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1.
BMC Med Genomics ; 17(1): 174, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38951840

RESUMO

BACKGROUND: This study investigates the distribution and characteristics of linezolid and vancomycin susceptibilities among Enterococcus faecalis (E. faecalis) and Enterococcus faecium (E. faecium) and explores the underlying resistance mechanisms. METHODS: A total of 2842 Enterococcus clinical isolates from patients were retrospectively collected, and their clinical data were further analyzed. The minimum inhibitory concentrations (MICs) of vancomycin and linezolid were validated by broth dilution method. The resistance genes optrA, cfr, vanA, vanB and vanM were investigated using polymerase chain reaction (PCR). Housekeeping genes and resistance genes were obtianed through whole-genome sequencing (WGS). RESULTS: Of the 2842 Enterococcus isolates, 88.5% (2516) originated from urine, with E. faecium accounted for 60.1% of these. The vanA gene was identified in 27/28 vancomycin resistant Enterococcus (VRE) isolates, 4 of which carried both vanA and vanM genes. The remaining strain was vanM positive. The optrA gene was identified in all E. faecalis isolates among linezolid resistant Enterococcus (LRE). E. faecium showed a higher multiple antibiotic resistance index (MAR index) compared to E. faecalis. The multi-locus sequence typing (MLST) showed the sequence type of E. faecium mainly belongs to clonal complex (CC) 17, nearly E. faecalis isolates analyzed were differentiated into 7 characteristics of sequence types (STs), among which ST16 of CC16 were the major lineage. CONCLUSION: Urine was the primary source of VRE and LRE isolates in this study. E. faecium showed higher levels of resistance compared to E. faecalis. OptrA gene was detected in 91.6% of LRE, which could explain linezolid resistance, and van genes were detected in all vancomycin resistant Enterococcus strains, while vanA was a key resistance mechanism in VRE identified in this study.


Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Linezolida , Testes de Sensibilidade Microbiana , Linezolida/farmacologia , Humanos , China/epidemiologia , Enterococcus faecium/genética , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/isolamento & purificação , Infecções por Bactérias Gram-Positivas/microbiologia , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Enterococcus faecalis/genética , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/isolamento & purificação , Feminino , Vancomicina/farmacologia , Antibacterianos/farmacologia , Epidemiologia Molecular , Adulto , Resistência a Vancomicina/genética , Idoso , Estudos Retrospectivos , Enterococos Resistentes à Vancomicina/genética , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Enterococos Resistentes à Vancomicina/isolamento & purificação , Adulto Jovem , Enterococcus/genética , Enterococcus/efeitos dos fármacos , Enterococcus/isolamento & purificação
2.
BMC Biotechnol ; 24(1): 47, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38978013

RESUMO

The threat of methicillin-resistant Staphylococcus aureus (MRSA) is increasing worldwide, making it significantly necessary to discover a novel way of dealing with related infections. The quick spread of MRSA isolates among infected individuals has heightened public health concerns and significantly limited treatment options. Vancomycin (VAN) can be applied to treat severe MRSA infections, and the indiscriminate administration of this antimicrobial agent has caused several concerns in medical settings. Owing to several advantageous characteristics, a niosomal drug delivery system may increase the potential of loaded antimicrobial agents. This work aims to examine the antibacterial and anti-biofilm properties of VAN-niosome against MRSA clinical isolates with emphasis on cytotoxicity and stability studies. Furthermore, we aim to suggest an effective approach against MRSA infections by investigating the inhibitory effect of formulated niosome on the expression of the biofilm-associated gene (icaR). The thin-film hydration approach was used to prepare the niosome (Tween 60, Span 60, and cholesterol), and field emission scanning electron microscopy (FE-SEM), an in vitro drug release, dynamic light scattering (DLS), and entrapment efficiency (EE%) were used to investigate the physicochemical properties. The physical stability of VAN-niosome, including hydrodynamic size, polydispersity index (PDI), and EE%, was analyzed for a 30-day storage time at 4 °C and 25 °C. In addition, the human foreskin fibroblast (HFF) cell line was used to evaluate the cytotoxic effect of synthesized niosome. Moreover, minimum inhibitory and bactericidal concentrations (MICs/MBCs) were applied to assess the antibacterial properties of niosomal VAN formulation. Also, the antibiofilm potential of VAN-niosome was investigated by microtiter plate (MTP) and real-time PCR methods. The FE-SEM result revealed that synthesized VAN-niosome had a spherical morphology. The hydrodynamic size and PDI of VAN-niosome reported by the DLS method were 201.2 nm and 0.301, respectively. Also, the surface zeta charge of the prepared niosome was - 35.4 mV, and the EE% ranged between 58.9 and 62.5%. Moreover, in vitro release study revealed a sustained-release profile for synthesized niosomal formulation. Our study showed that VAN-niosome had acceptable stability during a 30-day storage time. Additionally, the VAN-niosome had stronger antibacterial and anti-biofilm properties against MRSA clinical isolates compared with free VAN. In conclusion, the result of our study demonstrated that niosomal VAN could be promising as a successful drug delivery system due to sustained drug release, negligible toxicity, and high encapsulation capacity. Also, the antibacterial and anti-biofilm studies showed the high capacity of VAN-niosome against MRSA clinical isolates. Furthermore, the results of real-time PCR exhibited that VAN-niosome could be proposed as a powerful strategy against MRSA biofilm via down-regulation of icaR gene expression.


Assuntos
Antibacterianos , Biofilmes , Sistemas de Liberação de Medicamentos , Lipossomos , Staphylococcus aureus Resistente à Meticilina , Vancomicina , Biofilmes/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/fisiologia , Vancomicina/farmacologia , Vancomicina/química , Antibacterianos/farmacologia , Antibacterianos/química , Lipossomos/química , Humanos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Liberação Controlada de Fármacos
3.
Ann Clin Microbiol Antimicrob ; 23(1): 62, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38978096

RESUMO

BACKGROUND: This study analyzed the genetic traits and fitness costs of vancomycin-resistant Enterococcus faecium (VREfm) blood isolates carrying Tn1546-type transposons harboring the vanA operon. METHODS: All E. faecium blood isolates were collected from eight general hospitals in South Korea during one-year study period. Antimicrobial susceptibility testing and vanA and vanB PCR were performed. Growth rates of E. faecium isolates were determined. The vanA-positive isolates were subjected to whole genome sequencing and conjugation experiments. RESULTS: Among 308 E. faecium isolates, 132 (42.9%) were positive for vanA. All Tn1546-type transposons harboring the vanA operon located on the plasmids, but on the chromosome in seven isolates. The plasmids harboring the vanA operon were grouped into four types; two types of circular, nonconjugative plasmids (Type A, n = 50; Type B, n = 46), and two types of putative linear, conjugative plasmids (Type C, n = 16; Type D, n = 5). Growth rates of vanA-positive E. faecium isolates were significantly lower than those of vanA-negative isolates (P < 0.001), and reduction in growth rate under vancomycin pressure was significantly larger in isolates harboring putative linear plasmids than in those harboring circular plasmids (P = 0.020). CONCLUSIONS: The possession of vanA operon was costly to bacterial hosts in antimicrobial-free environment, which provide evidence for the importance of reducing vancomycin pressure for prevention of VREfm dissemination. Fitness burden to bacterial hosts was varied by type and size of the vanA operon-harboring plasmid.


Assuntos
Antibacterianos , Proteínas de Bactérias , Carbono-Oxigênio Ligases , Elementos de DNA Transponíveis , Enterococcus faecium , Testes de Sensibilidade Microbiana , Óperon , Plasmídeos , Plasmídeos/genética , Enterococcus faecium/genética , Humanos , Proteínas de Bactérias/genética , República da Coreia , Carbono-Oxigênio Ligases/genética , Antibacterianos/farmacologia , Sequenciamento Completo do Genoma , Infecções por Bactérias Gram-Positivas/microbiologia , Enterococos Resistentes à Vancomicina/genética , Resistência a Vancomicina/genética , Aptidão Genética , Vancomicina/farmacologia , Conjugação Genética
4.
Arch Microbiol ; 206(7): 304, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38878097

RESUMO

The extension of multidrug-resistant strains of Staphylococcus aureus (S. aureus) is one of the main health challenges in the world, which requires serious solutions to deal with it. Combination therapies using conventional antibiotics and new antibacterial compounds that target different bacterial pathways are effective methods against resistant bacterial infections. Gallium is an iron-like metal that competes with iron for uptake into bacteria and has the potential to disrupt iron-dependent vital processes in bacteria. In this study, we explored the antibacterial effects of gallium nitrate (Ga(NO3)3) and vancomycin alone and in combination with each other on methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) using microdilution assay and checkerboard test, respectively. Then, their effect on the formation and destruction of biofilms was investigated. Finally, the amount of ROS production in the presence of these two compounds in bacteria was evaluated. The results indicated that the vancomycin/ Ga(NO3)3 combination reduced the MIC of vancomycin in the MRSA strain and had an additive effect on it. Vancomycin plus Ga(NO3)3 reduced the formation of biofilms and increased the destruction of biofilms formed in both strains, especially in the MRSA strain. ROS production was also higher in the combination of vancomycin with Ga(NO3)3 compared to vancomycin alone, especially in MRSA. Therefore, our results showed that Ga(NO3)3 enhances the antibacterial activity of vancomycin and this combination therapy can be considered as a new strategy for the treatment of MRSA infections.


Assuntos
Antibacterianos , Biofilmes , Gálio , Staphylococcus aureus Resistente à Meticilina , Testes de Sensibilidade Microbiana , Staphylococcus aureus , Vancomicina , Gálio/farmacologia , Vancomicina/farmacologia , Antibacterianos/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Sinergismo Farmacológico , Espécies Reativas de Oxigênio/metabolismo , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Humanos
5.
Front Cell Infect Microbiol ; 14: 1403289, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38915921

RESUMO

Staphylococcus aureus is a major causative pathogen of osteomyelitis. Intracellular infections of resident bone cells including osteocytes can persist despite gold-standard clinical intervention. The mechanisms by which intracellular S. aureus evades antibiotic therapy are unknown. In this study, we utilised an in vitro S. aureus infection model of human osteocytes to investigate whether antibiotic-mediated dysregulation of autophagy contributes to this phenomenon. Infected or non-infected osteocyte-like cells were exposed to combinations of rifampicin, vancomycin, and modulators of autophagy. Intracellular bacterial growth characteristics were assessed using colony-forming unit (CFU) analysis, viable bacterial DNA abundance, and the rate of escape into antibiotic-free medium, together with measures of autophagic flux. Rifampicin, alone or in combination with vancomycin, caused a rapid decrease in the culturability of intracellular bacteria, concomitant with stable or increased absolute bacterial DNA levels. Both antibiotics significantly inhibited autophagic flux. However, modulation of autophagic flux did not affect viable bacterial DNA levels. In summary, autophagy was shown to be a factor in the host-pathogen relationship in this model, as its modulation affected the growth state of intracellular S. aureus with respect to both their culturability and propensity to escape the intracellular niche. While rifampicin and vancomycin treatments moderately suppressed autophagic flux acutely, this did not explain the paradoxical response of antibiotic treatment in decreasing S. aureus culturability whilst failing to clear bacterial DNA and hence intracellular bacterial load. Thus, off-target effects of rifampicin and vancomycin on autophagic flux in osteocyte-like cells could not explain the persistent S. aureus infection in these cells.


Assuntos
Antibacterianos , Autofagia , Osteócitos , Rifampina , Infecções Estafilocócicas , Staphylococcus aureus , Vancomicina , Autofagia/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia , Osteócitos/efeitos dos fármacos , Osteócitos/microbiologia , Antibacterianos/farmacologia , Humanos , Vancomicina/farmacologia , Rifampina/farmacologia , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/tratamento farmacológico , Interações Hospedeiro-Patógeno , DNA Bacteriano/genética
6.
Int J Med Microbiol ; 315: 151624, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38838390

RESUMO

Staphylococcus aureus is a notorious pathogen responsible for various severe diseases. Due to the emergence of drug-resistant strains, the prevention and treatment of S. aureus infections have become increasingly challenging. Vancomycin is considered to be one of the last-resort drugs for treating most methicillin-resistant S. aureus (MRSA), so it is of great significance to further reveal the mechanism of vancomycin resistance. VraFG is one of the few important ABC (ATP-binding cassette) transporters in S. aureus that can form TCS (two-component systems)/ABC transporter modules. ABC transporters can couple the energy released from ATP hydrolysis to translocate solutes across the cell membrane. In this study, we obtained a strain with decreased vancomycin susceptibility after serial passaging and selection. Subsequently, whole-genome sequencing was performed on this laboratory-derived strain MWA2 and a novel single point mutation was discovered in vraF gene, leading to decreased sensitivity to vancomycin and daptomycin. Furthermore, the mutation reduces autolysis of S. aureus and downregulates the expression of lytM, isaA, and atlA. Additionally, we observed that the mutant has a less net negative surface charge than wild-type strain. We also noted an increase in the expression of the dlt operon and mprF gene, which are associated with cell surface charge and serve to hinder the binding of cationic peptides by promoting electrostatic repulsion. Moreover, this mutation has been shown to enhance hemolytic activity, expand subcutaneous abscesses, reflecting an increased virulence. This study confirms the impact of a point mutation of VraF on S. aureus antibiotic resistance and virulence, contributing to a broader understanding of ABC transporter function and providing new targets for treating S. aureus infections.


Assuntos
Transportadores de Cassetes de Ligação de ATP , Antibacterianos , Proteínas de Bactérias , Infecções Estafilocócicas , Staphylococcus aureus , Vancomicina , Virulência/genética , Infecções Estafilocócicas/microbiologia , Antibacterianos/farmacologia , Vancomicina/farmacologia , Animais , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Staphylococcus aureus/genética , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/patogenicidade , Staphylococcus aureus/metabolismo , Testes de Sensibilidade Microbiana , Resistência a Vancomicina/genética , Sequenciamento Completo do Genoma , Daptomicina/farmacologia , Camundongos , Autólise , Humanos , Mutação Puntual , Mutação , Feminino
7.
Euro Surveill ; 29(23)2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38847117

RESUMO

BackgroundVancomycin-resistant enterococci (VRE) are increasing in Denmark and Europe. Linezolid and vancomycin-resistant enterococci (LVRE) are of concern, as treatment options are limited. Vancomycin-variable enterococci (VVE) harbour the vanA gene complex but are phenotypically vancomycin-susceptible.AimThe aim was to describe clonal shifts for VRE and VVE in Denmark between 2015 and 2022 and to investigate genotypic linezolid resistance among the VRE and VVE.MethodsFrom 2015 to 2022, 4,090 Danish clinical VRE and VVE isolates were whole genome sequenced. We extracted vancomycin resistance genes and sequence types (STs) from the sequencing data and performed core genome multilocus sequence typing (cgMLST) analysis for Enterococcus faecium. All isolates were tested for the presence of mutations or genes encoding linezolid resistance.ResultsIn total 99% of the VRE and VVE isolates were E. faecium. From 2015 through 2019, 91.1% of the VRE and VVE were vanA E. faecium. During 2020, to the number of vanB E. faecium increased to 254 of 509 VRE and VVE isolates. Between 2015 and 2022, seven E. faecium clusters dominated: ST80-CT14 vanA, ST117-CT24 vanA, ST203-CT859 vanA, ST1421-CT1134 vanA (VVE cluster), ST80-CT1064 vanA/vanB, ST117-CT36 vanB and ST80-CT2406 vanB. We detected 35 linezolid vancomycin-resistant E. faecium and eight linezolid-resistant VVEfm.ConclusionFrom 2015 to 2022, the numbers of VRE and VVE increased. The spread of the VVE cluster ST1421-CT1134 vanA E. faecium in Denmark is a concern, especially since VVE diagnostics are challenging. The finding of LVRE, although in small numbers, ia also a concern, as treatment options are limited.


Assuntos
Antibacterianos , Proteínas de Bactérias , Carbono-Oxigênio Ligases , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Linezolida , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Resistência a Vancomicina , Enterococos Resistentes à Vancomicina , Enterococos Resistentes à Vancomicina/genética , Enterococos Resistentes à Vancomicina/isolamento & purificação , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Enterococcus faecium/genética , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/isolamento & purificação , Humanos , Dinamarca/epidemiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Carbono-Oxigênio Ligases/genética , Linezolida/farmacologia , Resistência a Vancomicina/genética , Sequenciamento Completo do Genoma , Vancomicina/farmacologia , Vancomicina/uso terapêutico , Genótipo
8.
Antimicrob Agents Chemother ; 68(7): e0037224, 2024 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-38884456

RESUMO

Peptidoglycan (PG) is an important architectural element that imparts physical toughness and rigidity to the bacterial envelope. It is also a dynamic structure that undergoes continuous turnover or autolysis. Escherichia coli possesses redundant PG degradation enzymes responsible for PG turnover; however, the advantage afforded by the existence of numerous PG degradation enzymes remains incompletely understood. In this study, we elucidated the physiological roles of MltE and MltC, members of the lytic transglycosylase (LTG) family that catalyze the cleavage of glycosidic bonds between disaccharide subunits within PG strands. MltE and MltC are acidic LTGs that exhibit increased enzymatic activity and protein levels under acidic pH conditions, respectively, and deletion of these two LTGs results in a pronounced growth defect at acidic pH. Furthermore, inactivation of these two LTGs induces increased susceptibility at acidic pH against various antibiotics, particularly vancomycin, which seems to be partially caused by elevated membrane permeability. Intriguingly, inactivation of these LTGs induces a chaining morphology, indicative of daughter cell separation defects, only under acidic pH conditions. Simultaneous deletion of PG amidases, known contributors to daughter cell separation, exacerbates the chaining phenotype at acidic pH. This suggests that the two LTGs may participate in the cleavage of glycan strands between daughter cells under acidic pH conditions. Collectively, our findings highlight the role of LTG repertoire diversity in facilitating bacterial survival and antibiotic resistance under stressful conditions.


Assuntos
Antibacterianos , Proteínas de Escherichia coli , Escherichia coli , Glicosiltransferases , Peptidoglicano , Escherichia coli/genética , Escherichia coli/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Antibacterianos/farmacologia , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Peptidoglicano/metabolismo , Testes de Sensibilidade Microbiana , Vancomicina/farmacologia , Farmacorresistência Bacteriana/genética , Parede Celular/metabolismo , Parede Celular/efeitos dos fármacos , Estresse Fisiológico , Peptidoglicano Glicosiltransferase/genética , Peptidoglicano Glicosiltransferase/metabolismo
9.
Antimicrob Agents Chemother ; 68(7): e0056124, 2024 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-38899926

RESUMO

Staphylococcus aureus is a pathogenic bacterium responsible for a broad spectrum of infections, including cutaneous, respiratory, osteoarticular, and systemic infections. It poses a significant clinical challenge due to its ability to develop antibiotic resistance. This resistance limits therapeutic options, increases the risk of severe complications, and underscores the urgent need for new strategies to address this threat, including the investigation of treatments complementary to antibiotics. The evaluation of novel antimicrobial agents often employs animal models, with the zebrafish embryo model being particularly interesting for studying host-pathogen interactions, establishing itself as a crucial tool in this field. For the first time, this study presents a zebrafish embryo model for the in vivo assessment of bacteriophage efficacy against S. aureus infection. A localized infection was induced by microinjecting either methicillin-resistant S. aureus (MRSA) or methicillin-susceptible S. aureus (MSSA). Subsequent treatments involved administering either bacteriophage, vancomycin (the reference antibiotic for MRSA), or a combination of both via the same route to explore potential synergistic effects. Our findings indicate that the bacteriophage was as effective as vancomycin in enhancing survival rates, whether used alone or in combination. Moreover, bacteriophage treatment appears to be even more effective in reducing the bacterial load in S. aureus-infected embryos post-treatment than the antibiotic. Our study validates the use of the zebrafish embryo model and highlights its potential as a valuable tool in assessing bacteriophage efficacy treatments in vivo.


Assuntos
Antibacterianos , Staphylococcus aureus Resistente à Meticilina , Terapia por Fagos , Infecções Estafilocócicas , Vancomicina , Peixe-Zebra , Animais , Peixe-Zebra/microbiologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/virologia , Terapia por Fagos/métodos , Vancomicina/farmacologia , Vancomicina/uso terapêutico , Infecções Estafilocócicas/terapia , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/tratamento farmacológico , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Modelos Animais de Doenças , Embrião não Mamífero/microbiologia , Testes de Sensibilidade Microbiana
10.
Biomacromolecules ; 25(7): 4156-4167, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38922325

RESUMO

Polymeric drugs containing up to 60% by weight of the antibiotic vancomycin were synthesized based on dextran carriers activated with epichlorohydrin. Vancomycin was covalently bound, involving the primary amino group of the molecule through the hydroxypropyl radical to the C6 position of the anhydroglucose units of the dextran main chain. Covalent binding is necessary to prevent spontaneous release of the antibiotic from the gel, thereby reducing the risk of bacterial multiresistance. Antibacterial depot gels were obtained from those polymers, containing up to 17.5% by weight of polysaccharide with a cross-linking density of q = 3-5 nodes per macromolecule for the deposition of another type of drugs not covalently bound to the polymer gel. They were used to coat the surface of the internal pores of biocomposite bone implants based on bovine cancellous bone used in orthopedics. The chemical structure of the polymer was studied using 13C NMR spectroscopy and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. The stiffness of the gels was evaluated by the values of the accumulation modulus G' = 170-270 kPa and the loss modulus G″ = 3.7-4.2 kPa determined on a rheometer. Their values are close to those typical for materials used to replace soft tissue in plastic surgery. The minimum inhibitory concentration of the gels against Staphylococcus aureus P209 depends on the antibiotic content in the polymer. It equals 2.5 mg/L for vancomycin we used and 100 mg/L for a polymer containing 50% by weight of covalently bound antibiotic. The cytotoxic concentration measured with cell culture HEK 293T exceeds 1200 mg/L in 24 h exposure. The release dynamics of drugs not covalently bound to dextran from the depot gel were studied using fluorescein as a model. The release time is independent of the gel density and lasts up to 6 days for a 2 mm thick layer. Both the gel and the bone implants impregnated with it maintained consistently high antibacterial activity throughout the experiment, up to its completion after 168 h, with the local concentration of the released antibiotic at the site of bacterial attack exceeding the therapeutic level by 200 times.


Assuntos
Antibacterianos , Géis , Vancomicina , Vancomicina/farmacologia , Vancomicina/química , Antibacterianos/farmacologia , Antibacterianos/química , Humanos , Géis/química , Animais , Staphylococcus aureus/efeitos dos fármacos , Bovinos , Dextranos/química , Dextranos/farmacologia , Células HEK293 , Testes de Sensibilidade Microbiana , Próteses e Implantes
11.
Nat Chem Biol ; 20(7): 924-933, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38942968

RESUMO

Keratinicyclins and keratinimicins are recently discovered glycopeptide antibiotics. Keratinimicins show broad-spectrum activity against Gram-positive bacteria, while keratinicyclins form a new chemotype by virtue of an unusual oxazolidinone moiety and exhibit specific antibiosis against Clostridioides difficile. Here we report the mechanism of action of keratinicyclin B (KCB). We find that steric constraints preclude KCB from binding peptidoglycan termini. Instead, KCB inhibits C. difficile growth by binding wall teichoic acids (WTAs) and interfering with cell wall remodeling. A computational model, guided by biochemical studies, provides an image of the interaction of KCB with C. difficile WTAs and shows that the same H-bonding framework used by glycopeptide antibiotics to bind peptidoglycan termini is used by KCB for interacting with WTAs. Analysis of KCB in combination with vancomycin (VAN) shows highly synergistic and specific antimicrobial activity, and that nanomolar combinations of the two drugs are sufficient for complete growth inhibition of C. difficile, while leaving common commensal strains unaffected.


Assuntos
Antibacterianos , Clostridioides difficile , Testes de Sensibilidade Microbiana , Clostridioides difficile/efeitos dos fármacos , Antibacterianos/farmacologia , Antibacterianos/química , Vancomicina/farmacologia , Vancomicina/química , Parede Celular/efeitos dos fármacos , Parede Celular/metabolismo , Ácidos Teicoicos/metabolismo , Peptidoglicano/metabolismo , Peptidoglicano/química , Quimioterapia Combinada , Peptídeos Cíclicos , Lipopeptídeos
12.
Int J Nanomedicine ; 19: 3991-4005, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38720939

RESUMO

Purpose: Surgical site infections pose a significant challenge for medical services. Systemic antibiotics may be insufficient in preventing bacterial biofilm development. With the local administration of antibiotics, it is easier to minimize possible complications, achieve drugs' higher concentration at the injured site, as well as provide their more sustained release. Therefore, the main objective of the proposed herein studies was the fabrication and characterization of innovative hydrogel-based composites for local vancomycin (VAN) therapy. Methods: Presented systems are composed of ionically gelled chitosan particles loaded with vancomycin, embedded into biomimetic collagen/chitosan/hyaluronic acid-based hydrogels crosslinked with genipin and freeze-dried to serve in a flake/disc-like form. VAN-loaded carriers were characterized for their size, stability, and encapsulation efficiency (EE) using dynamic light scattering technique, zeta potential measurements, and UV-Vis spectroscopy, respectively. The synthesized composites were tested in terms of their physicochemical and biological features. Results: Spherical structures with sizes of about 200 nm and encapsulation efficiencies reaching values of approximately 60% were obtained. It was found that the resulting particles exhibit stability over time. The antibacterial activity of the developed materials against Staphylococcus aureus was established. Moreover, in vitro cell culture study revealed that the surfaces of all prepared systems are biocompatible as they supported the proliferation and adhesion of the model MG-63 cells. In addition, we have demonstrated significantly prolonged VAN release while minimizing the initial burst effect for the composites compared to bare nanoparticles and verified their desired physicochemical features during swellability, and degradation experiments. Conclusion: It is expected that the developed herein system will enable direct delivery of the antibiotic at an exposed to infections surgical site, providing drugs sustained release and thus will reduce the risk of systemic toxicity. This strategy would both inhibit biofilm formation and accelerate the healing process.


Assuntos
Antibacterianos , Quitosana , Hidrogéis , Staphylococcus aureus , Vancomicina , Vancomicina/química , Vancomicina/farmacologia , Vancomicina/administração & dosagem , Vancomicina/farmacocinética , Antibacterianos/química , Antibacterianos/farmacologia , Antibacterianos/administração & dosagem , Hidrogéis/química , Hidrogéis/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Humanos , Quitosana/química , Ácido Hialurônico/química , Ácido Hialurônico/farmacologia , Portadores de Fármacos/química , Colágeno/química , Colágeno/farmacologia , Tamanho da Partícula , Liberação Controlada de Fármacos , Infecção da Ferida Cirúrgica/prevenção & controle , Infecção da Ferida Cirúrgica/tratamento farmacológico , Testes de Sensibilidade Microbiana , Biofilmes/efeitos dos fármacos
13.
Biomacromolecules ; 25(5): 3190-3199, 2024 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-38693753

RESUMO

Intracellular bacteria in dormant states can escape the immune response and tolerate high-dose antibiotic treatment, leading to severe infections. To overcome this challenge, cascade-targeted nanoplatforms that can target macrophages and intracellular bacteria, exhibiting synergetic antibiotic/reactive oxygen species (ROS)/nitric oxide (NO)/immunotherapy, were developed. These nanoplatforms were fabricated by encapsulating trehalose (Tr) and vancomycin (Van) into phosphatidylserine (PS)-coated poly[(4-allylcarbamoylphenylboric acid)-ran-(arginine-methacrylamide)-ran-(N,N'-bisacryloylcystamine)] nanoparticles (PABS), denoted as PTVP. PS on PTVP simulates a signal of "eat me" to macrophages to promote cell uptake (the first-step targeting). After the uptake, the nanoplatform in the acidic phagolysosomes could release Tr, and the exposed phenylboronic acid on the nanoplatform could target bacteria (the second-step targeting). Nanoplatforms can release Van in response to infected intracellular overexpressed glutathione (GSH) and weak acid microenvironment. l-arginine (Arg) on the nanoplatforms could be catalyzed by upregulated inducible nitric oxide synthase (iNOS) in the infected macrophages to generate nitric oxide (NO). N,N'-Bisacryloylcystamine (BAC) on nanoplatforms could deplete GSH, allow the generation of ROS in macrophages, and then upregulate proinflammatory activity, leading to the reinforced antibacterial capacity. This nanoplatform possesses macrophage and bacteria-targeting antibiotic delivery, intracellular ROS, and NO generation, and pro-inflammatory activities (immunotherapy) provides a new strategy for eradicating intracellular bacterial infections.


Assuntos
Antibacterianos , Nanopartículas , Óxido Nítrico , Espécies Reativas de Oxigênio , Espécies Reativas de Oxigênio/metabolismo , Óxido Nítrico/metabolismo , Antibacterianos/farmacologia , Antibacterianos/química , Camundongos , Animais , Células RAW 264.7 , Nanopartículas/química , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/imunologia , Imunoterapia/métodos , Vancomicina/farmacologia , Vancomicina/química , Vancomicina/administração & dosagem , Infecções Bacterianas/tratamento farmacológico , Trealose/química , Trealose/farmacologia
14.
J Antimicrob Chemother ; 79(7): 1628-1636, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38785365

RESUMO

BACKGROUND: The glycopeptide vancomycin is the antimicrobial agent-of-choice for the treatment of severe non-gastrointestinal infections with members of Bacillus cereus sensu lato (s.l.). Recently, sporadic detection of vancomycin-resistant phenotypes emerged, mostly for agar diffusion testing such as the disc diffusion method or gradient test (e.g. Etest®) method. RESULTS: In this work, we were able to disprove a preliminarily assumed high resistance to vancomycin in an isolate of B. cereus s.l. using broth microdilution and agar dilution. Microscopic imaging during vancomycin susceptibility testing showed spreading towards the inhibition zone, which strongly suggested sliding motility. Furthermore, transcriptomic analysis using RNA-Seq on the nanopore platform revealed several key genes of biofilm formation (e.g. calY, tasA, krsEABC) to be up-regulated in pseudo-resistant cells, substantiating that bacterial sliding is responsible for the observed mobility. Down-regulation of virulence (e.g. hblABCD, nheABC, plcR) and flagellar genes compared with swarming cells also confirmed the non-swarming phenotype of the pseudo-resistant isolate. CONCLUSIONS: The results highlight an insufficiency of agar diffusion testing for vancomycin susceptibility in the B. cereus group, and reference methods like broth microdilution are strongly recommended. As currently no guideline mentions interfering phenotypes in antimicrobial susceptibility testing of B. cereus s.l., this knowledge is essential to obtain reliable results on vancomycin susceptibility. In addition, this is the first report of sliding motility undermining accurate antimicrobial susceptibility testing in B. cereus s.l. and may serve as a basis for future studies on bacterial motility in susceptibility testing and its potential impact on treatment efficacy.


Assuntos
Antibacterianos , Bacillus cereus , Testes de Sensibilidade Microbiana , Resistência a Vancomicina , Vancomicina , Bacillus cereus/efeitos dos fármacos , Bacillus cereus/genética , Testes de Sensibilidade Microbiana/métodos , Vancomicina/farmacologia , Antibacterianos/farmacologia , Resistência a Vancomicina/genética , Biofilmes/efeitos dos fármacos , Humanos , Perfilação da Expressão Gênica
15.
Mol Microbiol ; 121(6): 1182-1199, 2024 06.
Artigo em Inglês | MEDLINE | ID: mdl-38690761

RESUMO

The dipeptide D-Ala-D-Ala is an essential component of peptidoglycan and the target of vancomycin. Most Clostridioides difficile strains possess the vanG operon responsible for the synthesis of D-Ala-D-Ser, which can replace D-Ala-D-Ala in peptidoglycan. The C. difficile vanG operon is regulated by a two-component system, VanRS, but is not induced sufficiently by vancomycin to confer resistance to this antibiotic. Surprisingly, in the absence of the VanS histidine kinase (HK), the vanG operon is still induced by vancomycin and also by another antibiotic, ramoplanin, in a VanR-dependent manner. This suggested the cross-regulation of VanR by another HK or kinases that are activated in the presence of certain lipid II-targeting antibiotics. We identified these HKs as CD35990 and CD22880. However, mutations in either or both HKs did not affect the regulation of the vanG operon in wild-type cells suggesting that intact VanS prevents the cross-activation of VanR by non-cognate HKs. Overproduction of VanR in the absence of VanS, CD35990, and CD22880 led to high expression of the vanG operon indicating that VanR can potentially utilize at least one more phosphate donor for its activation. Candidate targets of CD35990- and CD22880-mediated regulation in the presence of vancomycin or ramoplanin were identified by RNA-Seq.


Assuntos
Antibacterianos , Proteínas de Bactérias , Clostridioides difficile , Regulação Bacteriana da Expressão Gênica , Histidina Quinase , Óperon , Resistência a Vancomicina , Vancomicina , Óperon/genética , Clostridioides difficile/genética , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/metabolismo , Histidina Quinase/metabolismo , Histidina Quinase/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Vancomicina/farmacologia , Resistência a Vancomicina/genética , Antibacterianos/farmacologia , Depsipeptídeos/farmacologia , Fatores de Transcrição
16.
Biomater Sci ; 12(13): 3411-3422, 2024 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-38809118

RESUMO

Bacteria have evolved survival mechanisms that enable them to live within host cells, triggering persistent intracellular infections that present significant clinical challenges due to the inability for conventional antibiotics to permeate cell membranes. In recent years, antibiotic nanocarriers or 'nanoantibiotics' have presented a promising strategy for overcoming intracellular infections by facilitating cellular uptake of antibiotics, thus improving targeting to the bacteria. However, prior to reaching host cells, nanocarriers experience interactions with proteins that form a corona and alter their physiological response. The influence of this protein corona on the cellular uptake, drug release and efficacy of nanoantibiotics for intracellular infections is poorly understood and commonly overlooked in preclinical studies. In this study, protein corona influence on cellular uptake was investigated for two nanoparticles; liposomes and cubosomes in macrophage and epithelial cells that are commonly infected with pathogens. Studies were conducted in presence of fetal bovine serum (FBS) to form a biologically relevant protein corona in an in vitro setting. Protein corona impact on cellular uptake was shown to be nanoparticle-dependent, where reduced internalization was observed for liposomes, the opposite was observed for cubosomes. Subsequently, vancomycin-loaded cubosomes were explored for their drug delivery performance against intracellular small colony variants of Staphylococcus aureus. We demonstrated improved bacterial killing in macrophages, with greater reduction in bacterial viability upon internalization of cubosomes mediated by the protein corona. However, no differences in efficacy were observed in epithelial cells. Thus, this study provides insights and evidence to the role of protein corona in modulating the performance of nanoparticles in a dynamic manner; these findings will facilitate improved understanding and translation of future investigations from in vitro to in vivo.


Assuntos
Antibacterianos , Lipossomos , Nanopartículas , Coroa de Proteína , Staphylococcus aureus , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/administração & dosagem , Coroa de Proteína/química , Coroa de Proteína/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Animais , Humanos , Lipossomos/química , Nanopartículas/química , Vancomicina/farmacologia , Vancomicina/química , Vancomicina/administração & dosagem , Camundongos , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Células RAW 264.7 , Testes de Sensibilidade Microbiana , Lipídeos/química , Portadores de Fármacos/química
17.
Gut Microbes ; 16(1): 2342583, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38722061

RESUMO

Vancomycin and metronidazole are commonly used treatments for Clostridioides difficile infection (CDI). However, these antibiotics have been associated with high levels of relapse in patients. Fidaxomicin is a new treatment for CDI that is described as a narrow spectrum antibiotic that is minimally active on the commensal bacteria of the gut microbiome. The aim of this study was to compare the effect of fidaxomicin on the human gut microbiome with a number of narrow (thuricin CD) and broad spectrum (vancomycin and nisin) antimicrobials. The spectrum of activity of each antimicrobial was tested against 47 bacterial strains by well-diffusion assay. Minimum inhibitory concentrations (MICs) were calculated against a select number of these strains. Further, a pooled fecal slurry of 6 donors was prepared and incubated for 24 h with 100 µM of each antimicrobial in a mini-fermentation system together with a no-treatment control. Fidaxomicin, vancomycin, and nisin were active against most gram positive bacteria tested in vitro, although fidaxomicin and vancomycin produced larger zones of inhibition compared to nisin. In contrast, the antimicrobial activity of thuricin CD was specific to C. difficile and some Bacillus spp. The MICs showed similar results. Thuricin CD exhibited low MICs (<3.1 µg/mL) for C. difficile and Bacillus firmus, whereas fidaxomicin, vancomycin, and nisin demonstrated lower MICs for all other strains tested when compared to thuricin CD. The narrow spectrum of thuricin CD was also observed in the gut model system. We conclude that the spectrum of activity of fidaxomicin is comparable to that of the broad-spectrum antibiotic vancomycin in vitro and the broad spectrum bacteriocin nisin in a complex community.


Assuntos
Antibacterianos , Fezes , Fidaxomicina , Microbioma Gastrointestinal , Testes de Sensibilidade Microbiana , Nisina , Vancomicina , Nisina/farmacologia , Antibacterianos/farmacologia , Humanos , Fidaxomicina/farmacologia , Vancomicina/farmacologia , Microbioma Gastrointestinal/efeitos dos fármacos , Fezes/microbiologia , Bactérias/efeitos dos fármacos , Bactérias/classificação , Clostridioides difficile/efeitos dos fármacos , Infecções por Clostridium/tratamento farmacológico , Infecções por Clostridium/microbiologia , Bacteriocinas/farmacologia
18.
BMC Microbiol ; 24(1): 177, 2024 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-38783194

RESUMO

BACKGROUND: Clostridioides difficile is the main pathogen of antimicrobial-associated diarrhoea and health care facility-associated infectious diarrhoea. This study aimed to investigate the prevalence, toxin genotypes, and antibiotic resistance of C. difficile among hospitalized patients in Xi'an, China. RESULTS: We isolated and cultured 156 strains of C. difficile, representing 12.67% of the 1231 inpatient stool samples collected. Among the isolates, tcdA + B + strains were predominant, accounting for 78.2% (122/156), followed by 27 tcdA-B + strains (27/156, 17.3%) and 6 binary toxin gene-positive strains. The positive rates of three regulatory genes, tcdC, tcdR, and tcdE, were 89.1% (139/156), 96.8% (151/156), and 100%, respectively. All isolates were sensitive to metronidazole, and the resistance rates to clindamycin and cephalosporins were also high. Six strains were found to be resistant to vancomycin. CONCLUSION: Currently, the prevalence rate of C. difficile infection (CDI) in Xi'an is 12.67% (156/1231), with the major toxin genotype of the isolates being tcdA + tcdB + cdtA-/B-. Metronidazole and vancomycin were still effective drugs for the treatment of CDI, but we should pay attention to antibiotic management and epidemiological surveillance of CDI.


Assuntos
Antibacterianos , Toxinas Bacterianas , Clostridioides difficile , Infecções por Clostridium , Fezes , Genótipo , Hospitais , Clostridioides difficile/genética , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/isolamento & purificação , Clostridioides difficile/classificação , Humanos , China/epidemiologia , Antibacterianos/farmacologia , Infecções por Clostridium/microbiologia , Infecções por Clostridium/epidemiologia , Toxinas Bacterianas/genética , Hospitais/estatística & dados numéricos , Fezes/microbiologia , Farmacorresistência Bacteriana/genética , Prevalência , Testes de Sensibilidade Microbiana , Feminino , Pessoa de Meia-Idade , Masculino , Idoso , Adulto , Proteínas de Bactérias/genética , Diarreia/microbiologia , Diarreia/epidemiologia , Metronidazol/farmacologia , Adulto Jovem , Enterotoxinas/genética , Adolescente , Vancomicina/farmacologia , Clindamicina/farmacologia , Idoso de 80 Anos ou mais
19.
BMC Infect Dis ; 24(1): 494, 2024 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-38745289

RESUMO

BACKGROUND: Brain-heart infusion agar supplemented with 4 µg/mL of vancomycin (BHI-V4) was commonly used for the detection of heterogeneous (hVISA) and vancomycin-intermediate Staphylococcus aureus (VISA). However, its diagnostic value remains unclear. This study aims to compare the diagnostic accuracy of BHI-V4 with population analysis profiling with area under the curve (PAP-AUC) in hVISA/VISA. METHODS: The protocol of this study was registered in INPLASY (INPLASY2023120069). The PubMed and Cochrane Library databases were searched from inception to October 2023. Review Manager 5.4 was used for data visualization in the quality assessment, and STATA17.0 (MP) was used for statistical analysis. RESULTS: In total, eight publications including 2153 strains were incorporated into the meta-analysis. Significant heterogeneity was evident although a threshold effect was not detected across the eight studies. The summary receiver operating characteristic (SROC) was 0.77 (95% confidence interval [CI], 0.74-0.81). The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, diagnostic score and diagnostic odds ratio were 0.59 (95% CI: 0.46-0.71), 0.96 (95%CI: 0.83-0.99), 14.0 (95% CI, 3.4-57.1), 0.43 (95%CI, 0.32-0.57), 3.48(95%CI, 2.12-4.85) and 32.62 (95%CI, 8.31-128.36), respectively. CONCLUSION: Our study showed that BHI-V4 had moderate diagnostic accuracy for diagnosing hVISA/VISA. However, more high-quality studies are needed to assess the clinical utility of BHI-V4.


Assuntos
Antibacterianos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas , Staphylococcus aureus , Vancomicina , Humanos , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/diagnóstico , Vancomicina/farmacologia , Antibacterianos/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/isolamento & purificação , Sensibilidade e Especificidade , Resistência a Vancomicina , Meios de Cultura , Área Sob a Curva
20.
Emerg Microbes Infect ; 13(1): 2361030, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38801248

RESUMO

BACKGROUND: Surveillance systems revealed that the prevalence of vancomycin-resistant Enterococcus faecium (VREfm) has increased. We aim to investigate the epidemiological and genomic characteristics of VREfm in China. METHODS: We collected 20,747 non-redundant E. faecium isolates from inpatients across 19 hospitals in six provinces between January 2018 and June 2023. VREfm was confirmed by antimicrobial susceptibility testing. The prevalence was analyzed using changepoint package in R. Genomic characteristics were explored by whole-genome sequencing. RESULTS: 5.59% (1159/20,747) of E. faecium isolates were resistant to vancomycin. The prevalence of VREfm increased in Guangdong province from 5% before 2021 to 20-50% in 2023 (p < 0.0001), but not in the other five provinces. Two predominant clones before 2021, ST17 and ST78, were substituted by an emerging clone, ST80, from 2021 to 2023 (88.63%, 195/220). All ST80 VREfm from Guangdong formed a single lineage (SC11) and were genetically distant from the ST80 VREfm from other countries, suggesting a regional outbreak. All ST80 VREfm in SC11 carried a new type of plasmid harbouring a vanA cassette, which was embedded in a Tn1546-like structure flanked by IS1678 and ISL3. However, no conjugation-related gene was detected and no transconjugant was obtained in conjugation experiment, indicating that the outbreak of ST80 VREfm could be attributed to clonal transmission. CONCLUSIONS: We revealed an ongoing outbreak of ST80 VREfm with a new vanA-harbouring plasmid in Guangdong, China. This clone has also been identified in other provinces and countries, foreboding a risk of wider spreading shortly. Continuous surveillance is needed to inform public health interventions.


Assuntos
Surtos de Doenças , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Enterococos Resistentes à Vancomicina , Sequenciamento Completo do Genoma , China/epidemiologia , Humanos , Enterococcus faecium/genética , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/isolamento & purificação , Enterococcus faecium/classificação , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Enterococos Resistentes à Vancomicina/genética , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Enterococos Resistentes à Vancomicina/isolamento & purificação , Masculino , Testes de Sensibilidade Microbiana , Antibacterianos/farmacologia , Feminino , Pessoa de Meia-Idade , Adulto , Idoso , Genoma Bacteriano , Prevalência , Criança , Adulto Jovem , Filogenia , Vancomicina/farmacologia , Adolescente
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