Vancomycin-resistant E. Faecalis Research Articles

Critical view on antimicrobial, antibiofilm and cytotoxic activities of quinazolin-4(3H)-one derived schiff bases and their Cu(II) complexes

A series of nine 2,3-disubstituted-quinazolin-4(3H)-one derived Schiff bases and their three Cu(II) complexes was prepared and tested for their antimicrobial activities against reference strains Staphylococcus aureus ATCC 29213 and Enterococcus faecalis ATCC 29212 and resistant clinical isolates of methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). All the substances were tested in vitro against Mycobacterium tuberculosis H37Ra ATCC 25177, M. kansasii DSM 44162 and M. smegmatis ATCC 700084. While anti-enterococcal and antimycobacterial activities were insignificant, 3-[(E)-(2-hydroxy-5-nitrobenzylidene)amino]-2-(2-hydroxy-5-nitrophenyl)-2,3-dihydroquinazolin-4(1H)-one (SB3) and its Cu(II) complex (SB3–Cu) demonstrated bacteriostatic antistaphylococcal activity. In addition, both compounds, as well as the other two prepared complexes, showed antibiofilm activity, which resulted in a reduction of biofilm formation and eradication of mature S. aureus biofilm by 80% even at concentrations lower than the values of their minimum inhibitory concentrations. In addition, the compounds were tested for their cytotoxic effect on the human monocytic leukemia cell line THP-1. The antileukemic efficiency was improved by the preparation of Cu(II) complexes from the corresponding non-chelated Schiff base ligands.

Synthesis, Characterization, and Bioactivity of Mesoporous Bioactive Glass Codoped with Zinc and Silver.

Due to the overconsumption of antimicrobials, antibiotic-resistant bacteria have become a critical health issue worldwide, especially methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). Recently, many efforts have been made to load metals into bioactive glasses to enhance the multifunctionality of materials, such as antibacterial and osteoinductive functions. Zinc has been documented to stimulate the gene expression of various regulatory factors in bone cells. Meanwhile, previous studies have reported that silver and zinc could be a promising antibacterial combination with synergistic antimicrobial effects. Here, we sought to develop a biomaterial coreleasing zinc and silver, designated 80S-ZnAg, and to evaluate its antibacterial activity and biocompatibility. The textural analyses demonstrated different coreleasing patterns of zinc and silver for the materials. The chemical characterization revealed that the zinc in 80S-ZnAg could be the network modifier when its molar ratio was high, releasing more zinc; zinc could also be the network former when its molar ratio was low, showing an extremely low rate of release. However, the ICP results for 80S-Zn3Ag2 demonstrated up to 7.5 ppm of zinc and 67.6 ppm of silver. Among all the 80S-ZnAg materials, 80S-Zn3Ag2 demonstrated more marked antibacterial activity against MRSA and VRE than the others, with inhibition zones of 11.5 and 13.4 mm, respectively. The cytotoxicity assay exhibited nearly 90% cell viability at 20 mg/mL of 80-Zn3Ag2. Further clinical study is needed to develop an innovative biomaterial to address the issue of antibiotic resistance.

New Hybrid Phenalenone Dimer, Highly Conjugated Dihydroxylated C28 Steroid and Azaphilone from the Culture Extract of a Marine Sponge-Associated Fungus, Talaromyces pinophilus KUFA 1767

An undescribed hybrid phenalenone dimer, talaropinophilone (3), an unreported azaphilone, 7-epi-pinazaphilone B (4), an unreported phthalide dimer, talaropinophilide (6), and an undescribed 9R,15S-dihydroxy-ergosta-4,6,8 (14)-tetraen-3-one (7) were isolated together with the previously reported bacillisporins A (1) and B (2), an azaphilone derivative, Sch 1385568 (5), 1-deoxyrubralactone (8), acetylquestinol (9), piniterpenoid D (10) and 3,5-dihydroxy-4-methylphthalaldehydic acid (11) from the ethyl acetate extract of the culture of a marine sponge-derived fungus, Talaromyces pinophilus KUFA 1767. The structures of the undescribed compounds were elucidated by 1D and 2D NMR as well as high-resolution mass spectral analyses. The absolute configuration of C-9' of 1 and 2 was revised to be 9'S using the coupling constant value between C-8' and C-9' and was confirmed by ROESY correlations in the case of 2. The absolute configurations of the stereogenic carbons in 7 and 8 were established by X-ray crystallographic analysis. Compounds 1,2, 4-8, 10 and 11 were tested for antibacterial activity against four reference strains, viz. two Gram-positive (Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212) and two Gram-negative (Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853), as well as three multidrug-resistant strains, viz. an extended-spectrum β-lactamase (ESBL)-producing E. coli, a methicillin-resistant S. aureus (MRSA) and a vancomycin-resistant E. faecalis (VRE). However, only 1 and 2 exhibited significant antibacterial activity against both S. aureus ATCC 29213 and MRSA. Moreover, 1 and 2 also significantly inhibited biofilm formation in S. aureus ATCC 29213 at both MIC and 2xMIC concentrations.

Trifluoromethylcinnamanilide Michael Acceptors for Treatment of Resistant Bacterial Infections

A series of thirty-two anilides of 3-(trifluoromethyl)cinnamic acid (series 1) and 4-(trifluoromethyl)cinnamic acid (series 2) was prepared by microwave-assisted synthesis. All the compounds were tested against reference strains Staphylococcus aureus ATCC 29213 and Enterococcus faecalis ATCC 29212 and resistant clinical isolates of methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). All the compounds were evaluated in vitro against Mycobacterium smegmatis ATCC 700084 and M. marinum CAMP 5644. (2E)-3-[3-(Trifluoromethyl)phenyl]-N-[4-(trifluoromethyl)phenyl]prop-2-enamide (1j), (2E)-N-(3,5-dichlorophenyl)-3-[3-(trifluoromethyl)phenyl]prop-2-enamide (1o) and (2E)-N-[3-(trifluoromethyl)phenyl]-3-[4-(trifluoromethyl)-phenyl]prop-2-enamide (2i), (2E)-N-[3,5-bis(trifluoromethyl)phenyl]-3-[4-(trifluoromethyl)phenyl]-prop-2-enamide (2p) showed antistaphylococcal (MICs/MBCs 0.15-5.57 µM) as well as anti-enterococcal (MICs/MBCs 2.34-44.5 µM) activity. The growth of M. marinum was strongly inhibited by compounds 1j and 2p in a MIC range from 0.29 to 2.34 µM, while all the agents of series 1 showed activity against M. smegnatis (MICs ranged from 9.36 to 51.7 µM). The performed docking study demonstrated the ability of the compounds to bind to the active site of the mycobacterial enzyme InhA. The compounds had a significant effect on the inhibition of bacterial respiration, as demonstrated by the MTT assay. The compounds showed not only bacteriostatic activity but also bactericidal activity. Preliminary in vitro cytotoxicity screening was assessed using the human monocytic leukemia cell line THP-1 and, except for compound 2p, all effective agents did show insignificant cytotoxic effect. Compound 2p is an interesting anti-invasive agent with dual (cytotoxic and antibacterial) activity, while compounds 1j and 1o are the most interesting purely antibacterial compounds within the prepared molecules.

Design, Synthesis and Antimicrobial Properties of New Tetracyclic Quinobenzothiazine Derivatives.

A new method for modifying the structure of tetracyclic quinobenzothiazinium derivatives has been developed, allowing introduction of various substituents at different positions of the benzene ring. The method consists of reacting appropriate aniline derivatives with 5,12-(dimethyl)thioquinantrenediinium bis-chloride. A series of new quinobenzothiazine derivatives was obtained with propyl, allyl, propargyl and benzyl substituents in 9, 10 and 11 positions, respectively. The structure of the obtained compounds was analyzed by 1H and 13C NMR (HSQC, HMBC) and X-ray analysis. All the compounds were tested against reference strains Staphylococcus aureus ATCC 29213 and Enterococcus faecalis ATCC 29212, and representatives of multidrug-resistant clinical isolates of methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). In addition, all the compounds were evaluated in vitro against Mycobacterium smegmatis ATCC 700084 and M. marinum CAMP 5644. 9-Benzyloxy-5-methyl-12H-quino [3,4-b][1,4]benzothiazinium chloride (6j), 9-propoxy-5-methyl-12H-quino[3,4-b][1,4]benzothiazinium chloride (6a) and 9-allyloxy-5-methyl-12H-quino[3,4-b][1,4]benzothiazinium chloride (6d) demonstrated high activity against the entire tested microbial spectrum. The activities of the compounds were comparable with oxacillin, tetracycline and ciprofloxacinagainst staphylococcal strains and with rifampicin against both mycobacterial strains. Compound 6j had a significant effect on the inhibition of bacterial respiration as demonstrated by the MTT assay. The compounds showed not only bacteriostatic activity, but also bactericidal activity. Preliminary in vitro cytotoxicity screening of the compounds performed using normal human dermal fibroblasts (NHDF) proved that the tested compounds showed an insignificant cytotoxic effect on human cells (IC50 > 37 µM), making these compounds interesting for further investigation. Moreover, the intermolecular similarity of novel compounds was analyzed in the multidimensional space (mDS) of the structure/property-related in silico descriptors by means of principal component analysis (PCA) and hierarchical clustering analysis (HCA), respectively. The distance-oriented structure/property distribution was related with the experimental lipophilic data.

Development of novel isatin thiazolyl-pyrazoline hybrids as promising antimicrobials in MDR pathogens.

Microbial Multidrug Resistance (MDR) is an emerging global crisis. Derivatization of natural or synthetic scaffolds is among the most reliable strategies to search for and obtain novel antimicrobial agents for the treatment of MDR infections. Here, we successfully manipulated the synthetically flexible isatin moieties to synthesize 22 thiazolyl-pyrazolines hybrids, and assessed their potential antimicrobial activities in vitro against various MDR pathogens, using the broth microdilution calorimetric XTT reduction method. We chose 5 strains to represent the major MDR microorganisms, viz: Methicillin-resistant S. aureus (MRSA), and Vancomycin-resistant E. faecalis (VRE) as Gram-positive bacteria; Carbapenem-resistant K. pneumonia (CRKP), and Extended-spectrum beta-lactamase E. coli (ESBL-E), as Gram-negative bacteria; and Fluconazole-resistant C. albicans (FRCA), as a yeast-like unicellular fungus. The cytotoxicity of compounds 9f and 10h towards mammalian lung fibroblast (MRC-5) cells demonstrated their potential satisfactory safety margin as represented by their relatively high IC50 values. The target compounds showed promising anti-MDR activities, suggesting they are potential leads for further development and in vivo studies.

Functionalized Cyclopentenones and an Oxime Ether as Antimicrobial Agents.

Several naturally occurring cyclopentenones, such as palmenones and nigrosporiones, exhibit antimicrobial activity. Herein we describe the antimicrobial activity of cyclopentenones and derivatives that can be easily accessed from biomass derivatives furfural and 5‐hydroxymethylfurfural. Upon screening a range of functionalized trans‐diamino‐cyclopentenones (DCPs) and δ‐lactone‐fused cyclopentenones (LCPs), an oxime ether derivative of DCP was identified that exhibited remarkable antimicrobial activity against Gram‐positive bacteria, including resistant strains such as methicillin‐resistant S. aureus (MRSA) and vancomycin‐resistant E. faecalis (VRE) strains.

VB_EfaS-DELF1, a novel Siphoviridae bacteriophage with highly effective lytic activity against vancomycin-resistant Enterococcus faecalis

Enterococcus faecalis is an environmental agent of bovine mastitis in cows and has many cytopathic effects on the urinary tract in both humans and animals. In this study, a novel lytic bacteriophage, vB_EfaS-DELF1, was isolated against 21 E. faecalis isolated from bovine mastitis, including vancomycin-resistant E. faecalis (VRE). vB_EfaS-DELF1 bacteriophage was specific for E. faecalis and showed no lytic effects against other tested Enterococcus spp., Gram-negative, or Gram-positive bacteria. Moreover, no activity was observed against yogurt starters. The phage suspension was stable in a wide range of pH, salinity, and temperature. It retained its activity in 3.5 % fat milk. vB_EfaS-DELF1 has the common phenotypic features of Siphoviridae with a double-strand DNA of 40,248 bp in length and a G + C content of 34.9 %. The genome encodes 62 putative ORFs and no tRNA. No undesirable genes such as lysogenic mediators, antibiotic resistance, or virulence factor genes were detected in the genome. The comparative genomic analysis demonstrated similarity to the other available phage genomes. The highest similarity was observed with two other phages (50 % coverage and 82.38 % identities with IME-EFm1; 35 % coverage and 86.22 % identities with IME-EFm5) that were placed in the same clade. The differences with the other aligned phages were high and were placed in distant clusters.Regarding the specificity of this new bacteriophage against all of the tested E. faecalis isolates and, in particular, against the vancomycin-resistant ones, and also the absence of antibiotic resistance or virulence genes in its genome, vB_EfaS-DELF1 is suggested as a potential candidate for biocontrol of E. faecalis infections.

Comparative genomics of vancomycin-resistant Enterococcus spp. revealed common resistome determinants from hospital wastewater to aquatic environments

The rise of vancomycin-resistant Enterococcus spp. (VRE) has led to treatment challenges in hospital settings worldwide. Hospital wastewater (HW) might disseminate this threat to the aquatic environment. Thus, this study elucidates the VRE resistance quotient (RQ) of different environmental matrixes in wastewater and compares genomic determinants of VRE strains recovered from HW to water resources. Presumptive Enterococcus spp. and VRE were quantified and isolated using standard microbiological procedures. Fourteen VRE genomes were then sequenced using an Illumina HiSeq X™ Ten platform. Subsequently, VRE genomes were compared based on antibiotic resistance genes, plasmids, bacteriophages, insertion sequences, transposons, virulence and pathogenicity. Wastewater effluent showed the highest RQ among all sampled matrixes. The phylogeny of vancomycin-resistant E. faecalis (VREfs) and E. faecium (VREfm) revealed a tree structure based on their respective sequence type. A comparative genomic analysis of 14 genomes highlighted regions encoding phage protein, phage holin, phage integrase, integrase and transposase on both query genomes and the reference genome. Acquired resistance to vancomycin was conferred by vanA, vanN, vanL, vanG and the intrinsic resistance vanC operons. Plasmids were dominated by the presence of conserved areas of the replication initiating genes (rep). The Tn3-like and Tn917 transposons were present in all erythromycin-carrying erm(B) isolated VRE genomes. All VRE genomes expect one were putatively predicted as human pathogens with varying degrees of virulence. The presence of such resistant bacteria in African water resource is of great public health concern. It is, therefore, recommended that these bacteria be tracked and characterised from different environments to contribute to improved epidemiological containment action.

Molecular characterization of vancomycin-resistant enterococci isolated from a hospital in Beijing, China.

To investigate the occurrence of vancomycin-resistant enterococci (VRE) isolated from patients in Peking Union Medical College Hospital, Beijing, China from 2011 to 2017, and to evaluate their resistance mechanisms and genetic relatedness. All isolates were identified using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF). Antibiotic susceptibility testing was performed using the broth microdilution method. Molecular characterization were detected by PCR and sequencing. Genotyping of VRE isolates was performed by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) analysis. Virulence genes were detected by multiplex PCR. A total of 87 consecutive VRE were collected, including 84 isolates of vancomycin resistant Enterococcus faecium (VREfm) and 3 isolates of Enterococcus faecalis (VREfs). Urine (40.2%, 35/87) and blood (17.2%, 15/87) were the most commonly specimens. All VREfm isolates were resistant to ampicillin, and were susceptible to daptomycin, linezolid and tigecycline. The resistant rate of teicoplanin was 47.6%. All of the VREfm isolates carried the vanA gene, no isolates carried vanB. 11.9% (10/84) VREfm isolates carried both vanA and vanM. Among them, 76.2% (64/84) and 66.7% (56/84) carried esp and hyl, respectively. The 3 vancomycin resistant E. faecalis (VREfs) isolates were varied, and only one carried vanB. A total of 3 and 18 STs were detected among VREfs and VREfm strains, respectively. PFGE results indicated a genetic diversity among VREfm isolates. This study confirms that VREfm isolates associated with ST78 were the main epidemic lineage responsible for nosocomial infections in China, as were also observed in other nations worldwide.

Антибіотикорезистентність клінічних штамів Enterococcus faecalis у хірургічних стаціонарах України: результати багатоцентрового дослідження (2011-2015)

Мета роботи — вивчити антибіотикорезистентність клінічних штамів Enterococcus faecalis, виділених у пацієнтів, госпіталізованих у хірургічні стаціонари в різних регіонах України.
 Матеріали і методи. Досліджено 87 273 клінічних штами E. faecalis, виділені від пацієнтів, госпіталізованих у хірургічні відділення 47 багатопрофільних стаціонарів у м. Києві та 24 областях України у 2011—2015 рр. Вивчено чутливість штамів E. faecalis до 34 антибіотиків (пеніцилін, ампіцилін, амоксицилін, оксацилін, ампіцилін/сульбактам, амоксицилін/клавуланат, цефазолін, цефалексин, цефуроксим, цефоперазон, цефотаксим, цефтріаксон, цефтазидим, цефепім, іміпенем, меропенем, гентаміцин, амікацин, нетилміцин, еритроміцин, кларитроміцин, азитроміцин, лінкоміцин, кліндаміцин, тетрациклін, доксициклін, ванкоміцин, офлоксацин, лефлоцин, норфлоксацин, гатифлоксацин, ципрофлоксацин, пефлоксацин, лінезолід) відповідно до рекомендацій Інституту клінічних та лабораторних стандартів США (Clinical and Laboratory Standards Institute (CLSI)).
 Результати та обговорення. У досліджених хірургічних стаціонарах 37 % штамів E. faecalis були резистентними до тестованих антибіотиків. Найактивнішими щодо штамів E. faecalis були лінезолід, ванкоміцин, меропенем, лефлоцин та іміпенем. Високу частоту резистентності відзначено до цефалексину (56,4 %), пеніциліну (47,9 %), цефуроксиму (47,4 %), цефоперазону (47,0 %), норфлоксацину (43,1 %), кларитроміцину (42,6 %), цефтазидиму (41,6 %), доксицикліну (40,5 %), кліндаміцину (39,7 %), цефотаксиму (39,5 %), амікацину (38,1 %), гентаміцину (37,4 %) та азитроміцину (37,6 %). Резистентність штамів E. faecalis до ванкоміцину становила від 8,0 до 49,4 % (у середньому — 20,8 %).
 Висновки. Антибіотикорезистентність клінічних штамів E. faecalis у досліджених стаціонарах — важлива терапевтична та епідеміологічна проблема. Найбільшою активністю щодо нозокоміальних штамів E. faecalis характеризуються лінезолід, ванкоміцин, меропенем, лефлоцин та іміпенем. З огляду на постійні зміни рівня резистентності нозокоміальних штамів E. faecalis у регіонах України необхідно проводити постійний моніторинг за резистентністю до антибіотиків у кожному стаціонарі та на підставі отриманих локальних даних розробити лікарняний формуляр антибіотиків. Політику використання антибіотиків у кожному хірургічному стаціонарі слід визначати залежно від локальних даних щодо резистентності до протимікробних препаратів. Необхідно налагодити систему епідеміологічного нагляду за мікробною резистентністю на локальному, регіональному та національному рівнях.

Detection of vancomycin resistance in enterococcus species isolated from clinical samples and feces of colonized patients by phenotypic and genotypic methods.

The aim of this study was to find out the clinical correlation between the presence of vancomycin-resistant genes (van A and van B) and their expression as detected by phenotypic tests in colonized patients and in clinical isolates. Enterococci were isolated from various clinical samples and also from fecal specimens of colonized patients at the time of admission, after 48 h and after 5 days of admission. Identification to species level was done using standard methods. Vancomycin susceptibility in Enterococci was detected by disc diffusion test. Minimum inhibitory concentration was determined by agar dilution method. Multiplex polymerase chain reaction (PCR) was used to detect the presence of van genes. Out of all the clinical and fecal samples processed, 12.0% isolates were either vancomycin resistant or vancomycin intermediate. Further, these isolates carried van A or van B genes as confirmed by PCR methods. Expression of van A gene was found to be more in Enterococcus faecalis (28.3%) as compared to Enterococcus faecium (25.0%) in both clinical and fecal isolates. 16.6% strains of E. faecium and 15.0% strains each of E. faecalis and Enterococcus gallinarum were found to carry van B genes. The overall prevalence of vancomycin resistant Enterococci (VRE) in colonized patients was about 9.6%. Prior administration of antibiotics had significant effect (P = 0.001) on VRE carriage. Urinary tract infection was the most common infection caused by vancomycin susceptible Enterococci (VSE), 105/214 (49.0%) and VRE, 13/36 (36.1%). There was no significant difference (P = 0.112) in the distribution of VRE and VSE in different infection types. Both clinical and fecal VRE showed maximum resistance to penicillin, ampicillin, and piperacillin. Resistance to linezolid was 2.8% in clinically isolated VRE. VRE in our study were found to be resistant to a number of commonly used antibiotics. The frequency of isolation of vancomycin resistant E. faecalis (VRE.fs), which is highly virulent, and the number of strains harboring van A gene in our hospital setup is high and needs to be addressed.

Analysis of 5-year trends in daptomycin activity tested against Staphylococcus aureus and enterococci from European and US hospitals (2009-2013).

This study evaluated daptomycin activity trends among meticillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE) over a 5-year period (2009-2013). Consecutive, unique patient isolates of clinical significance were collected in 67 European (EU) (17 nations) and 145 US medical centres and were susceptibility tested in a central reference laboratory against daptomycin and various comparators by CLSI broth microdilution methods. MIC results were interpreted according to EUCAST and CLSI breakpoint criteria (2014). A total of (EU/USA) 14245/22967 S. aureus (26.3/49.8% MRSA), 3043/2848 Enterococcus faecalis (1.6/3.5% vancomycin-resistant) and 1827/1537 Enterococcus faecium (29.5/77.7% vancomycin-resistant) were evaluated. Isolates were mainly from complicated skin and skin-structure infections (40%) and bacteraemia (32%). The highest MRSA rates were found in Portugal (66.3%), Russia (52.2%) and the USA (49.8%), whereas the highest rates of vancomycin-resistant E. faecium (VREFM) were observed in the USA (77.7%), Poland (63.1%) and Ireland (54.5%). VREFM and vancomycin-resistant E. faecalis (VREF) were observed in 15 and 8 EU nations, respectively. Daptomycin susceptibility rates were (EU/USA) 99.94/99.97%, 100.0/99.7% and 100.0/99.96% for S. aureus, E. faecium and E. faecalis, respectively. Daptomycin was very active against MRSA (MIC50/90, 0.25/0.5mg/L in Europe and USA), VREF (MIC50/90 of 0.5/1mg/L and 1/2mg/L in Europe and USA, respectively) and VREFM (MIC50/90, 2/2mg/L in Europe and USA). Among MRSA, only 6/6 (0.19/0.05%) daptomycin-non-susceptible isolates were observed in Europe (six cities in four countries)/USA (six states), with no increasing trend over the study period. Only two daptomycin-non-susceptible VRE were identified, both E. faecium from the USA.

Epidemiology and Molecular Characterization of Vancomycin-ResistantEnterococcus faecalis

Background: Vancomycin-resistant Enterococci (VRE) infections are caused by Enterococcus faecium in about 90% of the cases but can also be caused by Enterococcus faecalis. Thus, this study investigates factors that cause a low isolation rate of vancomycin-resistant E. faecalis (VREfs). To this end, the authors study the clinical traits, resistant gene structure, genomic classification, and molecular characteristics of the virulent factor. Methods: From January 2001 through September 2011, 17 vanA-containing E. faecalis isolates were collected from hospitalized patients at Ajou University Hospital in Korea. Identification, antimicrobial susceptibility testing, and PCR of van and esp genes were performed. Pulsed-field gel electrophoresis (PFGE) was used for strain typing. PCR and sequencing of the internal regions of Tn1546 were performed for structural analysis of the van gene. Results: Of 4,235 VRE infections, 3,918 (92.5%) were caused by E. faecium, and 95 (2.2%) were caused by E. faecalis. In 67% of VREfs infections, there was a preceding occurrence of E. faecium infection. All isolates were of genotype vanA. Our isolates were divided into three types according to the distribution of IS elements integrated into Tn1546 (types I to IIb). The PFGE results showed no clonal relatedness among isolates. Conclusion: Our study found that VREfs infections affect patients who have experienced vancomycin-resistant E. faecium. (VREfm) infection or undergo invasive procedures. The VREfs seems to involve the horizontal transfer of Tn1546 transposon from VREfm. (Ann Clin Microbiol 2015;18:76-81)

Clonal spread of vancomycin resistance Enterococcus faecalis in an Iranian referral pediatrics center.

Vancomycin-resistant enterococci (VRE) represent as an immediate threat to public health. Since few active compounds are available for VRE infections, rapid identification of these isolates are essential. In the absence of any report on the genetic relatedness of Enterococcus faecalis especially Vancomycin-resistant E. faecalis (VREF) isolates in Iran, we undertook this study to characterize these isolates using random amplification of polymorphic DNA (RAPD-PCR) genotyping method. In this study, E. faecalis strains isolated from various samples collected from different wards of Children Medical Hospital (Tehran, Iran). These isolates were identified by standard laboratory procedures and tested for antimicrobial resistance to vancomycin and teicoplanin. The genetic similarity of the strains was investigated by amplification of the RAPD-PCR. In our study among 91 E. faecalis isolates, 15 (16%) were identified as VREF. The similarity pattern built for E. faecalis isolates by RAPD-PCR, demonstrated the presence of four distinct clusters (A, B, C, D). It is of interest to note that 100% of VREF isolates belonged to Clusters A, indicating that there may have occurred horizontal transmission of the same strain between patients. In conclusion, rapid spread of VREF from a clonal origin calls for implementation of careful isolation and infection control measures. Therefore, environmental control by routine disinfection of patient area as well as screening of high risk patients and isolation of colonized patients should be imposed in order to diminish risk of acquiring nosocomial VRE.

3-Phenyl substituted 6,7-dimethoxyisoquinoline derivatives as FtsZ-targeting antibacterial agents

The emergence of multidrug-resistant bacteria has created an urgent need for antibiotics with a novel mechanism of action. The bacterial cell division protein FtsZ is an attractive target for the development of novel antibiotics. The benzo[c]phenanthridinium sanguinarine and the dibenzo[a,g]quinolizin-7-ium berberine are two structurally similar plant alkaloids that alter FtsZ function. The presence of a hydrophobic functionality at either the 1-position of 5-methylbenzo[c]phenanthridinium derivatives or the 2-position of dibenzo[a,g]quinolizin-7-ium derivatives is associated with significantly enhanced antibacterial activity. 3-Phenylisoquinoline represents a subunit within the ring-systems of both of these alkaloids. Several 3-phenylisoquinolines and 3-phenylisoquinolinium derivatives have been synthesized and evaluated for antibacterial activity against Staphylococcus aureus and Enterococcus faecalis, including multidrug-resistant strains of methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). A number of derivatives were found to have activity against both MRSA and VRE. The binding of select compounds to S. aureus FtsZ (SaFtsZ) was demonstrated and characterized using fluorescence spectroscopy. In addition, the compounds were shown to act as stabilizers of SaFtsZ polymers and concomitant inhibitors of SaFtsZ GTPase activity. Toxicological assessment of select compounds revealed minimal cross-reaction mammalian β-tubulin as well as little or no human cytotoxicity.

Screening of Flavonoids as Candidate Antibiotics against Enterococcus faecalis

beta-Ketoacyl acyl carrier protein synthase (KAS) III, the most divergent member of the condensing enzyme family, is a key catalyst in bacterial fatty acid biosynthesis and, thus, an attractive target for novel antibiotics. Here, we perform docking studies between Enterococcus faecalis KAS III (efKAS III) and one flavanone and 11 hydroxyflavanones with hydroxy groups at various positions. The MIC values of these flavanones for E. faecalis and vancomycin-resistant E. faecalis (VREF) were measured, and binding affinities to efKAS III were determined. Naringenin (9), eriodictyol (10), and taxifolin (12), with high-scoring functions and good binding affinities, docked well with efKAS III, resulting in MIC values in the range 128-512 microg/mL. Our results indicate that hydrogen bonds between the 5- and 4'-hydroxy groups and the side-chain of Arg38 and the backbone carbonyl of Phe308 are the key interactions for efKAS III inhibition. These flavanones are good candidate KAS III inhibitors and may be utilized as effective antimicrobials.

Virulence factors in vancomycin-resistant and vancomycin- susceptible Enterococcus faecalis from Brazil.

Enterococci are members of commensal flora of animals and insects, but are also important opportunistic pathogens. Our objective was to observe if there was any difference of virulence in several groups of E. faecalis, mainly between vancomycin-resistant E. faecalis (VREFS) of colonization and infection. VREFS and vancomycin-sensitive E. faecalis from Brazil were screened for the presence of virulence factor genes. Phenotypic assays were used to assess in vitro expression, to understand the pathogenic potential of these isolates and to determine whether a correlation exists between virulence and antibiotic resistance. Different virulence profiles were found suggesting that the disseminating clone may have generated several variations. However, our study showed that one constellation of traits appeared most commonly: gelatinase, aggregation substance and esp (GEA). These factors are important because they have been implicated in cell aggregation and biofilm formation. Biofilm formation may promote the conjugation of plasmids harboring resistance and virulence genes, enhancing the probability of entry of new resistance genes into species. Curiously, the profile GEA was not exclusive to VREFS, it was the second most observed in VSEFS isolates from colonization and infection in hospitalized patients and also from rectal swabs of healthy volunteers. Such strains appear to represent the entry gateway to new resistance genes into E. faecalis and may contribute to the spreading of E. faecalis mainly in hospitals.

ANTIBACTERIAL ACTIVITY OF THYMUS PUBESCENS METHANOLIC EXTRACT

During the last two decades, various medicinal plants have been studied for their possible antimicrobial activity to discover new antimicrobial agents capable of resolving problems such as the development of drug resistance in pathogenic microorganisms as well as the side effects of some present antibiotics. In this study, the antibacterial activity of methanolic extract of T. pubescens (rich in flavonoids) was investigated. The aerial parts of the plant were collected from Alvand mountainside (Hamadan, Iran) in May 2005 and identified by Hamadan Natural Sources Organization. The air-dried plant materials were ground to fine powder and then extracted by soxhelet apparatus using methanol. The extract was tested at a concentration of 100 mg/ml against a panel of Grampositive and Gram-negative bacteria using the disk diffusion technique. This methanolic extract demonstrated antibacterial activity against Gram-positive bacteria including Staphylococcus aureus, Methicillin-resistant S. aureus (MRSA), Streptococcus pyogenes, Enterococcus faecalis, Vancomycin-resistant E. faecalis (VRE) and Micrococcus luteus, and produced inhibition zones with 8-16 mm diameters. However, it showed no activity against Gram-negative bacteria, such as Escherichia coli, Pseudomonas aeruginosa and Salmonella spp. Minimum concentrations (MC) of the extract forming a clear zone were determined against susceptible bacteria. MC 90 of the extract against 10 MRSA strains tested was 1.56 mg/ml, indicating its good activity against this important nosocomial pathogen. T. pubescens seems to be a good candidate for further phythochemical studies in an attempt to find new chemical entities combating resistant bacteria.