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1.
Int J Nanomedicine ; 15: 5813-5824, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32821103

RESUMO

Introduction: This paper presents a novel technique for the synthesis of graphene oxide (GO) with various surface features using high-density atmospheric plasma deposition. Furthermore, to investigate the use of hydrophobic, super-hydrophobic, and hydrophilic graphene in biological applications, we synthesized hydrophobic, super-hydrophobic, and hydrophilic graphene oxides by additional heat treatment and argon plasma treatment, respectively. In contrast to conventional fabrication procedures, reduced graphene oxide (rGO) formed under low pressure and high-temperature environment using a new synthesis method-developed and described in this study-offers a convenient deposition method on any kind surface with controlled wettability. Methods: High density at atmospheric plasma is used for the synthesis of rGO and GO and its biocompatibility based on various wetting properties was evaluated using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, and the viability of cells in response to rGO and GO with various surface features was investigated. Structural integrity was characterized by Raman spectroscopy, FESEM and FE-TEM. Wettability was measured via contact angle method and confirmed with XPS analysis. Results: We found that GO coating with a hydrophilic feature is more biocompatible than other surfaces as observed in case of fibroblast cells. We have shown that wettability-controlled by GO deposition-influences biocompatibilities and antibacterial effect of biomaterial surfaces. Discussion: Measuring the contact angle, it is found that contact angle for hydrophobic is increased to 150.590 and reduced to 11.580 by heat and argon plasma treatment, respectively, from 75.880 that was initially in the case of hydrophobic surface. XPS analysis confirmed various oxygen-containing functional groups transforming as deposited hydrophobic surface into superhydrophobic and hydrophilic surface. Thus, we have proposed a new, direct, cost-effective, and highly productive method for the synthesis of rGO and GO-with various surface properties-for biological applications. Similarly, for the dental implant application, the Streptococcus mutans was used as an antibacterial effect and found that S. mutans grows slowly on hydrophilic surface. Thus, antibacterial effect was prominent on GO with hydrophilic surface.


Assuntos
Atmosfera/química , Grafite/síntese química , Gases em Plasma/farmacologia , Animais , Morte Celular/efeitos dos fármacos , Linhagem Celular , Grafite/química , Camundongos , Viabilidade Microbiana/efeitos dos fármacos , Oxirredução , Streptococcus mutans/efeitos dos fármacos , Água , Molhabilidade
2.
Nature ; 584(7821): 479-483, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32788728

RESUMO

Lipopolysaccharide (LPS) resides in the outer membrane of Gram-negative bacteria where it is responsible for barrier function1,2. LPS can cause death as a result of septic shock, and its lipid A core is the target of polymyxin antibiotics3,4. Despite the clinical importance of polymyxins and the emergence of multidrug resistant strains5, our understanding of the bacterial factors that regulate LPS biogenesis is incomplete. Here we characterize the inner membrane protein PbgA and report that its depletion attenuates the virulence of Escherichia coli by reducing levels of LPS and outer membrane integrity. In contrast to previous claims that PbgA functions as a cardiolipin transporter6-9, our structural analyses and physiological studies identify a lipid A-binding motif along the periplasmic leaflet of the inner membrane. Synthetic PbgA-derived peptides selectively bind to LPS in vitro and inhibit the growth of diverse Gram-negative bacteria, including polymyxin-resistant strains. Proteomic, genetic and pharmacological experiments uncover a model in which direct periplasmic sensing of LPS by PbgA coordinates the biosynthesis of lipid A by regulating the stability of LpxC, a key cytoplasmic biosynthetic enzyme10-12. In summary, we find that PbgA has an unexpected but essential role in the regulation of LPS biogenesis, presents a new structural basis for the selective recognition of lipids, and provides opportunities for future antibiotic discovery.


Assuntos
Membrana Celular/química , Escherichia coli/química , Escherichia coli/patogenicidade , Lipopolissacarídeos/química , Lipopolissacarídeos/metabolismo , Amidoidrolases/química , Amidoidrolases/metabolismo , Motivos de Aminoácidos , Membrana Externa Bacteriana/química , Membrana Externa Bacteriana/metabolismo , Sítios de Ligação , Membrana Celular/metabolismo , Estabilidade Enzimática , Escherichia coli/citologia , Escherichia coli/efeitos dos fármacos , Genes Essenciais , Hidrolases/química , Hidrolases/metabolismo , Lipídeo A/química , Lipídeo A/metabolismo , Lipopolissacarídeos/biossíntese , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Modelos Moleculares , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Periplasma/química , Periplasma/metabolismo , Ligação Proteica , Virulência
3.
Nat Commun ; 11(1): 4157, 2020 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-32814767

RESUMO

Swarming is a form of collective bacterial motion enabled by flagella on the surface of semi-solid media. Swarming populations exhibit non-genetic or adaptive resistance to antibiotics, despite sustaining considerable cell death. Here, we show that antibiotic-induced death of a sub-population benefits the swarm by enhancing adaptive resistance in the surviving cells. Killed cells release a resistance-enhancing factor that we identify as AcrA, a periplasmic component of RND efflux pumps. The released AcrA interacts on the surface of live cells with an outer membrane component of the efflux pump, TolC, stimulating drug efflux and inducing expression of other efflux pumps. This phenomenon, which we call 'necrosignaling', exists in other Gram-negative and Gram-positive bacteria and displays species-specificity. Given that adaptive resistance is a known incubator for evolving genetic resistance, our findings might be clinically relevant to the rise of multidrug resistance.


Assuntos
Antibacterianos/metabolismo , Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Resistência Microbiana a Medicamentos/fisiologia , Transdução de Sinais/fisiologia , Adaptação Fisiológica/fisiologia , Antibacterianos/farmacologia , Bactérias/classificação , Proteínas da Membrana Bacteriana Externa/metabolismo , Bactérias Gram-Negativas/classificação , Bactérias Gram-Negativas/metabolismo , Bactérias Gram-Positivas/classificação , Bactérias Gram-Positivas/metabolismo , Viabilidade Microbiana/efeitos dos fármacos , Periplasma/metabolismo , Especificidade da Espécie
4.
Proc Natl Acad Sci U S A ; 117(32): 19455-19464, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32703812

RESUMO

A better understanding of how antibiotic exposure impacts the evolution of resistance in bacterial populations is crucial for designing more sustainable treatment strategies. The conventional approach to this question is to measure the range of concentrations over which resistant strain(s) are selectively favored over a sensitive strain. Here, we instead investigate how antibiotic concentration impacts the initial establishment of resistance from single cells, mimicking the clonal expansion of a resistant lineage following mutation or horizontal gene transfer. Using two Pseudomonas aeruginosa strains carrying resistance plasmids, we show that single resistant cells have <5% probability of detectable outgrowth at antibiotic concentrations as low as one-eighth of the resistant strain's minimum inhibitory concentration (MIC). This low probability of establishment is due to detrimental effects of antibiotics on resistant cells, coupled with the inherently stochastic nature of cell division and death on the single-cell level, which leads to loss of many nascent resistant lineages. Our findings suggest that moderate doses of antibiotics, well below the MIC of resistant strains, may effectively restrict de novo emergence of resistance even though they cannot clear already-large resistant populations.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Relação Dose-Resposta a Droga , Farmacorresistência Bacteriana/genética , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Modelos Teóricos , Plasmídeos/genética , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/crescimento & desenvolvimento , Análise de Célula Única , Processos Estocásticos
5.
PLoS One ; 15(7): e0236059, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32716948

RESUMO

Most cosmetic products are susceptible to microbiological spoilage due to contaminations that could happen during fabrication or by consumer's repetitive manipulation. The composition of cosmetic products must guarantee efficient bacterial inactivation all along with the product shelf life, which is usually assessed by challenge-tests. A challenge-test consists in inoculating specific bacteria, i.e. Staphylococcus aureus, in the formula and then investigating the bacterial log reduction over time. The main limitation of this method is relative to the time-consuming protocol, where 30 days are needed to obtain results. In this study, we have proposed a rapid alternative method coupling High Content Screening-Confocal Laser Scanning Microscopy (HCS-CLSM), image analysis and modeling. It consists in acquiring real-time S. aureus inactivation kinetics on short-time periods (typically 4h) and in predicting the efficiency of preservatives on longer scale periods (up to 7 days). The action of two preservatives, chlorphenesin and benzyl alcohol, was evaluated against S. aureus at several concentrations in a cosmetic matrix. From these datasets, we compared two secondary models to determine the logarithm reduction time (Dc) for each preservative concentration. Afterwards, we used two primary inactivation models to predict log reductions for up to 7 days and we compared them to observed log reductions. The IQ model better fits datasets and the Q value gives information about the matrix level of interference.


Assuntos
Cosméticos/química , Microscopia Confocal , Conservantes Farmacêuticos/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Cinética , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Staphylococcus aureus/fisiologia , Fatores de Tempo
6.
PLoS One ; 15(7): e0236185, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32730344

RESUMO

Fluorescent markers are a powerful tool and have been widely applied in biology for different purposes. The genome sequence of Xanthomonas citri subsp. citri (X. citri) revealed that approximately 30% of the genes encoded hypothetical proteins, some of which could play an important role in the success of plant-pathogen interaction and disease triggering. Therefore, revealing their functions is an important strategy to understand the bacterium pathways and mechanisms involved in plant-host interaction. The elucidation of protein function is not a trivial task, but the identification of the subcellular localization of a protein is key to understanding its function. We have constructed an integrative vector, pMAJIIc, under the control of the arabinose promoter, which allows the inducible expression of red fluorescent protein (mCherry) fusions in X. citri, suitable for subcellular localization of target proteins. Fluorescence microscopy was used to track the localization of VrpA protein, which was visualized surrounding the bacterial outer membrane, and the GyrB protein, which showed a diffused cytoplasmic localization, sometimes with dots accumulated near the cellular poles. The integration of the vector into the amy locus of X. citri did not affect bacterial virulence. The vector could be stably maintained in X. citri, and the disruption of the α-amylase gene provided an ease screening method for the selection of the transformant colonies. The results demonstrate that the mCherry-containing vector here described is a powerful tool for bacterial protein localization in cytoplasmic and periplasmic environments.


Assuntos
Proteínas de Bactérias/metabolismo , Citoplasma/metabolismo , Periplasma/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Xanthomonas/metabolismo , Arabinose/farmacologia , Cromossomos Bacterianos/genética , Vetores Genéticos/metabolismo , Viabilidade Microbiana/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Amido/metabolismo , Frações Subcelulares/efeitos dos fármacos , Xanthomonas/patogenicidade
7.
PLoS One ; 15(6): e0220350, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32544163

RESUMO

Mycoplasma hyopneumoniae is the major pathogenic microorganism causing enzootic pneumonia in pigs. With increasing resistance of M. hyopneumoniae to conventional antibiotics, treatment is becoming complicated. Herein, we investigated the mutant selection window (MSW) of doxycycline, tylosin, danofloxacin, tiamulin, and valnemulin for treating the M. hyopneumoniae type strain (ATCC 25934) to determine the likelihood of promoting resistance with continued use of these antibiotics. Minimum inhibitory concentration (MIC) values against M. hyopneumoniae were determined for each antimicrobial agent based on microdilution broth and agar dilution methods (bacterial numbers ranged from 105 colony-forming units (CFU)/mL to 109 CFU/mL). The minimal concentration inhibiting colony formation by 99% (MIC99) and the mutant prevention concentration (MPC) were determined by the agar dilution method with three inoculum sizes. Antimicrobial killing was determined based on MIC99 and MPC values for all five agents. MIC values ranged from 0.001 to 0.25 µg/mL based on the microdilution broth method, and from 0.008 to 1.0 µg/mL based on the agar dilution method. MPC values ranged from 0.0016 to 10.24 µg/mL. MPC/MIC99 values were ordered tylosin > doxycycline > danofloxacin > tiamulin > valnemulin. MPC achieved better bactericidal action than MIC99. Based on pharmacodynamic analyses, danofloxacin, tylosin, and doxycycline are more likely to select resistant mutants than tiamulin and valnemulin.


Assuntos
Antibacterianos/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Mutação , Mycoplasma hyopneumoniae/efeitos dos fármacos , Mycoplasma hyopneumoniae/genética , Diterpenos/farmacologia , Doxiciclina/farmacologia , Fluoroquinolonas/farmacologia , Cinética , Testes de Sensibilidade Microbiana , Mycoplasma hyopneumoniae/fisiologia , Tilosina/farmacologia
8.
Artigo em Inglês | MEDLINE | ID: mdl-32517236

RESUMO

The use of bacterial strains as agents in bioremediation processes could reduce the harmfulness of potential toxic elements (PTEs) from water and soil with low or even no impact on the natural ecosystems. In this study, two new metal resistant-bacterial strains (Q3 and Q5) of Bacillus sp. were isolated from a sulfurous spring and their potential (as pure cultures or mixed) to remove Pb(II) and Cd(II) from an aqueous matrix was evaluated and optimized using response surface methodology (RSM). The optimal conditions for Cd(II) removal from all tested strains combinations were observed at an initial pH 5, a temperature of 38 °C, and an initial Cd(II) concentration of 50 mg L-1, while the performance of bacterial strains on Pb(II) removal was strongly correlated to initial pH and temperature conditions. Moreover, the efficiency of bacterial strains in removing both PTEs, Pb(II) and Cd(II), from an aqueous matrix was considerably higher when they were used as a mixed culture rather than pure. According to field emission SEM (FESEM) and EDS analysis, the two bacterial strains showed different mechanisms in removing Cd(II): Bacillus sp. Q5 bio-accumulated Cd(II) in its periplasmic space, whereas Bacillus sp. Q3 bio-accumulated Cd(II) on its cell surface. On the other hand, Pb(II) is removed by chemical precipitation (lead sulfide) induced by both Bacillus sp. Q3 and Q5. This study discloses new aspects of Pb(II) and Cd(II) bioremediation mechanisms in Bacillus species that can be extremely useful for designing and operating novel PTEs bioremediation processes.


Assuntos
Bacillus/isolamento & purificação , Biodegradação Ambiental , Cádmio/metabolismo , Chumbo/metabolismo , Metais Pesados/metabolismo , Desintoxicação por Sorção , Bacillus/metabolismo , Biomassa , Ecossistema , Humanos , Metais Pesados/toxicidade , Viabilidade Microbiana/efeitos dos fármacos , Desintoxicação por Sorção/métodos
9.
J Med Microbiol ; 69(5): 689-696, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32375980

RESUMO

Introduction. Rhein (4, 5-dihydroxyanthraquinone-2-carboxylic acid) has various properties, including anti-inflammatory, antioxidant and anticancer activities. However, the mechanism underlying the role of rhein in antimicrobial activity remains largely unknown.Aim. This study aims to identify potential natural compounds of rhein that are capable of inhibiting Cutibacterium acnes and elucidate the effects of rhein on NADH dehydrogenase-2 activity in C. acnes.Methodology. The anti-C. acnes activity of compounds was analysed using minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), the paper disc diffusion test and the checkerboard dilution test. To check whether rhein was inhibitory, putative type II NADH dehydrogenase (NDH-2) of C. acnes was analysed, cloned and expressed in Escherichia coli, and then NDH-2 purification was assessed with Ni-NTA before rhein inhibition of NADH dehydrogenase-2 activity was checked with ferricyanide [K3Fe(CN)6] as a substrate.Results. The results showed that the MIC of rhein against C. acnes was 6.25 µg ml-1, while the MBC was 12.5 µg ml-1, and there was a 38 mm inhibition zone in the paper disc diffusion test. Rhein showed an additive two- to fourfold reduction of the MIC value with four antibiotics on the checkerboard dilution test. The purified NADH dehydrogenase gene product showed a size of approximately 51 kDa and had a V max of 23 µmol and a K m of 280 µm. The inhibitory effect of rhein against NADH dehydrogenase-2 activity was non-competitive with ferricyanide [K3Fe(CN)6] with a K i value of 3.5-4.5 µm.Conclusion. This study provided evidence of the inhibitory effects of rhein on the growth of C. acnes by blocking of NADH dehydrogenase-2 activity. This mechanism of inhibitory activity in the reduction of ROS formation and ATP productivity should be further tested in C. acnes and the question of whether rhein inhibits the natural growth of C. acnes should be investigated.


Assuntos
Antraquinonas/farmacologia , Antibacterianos/farmacologia , Inibidores Enzimáticos/farmacologia , Infecções por Bactérias Gram-Positivas/microbiologia , NADH Desidrogenase/antagonistas & inibidores , Propionibacterium acnes/efeitos dos fármacos , Propionibacterium acnes/enzimologia , Antraquinonas/uso terapêutico , Antibacterianos/uso terapêutico , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/uso terapêutico , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Humanos , Cinética , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Propionibacterium acnes/crescimento & desenvolvimento , Espécies Reativas de Oxigênio/metabolismo , Proteínas Recombinantes
10.
Ecotoxicol Environ Saf ; 199: 110639, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32408033

RESUMO

Graphene Oxide (GO) has wide applications in many fields which has caused a large expected quantity of the graphene-based wastes. It is necessary to understand the toxic effects of the GO on the activated sludge (AS) considering its inevitable discharge to the wastewater treatment plants as the ultimate repositories for these wastes. In this study, the acute exposures of the multilayer Nano-graphene oxide (MNGO) at different dosages were conducted in order to investigate its integrated effects on the formation of the biofilm, mature biofilm and the microbial activity of the activated sludge. Raman spectroscopy and laser scanning confocal microscopy (LSCM) were adopted for the in-situ characterization of the biofilm with the exposure of the MNGO. The results showed that the activated sludge was tolerable to the acute exposure of the less than 100 mg/L of the MNGO, especially for the mature biofilm, and only a subtle decrease was found in the size and thickness during the formation of the biofilm, while the amount of 300 mg/L of the MNGO caused the sever deterioration on the activated sludge system. The microbial metabolic activity, viability, and the biological removal of the nutrients were significantly affected with the more than 100 mg/L of the MNGO. It was also demonstrated by the microbial cytotoxicity tests that the increase in the exposure of the MNGO was related to the increase in the reactive oxygen species (ROS) and the damaging degree of the cell membrane.


Assuntos
Biofilmes/efeitos dos fármacos , Grafite/toxicidade , Viabilidade Microbiana/efeitos dos fármacos , Esgotos/microbiologia , Águas Residuárias/microbiologia , Poluentes Químicos da Água/toxicidade , China , Espécies Reativas de Oxigênio/metabolismo , Esgotos/química , Águas Residuárias/química , Purificação da Água/métodos
11.
Nat Commun ; 11(1): 2200, 2020 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-32366839

RESUMO

Bacterial persister cells are phenotypic variants that exhibit a transient non-growing state and antibiotic tolerance. Here, we provide in vitro evidence of Staphylococcus aureus persisters within infected host cells. We show that the bacteria surviving antibiotic treatment within host cells are persisters, displaying biphasic killing and reaching a uniformly non-responsive, non-dividing state when followed at the single-cell level. This phenotype is stable but reversible upon antibiotic removal. Intracellular S. aureus persisters remain metabolically active but display an altered transcriptomic profile consistent with activation of stress responses, including the stringent response as well as cell wall stress, SOS and heat shock responses. These changes are associated with multidrug tolerance after exposure to a single antibiotic. We hypothesize that intracellular S. aureus persisters may constitute a reservoir for relapsing infection and could contribute to therapeutic failures.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Viabilidade Microbiana/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Células A549 , Animais , Linhagem Celular , Linhagem Celular Tumoral , Células Cultivadas , Farmacorresistência Bacteriana Múltipla/genética , Metabolismo Energético/efeitos dos fármacos , Metabolismo Energético/genética , Perfilação da Expressão Gênica/métodos , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Macrófagos/efeitos dos fármacos , Macrófagos/microbiologia , Camundongos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/genética , Microscopia Confocal , Staphylococcus aureus/genética , Staphylococcus aureus/fisiologia , Células THP-1
12.
PLoS One ; 15(5): e0232775, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32374766

RESUMO

Antibacterial photodynamic therapy (aPDT) and antibacterial blue light (aBL) are emerging treatment methods auxiliary to mechanical debridement for periodontitis. APDT provided with near-infrared (NIR) light in conjunction with an indocyanine green (ICG) photosensitizer has shown efficacy in several dental in-office-treatment protocols. In this study, we tested Streptococcus mutans biofilm sensitivity to either aPDT, aBL or their combination dual-light aPDT (simultaneous aPDT and aBL) exposure. Biofilm was cultured by pipetting diluted Streptococcus mutans suspension with growth medium on the bottom of well plates. Either aPDT (810 nm) or aBL (405 nm) or a dual-light aPDT (simultaneous 810 nm aPDT and 405 nm aBL) was applied with an ICG photosensitizer in cases of aPDT or dual-light, while keeping the total given radiant exposure constant at 100 J/cm2. Single-dose light exposures were given after one-day or four-day biofilm incubations. Also, a model of daily treatment was provided by repeating the same light dose daily on four-day and fourteen-day biofilm incubations. Finally, the antibacterial action of the dual-light aPDT with different energy ratios of 810 nm and 405 nm of light were examined on the single-day and four-day biofilm protocols. At the end of each experiment the bacterial viability was assessed by colony-forming unit method. Separate samples were prepared for confocal 3D biofilm imaging. On a one-day biofilm, the dual-light aPDT was significantly more efficient than aBL or aPDT, although all modalities were bactericidal. On a four-day biofilm, a single exposure of aPDT or dual-light aPDT was more efficient than aBL, resulting in a four logarithmic scale reduction in bacterial counts. Surprisingly, when the same amount of aPDT was repeated daily on a four-day or a fourteen-day biofilm, bacterial viability improved significantly. A similar improvement in bacterial viability was observed after repetitive aBL application. This viability improvement was eliminated when dual-light aPDT was applied. By changing the 405 nm to 810 nm radiant exposure ratio in dual-light aPDT, the increase in aBL improved the antibacterial action when the biofilm was older. In conclusion, when aPDT is administered repeatedly to S. mutans biofilm, a single wavelength-based aBL or aPDT leads to a significant biofilm adaptation and increased S. mutans viability. The combined use of aBL light in synchrony with aPDT arrests the adaptation and provides significantly improved and sustained antibacterial efficacy.


Assuntos
Adaptação Biológica/efeitos dos fármacos , Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Verde de Indocianina/farmacologia , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/farmacologia , Streptococcus mutans/efeitos dos fármacos , Adaptação Biológica/efeitos da radiação , Carga Bacteriana/efeitos dos fármacos , Carga Bacteriana/efeitos da radiação , Biofilmes/efeitos da radiação , Humanos , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Higiene Bucal/métodos , Periodontite/tratamento farmacológico , Streptococcus mutans/efeitos da radiação
14.
J Appl Microbiol ; 129(3): 601-611, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32281733

RESUMO

AIMS: To study the mechanism of the antibacterial action of tea polyphenols such as catechins and theaflavins against Bacillus coagulans, and the interaction of epigallocatechin gallate (EGCg) or theaflavin 3,3'-di-O-gallate (TFDG) with the surface of B. coagulans cells was investigated. METHODS AND RESULTS: The antibacterial activities of EGCg and TFDG against B. coagulans cells were measured by counting of the viable cells after the mixing with each polyphenol. Bactericidal effect of TFDG was shown at the concentration of greater than or equal to 62·5 mg l-1 ; however, at the same concentration, EGCg did not. According to the results of two dimensional (2D)-electrophoresis analysis, TFDG seemed to interact with cytoplasmic membrane proteins. The activity of the glucose transporters of the cells decreased 40% following the treatment with TFDG of 62·5 mg l-1 ; however, this decrease was only slight in case of EGCg. This result was in accordance with the strength of their bactericidal activities. CONCLUSION: Our results suggest that the direct interaction between membrane proteins and TFDG is an important factor in the antibacterial activity of polymerized catechins, affecting their functions and leading to cell death. SIGNIFICANCE AND IMPACT OF THE STUDY: Tea polyphenols can effectively use the prevention of product spoilage in the food and beverage industry.


Assuntos
Antibacterianos/farmacologia , Bacillus coagulans/efeitos dos fármacos , Biflavonoides/farmacologia , Catequina/análogos & derivados , Bacillus coagulans/metabolismo , Biflavonoides/metabolismo , Catequina/metabolismo , Catequina/farmacologia , Membrana Celular/efeitos dos fármacos , Glucose/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Viabilidade Microbiana/efeitos dos fármacos , Polifenóis/química , Polifenóis/farmacologia , Chá/química
15.
PLoS One ; 15(3): e0230812, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32214399

RESUMO

The aim of this study was to assess the efficacy of lactic acid (LA), caprylic acid (CA), high- (HDI) and low- (LDI) dose gamma irradiation and LDI combined with LA or CA on the inactivation of a pool of Shiga toxin-producing Escherichia coli (STEC) strains inoculated on beef trimmings. The three most efficacious treatments were selected to study their effect on meat quality parameters and sensory attributes. The inoculum included five native STEC serogroups (O26, O103, O111, O145 and O157). The treatments applied were 0.5% LA, 0.04% CA, 0.5 kGy LDI, 2 kGy HDI, LDI+LA and LDI+CA. Beef trimmings were divided into two groups; one was inoculated with high (7 log CFU/g) and the other with low (1 log CFU/g) level of inoculum. Efficacy was assessed by estimating log reduction and reduction of stx- and eae-positive samples after enrichment, respectively. Results showed that treatments with organic acids alone were not effective in reducing STEC populations. For high inoculum samples, the most effective treatment was HDI followed by LDI+LA and LDI alone or combined with CA. For low inoculum samples, the most effective treatment was HDI followed by LDI alone or combined with organic acids. Concerning meat quality parameters and sensory attributes, irradiation treatments (LDI and HDI) caused minimal changes, while LDI+LA modified them significantly compared with the control. Therefore, based on our results, no benefits were observed after combining organic acids with gamma irradiation.


Assuntos
Caprilatos/farmacologia , Raios gama , Ácido Láctico/farmacologia , Carne Vermelha/microbiologia , Escherichia coli Shiga Toxigênica/efeitos dos fármacos , Escherichia coli Shiga Toxigênica/efeitos da radiação , Relação Dose-Resposta à Radiação , Qualidade dos Alimentos , Inocuidade dos Alimentos , Concentração de Íons de Hidrogênio , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/efeitos da radiação , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Oxirredução/efeitos dos fármacos , Oxirredução/efeitos da radiação , Escherichia coli Shiga Toxigênica/fisiologia , Paladar
16.
Microbiol Immunol ; 64(6): 424-434, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32196736

RESUMO

Streptococcus mutans is a major cause of tooth decay due to its promotion of biofilm formation and acid production. Several plant extracts have been reported to have multiple biological activities such as anti-inflammation and antibacterial effects. This study investigated the antibacterial activity of three plant extracts, phellodendron bark (PB), yucca, and black ginger, and found that PB had a stronger effect than the other extracts. Then, the minimum inhibitory concentration (MIC) of PB against 100 S. mutans strains was investigated. The MIC range of PB was 9.8-312.5 µg/mL. PB suppressed the growth kinetics of S. mutans in a dose-dependent manner, even at sub-MICs of PB. Then, we investigated the effect of PB on S. mutans virulence. The PB suppressed biofilm formation at high concentrations, although PB did not affect the expression of glucosyltransferase genes. Additionally, PB suppressed the decrease in pH from adding an excess of glucose. The expression of genes responsible for acid production was increased by the addition of excess glucose without PB, whereas their expression levels were not increased in the presence of 1× and 2× MIC of PB. Although PB showed a bacteriostatic effect on planktonic S. mutans cells, it was found that more than 2× MIC of PB showed a partial bactericidal effect on biofilm cells. In conclusion, PB not only showed antibacterial activity against S. mutans but also decreased the cariogenic activity in S. mutans.


Assuntos
Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Viabilidade Microbiana/efeitos dos fármacos , Extratos Vegetais/farmacologia , Streptococcus mutans/efeitos dos fármacos , Gengibre/metabolismo , Testes de Sensibilidade Microbiana/métodos , Phellodendron/metabolismo , Casca de Planta/metabolismo , Streptococcus mutans/fisiologia , Yucca/metabolismo
17.
Transfusion ; 60(4): 799-805, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32129497

RESUMO

BACKGROUND: Risk of transfusion-transmitted (TT) malaria is mainly associated with whole blood (WB) or red blood cell (RBC) transfusion. Risk mitigation relies mostly on donor deferral while a limited number of countries perform blood testing, both negatively impacting blood availability. This study investigated the efficacy of the pathogen reduction system using amustaline and glutathione (GSH) to inactivate Plasmodium falciparum in WB. STUDY DESIGN AND METHODS: WB units were spiked with ring stage P. falciparum infected RBCs. Parasite loads were measured in samples at time of infection, after 24 hours at room temperature (RT), and after a 24-hour incubation at RT post-treatment with 0.2 mM amustaline and 2 mM GSH. Serial 10-fold dilutions of the samples were inoculated to RBC cultures and maintained up to 4 weeks. Parasitemia was quantified by cytometry. RESULTS: The P. falciparum viability assay has a limit of detection of a single live parasite per sample. Input parasite titer was >5.7 log10 TCID50 per mL. A 24-hour incubation at RT paused parasite development in controls, but they retained viability and infectivity when tested in culture. In contrast, no infectious parasites were detected in the amustaline/GSH-treated sample after 4 weeks of culture. CONCLUSION: A robust level of P. falciparum inactivation was achieved in WB using amustaline/GSH treatment. Parasite log reduction was >5.7 log10 TCID50 per mL. Development of such a pathogen reduction system may provide an opportunity to reduce the risk of TT malaria and improve blood availability.


Assuntos
Acridinas/farmacologia , Glutationa/farmacologia , Malária Falciparum/prevenção & controle , Viabilidade Microbiana/efeitos dos fármacos , Compostos de Mostarda Nitrogenada/farmacologia , Segurança do Sangue/métodos , Eritrócitos/microbiologia , Eritrócitos/parasitologia , Humanos , Malária Falciparum/sangue , Malária Falciparum/transmissão , Carga Parasitária , Parasitemia/tratamento farmacológico , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/crescimento & desenvolvimento
18.
Transfusion ; 60(5): 1050-1059, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32187695

RESUMO

BACKGROUND: Our previous study showed that ultraviolet C (UVC) from xenon (Xe) flash without any photoreactive compounds inactivated bacteria in platelet concentrates (PCs) with less damage to platelets (PLTs) as compared with Xe flash containing ultraviolet A, ultraviolet B, and visible light. Here, we report a UVC irradiation system for PCs under flow conditions consisting of a flow path-irradiation sheet, a peristaltic pump, and a collection bag. STUDY DESIGN AND METHODS: Platelet concentrates containing Ringer's solution (R-PCs) inoculated with bacteria were injected into a flow path sheet using a peristaltic pump, being irradiated with UVC from Xe flash. The quality of the irradiated PCs containing platelet additive solution (PAS-PCs) was assessed based on PC variables, PLT surface markers, and aggregation ability. RESULTS: Streptococcus dysgalactiae (12 tests) and Escherichia coli (11) were all negative on bacterial culture, while Staphylococcus aureus (12) and Klebsiella pneumoniae (14) grew in one and two R-PCs, respectively. Bacillus cereus spores were inactivated in 7 of 12 R-PCs. PC variables became significantly different between irradiated and nonirradiated PAS-PCs. P-selectin, first procaspase-activating compound (PAC-1) binding, and phosphatidylserine increased by irradiation. Aggregability stimulated by adenosine diphosphate, collagen, or thromboxane A2 increased in the irradiated PAS-PCs, while that by thrombin became smaller compared with nonirradiated controls. CONCLUSION: This newly developed system inactivated bacteria including spores in R-PCs. PAS-PCs irradiated by this system retained acceptable in vitro quality and aggregability. Usage of a peristaltic pump instead of agitator during irradiation may enable this system to be directly combined with an apheresis blood cell separator.


Assuntos
Plaquetas/citologia , Preservação de Sangue , Desinfecção/instrumentação , Viabilidade Microbiana , Raios Ultravioleta , Xenônio/farmacologia , Bacillus cereus/efeitos dos fármacos , Bacillus cereus/fisiologia , Bacillus cereus/efeitos da radiação , Bactérias/efeitos dos fármacos , Bactérias/efeitos da radiação , Remoção de Componentes Sanguíneos , Plaquetas/efeitos dos fármacos , Plaquetas/efeitos da radiação , Preservação de Sangue/instrumentação , Preservação de Sangue/métodos , Segurança do Sangue/instrumentação , Segurança do Sangue/métodos , Desinfecção/métodos , Contaminação de Medicamentos/prevenção & controle , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Escherichia coli/efeitos da radiação , Humanos , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/fisiologia , Klebsiella pneumoniae/efeitos da radiação , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Soluções para Preservação de Órgãos/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Agregação Plaquetária/fisiologia , Agregação Plaquetária/efeitos da radiação , Controle de Qualidade , Solução de Ringer/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia , Staphylococcus aureus/efeitos da radiação , Streptococcus/efeitos dos fármacos , Streptococcus/fisiologia , Streptococcus/efeitos da radiação
19.
J Appl Microbiol ; 129(3): 532-540, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32160376

RESUMO

AIM: To examine the synergistic effect of calycosin combined with polymyxin B against various mcr-1-positive bacterial strains. METHODS AND RESULTS: In this study, we found a potential inhibitor of MCR-1, calycosin, that could significantly restore the antibacterial activity of polymyxin B. The synergistic effect of calycosin combined with polymyxin B against various mcr-1-positive bacterial strains was confirmed by checkerboard minimum inhibitory concentration assays, time-kill curve assays and disk diffusion assays. The fractional inhibitory concentration indexes ranged from 0·15 ± 0·03 to 0·28 ± 0·05, and the zones of inhibition increased from 13·33 ± 0·47 to 17·67 ± 0·47 mm with the combined therapy of calycosin and polymyxin B. In addition, the combined therapy significantly reduced the number of bacteria in the medium. However, at the concentrations required for the synergistic effect with polymyxin B, calycosin alone showed no effect on bacterial growth or MCR-1 production. Calycosin treatment exhibited no cytotoxicity to HeLa cells or A549 cells at calycosin concentrations below 32 µg ml-1 . CONCLUSIONS: Therefore, our results suggested that calycosin could be used as a potential MCR-1 inhibitor to restore the bactericidal effect of polymyxin B without affecting bacterial viability or existing cytotoxicity. SIGNIFICANCE AND IMPACT OF THE STUDY: The synergistic effect of calycosin combined with polymyxin B against various mcr-1-positive bacterial strains paves the way for future pharmaceutical applications of calycosin in fighting mcr-1-positive bacterial infections.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Isoflavonas/farmacologia , Polimixina B/farmacologia , Células A549 , Sinergismo Farmacológico , Células HeLa , Humanos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos
20.
Biochemistry (Mosc) ; 85(2): 248-256, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32093601

RESUMO

Streptococcus iniae is a pathogenic and zoonotic bacterium responsible for human diseases and mortality of many fish species. Recently, this bacterium has demonstrated an increasing trend for antibiotics resistance, which has warranted a search for new approaches to tackle its infection. Glutamate racemase (MurI) is a ubiquitous enzyme of the peptidoglycan synthesis pathway that plays an important role in the cell wall integrity maintenance; however, the significance of this enzyme differs in different species. In this study, we knocked out the MurI gene in S. iniae in order to elucidate the role of glutamate racemase in maintaining cell wall integrity in this bacterial species. We also cloned, expressed, and purified MurI and determined its biochemical characteristics. Biochemical analysis revealed that the MurI gene in S. iniae encodes a functional enzyme with a molecular weight of 30 kDa, temperature optimum at 35°C, and pH optimum at 8.5. Metal ions, such as Cu2+, Mn2+, Co2+ and Zn2+, inhibited the enzyme activity. MurI was found to be essential for the viability and cell wall integrity of S. iniae. The optimal growth of the MurI-deficient S. iniae mutant can be achieved only by adding a high concentration of D-glutamate to the medium. Membrane permeability assay of the mutant showed an increasing extent of the cell wall damage with time upon D-glutamate starvation. Moreover, the mutant lost its virulence when incubated in fish blood. Our results demonstrated that the MurI knockout leads to the generation of S. iniae auxotroph with damaged cell walls.


Assuntos
Isomerases de Aminoácido/metabolismo , Parede Celular , Viabilidade Microbiana , Streptococcus iniae/enzimologia , Isomerases de Aminoácido/antagonistas & inibidores , Isomerases de Aminoácido/genética , Parede Celular/efeitos dos fármacos , Parede Celular/metabolismo , Concentração de Íons de Hidrogênio , Metais Pesados/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Mutação , Streptococcus iniae/efeitos dos fármacos , Streptococcus iniae/metabolismo
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