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1.
Proc Natl Acad Sci U S A ; 117(37): 22967-22973, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32868444

RESUMO

Hospital-acquired infections are a global health problem that threatens patients' treatment in intensive care units, causing thousands of deaths and a considerable increase in hospitalization costs. The endotracheal tube (ETT) is a medical device placed in the patient's trachea to assist breathing and delivering oxygen into the lungs. However, bacterial biofilms forming at the surface of the ETT and the development of multidrug-resistant bacteria are considered the primary causes of ventilator-associated pneumonia (VAP), a severe hospital-acquired infection for significant mortality. Under these circumstances, there has been a need to administrate antibiotics together. Although necessary, it has led to a rapid increase in bacterial resistance to antibiotics. Therefore, it becomes necessary to develop alternatives to prevent and combat these bacterial infections. One possibility is to turn the ETT itself into a bactericide. Some examples reported in the literature present drawbacks. To overcome those issues, we have designed a photosensitizer-containing ETT to be used in photodynamic inactivation (PDI) to avoid bacteria biofilm formation and prevent VAP occurrence during tracheal intubation. This work describes ETT's functionalization with curcumin photosensitizer, as well as its evaluation in PDI against Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli A significant photoinactivation (up to 95%) against Gram-negative and Gram-positive bacteria was observed when curcumin-functionalized endotracheal (ETT-curc) was used. These remarkable results demonstrate this strategy's potential to combat hospital-acquired infections and contribute to fighting antimicrobial resistance.


Assuntos
Antibacterianos/farmacologia , Curcumina/farmacologia , Intubação Intratraqueal/instrumentação , Pneumonia Associada à Ventilação Mecânica/prevenção & controle , Antibacterianos/química , Biofilmes/efeitos dos fármacos , Curcumina/química , Humanos , Intubação Intratraqueal/efeitos adversos , Pneumonia Associada à Ventilação Mecânica/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia
2.
Med Microbiol Immunol ; 209(6): 669-680, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32880037

RESUMO

The ability of bacteria to aggregate and form biofilms impairs phagocytosis by polymorphonuclear leukocytes (PMNs). The aim of this study was to examine if the size of aggregates is critical for successful phagocytosis and how bacterial biofilms evade phagocytosis. We investigated the live interaction between PMNs and Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli and Staphylococcus epidermidis using confocal scanning laser microscopy. Aggregate size significantly affected phagocytosis outcome and larger aggregates were less likely to be phagocytized. Aggregates of S. epidermidis were also less likely to be phagocytized than equally-sized aggregates of the other three species. We found that only aggregates of approx. 5 µm diameter or smaller were consistently phagocytosed. We demonstrate that planktonic and aggregated cells of all four species significantly reduced the viability of PMNs after 4 h of incubation. Our results indicate that larger bacterial aggregates are less likely to be phagocytosed by PMNs and we propose that, if the aggregates become too large, circulating PMNs may not be able to phagocytose them quickly enough, which may lead to chronic infection.


Assuntos
Biofilmes , Escherichia coli/fisiologia , Neutrófilos/fisiologia , Fagocitose , Pseudomonas aeruginosa/fisiologia , Staphylococcus aureus/fisiologia , Staphylococcus epidermidis/fisiologia , Escherichia coli/ultraestrutura , Humanos , Pseudomonas aeruginosa/ultraestrutura , Pele/microbiologia , Staphylococcus aureus/ultraestrutura , Staphylococcus epidermidis/ultraestrutura
3.
PLoS One ; 15(8): e0238006, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32857814

RESUMO

This study aimed to evaluate the effects of two prebiotics in different concentrations on nutrient digestibility, fermentative products and immunological variables in adult dogs. Twenty-four adult dogs were randomly divided into six blocks according to their metabolic body weights (BW0.75); within these groups, dogs were randomized to four treatments: control without prebiotics (CO); inclusion of 0.5% prebiotic blend Yes-Golf (B1); inclusion of 1.0% galactooligosaccharide (GOS); and inclusion of 1.0% prebiotic blend Yes-Golf (B2). The experiment lasted 30 days, with 20 days adaptation and 10 days stool and blood collection. Results were analyzed for normality and means were separated by ANOVA and adjusted by the Tukey test at the significance level of 5.0%. Prebiotic supplementation had no effect on apparent digestibility coefficients (ADC), total stool production and fecal scores (p > 0.05). Prebiotics evaluated also did not alter fecal pH, nor the concentrations of ammonia, lactic acid, short chain fatty acids (SCFA) and most fecal branched chain fatty acids (BCFA) (p > 0.05). The addition of GOS decreased the concentration of iso-valeric acid (p = 0.0423). Regarding immunological variables, concentrations of fecal IgA were not influenced by the treatments. Treatments GOS and B2 increased the total number of polymorphonuclear cells, as well as the oxidative burst in relation to treatments B1 and CO (p < 0.0001). Treatment B2 improved the rate of S. aureus phagocytosis in relation to CO (p = 0.0111), and both the GOS and B2 treatments had a better index for E. coli phagocytosis than the CO treatment (p = 0.0067). In conclusion, there was indication that both prebiotics GOS and B2 at 1.0% inclusion improved the immunity of healthy dogs.


Assuntos
Colo/efeitos dos fármacos , Oligossacarídeos/farmacologia , Prebióticos , Animais , Colo/imunologia , Colo/microbiologia , Dieta/veterinária , Cães , Ácidos Graxos Voláteis/análise , Fezes/química , Microbioma Gastrointestinal/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Leucócitos/citologia , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Fagocitose/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Staphylococcus aureus/fisiologia
4.
Nat Commun ; 11(1): 3334, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620760

RESUMO

TH17 cells exemplify environmental immune adaptation: they can acquire both a pathogenic and an anti-inflammatory fate. However, it is not known whether the anti-inflammatory fate is merely a vestigial trait, or whether it serves to preserve the integrity of the host tissues. Here we show that the capacity of TH17 cells to acquire an anti-inflammatory fate is necessary to sustain immunological tolerance, yet it impairs immune protection against S. aureus. Additionally, we find that TGF-ß signalling via Smad3/Smad4 is sufficient for the expression of the anti-inflammatory cytokine, IL-10, in TH17 cells. Our data thus indicate a key function of TH17 cell plasticity in maintaining immune homeostasis, and dissect the molecular mechanisms explaining the functional flexibility of TH17 cells with regard to environmental changes.


Assuntos
Homeostase/imunologia , Inflamação/imunologia , Interleucina-10/imunologia , Intestinos/imunologia , Células Th17/imunologia , Animais , Plasticidade Celular/imunologia , Resistência à Doença/genética , Resistência à Doença/imunologia , Células HEK293 , Humanos , Interleucina-10/genética , Interleucina-10/metabolismo , Interleucina-17/genética , Interleucina-17/imunologia , Interleucina-17/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/imunologia , Staphylococcus aureus/fisiologia , Células Th17/metabolismo , Fator de Crescimento Transformador beta/imunologia , Fator de Crescimento Transformador beta/metabolismo
5.
Mikrobiyol Bul ; 54(2): 223-234, 2020 Apr.
Artigo em Turco | MEDLINE | ID: mdl-32723278

RESUMO

Biofilm-related infections are considered as among the foremost causes of treatment failure nowadays. One of the most common causes of biofilm-related infections is Staphylococcus aureus. It becomes extremely difficult to determine the appropriate treatment protocol while biofilm-related infections are coexisting with bacterial methicillin resistance. The aim of this study was to observe the potential of biofilm formation of methicillin-sensitive and -resistant S.aureus strains isolated from different clinical specimens and to determine reliable and effective methods for biofilm detection. A total of 200 S.aureus strains (100 methicillin-resistant and 100 methicillin-susceptible) isolated from 107 wound, 93 blood and catheter specimens, which were accepted as causative agents, included in the study. In order to determine the methicillin sensitivity, oxacillin minimal inhibitory concentration value obtained by an automated system and cefoxitin disc diffusion method were evaluated together. Biofilm formation was investigated by modified Christensen (MC), MTT, BioTimer and Congo Red Agar (CRA) methods, and the presence of ica operon responsible for biofilm formation was also observed by polymerase chain reaction. It has been shown that methicillin-resistant isolates produce biofilms in a shorter time and higher rate, and their biofilm structure is denser than methicillin-sensitive isolates in all MC, MTT and BioTimer methods. There was no difference between blood and wound isolates in biofilm formation. The most sensitive and specific conventional methods were MTT and BioTimer methods respectively. There was no significant difference between the isolates containing a gene region of icaADBC operon and the biofilm forming isolates according to MC, MTT, BioTimer and CCA methods. There was a high correlation between the presence of biofilm and ica positivity, and the tendency to form biofilm augmented as the number of ica genes increased. It has been emphasized that more virulent strains such as methicillin-resistant S.aureus have a higher tendency to form biofilm, and these two resistance mechanisms have been shown to support each other as cascade. ica detection may be an important reagent in itself for the detection of virulent strains, thus detection of the ica presence may be an early marker of treatment decisions, determination of protection strategies, and struggle with biofilm-related infections. In cases where molecular methods are not available, the existence of quick, easy-to-apply and reliable conventional methods to detect biofilm formation is extremely important. All conventional methods used in this study seem to be sufficient in this respect. MC and MTT methods stand out in terms of biofilm quantitation. BioTimer method is a very new and remarkable test used to detect biofilm formation. In conclusion, determining the potential of biofilm formation of colonizing or causative agents and taking essential precautions before interventional procedures will decrease biofilm related infections and related morbidity and mortality.


Assuntos
Biofilmes , Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Staphylococcus aureus , Antibacterianos/farmacologia , Humanos , Meticilina/farmacologia , Staphylococcus aureus Resistente à Meticilina/fisiologia , Testes de Sensibilidade Microbiana , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia
6.
7.
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
8.
Proc Natl Acad Sci U S A ; 117(29): 17228-17239, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32616573

RESUMO

The ability of Staphylococcus aureus to infect many different tissue sites is enabled, in part, by its transcriptional regulatory network (TRN) that coordinates its gene expression to respond to different environments. We elucidated the organization and activity of this TRN by applying independent component analysis to a compendium of 108 RNA-sequencing expression profiles from two S. aureus clinical strains (TCH1516 and LAC). ICA decomposed the S. aureus transcriptome into 29 independently modulated sets of genes (i-modulons) that revealed: 1) High confidence associations between 21 i-modulons and known regulators; 2) an association between an i-modulon and σS, whose regulatory role was previously undefined; 3) the regulatory organization of 65 virulence factors in the form of three i-modulons associated with AgrR, SaeR, and Vim-3; 4) the roles of three key transcription factors (CodY, Fur, and CcpA) in coordinating the metabolic and regulatory networks; and 5) a low-dimensional representation, involving the function of few transcription factors of changes in gene expression between two laboratory media (RPMI, cation adjust Mueller Hinton broth) and two physiological media (blood and serum). This representation of the TRN covers 842 genes representing 76% of the variance in gene expression that provides a quantitative reconstruction of transcriptional modules in S. aureus, and a platform enabling its full elucidation.


Assuntos
Regulação Bacteriana da Expressão Gênica , Redes Reguladoras de Genes/genética , Staphylococcus aureus/genética , Staphylococcus aureus/fisiologia , Transcriptoma , Proteínas de Bactérias/genética , Proteínas de Ligação a DNA/genética , Redes e Vias Metabólicas , Proteínas Repressoras/genética , Análise de Sequência de RNA , Fator sigma/genética , Infecções Estafilocócicas , Virulência/genética , Fatores de Virulência/genética
9.
J Vis Exp ; (160)2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32658180

RESUMO

Spine implant infections portend poor outcomes as diagnosis is challenging and surgical eradication is at odds with mechanical spinal stability. The purpose of this method is to describe a novel mouse model of spinal implant infection (SII) that was created to provide an inexpensive, rapid, and accurate in vivo tool to test potential therapeutics and treatment strategies for spinal implant infections. In this method, we present a model of posterior-approach spinal surgery in which a stainless-steel k-wire is transfixed into the L4 spinous process of 12-week old C57BL/6J wild-type mice and inoculated with 1 x 103 CFU of a bioluminescent strain of Staphylococcus aureus Xen36 bacteria. Mice are then longitudinally imaged for bioluminescence in vivo on post-operative days 0, 1, 3, 5, 7, 10, 14, 18, 21, 25, 28, and 35. Bioluminescence imaging (BLI) signals from a standardized field of view are quantified to measure in vivo bacterial burden. To quantify bacteria adhering to implants and peri-implant tissue, mice are euthanized and the implant and surrounding soft tissue are harvested. Bacteria are detached from the implant by sonication, cultured overnight and then colony forming units (CFUs) are counted. The results acquired from this method include longitudinal bacterial counts as measured by in vivo S. aureus bioluminescence (mean maximum flux) and CFU counts following euthanasia. While prior animal models of instrumented spine infection have involved invasive, ex vivo tissue analysis, the mouse model of SII presented in this paper leverages noninvasive, real time in vivo optical imaging of bioluminescent bacteria to replace static tissue study. Applications of the model are broad and may include utilizing alternative bioluminescent bacterial strains, incorporating other types of genetically engineered mice to contemporaneously study host immune response, and evaluating current or investigating new diagnostic and therapeutic modalities such as antibiotics or implant coatings.


Assuntos
Próteses e Implantes/microbiologia , Infecções Relacionadas à Prótese/microbiologia , Coluna Vertebral , Infecções Estafilocócicas , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Endogâmicos C57BL , Staphylococcus aureus/fisiologia
10.
Int J Infect Dis ; 96: 601-606, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32505877

RESUMO

BACKGROUND: Despite high mortality rates, physicians can alter the course of the Staphylococcus aureus bacteraemia (SAB) by following recommended standards of care. We aim to assess the adherence of these guidelines and their impact on mortality. METHODS: Substudy from a prospective cohort of hospitalized patients with SAB from three hospitals from Peru. Hazard ratios were calculated using Cox proportional regression to evaluate the association between 30-day mortality and the performance of standards of care: removal of central venous catheters (CVC), follow-up blood cultures, echocardiography, correct duration, and appropriate definitive antibiotic therapy. RESULTS: 150 cases of SAB were evaluated; 61.33% were MRSA. 30-day attributable mortality was 22.39%. CVC removal was done in 42.86% of patients. Follow-up blood cultures and echocardiograms were performed in 8% and 29.33% of cases, respectively. 81.33% of cases had appropriate empirical treatment, however, only 22.41% of MSSA cases were given appropriate definitive treatment, compared to 93.47% of MRSA. The adjusted regression for all-cause mortality found a substantial decrease in hazards when removing CVC (aHR 0.28, 95% CI: 0.10 - 0.74) and instituting appropriate definitive treatment (aHR 0.27, 95% CI: 0.08 - 0.86), while adjusting for standards of care, qPitt bacteraemia score, comorbidities, and methicillin susceptibility; similar results were found in the attributable mortality model (aHR 0.24, 95% CI: 0.08 - 0.70 and aHR 0.21, 95% CI: 0.06 - 0.71, respectively). CONCLUSIONS: Deficient adherence to standards of care was observed, especially definitive treatment for MSSA. CVC removal and the use of appropriate definitive antibiotic therapy reduced the hazard mortality of SAB.


Assuntos
Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Bacteriemia/mortalidade , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/mortalidade , Adulto , Idoso , Bacteriemia/epidemiologia , Bacteriemia/microbiologia , Estudos de Coortes , Comorbidade , Feminino , Fidelidade a Diretrizes , Humanos , Masculino , Pessoa de Meia-Idade , Peru/epidemiologia , Estudos Prospectivos , Padrão de Cuidado , Infecções Estafilocócicas/epidemiologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/fisiologia
11.
PLoS One ; 15(6): e0235093, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32584878

RESUMO

Bacterial biofilms are one of the major issues in the treatment of chronic infections such as chronic wounds, where biofilms are typically polymicrobial. The synergy between species can occur during most polymicrobial infections, where antimicrobial resistance enhances as a result. Furthermore, self-produced extracellular polymeric substance (EPS) in biofilms results in a high tolerance to antibiotics that complicates wound healing. Since most antibiotics fail to remove biofilms in chronic infections, new therapeutic modalities may be required. Disruption of EPS is one of the effective approaches for biofilm eradication. Therefore, degradation of EPS using enzymes may result in improved chronic wounds healing. In the current study, we investigated the efficacy of trypsin, ß-glucosidase, and DNase I enzymes on the degradation of dual-species biofilms of Pseudomonas aeruginosa and Staphylococcus aureus in a wound-like medium. These species are the two most common bacteria associated with biofilm formation in chronic wounds. Moreover, the reduction of minimum biofilm eradication concentration (MBEC) of meropenem and amikacin was evaluated when combined with enzymes. The minimum effective concentrations of trypsin, ß-glucosidase, and DNase I enzymes to degrade biofilms were 1 µg/ml, 8 U/ml, and 150 U/ml, respectively. Combination of 0.15 µg/ml trypsin and 50 U/ml DNase I had a significant effect on S. aureus-P. aeruginosa biofilms which resulted in the dispersal and dissolution of all biofilms. In the presence of the enzymatic mixture, MBECs of antibiotics showed a significant decrease (p < 0.05), at least 2.5 fold. We found that trypsin/DNase I mixture can be used as an anti-biofilm agent against dual-species biofilms of S. aureus-P. aeruginosa.


Assuntos
Antibacterianos/farmacologia , Biofilmes , Infecções por Pseudomonas/tratamento farmacológico , Pseudomonas aeruginosa/fisiologia , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/fisiologia , Infecção dos Ferimentos/tratamento farmacológico , Ferimentos e Lesões/microbiologia , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Quimioterapia Combinada , Humanos , Infecção dos Ferimentos/microbiologia
12.
PLoS One ; 15(6): e0234239, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32525884

RESUMO

Hummingbirds are essential pollinators in many ecosystems, making their conservation critical. As is the case with many species, hummingbirds are now facing a variety of challenges resulting from anthropogenic changes. As populations shift and species interactions change, disease is likely to pose a significant threat. There is a basic understanding of which pathogens currently affect a variety of hummingbird species, however there is a paucity of information about their immune systems capacity to kill pathogens and what specific factors may affect immunity. The objective of this study was to gain a basic understanding of the effect of age, sex, and molt on the constitutive innate immunity of hummingbirds. An in vitro assay was used to assess the microbiocidal capacity of the whole blood of Anna's Hummingbirds (Calypte anna) against three different microbes: Escherichia coli (E. coli), Staphylococcus aureus (S. aureus) and Candida albicans (C. albicans). The effect of age, sex and molt on anti-microbial capacity varied based on the microbe type. After-hatch-year birds tended to have better anti-microbial capacity compared to hatch-year birds. Male birds had higher anti-microbial activity than female birds, although this was not observed against C. albicans. Molting birds had a weaker antimicrobial activity against E. coli and S. aureus than birds that were not molting. These results represent an important first step towards defining the parameters of constitutive innate immunity of Anna's Hummingbirds as well as providing important knowledge about factors that should be considered when evaluating the health of wild populations.


Assuntos
Aves/sangue , Plasma/metabolismo , Animais , Aves/crescimento & desenvolvimento , Candida albicans/fisiologia , Escherichia coli/fisiologia , Feminino , Masculino , Viabilidade Microbiana , Muda , Caracteres Sexuais , Staphylococcus aureus/fisiologia
13.
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
14.
Nat Commun ; 11(1): 2211, 2020 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-32371860

RESUMO

Microbial biofilm formation on indwelling medical devices causes persistent infections that cannot be cured with conventional antibiotics. To address this unmet challenge, we engineer tunable active surface topographies with micron-sized pillars that can beat at a programmable frequency and force level in an electromagnetic field. Compared to the flat and static controls, active topographies with the optimized design prevent biofilm formation and remove established biofilms of uropathogenic Escherichia coli (UPEC), Pseudomonas aeruginosa, and Staphylococcus aureus, with up to 3.7 logs of biomass reduction. In addition, the detached biofilm cells are found sensitized to bactericidal antibiotics to the level comparable to exponential-phase planktonic cells. Based on these findings, a prototype catheter is engineered and found to remain clean for at least 30 days under the flow of artificial urine medium, while the control catheters are blocked by UPEC biofilms within 5 days.


Assuntos
Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Pseudomonas aeruginosa/fisiologia , Staphylococcus aureus/fisiologia , Escherichia coli Uropatogênica/fisiologia , Infecções Bacterianas/microbiologia , Infecções Bacterianas/prevenção & controle , Biofilmes/efeitos dos fármacos , Biomassa , Campos Eletromagnéticos , Testes de Sensibilidade Microbiana/métodos , Infecções Relacionadas à Prótese/microbiologia , Infecções Relacionadas à Prótese/prevenção & controle , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/metabolismo , Fatores de Tempo , Escherichia coli Uropatogênica/efeitos dos fármacos , Escherichia coli Uropatogênica/metabolismo
15.
PLoS One ; 15(5): e0231168, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32365127

RESUMO

Staphylococcus aureus is one of the most important pathogens causing mastitis in dairy cows. The objective of this study was to establish a rat model of mastitis induced by S. aureus infection and to explore changes in the proteomes of mammary tissue in different udder states, providing a better understanding of the host immune response to S. aureus mastitis. On day 3 post-partum, 6 rats were randomly divided into two groups (n = 3), with either 100 µL of PBS (blank group) or a S. aureus suspension containing 2×107 CFU·mL-1 (challenge group) infused into the mammary gland duct. After 24 h of infection, the rats were sacrificed, and mammary gland tissue was collected. Tandem mass tag (TMT)-based technology was applied to compare the proteomes of healthy and mastitic mammary tissues. Compared with the control group, the challenge group had 555 proteins with significant differences in expression, of which 428 were significantly upregulated (FC>1.2 and p<0.05) and 127 were downregulated (FC>0.83 and p<0.05 or p<0.01). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that upregulated differentially significant expressed proteins (DSEPs) were associated with mainly immune responses, including integrin alpha M, inter-α-trypsin inhibitor heavy chain 4, and alpha-2-macroglobulin. This study is the first in which a rat model of S. aureus-induced mastitis was used to explore the proteins related to mastitis in dairy cows by TMT technology, providing a model for replication of dairy cow S. aureus-induced mastitis experiments.


Assuntos
Glândulas Mamárias Animais/metabolismo , Mastite/metabolismo , Proteoma/análise , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/fisiologia , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia Líquida , Feminino , Glândulas Mamárias Animais/microbiologia , Glândulas Mamárias Animais/patologia , Mastite/microbiologia , Mastite/patologia , Gravidez , Proteoma/metabolismo , Proteômica/métodos , Ratos , Ratos Sprague-Dawley , Infecções Estafilocócicas/patologia , Espectrometria de Massas em Tandem
16.
PLoS One ; 15(5): e0233973, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32470063

RESUMO

Bacterial biofilms are organized sessile communities of bacteria enclosed in extracellular polymeric substances (EPS). To analyze organization of bacteria and EPS in high resolution and high magnification by scanning electron microscopy (SEM), it is important to preserve the complex architecture of biofilms. Therefore, fixation abilities of formalin, glutaraldehyde, and Methacarn (methanol/chloroform/acetic acid-6:3:1) fixatives were evaluated to identify which fixative would best preserve the complex structure of bacterial biofilms. Economically important Gram-negative Mannheimia haemolytica, the major pathogen associated with bovine respiratory disease complex, and Gram-positive Staphylococcus aureus, the major cause of chronic mastitis in cattle, bacteria were selected since both form biofilms on solid-liquid interface. For SEM analysis, round glass coverslips were placed into the wells of 24-well plates and diluted M. haemolytica or S. aureus cultures were added, and incubated at 37°C for 48-72 h under static growth conditions. Culture media were aspirated and biofilms were fixed with an individual fixative for 48 h. SEM examination revealed that all three fixatives were effective preserving the bacterial cell morphology, however only Methacarn fixative could consistently preserve the complex structure of biofilms. EPS layers were clearly visible on the top, in the middle, and in the bottom of the biofilms with Methacarn fixative. Biomass and three-dimensional structure of the biofilms were further confirmed spectrophotometrically following crystal violet staining and by confocal microscopy after viability staining. These findings demonstrate that Methacarn fixative solution is superior to the other fixatives evaluated to preserve the complex architecture of biofilms grown on glass coverslips for SEM evaluation.


Assuntos
Biofilmes , Mannheimia haemolytica/fisiologia , Mannheimia haemolytica/ultraestrutura , Microscopia Eletrônica de Varredura , Staphylococcus aureus/fisiologia , Staphylococcus aureus/ultraestrutura , Biomassa , Viabilidade Microbiana
17.
Proc Natl Acad Sci U S A ; 117(23): 12598-12605, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32457154

RESUMO

The mechano-bactericidal activity of nanostructured surfaces has become the focus of intensive research toward the development of a new generation of antibacterial surfaces, particularly in the current era of emerging antibiotic resistance. This work demonstrates the effects of an incremental increase of nanopillar height on nanostructure-induced bacterial cell death. We propose that the mechanical lysis of bacterial cells can be influenced by the degree of elasticity and clustering of highly ordered silicon nanopillar arrays. Herein, silicon nanopillar arrays with diameter 35 nm, periodicity 90 nm and increasing heights of 220, 360, and 420 nm were fabricated using deep UV immersion lithography. Nanoarrays of 360-nm-height pillars exhibited the highest degree of bactericidal activity toward both Gram stain-negative Pseudomonas aeruginosa and Gram stain-positive Staphylococcus aureus bacteria, inducing 95 ± 5% and 83 ± 12% cell death, respectively. At heights of 360 nm, increased nanopillar elasticity contributes to the onset of pillar deformation in response to bacterial adhesion to the surface. Theoretical analyses of pillar elasticity confirm that deflection, deformation force, and mechanical energies are more significant for the substrata possessing more flexible pillars. Increased storage and release of mechanical energy may explain the enhanced bactericidal action of these nanopillar arrays toward bacterial cells contacting the surface; however, with further increase of nanopillar height (420 nm), the forces (and tensions) can be partially compensated by irreversible interpillar adhesion that reduces their bactericidal effect. These findings can be used to inform the design of next-generation mechano-responsive surfaces with tuneable bactericidal characteristics for antimicrobial surface technologies.


Assuntos
Antibacterianos/farmacologia , Nanoestruturas/química , Estresse Mecânico , Antibacterianos/química , Aderência Bacteriana , Elasticidade , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia , Silício/química , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia
18.
Int J Food Microbiol ; 326: 108653, 2020 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-32449679

RESUMO

Staphylococcus xylosus, a coagulase-negative Staphylococcus, is frequently isolated from food products of animal origin and used as a starter culture in these products in which it contributes to their flavour, while Staphylococcus aureus, a coagulase-positive bacterium, causes foodborne intoxication and is implicated in a broad diversity of infections in medical sector, notably in nosocomial infections. S. xylosus and S. aureus are both capable of forming a biofilm and share the same ecological niches, thus we explored their interaction in biofilms with a view to limiting the risks associated with S. aureus. Cell-free supernatants of different strains of S. xylosus were able to inhibit the biofilm formation of S. aureus. The S. xylosus C2a strain released into the supernatant a molecule of molecular weight above 30 kDa that is resistant to proteolytic enzymes and inhibits the formation of S. aureus MW2 biofilm, though the mechanism involved has yet to be elucidated. Furthermore, S. xylosus C2a modified the architecture of S. aureus MW2 in co-culture biofilm. Confocal laser scanning microscopy revealed that S. aureus formed a biofilm with a flat and compact structure while in co-culture with S. xylosus the two species formed large juxtaposed aggregates throughout the period of incubation. This architecture made the S. aureus biofilm more susceptible to detachment.


Assuntos
Antibiose/fisiologia , Biofilmes/crescimento & desenvolvimento , Staphylococcus aureus/fisiologia , Staphylococcus/fisiologia , Animais , Coagulase , Microbiologia de Alimentos , Doenças Transmitidas por Alimentos/microbiologia , Doenças Transmitidas por Alimentos/prevenção & controle , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/prevenção & controle
19.
PLoS One ; 15(5): e0232987, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32407399

RESUMO

Escherichia coli and Staphylococcus aureus are important agents of urinary tract infections that can often evolve to severe infections. The rise of antibiotic-resistant strains has driven the search for novel therapies to replace the use or act as adjuvants of antibiotics. In this context, plant-derived compounds have been widely investigated. Cuminaldehyde is suggested as the major antimicrobial compound of the cumin seed essential oil. However, this effect is not fully understood. Herein, we investigated the in silico and in vitro activities of cuminaldehyde, as well as its ability to potentiate ciprofloxacin effects against S. aureus and E. coli. In silico analyses were performed by using different computational tools. The PASS online and SwissADME programmes were used for the prediction of biological activities and oral bioavailability of cuminaldehyde. For analysis of the possible toxic effects and the theoretical pharmacokinetic parameters of the compound, the Osiris, SwissADME and PROTOX programmes were used. Estimations of cuminaldehyde gastrointestinal absorption, blood brain barrier permeability and skin permeation by using SwissADME; and drug likeness and score by using Osiris, were also evaluated The in vitro antimicrobial effects of cuminaldehyde were determined by using microdilution, biofilm formation and time-kill assays. In silico analysis indicated that cuminaldehyde may act as an antimicrobial and as a membrane permeability enhancer. It was suggested to be highly absorbable by the gastrointestinal tract and likely to cross the blood brain barrier. Also, irritative and harmful effects were predicted for cuminaldehyde if swallowed at its LD50. Good oral bioavailability and drug score were also found for this compound. Cuminaldehyde presented antimicrobial and anti-biofilm effects against S. aureus and E. coli.. When co-incubated with ciprofloxacin, it enhanced the antibiotic antimicrobial and anti-biofilm actions. We suggest that cuminaldehyde may be useful as an adjuvant therapy to ciprofloxacin in S. aureus and E. coli-induced infections.


Assuntos
Antibacterianos/administração & dosagem , Benzaldeídos/administração & dosagem , Ciprofloxacino/administração & dosagem , Cimenos/administração & dosagem , Escherichia coli/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Adjuvantes Farmacêuticos/administração & dosagem , Adjuvantes Farmacêuticos/farmacocinética , Adjuvantes Farmacêuticos/toxicidade , Administração Oral , Benzaldeídos/farmacocinética , Benzaldeídos/toxicidade , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Disponibilidade Biológica , Simulação por Computador , Cimenos/farmacocinética , Cimenos/toxicidade , Sinergismo Farmacológico , Escherichia coli/patogenicidade , Escherichia coli/fisiologia , Infecções por Escherichia coli/tratamento farmacológico , Humanos , Técnicas In Vitro , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/patogenicidade , Staphylococcus aureus/fisiologia , Infecções Urinárias/tratamento farmacológico
20.
J Med Chem ; 63(10): 5287-5296, 2020 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-32343145

RESUMO

We report herein the syntheses of 79 derivatives of the 4(3H)-quinazolinones and their structure-activity relationship (SAR) against methicillin-resistant Staphylococcus aureus (MRSA). Twenty one analogs were further evaluated in in vitro assays. Subsequent investigation of the pharmacokinetic properties singled out compound 73 ((E)-3-(5-carboxy-2-fluorophenyl)-2-(4-cyanostyryl)quinazolin-4(3H)-one) for further study. The compound synergized with piperacillin-tazobactam (TZP) both in vitro and in vivo in a clinically relevant mouse model of MRSA infection. The TZP combination lacks activity against MRSA, yet it synergized with compound 73 to kill MRSA in a bactericidal manner. The synergy is rationalized by the ability of the quinazolinones to bind to the allosteric site of penicillin-binding protein (PBP)2a, resulting in opening of the active site, whereby the ß-lactam antibiotic now is enabled to bind to the active site in its mechanism of action. The combination effectively treats MRSA infection, for which many antibiotics (including TZP) have faced clinical obsolescence.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Quinazolinonas/química , Quinazolinonas/farmacologia , Animais , Antibacterianos/uso terapêutico , Feminino , Camundongos , Camundongos Endogâmicos ICR , Testes de Sensibilidade Microbiana/métodos , Neutropenia/tratamento farmacológico , Neutropenia/microbiologia , Quinazolinonas/uso terapêutico , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia , Relação Estrutura-Atividade
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