyclic compounds containing a pyrazole moiety were synthesized with good to excellent yields (51 to 91%). Compounds ( 3d , 3m , 4a , 4b , 4d , and 4g ) showed potent antibacterial activity against MSSA (Methicillin-susceptible strains of Staphylococcus aureus ) and MRSA (Methicillin-resistant strains of Staphylococcus aureus ) with MIC (the minimum inhibitory concentration) ranging between 4 and 16 µg/mL as compared to ciprofloxacin (MIC = 8–16 µg/mL). Compounds ( 4a , 4h , 4i , and 4l ) showed potent antifungal activity against Aspergillus niger with MIC ranging between 16 and 32 µg/mL as compared to fluconazole (MIC = 128 µg/mL). In particular, compound 4a exhibited the strongest activity among the synthesized compounds in both bacterial and fungal strains with MIC ranging between 4 and 16 µg/mL. Furthermore, the nine most active compounds showed a good ADMET (absorption, distribution, metabolism, excretion, and toxicity) profile in comparison to ciprofloxacin and fluconazole as reference drugs. Molecular docking predicted that DHFR (dihydrofolate reductase) protein from Staphylococcus aureus and NMT ( N -myristoyl transferase) protein from Candida albicans are the most suitable targets for the antimicrobial activities of these potent compounds.

Rotating Magnetic Field Increases β-Lactam Antibiotic Susceptibility of Methicillin-Resistant Staphylococcus aureus Strains.

Methicillin-resistant strains of Staphylococcus aureus (MRSA) have developed resistance to most β-lactam antibiotics and have become a global health issue. In this work, we analyzed the impact of a rotating magnetic field (RMF) of well-defined and strictly controlled characteristics coupled with β-lactam antibiotics against a total of 28 methicillin-resistant and sensitive S. aureus strains. The results indicate that the application of RMF combined with β-lactam antibiotics correlated with favorable changes in growth inhibition zones or in minimal inhibitory concentrations of the antibiotics compared to controls unexposed to RMF. Fluorescence microscopy indicated a drop in the relative number of cells with intact cell walls after exposure to RMF. These findings were additionally supported by the use of SEM and TEM microscopy, which revealed morphological alterations of RMF-exposed cells manifested by change of shape, drop in cell wall density and cytoplasm condensation. The obtained results indicate that the originally limited impact of β-lactam antibiotics in MRSA is boosted by the disturbances caused by RMF in the bacterial cell walls. Taking into account the high clinical need for new therapeutic options, effective against MRSA, the data presented in this study have high developmental potential and could serve as a basis for new treatment options for MRSA infections.

The Effect of Rotating Magnetic Field on Susceptibility Profile of Methicillin-Resistant Staphylococcus aureus Strains Exposed to Activity of Different Groups of Antibiotics.

Methicillin-resistant strains of Staphylococcus aureus (MRSA) have become a global issue for healthcare systems due to their resistance to most β-lactam antibiotics, frequently accompanied by resistance to other classes of antibiotics. In this work, we analyzed the impact of combined use of rotating magnetic field (RMF) with various classes of antibiotics (β-lactams, glycopeptides, macrolides, lincosamides, aminoglycosides, tetracyclines, and fluoroquinolones) against nine S. aureus strains (eight methicillin-resistant and one methicillin-sensitive). The results indicated that the application of RMF combined with antibiotics interfering with cell walls (particularly with the β-lactam antibiotics) translate into favorable changes in staphylococcal growth inhibition zones or in minimal inhibitory concentration values compared to the control settings, which were unexposed to RMF. As an example, the MIC value of cefoxitin was reduced in all MRSA strains by up to 42 times. Apart from the β-lactams, the reduced MIC values were also found for erythromycin, clindamycin, and tetracycline (three strains), ciprofloxacin (one strain), gentamicin (six strains), and teicoplanin (seven strains). The results obtained with the use of in vitro biofilm model confirm that the disturbances caused by RMF in the bacterial cell walls increase the effectiveness of the antibiotics towards MRSA. Because the clinical demand for new therapeutic options effective against MRSA is undisputable, the outcomes and conclusions drawn from the present study may be considered an important road into the application of magnetic fields to fight infections caused by methicillin-resistant staphylococci.

Comparison of PCR and phenotypic methods for the detection of methicillin resistant Staphylococcus aureus.

Background and Objectives:Resistance to methicillin in methicillin resistant strains of Staphylococcus aureus (MRSA) is due to the presence of mec-A gene, which encodes a low affinity penicillin binding protein (PBP)-2a or PBP2. Accurate and rapid identification of MRSA in clinical specimens is essential for timely decision on effective treatment. The aim of the study was to compare three different methods for detection of MRSA namely cefoxitin disc diffusion, CHROM agar MRSA and VITEK-2 susceptibility with PCR which is the gold standard reference method and to find the antibiotic susceptibility pattern of these isolates by VITEK-2.Materials and Methods:A Total of 100 non-duplicate S. aureus isolates were collected from different clinical samples among both outpatient and inpatients. Detection of MRSA among these isolates was done by cefoxitin disc diffusion, VITEK-2, CHROM agar MRSA and PCR.Results:The sensitivity and specificity of cefoxitin disc diffusion and Vitek was found to be 97.2% and 100%, while that of CHROM agar was found to be 100% and 78.6%. The overall prevalence of MRSA in our study by PCR was 72%.Conclusion:Based on the findings in our study, isolates which show cefoxitin zone diameter < 22 mm can be reported as MRSA. However, those isolates which have a zone diameter between 22–24 mm, should ideally be confirmed by PCR.

Bio-synthesis of silver nanoparticles with the brackish water blue-green alga Oscillatoria princeps and antibacterial assessment

This study describes the morphology of a filamentous non-heterocystous blue-green alga (cyanobacterium) Oscillatoria princeps isolated from the Bhabakundalswara sea mouth, its use in biogenic synthesis of silver nanoparticles (AgNPs), along with antimicrobial studies using multidrug resistant (MDR) methicillin-resistant strains of Staphylococcus aureus (MRSA), Streptococcus pyogenes and Escherichia coli, isolated from clinical samples. The synthesized O. princeps -AgNPs were characterized with UV–visible spectrometry, scanning electron microscopy (SEM), X-ray diffraction (XRD), dynamic light scattering analysis (DLS) and Fourier transform infrared spectroscopy (FT-IR) analyses. The synthesized AgNPs had remarkable antibacterial activities against MDR strains of MRSA, S. pyogenes and E. coli with the inhibitory zone sizes, 14–16 mm as recorded from agar well diffusion method, while MIC values were 100, 80, 60 µg/ml, for the respective cited bacterial species. Thus, O. princeps -AgNPs in future would be druggable antibacterials.

Virulent gene detection and antibiogramic profile of methicillin resistant Staphylococcus aureus isolated from birds of a poultry farm

Methicillin-resistant strains of Staphylococcus aureus (MRSA) are gaining global attention, largely due to their potential public health significance. In the current study, we investigated the prevalence of MRSA in poultry birds from Benson Idahosa University farm. Twenty-five samples each from poultry droppings, cloacae and nostrils were collected aseptically and screened for total heterotrophic and Staphylococcus aureus counts usingstandard culture-based methods. Phenotypic identification of MRSA was carried out using mannitol-oxacillin agar, while the presence of virulence genes (mecA, entA, entB, entC, SCCmec (I, II, III) and tsstI) was investigated by polymerase chain reaction using specific primers. Also, the antibiogramic activities and multi drug-resistant index were investigated against 6 clinically relevant antibiotics (oxacillin (1 μg), vancomycin (30 μg), ceftazidime (30 μg), ceftriaxone (30 μg), amikacin (30 μg) and ofloxacin (5 μg)). The results showed that total heterotrophic bacterial and S. aureus are prevalent in poultry birds, with mean heterotrophic counts of 243.08 ± 21.01 × 105 cfu, 169.37 ± 25.30 × 105 cfu and 216 ± 15.89× 105 cfu for poultry droppings, cloacae and nostrils respectively and mean S. aureus counts of 176.30 ± 28.85 × 105 cfu for poultry droppings, 16.52 ± 11.92 × 105 cfu for cloacae and 5 2.72 ± 0.41 × 105 cfu for nostrils. Interestingly, 93.33%, 25% and 10.5% of S. aureus isolated from the droppings, cloacae and nostrils respectively were methicillin resistant. The antibiogramic activities showed the majority of isolates to be multi drug-resistant, while only one MRSA (from poultry droppings) had the mecA gene. The presence of these isolates in poultry birds is of a potential public health concern as they may travel through the food chain and efforts should be made by poultry owners to avoid the indiscriminate use of antibiotics.&#x0D; Keywords: Methicillin-Resistant Staphylococcus aureus; Poultry; Public Health; Virulence Gene

Marked Presence of Methicillin-Resistant Staphylococcus aureus in Wild Lagomorphs in Valencia, Spain.

Simple SummaryThe presence of bacteria resistant to antibiotics, such as Staphylococcus aureus resistant to methicillin (MRSA), is becoming an increasing everyday concern for their implications for human and animal health. This is even more alarming when such bacteria are isolated in wild animals which, in principle, should not come into contact with antibiotics. This work studied 353 rabbits and 10 hares hunted in rabbit high-density areas. Of these, 41.3% carried S. aureus in some sampled locations, of which 63.3% were MRSA. These are surprising results given the unexpected high presence of MRSA in the studied animals. This finding is very worrying because these animals tend to enter the food chain with no veterinary control, which implies a risk for human health. Thus it is necessary to extend this study to other, less-populated areas with other animal species (ruminants, rabbit predators, hunting dogs, etc.) or even water sources to obtain further knowledge about the origin of bacterial resistances in nature.The appearance of methicillin-resistant strains of Staphylococcus aureus (MRSA) in several animal species (including rabbits) has set off alarms for their capacity to act as reservoirs for this bacterium. This is especially important in wild animals given its epidemiological implications. The objectives of this study were to identify and characterize S. aureus, specifically MRSA, strains in wild lagomorph high-density areas. Ten hares and 353 wild rabbits from 14 towns with a high rabbit density in the Valencian region (eastern Spanish coast) were sampled. Swabs from the nasal cavity, ears, perineum and lesions (when present) were taken for microbiological studies. The detection of different genes and antibiotic susceptibility studies were also carried out. Of all the animals, 41.3% were positive for S. aureus, of which 63.3% were MRSA. Ears were the anatomical location with more S. aureus and MRSA strains. The more frequently identified MLST type was ST1945 (97.1%, 136/140). The mecA gene was found only in one sample. The rest (n = 139) carried the mecC gene and were included in CC130, except one. Penicillin resistance was detected in 28 mec-negative isolates and, in one case, bacitracin resistance. mecA isolate presented resistance to enrofloxacin and tetracycline, and 10 mecC isolates also showed bacitracin resistance. No MRSA isolate was positive for genes chp, sea, tst and PVL. Two ST1945 isolates contained IEC type E (comprising genes scn and sak). mecA-isolate was positive for blaZ. Of the 28 MSSA strains showing resistance to penicillin, 22 carried the blaZ gene. These surprising results highlight the marked presence of MRSA strains in wild rabbits in high-density areas.

The Phenotypic and Genetic Detection of the Methicillin-Resistant Staphylococcus aureus (MRSA)

Methicillin resistant strains of Staphylococcus aureus (MRSA) were identified shortly upon the introduction of methicillin into the clinical practice. The S. aureus samples were taken from patients hospitalized in General Hospital of Chania “Agios Georgios”. The strains were isolated from different pathological products, in the hospital laboratory. All isolates were tested using the cefoxitin disk diffusion, the oxacillin MIC methods and the PBP2’ latex agglutination test (bioMérieux), for the production of the Penicillin-Binding Protein 2a (PBP2a or PBP2’ protein). S. aureus ATCC 29213 was used as a negative control. We followed the detection of the mecA gene throughout the PCR method, as the standard “gold” method, in order to identify MRSA strains. Conventional methods for MRSA strains detection were compared with the PCR method. The antibiotic susceptibility testing was performed throughout the Kirby-Bauer disk diffusion method using antibiotic discs from Bioanalyse ltd. The mecA gene was found by PCR in 57.5 % of S. aureus strains, which allowed defining the isolated strains as MRSA strains. According to the oxacillin MIC values of the studied strains, 25 strains (53.2%) were identified as MRSA, 21 (44.68%) as MSSA and 1*strain (2.13%) as Borderline (BL) MRSA. The mecA gene is present in 24 of the MRSA strains with oxacillin MIC ≥ 4 being more common in strains showing the oxacillin MICs ≥ 256 (12/27). Adding the BL strains to the methicillin resistant strains, the rate of the MRSA strains increases to 26 (55.32%), the appropriate values of the MRSA strains percent, as determined by the PCR method (57.45%), which shows a concordance of 96.3% (26/27), between the results obtained by the two tests. Comparing the results obtained using the PCR method; with the oxacillin MICs and the PBP2’ latex agglutination test, concordant results were obtained for 89.36% of the strains (42/47) by oxacillin MICs and for 97.87% of the strains (46/47) by PBP2’ latex agglutination test. We conclude that the specificity of these methods is 100% for the mecA PCR method, 97.87% for the PBP2a latex method and 89.36% for the oxacillin MIC. The comparison of phenotypic methods (the PBP2a latex reaction, oxacillin MICs, the Cefoxitin disk diffusion test with the genotypic methods (the presence of the mecA gene), reveals that the PBP2a latex reaction has high sensitivity (97.87%), and can be used as an alternative method to PCR for the MRSA detection, in resource constraint settings.

Magnetite nanoparticles functionalized with propolis against methicillin resistant strains of Staphylococcus aureus

Abstract Magnetite nanoparticles (MNPs) have been evaluated for inhibiting microbial growth and biofilm formation. In this study the effect of the nanocomposite Moroccan propolis extract / MNPs acting against methicillin resistant strains of Staphylococcus aureus (MRSA) was evaluated. Chemical composition of propolis was established by pyrolysis coupled to gas chromatography and mass spectrometry method (pyrolysis GC/MS). MNPs were obtained through the co-precipitation method. The fabricated nanostructure was characterized by X-ray Diffraction (DRX), Transmission Electron Microscopy (TEM), and Fourier Transform-Infrared Spectroscopy (FTIR). TEM of MNPs provided a particle average size of 15 nm, FTIR spectral analysis enabled a fast way of identification of MNPs, attesting the occurrence of the different combinations. The use of MNPs loaded with propolis and the antibiotic chloramphenicol at Minimum Inhibitory Concentration (MIC) value inhibited the bacterial growth of MSSA (methicillin susceptible strain of S. aureus) and MRSA strains. After the treatment with MNPs-OA-P-CLo nanocomposite (MNPs with oleic acid, propolis and chloramphenicol), the disruption of the bacterial cell was observed by TEM. The combination of propolis and chloramphenicol in free form at MIC value largely impaired both MSSA and MRSA strains as, after 2 h of treatment, no viable cells of MRSA 2 and MRSA 16 were recovered. Hence, the results elucidated a new antibacterial nanocomposite synthesis, for possible applications as prospective nanoantibacterial agents or drug carriers.

Design of Bactericidal Peptides Against Escherichia coli O157:H7, Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus.

Antimicrobial peptides are on the first line of defense against pathogenic microorganisms of many living beings. These compounds are considered natural antibiotics that can overcome bacterial resistance to conventional antibiotics. Due to this characteristic, new peptides with improved properties are quite appealing for designing new strategies for fighting pathogenic bacteria. Sixteen designed peptides were synthesized using Fmoc chemistry; five of them are new cationic antimicrobial peptides (CAMPs) designed using a genetic algorithm that optimizes the antibacterial activity based on selected physicochemical descriptors and 11 analog peptides derived from these five peptides were designed and constructed by single amino acid substitutions. These 16 peptides were structurally characterized and their biological activity was determined against Escherichia coli O157:H7 (E. coli O157:H7), and methicillin-resistant strains of Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa (P. aeruginosa) were determined. These 16 peptides were folded into an α-helix structure in membrane-mimicking environment. Among these 16 peptides, GIBIM-P5S9K (ATKKCGLFKILKGVGKI) showed the highest antimicrobial activity against E. coli O157:H7 (MIC=10µM), methicillin-resistant Staphylococcus aureus (MRSA) (MIC=25µM) and Pseudomonas aeruginosa (MIC=10 µM). Peptide GIBIM-P5S9K caused permeabilization of the bacterial membrane at 25 µM as determined by the Sytox Green uptake assay and the labelling of these bacteria by using the fluoresceinated peptide. GIBIM-P5S9K seems to be specific for these bacteria because at 50 µM, it provoked lower than 40% of erythrocyte hemolysis. New CAMPs have been designed using a genetic algorithm based on selected physicochemical descriptors and single amino acid substitution. These CAMPs interacted quite specifically with the bacterial cell membrane, GIBIM-P5S9K exhibiting high antibacterial activity on Escherichia coli O157:H7, methicillin-resistant strains of Staphylococcus aureus and P. aeruginosa.

Antibacterial Activity of In Situ Prepared Chitosan/Silver Nanoparticles Solution Against Methicillin-Resistant Strains of Staphylococcus aureus

BackgroundInvestigation of new effective drugs against the methicillin-resistant strains of Staphylococcus aureus (MRSA) is an urgent issue of modern medicine. Antiseptics as an alternative of antibiotics are strong, sustained, and active preparations against resistant strains and do not violate microbiocenosis.Materials and MethodsThe activity of in situ prepared chitosan-Ag nanoparticles (Ag NPs) solution with different component ratio was tested against MRSA isolated from patients. Ag NPs were synthesized via chemical reduction method using green chemistry approach. In order to improve antimicrobial activity and dispersibility of Ag NPs, surface modification of Ag NPs by cetrimonium bromide (CTAB) was performed.Ag NPs and chitosan-Ag NPs solution were characterized using X-ray diffraction, transmission electron microscopy, infrared spectroscopy, and spectrophotometric measurements.Results and ConclusionsThe results of XRD, FTIR, UV–Vis, and TEM measurements confirmed the chemical composition of chitosan and Ag NPs and their high purity.Chitosan-AgNPs solutions have shown their superior antimicrobial efficacy compared to its pure forms. At the same time, in situ preparation of chitosan-Ag NPs solution (chitosan powder 6.0 μg/ml, Ag/CTAB NPs) was not possible due to the precipitation of the components. This result is very promising and may be considered as an effective solution in fighting against drug-resistant bacteria.