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1.
PLoS Biol ; 18(8): e3000814, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32797039

RESUMO

Plasmid-mediated horizontal gene transfer of antibiotic resistance and virulence in pathogenic bacteria underlies a major public health issue. Understanding how, in the absence of antibiotic-mediated selection, plasmid-bearing cells avoid being outnumbered by plasmid-free cells is key to developing counterstrategies. Here, we quantified the induction of the plasmidial sex pheromone pathway of Enterococcus faecalis to show that the integration of the stimulatory (mate-sensing) and inhibitory (self-sensing) signaling modules from the pCF10 conjugative plasmid provides a precise measure of the recipient-to-donor ratio, agnostic to variations in population size. Such ratiometric control of conjugation favors vertical plasmid transfer under low mating likelihood and allows activation of conjugation functions only under high mating likelihood. We further show that this strategy constitutes a cost-effective investment into mating effort because overstimulation produces unproductive self-aggregation and growth rate reduction. A mathematical model suggests that ratiometric control of conjugation increases plasmid fitness and predicts a robust long-term, stable coexistence of donors and recipients. Our results demonstrate how population-level parameters can control transfer of antibiotic resistance in bacteria, opening the door for biotic control strategies.


Assuntos
Proteínas de Bactérias/genética , Resistência Microbiana a Medicamentos/genética , Enterococcus faecalis/genética , Transferência Genética Horizontal , Feromônios/genética , Percepção de Quorum/efeitos dos fármacos , Antibacterianos/farmacologia , Carga Bacteriana , Proteínas de Bactérias/metabolismo , Conjugação Genética , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/crescimento & desenvolvimento , Enterococcus faecalis/metabolismo , Expressão Gênica , Aptidão Genética , Modelos Estatísticos , Feromônios/biossíntese , Plasmídeos/química , Plasmídeos/metabolismo , Percepção de Quorum/genética , Virulência
2.
Surg Clin North Am ; 100(4): 757-776, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32681875

RESUMO

This review of the literature concerning bacteria, antibiotics and tissue repair shows there are extensive data supporting microbial interference with wound healing once bacterial burden exceeds 104 CFU per unit of measure, The mechanism of bacterial interference lies largely in prolonging the inflammatory phase of tissue repair. Reducing the microbial bioburden allows tissue repair to continue. Systemic and topical antimicrobials appear critical to reducing the bioburden and facilitating repair. The current controversy over the use of antimicrobials in patients with chronically infected wounds, in particular, revolves around the definition of infection. The reliance on classic clinical signs of inflammation to support antimicrobial use in these patients is tenuous due to the lack of correlation of these signs with the microbial burden known to impair tissue repair.


Assuntos
Antibacterianos/uso terapêutico , Infecções Bacterianas/tratamento farmacológico , Cicatrização/fisiologia , Infecção dos Ferimentos/tratamento farmacológico , Infecções Bacterianas/fisiopatologia , Carga Bacteriana/fisiologia , Biofilmes , Hipóxia Celular/fisiologia , Humanos , Neutrófilos/fisiologia , Infecção dos Ferimentos/fisiopatologia
3.
BMC Infect Dis ; 20(1): 505, 2020 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-32660552

RESUMO

BACKGROUND: Meningococcal meningitis (MM) is a life-threatening disease associated with approximately 10% case fatality rates and neurological sequelae in 10-20% of the cases. Recently, we have shown that the matrix metalloproteinase (MMP) inhibitor BB-94 reduced brain injury in a mouse model of MM. The present study aimed to assess whether doxycycline (DOX), a tetracycline that showed a neuroprotective effect as adjuvant therapy in experimental pneumococcal meningitis (PM), would also exert a beneficial effect when given as adjunctive therapy to ceftriaxone (CRO) in experimental MM. METHODS: BALB/c mice were infected by the intracisternal route with a group C Neisseria meningitidis strain. Eighteen h post infection (hpi), animals were randomised for treatment with CRO [100 mg/kg subcutaneously (s.c.)], CRO plus DOX (30 mg/kg s.c.) or saline (control s.c.). Antibiotic treatment was repeated 24 and 40 hpi. Mouse survival and clinical signs, bacterial counts in cerebella, brain damage, MMP-9 and cyto/chemokine levels were assessed 48 hpi. RESULTS: Analysis of bacterial load in cerebella indicated that CRO and CRO + DOX were equally effective at controlling meningococcal replication. No differences in survival were observed between mice treated with CRO (94.4%) or CRO + DOX (95.5%), (p > 0.05). Treatment with CRO + DOX significantly diminished both the number of cerebral hemorrhages (p = 0.029) and the amount of MMP-9 in the brain (p = 0.046) compared to untreated controls, but not to CRO-treated animals (p > 0.05). Levels of inflammatory markers in the brain of mice that received CRO or CRO + DOX were not significantly different (p > 0.05). Overall, there were no significant differences in the parameters assessed between the groups treated with CRO alone or CRO + DOX. CONCLUSIONS: Treatment with CRO + DOX showed similar bactericidal activity to CRO in vivo, suggesting no antagonist effect of DOX on CRO. Combined therapy significantly improved mouse survival and disease severity compared to untreated animals, but addition of DOX to CRO did not offer significant benefits over CRO monotherapy. In contrast to experimental PM, DOX has no adjunctive activity in experimental MM.


Assuntos
Antibacterianos/uso terapêutico , Ceftriaxona/uso terapêutico , Doxiciclina/uso terapêutico , Meningite Meningocócica/tratamento farmacológico , Neisseria meningitidis Sorogrupo C , Animais , Antibacterianos/administração & dosagem , Carga Bacteriana/efeitos dos fármacos , Ceftriaxona/administração & dosagem , Hemorragia Cerebral/tratamento farmacológico , Quimiocinas/análise , Quimiocinas/metabolismo , Modelos Animais de Doenças , Doxiciclina/administração & dosagem , Quimioterapia Combinada , Feminino , Humanos , Estimativa de Kaplan-Meier , Metaloproteinase 9 da Matriz/análise , Metaloproteinase 9 da Matriz/metabolismo , Meningite Meningocócica/mortalidade , Camundongos , Camundongos Endogâmicos BALB C , Distribuição Aleatória , Resultado do Tratamento
4.
J Endod ; 46(9): 1228-1234, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32653532

RESUMO

INTRODUCTION: This randomized clinical study compared the in vivo antibacterial efficacy of Reciproc Blue (RB), XP-endo Shaper (XP-S), and XP-endo Shaper associated with XP-endo Finisher (XP-F) systems in infected oval-shaped root canals with primary apical periodontitis. METHODS: In this study, 28 human teeth with a single root and a single canal were randomly assigned to 2 groups according to the instrumentation technique: group 1, RB (n = 14) and group 2, XP-endo (XP-S and XP-F, n = 14). The single-rooted teeth were prepared by reciprocating and rotary nickel-titanium instruments with 5.25% sodium hypochlorite irrigation. Samples were collected from the canal at the baseline (S1), after chemomechanical preparation (S2), and after XP-F instrumentation (S3). The DNA extracts were subjected to quantitative analysis for total bacterial counts by quantitative real-time polymerase chain reaction. The data were analyzed using the analysis of variance test, and the level of significance was set at 5%. RESULTS: All samples tested positive for the presence of bacteria at baseline, and the bacterial counts substantially reduced after treatment procedures (P < .01). The results showed no statistical difference between RB and XP-S instrumentation with respect to the bacterial reduction (P > .05). A marked bacterial reduction was observed after the use of the XP-F instrument (P < .01). CONCLUSIONS: The XP-S and RB systems sharply reduced the bacterial load in oval-shaped root canals with primary apical periodontitis. XP-F used as a supplementary instrument to chemomechanical preparation promoted a significantly higher bacterial reduction.


Assuntos
Periodontite Periapical , Preparo de Canal Radicular , Carga Bacteriana , Cavidade Pulpar , Humanos , Irrigantes do Canal Radicular , Hipoclorito de Sódio
5.
J Dent Res ; 99(11): 1228-1238, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32660314

RESUMO

The aim of this systematic review and network meta-analysis was to identify and rank the effectiveness of different interventions used in dental practice to reduce microbial load in aerosolized compounds. Seven electronic databases were searched to April 6, 2020, for randomized controlled trials (RCTs) or nonrandomized prospective studies in the field. Study selection, data extraction, and risk-of-bias assessment were performed for all included studies, while the outcome of interest pertained to differences in bacterial load quantification through the use of different interventions prior to aerosol-generating procedures in dental practices. Random effects frequentist network meta-analysis was performed, with mean difference (MD) and 95% CI as the effect measure. Confidence in the documented evidence was assessed through the newly fueled CINeMA framework (Confidence in Network Meta-analysis) based on the GRADE approach (Grading of Recommendations, Assessment, Development and Evaluation). Twenty-nine clinical trials were deemed eligible, 21 RCTs and 8 nonrandomized studies, while 11 RCTs contributed to the network meta-analysis, comprising 10 competing interventions. Tempered chlorhexidine (CHX) 0.2% as compared with nonactive control mouth rinse, prior to routine ultrasonic scaling, was most effective toward reduced postprocedural bacterial load with an MD of -0.92 (95% CI, -1.54 to -0.29) in log10 bacterial CFUs (colony-forming units). For CHX 0.2%, an MD of -0.74 (95% CI, -1.07 to -0.40) was observed as compared with control. Tempered CHX 0.2% presented the highest probabilities of being ranked the most effective treatment (31.2%). Level of confidence varied from very low to moderate across all formulated comparisons. These findings summarize the current state of research evidence in the field of aerosolized bacteria in dentistry. Instigated by the era of SARS-CoV-2 pandemic, the stipulation of a broader evaluation of the aerosolized microbes, including viruses, potentially coupled with disinfectant-based prevention schemes should be prioritized.


Assuntos
Aerossóis , Infecções por Coronavirus/prevenção & controle , Odontologia/métodos , Desinfetantes , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Carga Bacteriana , Betacoronavirus , Humanos , Metanálise em Rede , Estudos Prospectivos , Ensaios Clínicos Controlados Aleatórios como Assunto , Carga Viral
6.
PLoS Biol ; 18(6): e3000644, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32511236

RESUMO

Mucosa-associated invariant T (MAIT) cells are abundant antimicrobial T cells in humans and recognize antigens derived from the microbial riboflavin biosynthetic pathway presented by the MHC-Ib-related protein (MR1). However, the mechanisms responsible for MAIT cell antimicrobial activity are not fully understood, and the efficacy of these mechanisms against antibiotic resistant bacteria has not been explored. Here, we show that MAIT cells mediate MR1-restricted antimicrobial activity against Escherichia coli clinical strains in a manner dependent on the activity of cytolytic proteins but independent of production of pro-inflammatory cytokines or induction of apoptosis in infected cells. The combined action of the pore-forming antimicrobial protein granulysin and the serine protease granzyme B released in response to T cell receptor (TCR)-mediated recognition of MR1-presented antigen is essential to mediate control against both cell-associated and free-living, extracellular forms of E. coli. Furthermore, MAIT cell-mediated bacterial control extends to multidrug-resistant E. coli primary clinical isolates additionally resistant to carbapenems, a class of last resort antibiotics. Notably, high levels of granulysin and granzyme B in the MAIT cell secretomes directly damage bacterial cells by increasing their permeability, rendering initially resistant E. coli susceptible to the bactericidal activity of carbapenems. These findings define the role of cytolytic effector proteins in MAIT cell-mediated antimicrobial activity and indicate that granulysin and granzyme B synergize to restore carbapenem bactericidal activity and overcome carbapenem resistance in E. coli.


Assuntos
Antígenos de Diferenciação de Linfócitos T/metabolismo , Carbapenêmicos/farmacologia , Citotoxicidade Imunológica , Farmacorresistência Bacteriana/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Granzimas/metabolismo , Células T Invariáveis Associadas à Mucosa/imunologia , Anti-Infecciosos/farmacologia , Carga Bacteriana/efeitos dos fármacos , Citotoxicidade Imunológica/efeitos dos fármacos , Células HeLa , Humanos , Cinética
7.
Niger J Clin Pract ; 23(6): 783-791, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32525112

RESUMO

Background: Microorganisms in the mouth are protected from negative environmental conditions by forming biofilms; however, the use of anti-plaque agents in children is not preferred due to toxic side effects. Green tea has been reported to have anti-microbial and anti-dental caries properties. Aims: The aim of this study was to assess the ability of green tea extract to prevent the formation of biofilm on the teeth of children using space maintainers. Methods: Bacteria were isolated from samples obtained from children aged between 8 and 10 years. The micro-titer plate method and Congo red agar were used to assay biofilm formation. Green tea leaves were obtained from Rize, Turkey. Methanol, hexane and distilled water were used for preparing the extracts. The effects of green tea extract and chlorhexidine on biofilm formation were examined using scanning electron microscopy. Results: Presence of S. mutans 3,3, S. anginosus 2.1.b, S. dysgalactie 6.1.4.1, and E. faecium 10.2. was measured in the biofilm samples. The extracts showed a bacteriostatic effect on the test bacteria, and among the green tea extracts, the methanol extract was found to exhibit the highest efficacy against biofilm formation by S. mutans 3.3. Conclusion: Green tea extract showed good efficacy in controlling bacterial growth, and is recommended as a better-tasting alternative for daily oral hygiene due to a lack of known side effects.


Assuntos
Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Clorexidina/farmacologia , Cárie Dentária/microbiologia , Braquetes Ortodônticos/microbiologia , Extratos Vegetais/farmacologia , Infecções Estreptocócicas/tratamento farmacológico , Streptococcus mutans/efeitos dos fármacos , Chá/química , Antioxidantes/farmacologia , Carga Bacteriana/efeitos dos fármacos , Criança , Cárie Dentária/tratamento farmacológico , Humanos , Metanol , Testes de Sensibilidade Microbiana , Boca , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Solventes/química , Streptococcus mutans/isolamento & purificação , Streptococcus mutans/fisiologia , Turquia
8.
PLoS Negl Trop Dis ; 14(6): e0008051, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32569298

RESUMO

BACKGROUND: In Japan, Buruli ulcer cases are often advanced, requiring surgical treatment. However, extensive debridement is often difficult because of cosmetic and functional sequelae. Moreover, the lesions are complicated and composed of edematous erythema, necrotic ulcer, and erythematous skin lesions caused by a paradoxical reaction, which also make it difficult to perform adequate debridement. METHODOLOGY/PRINCIPAL FINDINGS: We performed quantitative polymerase chain reaction (PCR) analysis for IS2404 using 29 samples taken from mapping biopsy. We evaluated the relationship among mycobacterial burden, histopathological findings, and clinical outcomes using 83 tissue samples taken from mapping biopsy and debrided Buruli ulcer. On quantitative PCR, the Cp values of IS2404 amplification were substantially different in each site. The major histological findings could be divided into massive subcutaneous necrosis with scant inflammatory cell infiltration and dense inflammatory cell infiltration. Of the 84 sites, 34 were subjected to repeated histological evaluations. In these sites, histological necrosis did not disappear over time despite standard antibiotic treatment. In contrast, the ulcers were cured and no recurrences were observed without resecting the 11 biopsied sites that lacked histological necrosis. Although quantitative PCR revealed that a lower Cp value of IS2404 was associated with histological massive necrosis, sites that showed lower Cp values clinically did not always need debridement. CONCLUSION/SIGNIFICANCE: Our descriptive study revealed that the histological findings and amounts of mycobacterial DNA differed according to the sites despite being found in one lesion. Our results showed that the need for surgical debridement in each site was correlated with histological necrosis without inflammatory cell infiltration, as the inflammation is supposed to represent an active host immune response rather than mycobacterial burden. We suggest that the debridement of lesions with histological necrosis in mapping biopsy may be useful for Japanese cases with unsuccessful standard antibiotic treatment to achieve sufficient clinical improvement.


Assuntos
Carga Bacteriana , Úlcera de Buruli/microbiologia , Úlcera de Buruli/patologia , Histocitoquímica , Mycobacterium ulcerans/isolamento & purificação , Adulto , Biópsia , Elementos de DNA Transponíveis , DNA Bacteriano/análise , DNA Bacteriano/genética , Humanos , Japão , Masculino , Mycobacterium ulcerans/genética , Reação em Cadeia da Polimerase em Tempo Real , Adulto Jovem
9.
PLoS One ; 15(6): e0235006, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32559258

RESUMO

Distal limb wounds are common injuries sustained by horses and their healing is fraught with complications due to equine anatomy, prevalence of infection, and challenges associated with wound management. Gallium is a semi-metallic element that has been shown to possess antimicrobial properties and aid in wound healing in various preclinical models. The effects of Gallium have not been studied in equine wound healing. Therefore, the objective of this study was to compare healing rates between gallium-treated and untreated wounds of equine distal limbs and to demonstrate the antimicrobial effects of gallium on wounds inoculated with S. aureus. Using an established model of equine wound healing we demonstrated beneficial effects of 0.5% topical gallium maltolate on equine wound healing. Specifically we documented reduced healing times, reduced bioburden, and reduced formation of exuberant granulation tissue in wounds treated with gallium maltolate as compared with untreated wounds. Gallium appeared to exert its beneficial effects via its well-described antimicrobial actions as well as by altering the expression of specific genes known to be involved in wound healing of horses and other animals. Specifically, gallium maltolate appeared to increase expression of transforming growth factor-ß in both infected and un-infected wounds. Further work is needed to document the effects of gallium on naturally occurring equine wounds and to compare the effects of gallium with other wound treatment options. These data, however, suggest that gallium may be an attractive and novel means of improving equine distal limb wound healing.


Assuntos
Antibacterianos/uso terapêutico , Doenças dos Cavalos/tratamento farmacológico , Traumatismos da Perna/tratamento farmacológico , Compostos Organometálicos/uso terapêutico , Pironas/uso terapêutico , Infecções Estafilocócicas/tratamento farmacológico , Administração Tópica , Animais , Antibacterianos/administração & dosagem , Carga Bacteriana , Citocinas/genética , Citocinas/metabolismo , Doenças dos Cavalos/metabolismo , Cavalos , Traumatismos da Perna/metabolismo , Traumatismos da Perna/veterinária , Compostos Organometálicos/administração & dosagem , Pironas/administração & dosagem , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/veterinária , Cicatrização
10.
PLoS One ; 15(6): e0234127, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32492060

RESUMO

Several studies have shown the ubiquitous presence of bacteria in hospital surfaces, staff, and patients. Frequently, these bacteria are related to HAI (healthcare-associated infections) and carry antimicrobial resistance (AMR). These HAI-related bacteria contribute to a major public health issue by increasing patient morbidity and mortality during or after hospital stay. Bacterial high-throughput amplicon gene sequencing along with identification of AMR genes, as well as whole genome sequencing (WGS), are biotechnological tools that allow multiple-sample screening for a diversity of bacteria. In this paper, we used these methods to perform a one-year cross sectional profiling of bacteria and AMR genes in adult and neonatal intensive care units (ICU and NICU) in a Brazilian public, tertiary hospital. Our results showed high abundances of HAI-related bacteria such as S. epidermidis, S. aureus, K. pneumoniae, A. baumannii complex, E. coli, E. faecalis, and P. aeruginosa in patients and hospital surfaces. Most abundant AMR genes detected throughout ICU and NICU were mecA, blaCTX-M-1 group, blaSHV-like, and blaKPC-like. We found that NICU environment and patients were more widely contaminated with pathogenic bacteria than ICU. Patient samples, despite the higher bacterial load, have lower bacterial diversity than environmental samples in both units. Finally, we also identified contamination hotspots in the hospital environment showing constant frequencies of bacterial and AMR contamination throughout the year. Whole genome sequencing (WGS), 16S rRNA oligotypes, and AMR identification allowed a high-resolution characterization of the hospital microbiome profile.


Assuntos
Bactérias/genética , Farmacorresistência Bacteriana/genética , Adulto , Antibacterianos/farmacologia , Anti-Infecciosos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/isolamento & purificação , Carga Bacteriana , Brasil , Infecção Hospitalar/microbiologia , Infecção Hospitalar/patologia , Estudos Transversais , Farmacorresistência Bacteriana/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Humanos , Recém-Nascido , Unidades de Terapia Intensiva , Unidades de Terapia Intensiva Neonatal , RNA Ribossômico 16S/química , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/isolamento & purificação , Centros de Atenção Terciária , Sequenciamento Completo do Genoma
11.
An Real Acad Farm ; 86(2): 117-124, abr.-jun. 2020. graf, ilus, tab
Artigo em Espanhol | IBECS | ID: ibc-193547

RESUMO

Uno de los quesos frescos tradicionales típicos del Ecuador es el queso de hoja, un queso de pasta hilada, que se elabora de manera artesanal envuelto en hoja de achira (Canna indica), mientras en la industria es empacado al vacío en bolsas de polietileno. En el presente estudio se comparó la calidad microbiológica de estos quesos, evaluando los indicadores de la calidad higiénico-sanitaria, además se cuantificaron y caracterizaron fenotípicamente bacterias ácido lácticas (BAL). Las muestras se recolectaron en queseras artesanales y plantas industriales de la ciudad de Latacunga. Se determinaron recuentos de aerobios mesófilos aplicando la Norma Técnica Ecuatoriana NTE INEN 1529 5, coliformes totales, Escherichia coli y Staphylococcus aureus se analizaron utilizando placas Petrifilm (AOAC 991.14 - AOAC 2003.07). El recuento de bacterias ácido lácticas empleó el método PRT-712.02-047. Los resultados de los indicadores de la calidad higiénico-sanitaria: coliformes, E. coli y S. aureus examinados en los quesos de hoja artesanales y de elaboración industrial, en todos los casos superaron los límites de aceptabilidad establecidos por la normativa ecuatoriana, sugiriendo deficiente calidad higiénica de los procesos o incorrecta manipulación de la leche empleada como materia prima. Los recuentos de Lactobacillus y Lactococcus en los quesos industriales presentaron una reducción estadísticamente significativa comparados con los recuentos de estos microorganismos en los quesos artesanales, esto corresponde a una diferencia en porcentaje de 18,15 % y 14,27 % respectivamente. Se aisló un total de 32 cepas de bacterias de ácido láctico, estas mostraron características fenotípicas similares, pero tuvieron una respuesta diferente a la tolerancia a niveles de pH (4.4; 9.4) y NaCl (6.5 %). Es importante complementar la evaluación microbiológica con un análisis sensorial para evaluar el efecto de las BAL sobre las características organolépticas de los quesos de hoja de procedencia artesanal e industrial


One of the traditional fresh cheeses in Ecuador is the artisanal leafcheese, a kind of stretched-curd cheese. The artisanal product is wrapped in achira leaves (Canna indica), while the industrial leafcheese is packed at vacuum in high density polyethylene bags. In this study the microbiological quality of both products was compared. The hygienic-sanitary microbial indicators and lactic acid bacteria (LAB) were quantified. The LAB isolated were characterized phenotypically. The samples were obtained from artisanal cheese-making and industrial located in Latacunga city province Cotopaxi. The total aerobic mesophilic count was made based on national regulations (NTE INEN 1529 5); total coliforms, Escherichia coli and Staphylococcus aureus was evaluated using petrifilm methods (AOAC 991.14 - AOAC 2003.07) and to LAB was used PRT-712.02-047. The results show high quantities of total coliforms, E. coli and S. aureus in both products, these data exceed the limits of acceptability established in Ecuadorian regulations, this evidence poor hygienic quality of the processes or incorrect controls of milk as raw material. The lactic acid bacteria count showed statistically significant differences, the industrial cheeses had a reduction of 18,15% of Lactobacillus and 14,27% of Lactococcus compared with artisanal cheeses. A total of 32 strains of lactic acid bacteria were isolated, these showed similar phenotypic characteristics, but these had a different response at the level of pH (4,4;9,4) and NaCl (6,5%). The sensory evaluation will be an important complement in this type of study


Assuntos
Queijo/microbiologia , Microbiologia de Alimentos/normas , Coliformes , Escherichia coli/isolamento & purificação , Staphylococcus aureus/isolamento & purificação , Carga Bacteriana , Equador , Queijo/normas , Escherichia coli/genética , Staphylococcus aureus/genética , Fenótipo
12.
BMC Infect Dis ; 20(1): 331, 2020 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-32393186

RESUMO

BACKGROUND: IL-6 was associated with the severity of mycoplasma pneumoniae pneumonia (MPP). But the relationship between IL-27 and MPP was unknown. METHODS: Ninety-eight patients with MPP < 14 years old were enrolled in this study and divided into groups by severity (mild cases and severe cases), infection types (MP single infection group and MP mixed infection group) and DNA loads (low MP DNA loads group and high MP DNA loads group), respectively. Fifteen children with foreign bodies in bronchus were also enrolled as control. IL-6 s and IL-27 s in bronchoalveolar lavage fluids (BALFs) from these children were measured by ELISA. RESULTS: There were significant differences in IL-6 s of BALFs from patients between mild cases and severe cases, MP single infection group and MP mixed infection group, and low MP DNA loads group and high MP DNA loads group, respectively (P < 0.05). Compared with IL-6 s of BALFs from control, IL-6 s in BALFs from the 6 patient groups were significantly higher (P < 0.05). IL-27 s in BALFs from MP mixed infection group were significantly lower than those from MP single infection group and control (P < 0.05) respectively. CONCLUSION: IL-6 was firmly associated with MPP and had potential application in clinical practice while IL-27 was not related to MP infection.


Assuntos
Líquido da Lavagem Broncoalveolar/imunologia , Interleucina-6/análise , Interleucinas/análise , Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/imunologia , Adolescente , Carga Bacteriana/genética , Criança , Pré-Escolar , DNA Bacteriano/genética , Feminino , Humanos , Lactente , Masculino , Mycoplasma pneumoniae/isolamento & purificação , Pneumonia por Mycoplasma/microbiologia , Reação em Cadeia da Polimerase em Tempo Real
13.
Int J Food Microbiol ; 325: 108643, 2020 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-32361054

RESUMO

Biofilms formed on food contact surfaces are frequently exposed to disinfectants at different concentrations. This study was designed to evaluate how S. Enteritidis in single species and dual species biofilms with P. fluorescens respond to quaternary ammonium compounds (QAC) residues on food contact surfaces. The 48 h-biofilms of S. Enteritidis and P. fluorescens in single/dual species were continuously exposed to 20 ppm QAC for 5 days, followed by QAC challenge at 200 ppm and 100 ppm for attached and detached cells, respectively. Biofilm structures were observed by confocal laser scanning microscopy (CLSM) and extracellular polymeric substances (EPS)-related gene expression was also evaluated. Results showed that QAC stress led to one log lower cell counts of S. Enteritidis and P. fluorescens single species biofilms. More cellulose observed by CLSM images and increased transcript levels of cellulose-related genes (csgD, bcsA and ardA) of S. Enteritidis were induced by QAC stress. Nevertheless, high percentage of membrane damaged cells in QAC pre-exposed biofilms might contribute to the increased sensitivity of S. Enteritidis in both attached and detached cells. Previous QAC exposure did not influence S. Enteritidis viable cell counts in dual specie biofilms, in which S. Enteritidis showed strong resistance to QAC with <2 log CFU/cm2 reductions. Decreased transcript levels of cellulose-related genes were observed of S. Enteritidis in dual species biofilms, but EPS-related gene expression of P. fluorescens was not affected by single/duals species. The dual species biofilm matrix which has big microcolonies extruding from bottom layers with great amounts of polysaccharides mainly produced by P. fluorescens could possibly protect S. Enteritidis against disinfection. Enhanced survival of S. Enteritidis in dual species biofilms was also found when they were detached from the coupons. Overall, our findings highlight that although repeated exposures to low dose of QAC sensitized S. Enteritidis, the presence of P. fluorescens in dual species biofilms could enhance QAC resistance of S. Enteritidis, probably contributing to survival of S. Enteritidis in food processing plants.


Assuntos
Biofilmes/efeitos dos fármacos , Desinfetantes/farmacologia , Pseudomonas fluorescens/efeitos dos fármacos , Compostos de Amônio Quaternário/farmacologia , Salmonella enteritidis/efeitos dos fármacos , Carga Bacteriana/efeitos dos fármacos , Desinfecção/métodos , Manipulação de Alimentos/métodos , Testes de Sensibilidade Microbiana , Pseudomonas fluorescens/crescimento & desenvolvimento , Salmonella enteritidis/crescimento & desenvolvimento
14.
Thorax ; 75(7): 606-608, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32354738

RESUMO

In this comparative biomarker study, we analysed 1768 serial sputum samples from 178 patients at 4 sites in Southeast Africa. We show that tuberculosis Molecular Bacterial Load Assay (TB-MBLA) reduces time-to-TB-bacillary-load-result from days/weeks by culture to hours and detects early patient treatment response. By day 14 of treatment, 5% of patients had cleared bacillary load to zero, rising to 58% by 12th week of treatment. Fall in bacillary load correlated with mycobacterial growth indicator tube culture time-to-positivity (Spearmans r=-0.51, 95% CI (-0.56 to -0.46), p<0.0001). Patients with high pretreatment bacillary burdens (above the cohort bacillary load average of 5.5log10eCFU/ml) were less likely to convert-to-negative by 8th week of treatment than those with a low burden (below cohort bacillary load average), p=0.0005, HR 3.1, 95% CI (1.6 to 5.6) irrespective of treatment regimen. TB-MBLA distinguished the bactericidal effect of regimens revealing the moxifloxacin-20 mg rifampicin regimen produced a shorter time to bacillary clearance compared with standard-of-care regimen, p=0.008, HR 2.9, 95% CI (1.3 to 6.7). Our data show that the TB-MBLA could inform clinical decision making in real-time and expedite drug TB clinical trials.


Assuntos
Antibióticos Antituberculose/uso terapêutico , Mycobacterium tuberculosis/crescimento & desenvolvimento , Escarro/microbiologia , Tuberculose Pulmonar/microbiologia , Adulto , Carga Bacteriana , Biomarcadores/metabolismo , Feminino , Seguimentos , Humanos , Masculino , Mycobacterium tuberculosis/isolamento & purificação , Prognóstico , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/metabolismo
15.
PLoS One ; 15(5): e0232485, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32357157

RESUMO

Various produce including cantaloupe, caramel-coated apples, and packaged salads, have been recognized in recent years as vehicles for listeriosis, a human foodborne disease caused by intracellular pathogen Listeria monocytogenes. Our knowledge regarding the role of these foods in L. monocytogenes virulence, however, is limited. Understanding their role in modulating L. monocytogenes virulence can be useful in risk assessments and for developing control measures. In this study, we employed the Galleria mellonella larvae model to evaluate virulence potential of fifteen clinical, environmental and food isolates of L. monocytogenes, related to three major outbreaks, after growth on different foods. The non-human pathogen Listeria innocua was also included in the panel. Strains were inoculated in parallel in 5ml of brain heart infusion (BHI) broth, and on the surfaces of cantaloupe and apple fragments (5g each) at about 105 colony forming units (CFU)/ml/fragment. One set of inoculated broth and food fragments was incubated at 10°C for 5 days while the second set was kept at 25°C for 3 days. L. monocytogenes cells were recovered from the fruits and BHI, washed twice, re-suspended in saline, and used to inoculate G. mellonella larvae at final concentrations of 106 and 105 CFU/larva. The larvae were incubated at 37°C and monitored for mortality (LT50-time taken to kill 50% of the larvae) and phenotypic changes over seven days. L. monocytogenes grown on cantaloupe and apple flesh surfaces resulted in higher virulence than when grown in BHI. L. monocytogenes infection at 106 CFU/larvae resulted in an average LT50 of ≤ 30, 36 and 47 hours on cantaloupe, apples and BHI, respectively. These results represent a 2.5-4-fold increased mortality compared with an LT50 ≥120 hours in larvae infected with the same doses of L. innocua grown in corresponding matrices. Similar trends were also recorded with doses of about 105 CFU /larvae.


Assuntos
Microbiologia de Alimentos , Listeria monocytogenes/patogenicidade , Animais , Carga Bacteriana , Cucumis melo/microbiologia , Meios de Cultura , Doenças Transmitidas por Alimentos/etiologia , Humanos , Larva/microbiologia , Listeria/crescimento & desenvolvimento , Listeria/patogenicidade , Listeria monocytogenes/crescimento & desenvolvimento , Listeriose/etiologia , Malus/microbiologia , Modelos Biológicos , Mariposas/microbiologia , Medição de Risco , Virulência
16.
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
17.
Nat Commun ; 11(1): 2607, 2020 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-32451375

RESUMO

Quantification of pathogen and host biomarkers is essential for the diagnosis, monitoring, and treatment of infectious diseases. Here, we demonstrate sensitive and rapid quantification of bacterial load and cytokines from human biological samples to generate actionable hypotheses. Our digital assay measures IL-6 and TNF-α proteins, gram-negative (GN) and gram-positive (GP) bacterial DNA, and the antibiotic-resistance gene blaTEM with femtomolar sensitivity. We use our method to characterize bronchoalveolar lavage fluid from patients with asthma, and find elevated GN bacteria and IL-6 levels compared to healthy subjects. We then analyze plasma from patients with septic shock and find that increasing levels of IL-6 and blaTEM are associated with mortality, while decreasing IL-6 levels are associated with recovery. Surprisingly, lower GN bacteria levels are associated with higher probability of death. Applying decision-tree analysis to our measurements, we are able to predict mortality and rate of recovery from septic shock with over 90% accuracy.


Assuntos
Citocinas/sangue , DNA Bacteriano/sangue , Choque Séptico/imunologia , Choque Séptico/microbiologia , Asma/imunologia , Asma/microbiologia , Carga Bacteriana , Biomarcadores/análise , Biomarcadores/sangue , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/imunologia , Líquido da Lavagem Broncoalveolar/microbiologia , Estudos de Casos e Controles , Citocinas/análise , DNA Bacteriano/genética , Árvores de Decisões , Genes Bacterianos , Bactérias Gram-Negativas/genética , Bactérias Gram-Negativas/isolamento & purificação , Bactérias Gram-Positivas/genética , Bactérias Gram-Positivas/isolamento & purificação , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Interleucina-6/análise , Interleucina-6/sangue , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase Multiplex/estatística & dados numéricos , Prognóstico , Sensibilidade e Especificidade , Choque Séptico/mortalidade , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/sangue , Resistência beta-Lactâmica/genética
18.
Invest Ophthalmol Vis Sci ; 61(5): 26, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32416603

RESUMO

Purpose: IFN-stimulated gene (ISG) 15 is a type 1 IFN-induced protein and known to modify target proteins in a manner similar to ubiquitylation (protein conjugation by ISG15 is termed ISGylation). We sought to determine the role of ISG15 and its underlying mechanisms in corneal innate immune defense against Pseudomonas aeruginosa keratitis. Methods: ISG15 expression in cultured human corneal epithelial cells (HCECs) and mouse corneas was determined by PCR and Western blot analysis. Gene knockout mice were used to define the role of ISG15 signaling in controlling the severity of P. aeruginosa keratitis, which was assessed with photographing, clinical scoring, bacterial counting, myeloperoxidase assay, and quantitative PCR determination of cytokine expression. Integrin LFA-1 inhibitor was used to assess its involvement of ISG15 signaling in P. aeruginosa-infected corneas. Results: Heat-killed P. aeruginosa induced ISG15 expression in cultured HCECs and accumulation in the conditioned media. Isg15 deficiency accelerated keratitis progress, suppressed IFNγ and CXCL10, and promoted IL-1ß while exhibiting no effects on IFNα expression. Moreover, exogenous ISG15 protected the corneas of wild-type mice from P. aeruginosa infection while markedly reducing the severity of P. aeruginosa keratitis in type 1 IFN-receptor knockout mice. Exogenous ISG15 increased bacteriostatic activity of B6 mouse corneal homogenates, and inhibition of LFA-1 exacerbated the severity of and abolished protective effects of ISG15 on P. aeruginosa keratitis. Conclusions: Type 1 INF-induced ISG15 regulates the innate immune response and greatly reduces the susceptibility of B6 mouse corneas to P. aeruginosa infection in an LFA-1-dependent manner.


Assuntos
Úlcera da Córnea/imunologia , Citocinas/fisiologia , Infecções Oculares Bacterianas/imunologia , Imunidade Inata/fisiologia , Infecções por Pseudomonas/imunologia , Ubiquitinas/fisiologia , Animais , Carga Bacteriana , Western Blotting , Células Cultivadas , Úlcera da Córnea/metabolismo , Úlcera da Córnea/fisiopatologia , Citocinas/metabolismo , Epitélio Anterior/metabolismo , Infecções Oculares Bacterianas/metabolismo , Infecções Oculares Bacterianas/fisiopatologia , Regulação da Expressão Gênica/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Peroxidase/metabolismo , Infecções por Pseudomonas/metabolismo , Infecções por Pseudomonas/fisiopatologia , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/fisiologia
19.
J Vis Exp ; (158)2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32420999

RESUMO

Tuberculosis is caused by Mycobacterium tuberculosis (Mtb), a pathogen classified by the United Nations (UN) as a dangerous category B biological substance. For the sake of the workers' safety, handling of all samples presumed to carry Mtb must be conducted in a containment level (CL) 3 laboratory. The TB molecular bacterial load assay (TB-MBLA) test is a reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) test that quantifies Mtb bacillary load using primers and dual-labelled probes for 16S rRNA. We describe the use of heat inactivation to render TB samples noninfectious while preserving RNA for the TB-MBLA. A 1 mL aliquot of the sputum sample in tightly closed 15 mL centrifuge tubes is boiled for 20 min at either 80 °C, 85 °C, or 95 °C to inactivate Mtb bacilli. Cultivation of the heat inactivated and control (live) samples for 42 days confirmed the death of TB. The inactivated sample is then spiked with 100 µL of the extraction control and RNA is extracted following the standard RNA isolation procedure. No growth was observed in the cultures of heat treated samples. The isolated RNA is subjected to real-time RT-qPCR, which amplifies a specific target in the Mtb 16S rRNA gene, yielding results in the form of quantification cycles (Cq). A standard curve is used to translate Cq into bacterial load, or estimated colony forming units per mL (eCFU/mL). There is an inverse relationship between Cq and the bacterial load of a sample. The limitation is that heat inactivation lyses some cells, exposing the RNA to RNases that cause a loss of <1 log10eCFU/mL (i.e., <10 CFU/mL). Further studies will determine the proportion of very low burden patients that cause false negative results due to heat inactivation.


Assuntos
Carga Bacteriana/métodos , Mycobacterium tuberculosis/isolamento & purificação , RNA Bacteriano/análise , Escarro/microbiologia , Tuberculose/diagnóstico , Carga Bacteriana/instrumentação , Carga Bacteriana/normas , Humanos , Mycobacterium tuberculosis/genética , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Reação em Cadeia da Polimerase em Tempo Real , Manejo de Espécimes , Tuberculose/microbiologia
20.
J Vis Exp ; (159)2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32449738

RESUMO

Cigarette smoking is the major etiological cause for lung emphysema and chronic obstructive pulmonary disease (COPD). Cigarette smoking also promotes susceptibility to bacterial infections in the respiratory system. However, the effects of cigarette smoking on bacterial infections in human lung epithelial cells have yet to be thoroughly studied. Described here is a detailed protocol for the preparation of cigarette smoking extracts (CSE), treatment of human lung epithelial cells with CSE, and bacterial infection and infection determination. CSE was prepared with a conventional method. Lung epithelial cells were treated with 4% CSE for 3 h. CSE-treated cells were, then, infected with Pseudomonas at a multiplicity of infection (MOI) of 10. Bacterial loads of the cells were determined by three different methods. The results showed that CSE increased Pseudomonas load in lung epithelial cells. This protocol, therefore, provides a simple and reproducible approach to study the effect of cigarette smoke on bacterial infections in lung epithelial cells.


Assuntos
Células Epiteliais/microbiologia , Células Epiteliais/patologia , Pulmão/patologia , Infecções por Pseudomonas/etiologia , Fumar/efeitos adversos , Carga Bacteriana/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Gentamicinas/farmacologia , Humanos , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia
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