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AIMS: Enterococcus faecalis (E. faecalis) is a leading cause of nosocomial infection and presents a wide spectrum of antibiotic resistance, being vancomycin-resistant Enterococcus (VRE) one of the most relevant. Synthetic antimicrobial peptides (SAMPs) are currently a promising option to overcome antimicrobial resistance. Thus, the purpose of this study was to assess the effect of eight SAMPs against vancomycin-resistant E. faecalis, as well as to investigate their mechanism of action and synergy with conventional antibiotics. METHODS AND RESULTS: Here, eight SAMPs, Mo-CBP3-PepI, Mo-CBP3-PepII, Mo-CBP3-PepIII, RcAlb-PepI, RcAlb-PepII, RcAlb-PepIII, PepGAT, and PepKAA, were tested for antibacterial activity in vitro against E. faecalis (ATCC® 51299) through broth microdilution. A maximum of 48% of E. faecalis growth inhibition was achieved by treatment with SAMPs alone. However, when these peptides were combined with the antibiotic chloramphenicol, assessed by checkerboard method, the inhibition increased to 55%-76% of inhibition, two to three-folds of increase if compared to the effects of the compounds alone. Microscopic analysis showed that E. faecalis cells treated with a combination of SAMPs and chloramphenicol resulted in bacterial membrane damage. The biofilm inhibition maximum was 22% for SAMPs alone, when combined with chloramphenicol, the maximum increased to 33%. CONCLUSIONS: SAMPs and their combination with chloramphenicol demonstrate antibacterial activity against E. faecalis, possibly by inducing bacterial membrane damage.
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Antibacterianos , Peptídeos Antimicrobianos , Cloranfenicol , Sinergismo Farmacológico , Enterococcus faecalis , Testes de Sensibilidade Microbiana , Enterococos Resistentes à Vancomicina , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/crescimento & desenvolvimento , Antibacterianos/farmacologia , Cloranfenicol/farmacologia , Peptídeos Antimicrobianos/farmacologia , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Vancomicina/farmacologiaRESUMO
Mangrove ecosystems are hotspots of biodiversity, but have been threatened by anthropogenic activities. Vancomycin-resistant enterococci (VRE) are nosocomial bacteria classified as high priority by the World Health Organization (WHO). Herein, we describe the identification and genomic characteristics of a vancomycin-resistant Enterococcus faecalis strain isolated from a highly impacted mangrove ecosystem of the northeastern Brazilian, in 2021. Genomic analysis confirmed the existence of the transposon Tn1546-vanA and clinically relevant antimicrobial resistance genes, such as streptogramins, tetracycline, phenicols, and fluoroquinolones. Virulome analysis identified several genes associated to adherence, immune modulation, biofilm, and exoenzymes production. The UFSEfl strain was assigned to sequence type (ST9), whereas phylogenomic analysis with publicly available genomes from a worldwide confirmed clonal relatedness with a hospital-associated Brazilian clone. Our findings highlight the successful expansion of hospital-associated VRE in a mangrove area and shed light on the need for strengthening genomic surveillance of WHO priority pathogens in these vital ecosystems.
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Ecossistema , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Enterococos Resistentes à Vancomicina , Humanos , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Brasil/epidemiologia , Células Clonais , Enterococcus faecalis/genética , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Testes de Sensibilidade Microbiana , Vancomicina , Resistência a Vancomicina/genética , Enterococos Resistentes à Vancomicina/genética , Infecção Hospitalar/microbiologiaRESUMO
This study describes the discovery of a variety of quinoline2-one derivatives with significant antibacterial action vs a spectrum of multidrug-resistant Gram-positive bacterial strains, especially methicillin-resistant Staphylococcus aureus (MRSA). Compounds 6c, 6l, and 6o exhibited significant antibacterial activity versus the Gram-positive bacterial pathogens evaluated. In comparison to the reference daptomycin, compound 6c demonstrated the most effective activity among the assessed derivatives, with MIC concentrations of 0.75 µg/mL versus MRSA and VRE and 2.50 µg/mL against MRSE. We also reported on these compounds' biofilm and dihydrofolate reductase inhibitory activities. Compound 6c showed the greatest antibiofilm action in a dose-dependent way and a substantial decrease of biofilm development in the MRSA ACL51 strain at concentrations of 0.5, 0.25, and 0.12 MIC, with reductions of 79%, 55%, and 38%, consecutively, whereas the corresponding values for vancomycin were 20%, 12%, and 9%. These findings imply that these quinoline compounds could be used to develop a new category of antibiotic representatives to prevent Gram-positive drug-resistant bacterial strains.
Assuntos
Daptomicina , Staphylococcus aureus Resistente à Meticilina , Quinolinas , Antibacterianos/farmacologia , Daptomicina/farmacologia , Vancomicina/farmacologia , Quinolinas/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
We analyzed the antimicrobial resistance (AMR) data of 6519 clinical isolates of Escherichia coli (n = 3985), Klebsiella pneumoniae (n = 775), Acinetobacter baumannii (n = 163), Pseudomonas aeruginosa (n = 781), Enterococcus faecium (n = 124), and Staphylococcus aureus (n = 691) from 43 centers in Mexico. AMR assays were performed using commercial microdilution systems (37/43) and the disk diffusion susceptibility method (6/43). The presence of carbapenemase-encoding genes was assessed using PCR. Data from centers regarding site of care, patient age, and clinical specimen were collected. According to the site of care, the highest AMR was observed in E. coli, K. pneumoniae, and P. aeruginosa isolates from ICU patients. In contrast, in A. baumannii, higher AMR was observed in isolates from hospitalized non-ICU patients. According to age group, the highest AMR was observed in the ≥60 years age group for E. coli, E. faecium, and S. aureus, and in the 19-59 years age group for A. baumannii and P. aeruginosa. According to clinical specimen type, a higher AMR was observed in E. coli, K. pneumoniae, and P. aeruginosa isolates from blood specimens. The most frequently detected carbapenemase-encoding gene in E. coli was blaNDM (84%).
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Enterococcus spp., including E. faecalis and E. faecium, pose risks to dairy farms as opportunistic pathogens. The study evaluates antimicrobial resistance (AMR) and virulence characteristics of Enterococcus spp. isolated from bovine milk. Bile esculin agar was used to assess 1471 milk samples, followed by colony identification, gram staining, catalase tests, and 45 °C incubation. PCR analysis targeted E. faecalis and E. faecium in characteristic Enterococcus spp. colonies, with MALDI-TOF used for negative samples. Multiple tests, including disk diffusion, chromogenic VRE agar for vancomycin resistance, Vancomycin Etest® for MIC determination, and PCR for virulence factors (cylA, esp, efaA, ace, asa1, gelE, and hyl genes), were performed. Out of 100 identified strains, E. durans (30.66%), E. faecium (26.28%), and E. faecalis (18.25%) were predominant. AMR in Enterococcus spp. varied, with the highest rates against rifampicin (27%), tetracycline (20%), and erythromycin (18%). Linezolid (5%), vancomycin, ciprofloxacin, and teicoplanin (3% each) had lower prevalence. E. faecium and E. faecalis showed high AMR to rifampicin, erythromycin, and tetracycline. Thirty-two strains (18.98%) grew on VRE Chromoselect agar, while 4 (2 E. faecalis and 2 E. faecium) showed vancomycin resistance by MIC values. E. faecalis carried gelE (45.5%) and asa1 (36%), and E. gallinarum had 9.1% with the asa1 gene. Detecting resistant Enterococcus in bovine milk supports control strategies for enterococci on dairy farms, highlighting AMR concerns in the food chain.
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The emergence of vancomycin-resistant Enterococcus faecium (Efm) harboring vanA gene and multidrug-resistant determinants is a relevant public health concern. It is an opportunistic pathogen responsible for nosocomial infections widely distributed in the environment, including wastewater treatment plants (WWTPs). Our study addresses a genomic investigation of vanA-carrying Efm from WWTPs in Brazil. Samples from five WWTPs supplied with sewage from different sources were evaluated. Here we present whole-genome sequencing of eight vanA-Efm isolates performed on Illumina MiSeq platform. All these isolates presented multidrug-resistant profile, and five strains were from treated wastewater. Multiple antimicrobial resistance genes (ARGs) were found, such as aph(3')-IIIa, ant(6')-Ia, erm(B), and msrC, some of them being allocated in plasmids. The virulence profile was predominantly constituted by efaAfm and acm genes and all isolates, except for one, were predicted as human pathogens. Multilocus sequence typing analysis revealed a new allele and five different STs, three previously described (ST32, ST168, and ST253) and two novel ones (ST1893 and ST1894). Six strains belonged to CC17, often associated with hospital outbreaks. As far as our knowledge, no genomic studies of vanA-Efm recovered from WWTPs revealed isolates belonging to CC17 in Brazil. Therefore, our findings point to the environmental spread of Efm carrying multiple ARGs.
Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Purificação da Água , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Infecções por Bactérias Gram-Positivas/epidemiologia , Humanos , Resistência a Vancomicina/genéticaRESUMO
In the present scenario of a major demand for new compounds with antimicrobial activity, bacteriocin and bacteriocin-like inhibitory substances (BLIS) are promising tools against deteriorating and pathogenic microorganisms, thus having potential applications in both the food industry and infectious disease control. In the present report, we describe the genetic and phenotypic characteristics of BLIS produced by Enterococcus faecium E86, a strain previously isolated and sequenced by our group, focusing on the structural genes of two bacteriocins identified: enterocin TW21 and enterocin P. Transcription of all four genes associated with the biosynthesis and immunity of enterocin P and enterocin TW21 were confirmed by RT-PCR. However, Sanger sequencing confirmed a truncation of the structural gene of enterocin TW21 due to one base pair deletion (A/T). Thus, although E. faecium E86 was shown to carry two bacteriocinogenic gene clusters, only one cluster encodes a functional bacteriocin, enterocin P. Enterocin P was able to inhibit different strains of Listeria monocytogenes and vancomycin-resistant enterococci (both Enterococcus faecalis and Enterococcus faecium), showing intense bacteriolytic activity, in most cases.
Assuntos
Bacteriocinas , Enterococcus faecium , Listeria monocytogenes , Enterococos Resistentes à Vancomicina , Bacteriocinas/genética , Bacteriocinas/farmacologia , Enterococcus faecium/genética , Listeria monocytogenes/efeitos dos fármacos , Vancomicina , Enterococos Resistentes à Vancomicina/efeitos dos fármacosRESUMO
Aim: To evaluate the activity of (-)-camphene-based thiosemicarbazide (TSC) and 4-hydroxy-thiosemicarbazone (4-OH-TSZ), alone and in combination against Gram-positive. Material & methods: MIC were determined for Staphylococcus aureus, Enterococcus spp. reference strains and clinical isolates. Drug combination, time-kill and cytotoxicity assays were also performed. Results: TSC and 4-OH-TSZ demonstrated potent inhibitory activity against S. aureus and Enterococcus spp., including multidrug-resistant isolates (MIC ranging from 1.9 to 31.2 µg/ml), and were bactericidal for the reference strains of both Gram-positive tested. The derivatives proved to be selective for the bacteria and synergistic with oxacillin and vancomycin. Conclusion: (-)-Camphene-based derivatives can represent promising drug candidates against critical pathogens, such as S. aureus and Enterococcus spp., including MRSA and vancomycin resistance Enterococcus spp. isolates.
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Antibacterianos/farmacologia , Monoterpenos Bicíclicos/farmacologia , Enterococcus/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Tiossemicarbazonas/farmacologia , Antibacterianos/química , Farmacorresistência Bacteriana Múltipla , Enterococcus/crescimento & desenvolvimento , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/crescimento & desenvolvimento , Tiossemicarbazonas/química , Vancomicina/farmacologiaRESUMO
The study's aim was to analyze the population structure of enterococci causing human invasive infections in a medium-sized Argentinian Hospital coincidental with a 5 year-period of increased recovery of antibiotic resistant enterococci (2010-2014). Species identification (biochemical testing/MALDI-TOF-MS), antimicrobial susceptibility (disk-diffusion) and clonal relatedness (PFGE/MLST/BAPS) were determined according to standard guidelines. ß-lactamase production was determined by a nitrocefin test and confirmed by PCR/sequencing. The isolates were identified as Enterococcus faecalis and Enterococcus faecium at a 2:1 ratio. Most of the E. faecalis isolates, grouped in 25 PFGE-types (ST9/ST179/ST236/ST281/ST388/ST604/ST720), were resistant to high-levels (HLR) of gentamicin/streptomycin. A ST9 clone (bla+/HLR-gentamicin) was detected in patients of different wards during 2014. E. faecium isolates were grouped in 10 PFGE-types (ST25/ST18/ST19/ST52/ST792), with a low rate of ampicillin resistance. Five vancomycin-resistant E. faecium, three vanA (ST792/ST25) and two vanB (ST25) were detected. The ST25 clone carried either vanA or vanB. The recovery of a bla+-ST9-E. faecalis clone similar to that described in the late 1980s in Argentina suggests the possibility of a local hidden reservoir. These results reflect the relevance of local epidemiology in understanding the population structure of enterococci as well as the emergence and spread of antimicrobial resistance in predominant enterococcal clonal lineages.
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We aimed to investigate the occurrence of CRISPR elements in the genomes of vancomycin-resistant (VRE) and vancomycin-susceptible (VSE) enterococci and their association with the presence of antimicrobial resistance and virulence genes. We analyzed 180 isolates, including 91 VRE and 89 VSE. Isolates were identified by PCR or MALDI-TOF. Antimicrobial susceptibility and MICs for vancomycin were determined by the disk-diffusion method and E-test®, respectively. The presence of resistance and virulence genes, as well as CRISPR elements, was investigated by PCR. We identified 95 (53%) E. faecalis, 78 (43%) E. faecium, five (2.8%) E. gallinarum, and one (0.6% each) E. casseliflavus and E. durans. The highest and the lowest non-susceptibility frequencies were observed for erythromycin (n = 152; 84.4%) and fosfomycin (n = 5; 2.8%), respectively. Most erythromycin-resistant isolates had the erm(B) gene (106/152; 69.7%). Of 118 (65.6%) isolates with high-level resistance to aminoglycoside, 69 (58.5%) had at least one aminoglycoside resistance gene, mostly ant(6)-Ia and aac(6')-Ie + aph(2â³)-Ia. We found at least one virulence gene among 135 (75%) isolates, mostly gelE (79/180; 43.9%). Ninety-two (51.1%) isolates had at least one CRISPR element, especially CRISPR3 (62/92; 67.4%). CRISPR elements were more common among E. faecalis, in which we observed a relationship between the absence of CRISPR and the presence of the vanA resistance gene, and the hyl and esp virulence genes. Among VRE. faecium, a relationship was found between the absence of CRISPR and the hyl gene. In conclusion, we found evident associations between the lack of CRISPR elements with species, multidrug resistance, and major resistance- and virulence-associated genes.
Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Farmacorresistência Bacteriana , Microbiologia de Alimentos , Infecções por Bactérias Gram-Positivas/microbiologia , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Enterococos Resistentes à Vancomicina/genética , Proteínas de Bactérias/genética , Humanos , Testes de Sensibilidade Microbiana , Virulência , Fatores de Virulência/genéticaRESUMO
We report the occurrence and genomic features of multidrug-resistant vancomycin-resistant Enterococcus faecium vanA belonging to a novel sequence type (designated ST1336), carrying a Tn1546-like element, in marine brown mussels (Perna perna) from anthropogenically affected coastal waters of the Atlantic coast of Brazil, highlighting a potential source of dissemination for related ecosystems, with additional consequences for seafood safety and quality.
Assuntos
Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/isolamento & purificação , Perna (Organismo)/microbiologia , Resistência a Vancomicina/genética , Animais , Proteínas de Bactérias/genética , Brasil , Carbono-Oxigênio Ligases/genética , Elementos de DNA Transponíveis , Ecossistema , Enterococcus faecium/genética , Humanos , Testes de Sensibilidade MicrobianaRESUMO
OBJECTIVES: The aim of this study was to report the draft genome sequence of the bacteriocinogenic strain Enterococcus faecium E86. Bacteriocins are prokaryotic peptides or proteins with antimicrobial activity. The genome information may contribute to the identification of enterocins produced by this strain that exhibit inhibitory activity against the foodborne pathogen Listeria monocytogenes and vancomycin-resistant enterococci (VRE) involved in human infections, among other bacterial genera and species. METHODS: An Illumina MiSeq platform was used for genome sequencing. De novo assembly of 5 735 838 paired-end reads was done using the A5-miseq pipeline, yielding >300-fold average genome coverage. Genome annotation was performed by the RAST server, and mining of the bacteriocinogenic gene clusters was done using the BAGEL3 and antiSMASH v.4 platforms. RESULTS: The total scaffold size was determined to be 2 689 107 bp, approximately 2.7 Mbp, featuring a G + C content of 38.1%. The genome contains 2858 coding sequences and 74 RNA genes. Genome analyses revealed the presence of: 30 genes involved in drug resistance; 2 bacteriocinogenic gene clusters (for enterocin P and enterocin TW21); EntiTW21, a novel bacteriocin immunity protein and a novel multilocus sequence type (ST1500). CONCLUSION: This work highlights the potential biotechnological application of this strain for the production of enterocin P, a bacteriocin that can be employed in the food industry as a biopreservative against L. monocytogenes and as an alternative to classical antibiotics against VRE.
Assuntos
Peptídeos Catiônicos Antimicrobianos/biossíntese , Bacteriocinas/biossíntese , Enterococcus faecium/genética , Genoma Bacteriano , Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bacteriocinas/genética , Bacteriocinas/farmacologia , Farmacorresistência Bacteriana/genética , Humanos , Listeria monocytogenes/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Família Multigênica , Análise de Sequência de DNA , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Sequenciamento Completo do GenomaRESUMO
Antibiotic resistance is one of the greatest challenges in the health system nowadays, representing a serious problem for public health. Initially, antibiotic-resistant strains were restricted to the hospital environment, but now they can be found everywhere. Globalization, excessive use of antibiotics in animal husbandry and aquaculture, use of multiple broad-spectrum agents, and lack of good antimicrobial stewardship can be listed as the factors most responsible for the spread of antibiotic resistance. The increase in the prevalence of antibiotic-resistant pathogens implies having fewer antimicrobial agents to treat infections. The estimate is that by 2050, there will be no effective antibiotic available, if no new drug is developed or discovered. This raises the need to search for alternative methods of controlling antibiotic-resistant pathogens. Considering this problem, the objective of this review is to outline the most frequent antibiotic-resistant bacteria and describe the advantageous and limitations of alternative methods that have been proposed to control them.
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Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Animais , HumanosRESUMO
BACKGROUND: Vancomycin-resistant enterococci (VRE) are an important agent of colonization and infection in haematology patients. However, the role of virulence on VRE colonization and infection is controversial. AIM: To characterize the lineage, virulence and resistance profile of VRE infection and colonization isolates; as well as their impact on outcome of haematology patients using a regression logistic model. METHODS: Eighty-six isolates (80 Enterococcus faecium and six E. faecalis) from 76 patients were evaluated. Polymerase chain reaction for resistance and virulence genes, and pulsed-field gel electrophoresis and whole genome sequencing of the major clusters, were performed. Bivariate and multivariate analyses were carried out to evaluate the role of virulence genes on outcome. FINDINGS: All isolates harboured the vanA gene. Regarding the virulence genes, 96.5% of isolates were positive for esp, 69.8% for gelE and asa1 genes. VRE infection isolates were more virulent than colonization isolates and harboured more often the gelE gene (P = 0.008). Infections caused by VRE carrying asa1 gene resulted more frequently in death (P = 0.004), but only the predominant clone remained as protector in the multivariate model. The E. faecium strains were assigned to seven STs (ST78, ST412, ST478, ST792, ST896, ST987, ST963) that belonged to CC17. The E. faecalis sequenced belonged to ST9 (CC9). CONCLUSION: E. faecium was predominant, and infection isolates were more virulent than colonization isolates and harboured more often the gene gelE. Infections caused by VRE carrying the asa1 gene appeared to be associated with a fatal outcome.
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Enterococcus faecalis/isolamento & purificação , Enterococcus faecium/isolamento & purificação , Infecções por Bactérias Gram-Positivas/epidemiologia , Doenças Hematológicas/complicações , Enterococos Resistentes à Vancomicina/isolamento & purificação , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Eletroforese em Gel de Campo Pulsado , Enterococcus faecalis/classificação , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/genética , Enterococcus faecium/classificação , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/genética , Feminino , Genes Bacterianos , Genótipo , Infecções por Bactérias Gram-Positivas/microbiologia , Infecções por Bactérias Gram-Positivas/mortalidade , Humanos , Lactente , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Epidemiologia Molecular , Reação em Cadeia da Polimerase , Prevalência , Estudos Retrospectivos , Análise de Sobrevida , Enterococos Resistentes à Vancomicina/classificação , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Enterococos Resistentes à Vancomicina/genética , Fatores de Virulência/análise , Fatores de Virulência/genética , Sequenciamento Completo do Genoma , Adulto JovemRESUMO
ABSTRACT Vancomycin-resistant Enterococcus faecium (VREfm) has emerged as an important global nosocomial pathogen, and this trend is associated with the spread of high-risk clones. Here, we determined the genetic and phenotypic features of 93 VREfm isolates that were obtained from patients in 13 hospitals in Vitória, Espírito Santo, Brazil, during 2012-2013. All the isolates were vancomycin-resistant and harbored the vanA gene. Only 6 (6.5%) of the VREfm isolates showed the ability to form biofilm. The 93 isolates analyzed belong to a single pulsed-field gel electrophoresis lineage and presented six subtypes. MLST genotyping showed that all VREfm belonged to ST412 (the high-risk clone, hospital-adapted). The present study describes the dissemination of ST412 clone in the local hospitals. The clonal spread of these ST412 isolates in the area we analyzed as well as other hospitals in southeastern Brazil supports the importance of identifying and controlling the presence of these microorganisms in health care-related services.
Assuntos
Humanos , Infecção Hospitalar/microbiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Enterococcus faecium/genética , Enterococos Resistentes à Vancomicina/genética , Proteínas de Bactérias , Brasil , Testes de Sensibilidade Microbiana , Técnicas de Tipagem Bacteriana , Enterococcus faecium/efeitos dos fármacos , Eletroforese em Gel de Campo Pulsado , Tipagem de Sequências Multilocus , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Antibacterianos/farmacologiaRESUMO
Gram-positive pathogens such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE) have been frequently associated with bacterial resistance mechanisms. These mechanisms, in turn, restrict a range of therapeutic opportunities for the treatment of infections caused by these micro-organisms. Faced with this problem, the present study aims to isolate and characterize molecules with antimicrobial activity derived from the fungus Penicillium citrinum isolated from Cerrado soil. Furthermore, we also tested possible antibacterial potential alone and in combination with commercial antimicrobial agents. In this context, citrinin was isolated and characterized by nuclear magnetic resonance and electrospray ionization. Functional analyses showed MIC of 128 µg ml-1 against S. aureus ATCC 25923, E. faecalis ATCC 29212 and a clinical isolate of vancomycin-resistant E. faecium (VRE01). However, for a clinical strain of methicillin-resistant S. aureus (MRSA01), the MIC was 256 µg ml-1. In order to avoid such high concentrations and reduce the collateral effects, additive effects were evidenced by combining citrinin with cefoxitin against MRSA01 (FIC index=0.5) and also citrinin with vancomycin toward VRE01 (FIC index=0.5). In vivo studies with BALB/c-tipe mice (MRSA assay) demonstrated a clinical ineffectiveness of cefoxitin associated with citrinin (9.8 mg kg-1 of cefoxitin +0.2 mg kg-1 of citrinin), with this combination being inefficient to increase animal survival. However, the combination used in the treatment of VRE (23.5 mg kg-1 of citrinin +1.5 mg kg-1 of vancomycin) sepsis model was extremely promising, leading to an animal survival rate of 80 percent. In summary, our data show, for the first time, the possible successful use of citrinin associated with vancomycin for pathogenic bacteria control.
Assuntos
Antibacterianos/farmacologia , Citrinina/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Penicillium/química , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Vancomicina/farmacologia , Animais , Antibacterianos/isolamento & purificação , Antibacterianos/toxicidade , Cefoxitina/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Citrinina/isolamento & purificação , Citrinina/toxicidade , Sinergismo Farmacológico , Feminino , Dose Letal Mediana , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Penicillium/citologia , Células RAW 264.7 , Testes de ToxicidadeRESUMO
The aim of this study was to determine the association between Clostridium difficile (C. difficile) and vancomycin-resistant Enterococcus (VRE) and efficacy of screening stools submitted for C. difficile toxin assay for prevalence of VRE. Between April 2012 and February 2014, 158 stool samples submitted for C. difficile toxin to the Marmara University Microbiology Laboratory, were included in the study. Stool samples were analyzed by enzyme immuno assay test; VIDAS (bioMerieux, France) for Toxin A&B. Samples were inoculated on chromID VRE (bioMerieux, France) and incubated 24 h at 37 °C. Manuel tests and API20 STREP (bioMerieux, France) test were used to identify the Enterococci species. After the species identification, vancomycin and teicoplanin MIC's were performed by E test and molecular resistance genes for vanA vs vanB were detected by polymerase chain reaction (PCR). Of the 158 stool samples, 88 were toxin positive. The prevalence of VRE was 17%(n:19) in toxin positives however, 11.4% in toxin negatives(n:70). All VRE isolates were identified as Enterococcus faecium. These results were evaluated according to Fischer's exact chi-square test and p value between VRE colonization and C. difficile toxin positivity was detected 0.047 (p 0.05). PPV and NPV were 79% and 47% respectively. In our study, the presence of VRE in C. difficile toxin positives is statistically significant compared with toxin negatives (p 0.05). Screening for VRE is both additional cost and work load for the laboratories. Therefore VRE screening among C. difficile toxin positive samples, will be cost effective for determination of high risk patients in the hospitals especially for developing countries.(AU)
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Clostridioides difficile , Enterococos Resistentes à Vancomicina , Técnicas Imunoenzimáticas , Infecção HospitalarRESUMO
Vancomycin-resistant Enterococcus faecium (VREfm) has emerged as an important global nosocomial pathogen, and this trend is associated with the spread of high-risk clones. Here, we determined the genetic and phenotypic features of 93 VREfm isolates that were obtained from patients in 13 hospitals in Vitória, Espírito Santo, Brazil, during 2012-2013. All the isolates were vancomycin-resistant and harbored the vanA gene. Only 6 (6.5%) of the VREfm isolates showed the ability to form biofilm. The 93 isolates analyzed belong to a single pulsed-field gel electrophoresis lineage and presented six subtypes. MLST genotyping showed that all VREfm belonged to ST412 (the high-risk clone, hospital-adapted). The present study describes the dissemination of ST412 clone in the local hospitals. The clonal spread of these ST412 isolates in the area we analyzed as well as other hospitals in southeastern Brazil supports the importance of identifying and controlling the presence of these microorganisms in health care-related services.
Assuntos
Infecção Hospitalar/microbiologia , Enterococcus faecium/genética , Infecções por Bactérias Gram-Positivas/microbiologia , Enterococos Resistentes à Vancomicina/genética , Antibacterianos/farmacologia , Proteínas de Bactérias , Técnicas de Tipagem Bacteriana , Brasil , Eletroforese em Gel de Campo Pulsado , Enterococcus faecium/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Enterococos Resistentes à Vancomicina/efeitos dos fármacosRESUMO
Abstract The aim of this study was to determine the association between Clostridium difficile (C. difficile) and vancomycin-resistant Enterococcus (VRE) and efficacy of screening stools submitted for C. difficile toxin assay for prevalence of VRE. Between April 2012 and February 2014, 158 stool samples submitted for C. difficile toxin to the Marmara University Microbiology Laboratory, were included in the study. Stool samples were analyzed by enzyme immuno assay test; VIDAS (bioMerieux, France) for Toxin A&B. Samples were inoculated on chromID VRE (bioMerieux, France) and incubated 24 h at 37 °C. Manuel tests and API20 STREP (bioMerieux, France) test were used to identify the Enterococci species. After the species identification, vancomycin and teicoplanin MIC's were performed by E test and molecular resistance genes for vanA vs vanB were detected by polymerase chain reaction (PCR). Of the 158 stool samples, 88 were toxin positive. The prevalence of VRE was 17%(n:19) in toxin positives however, 11.4% in toxin negatives(n:70). All VRE isolates were identified as Enterococcus faecium. These results were evaluated according to Fischer's exact chi-square test and p value between VRE colonization and C. difficile toxin positivity was detected 0.047 (p < 0.05). PPV and NPV were 79% and 47% respectively. In our study, the presence of VRE in C. difficile toxin positives is statistically significant compared with toxin negatives (p < 0.05). Screening for VRE is both additional cost and work load for the laboratories. Therefore VRE screening among C. difficile toxin positive samples, will be cost effective for determination of high risk patients in the hospitals especially for developing countries.