RESUMO
Resumen Staphylococcus aureus coloniza la nasofaringe en un tercio de los individuos sanos y además es causante de infecciones graves en pediatría, como endocarditis, neumonía e infecciones osteoarticulares. Posee varios mecanismos de virulencia, siendo la leucocidina de Panton Valentine (LPV) uno de ellos, una exotoxina que causa muerte celular. Su producción está comúnmente relacionada con Staphylococcus aureus resistente a meticilina (SARM) e infecciones pulmonares y musculo-esqueléticas graves. Sin embargo, la producción de LPV no es exclusiva de SARM. Se presentan dos casos clínicos de pacientes con infección por Staphylococcus aureus sensible a meticilina productora de esta exotoxina.
Abstract Staphylococcus aureus colonizes the nasopharynx in one third of healthy individuals and is also responsible for several infections in pediatrics such as endocarditis, pneumonia and osteoarticular infections. It has several virulence mechanisms, such as Panton Valentine leukocidin (PVL), which is an exotoxin that causes cell death. It is commonly related to methicillin-resistant Staphylococcus aureus (MRSA) and more serious pulmonary and musculoskeletal infections. However, PVL is not exclusive to MRSA. Two clinical cases of patients with infection by methicillin-sensitive Staphylococcus aureus producing this exotoxin are presented.
Assuntos
Humanos , Masculino , Criança , Adolescente , Osteomielite/tratamento farmacológico , Pediatria , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus Resistente à Meticilina , Staphylococcus aureus , Toxinas Bacterianas , Exotoxinas , Leucocidinas , Meticilina/farmacologiaRESUMO
Resumen Introducción: Clostridioides difficile causa diarrea y colitis pseudomembranosa. Su diagnóstico se realiza con la detección de glutamato-deshidrogenasa (GDH) o las toxinas A y B y se confirma con pruebas de amplificación de ácidos nucleicos. Objetivo: Definir si la determinación de GDH es redundante a la de las toxinas. Métodos: Estudio observacional retrospectivo de muestras fecales de pacientes con sospecha de infección por Clostridioides difficile. Las toxinas y GDH se determinaron mediante inmunocromatografía. Se realizó una simulación bayesiana con los cocientes de probabilidad; se consideró significativo un valor de p < 0.05. Resultados: Se analizaron 329 resultados de GDH y toxinas A y B. Se encontró una prevalencia de infección de Clostridioides difficile de 18.2 %. La sensibilidad y especificidad de la prueba de GDH fue de 0.90 y 0.89, respectivamente. El cociente de probabilidad positivo fue de 8.9 y el negativo, de 0.11. Conclusiones: Un resultado negativo de GDH disminuye considerablemente la probabilidad de infección, pero no la descarta. La detección de toxinas de Clostridioides difficile puede ser necesaria en instituciones donde la amplificación de ácidos nucleicos no es económica o accesible.
Abstract Introduction: Clostridioides difficile causes diarrhea and pseudomembranous colitis. Its diagnosis is made with glutamate dehydrogenase (GDH) or toxins A and B detection and is confirmed with nucleic acid amplification tests. Objective: To define if GDH determination is redundant to that of toxins. Methods: Retrospective, observational study in diarrheal stools of patients with suspected Clostridioides difficile infection. Toxins and GDH were determined by immunochromatography. Bayesian simulation was performed with likelihood ratios; a p-value < 0.05 was regarded as significant. Results: 329 GDH and toxin A and B results were analyzed. Clostridioides difficile infection prevalence was 18.2 %. Sensitivity and specificity of the GDH test were 0.90 and 0.89, respectively. Positive likelihood ratio was 8.9, and negative was 0.11. Conclusions: A negative GDH result considerably reduces the probability of infection but does not rule it out. Clostridioides difficile toxins detection may be necessary in institutions where nucleic acid amplification is not affordable or accessible.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Proteínas de Bactérias/análise , Toxinas Bacterianas/análise , Clostridioides difficile , Infecções por Clostridium/diagnóstico , Enterotoxinas/análise , Fezes/química , Biomarcadores/análise , Funções Verossimilhança , Prevalência , Estudos Retrospectivos , Teorema de Bayes , Sensibilidade e Especificidade , Infecções por Clostridium/epidemiologia , Diarreia/microbiologia , Fezes/enzimologia , Glutamato Desidrogenase/análiseRESUMO
RESUMEN La investigación se realizó a fin de evaluar el efecto del gas de formalina sobre el recuento de mesófilos en nailon comercial (poliamida) destinado a procedimientos quirúrgicos. En primer lugar, se evaluó la contaminación de una muestra del material comercial de 1 g expuesta al ambiente de quirófano por 72 horas, a través de la técnica de recuento en placa para mesófilos; se obtuvieron 850 UFC/g. Una vez comprobada la contaminación de la poliamida comercial, esta se sometió a gases de formalina en comprimidos. Se colocaron 5 muestras (n=5) de nailon de 1 g cada una en 5 recipientes herméticos de 1 litro con 1 gramo de formalina cada uno; estos recipientes se almacenaron en un mueble en sala de esterilización a un metro de altura del piso y posteriormente, fueron abiertos y cultivados a través de la técnica de recuento en placa para mesófilos, uno por día a lo largo de 5 días a intervalos de 24 horas. Los resultados obtenidos no registraron crecimientos de microorganismos a partir de las 24 horas y durante los 5 días posteriores y se encontraron diferencias estadísticamente significativas (p < 0,05). Se concluye que bajo las condiciones del presente estudio el tiempo de esterilización de la formalina sobre nailon comercial es de 24 horas.
ABSTRACT Hie present research work was carried out with the purpose of evaluating the effect of formalin gas on the count of mesophiles in commercial nylon (polyamide) destined to surgical procedures. Firstly, the contamination of a 1 g commercial nylon sample, exposed to the operating room environment for 72 hours, was evaluated through the plate counting technique for mesophiles; it yielded a result of 850 CFU / gram. Once the contamination of the commercial polyamide was checked, it was subjected to formalin gases in tablets. Five nylon samples (n=5) of 1 gram each were placed in 5 1 liter airtight containers containing 1 gram of formalin; Hese containers were stored in a cabinet in the sterilization room one meter above the floor, and later opened and cultivated through the plate counting technique for mesophiles, one per day for 5 days at 24 hour intervals. The results obtained after being exposed to formalin gases in tablets did not register growths of microorganisms after 24 hours and during the 5 days after the study, finding statistically significant differences (p < 0.05) in relation to the studied times concluding under the conditions of the present study that the sterilization time of the formalin on commercial nylon is equal to 24 hours.
Assuntos
Animais , Procedimentos Cirúrgicos Operatórios , Esterilização , Poluição Ambiental , Formaldeído , Gases , Resistência à Seca , Nylons , Salas Cirúrgicas , Pesquisa , Suturas , Comprimidos , Termômetros , Tempo , Bactérias , Toxinas Bacterianas , Contagem de Colônia Microbiana , Abscesso , Temperatura Alta , InflamaçãoRESUMO
Clostridium difficile causes intestinal inflammation, which increases adenosine. We compared the expression of adenosine receptors (AR) subtypes A1, A2A, A2B, and A3 in HCT-8, IEC-6 cells, and isolated intestinal epithelial cells, challenged or not with Clostridium difficile toxin A and B (TcdA and TcdB) or infection (CDI). In HCT-8, TcdB induced an early A2BR expression at 6 h and a late A2AR expression at 6 and 24 h. In addition, both TcdA and TcdB increased IL-6 expression at all time-points (peak at 6 h) and PSB603, an A2BR antagonist, decreased IL-6 expression and production. In isolated cecum epithelial cells, TcdA induced an early expression of A2BR at 2s and 6 h, followed by a late expression of A2AR at 6 and 24 h and of A1R at 24 h. In CDI, A2AR and A2BR expressions were increased at day 3, but not at day 7. ARs play a role in regulating inflammation during CDI by inducing an early pro-inflammatory and a late anti-inflammatory response. The timing of interventions with AR antagonist or agonists may be of relevance in treatment of CDI.
Assuntos
Animais , Toxinas Bacterianas , Clostridioides difficile , Infecções por Clostridium , Receptores Purinérgicos P1/metabolismo , Proteínas de Bactérias , Regulação para Cima , Interleucina-6 , Modelos Animais de Doenças , Enterotoxinas , Infecções , Anti-InflamatóriosRESUMO
BACKGROUND Bacillus anthracis is the aetiologic agent of anthrax, a re-emerging, septicaemic, haemorrhagic and lethal disease that affects humans, domestic ruminants and wildlife. Plasmids pXO1 and pXO2 are attributes that confer pathogenicity to B. anthracis strains. This bacterium was used as biological weapon in the World Wars and in the biological attack in the United States of America at 2001. B. anthracis is classified as a Tier 1 bioterrorism agent by the Centers for Diseases Control and Prevention. Anthrax is recognised as a re-emerging disease. Several studies concerning the dynamics of B. anthracis cycle in soil revealed that nonpathogenic B. anthracis strains due to lack of pXO2 plasmid are commonly found in some types of soil. OBJECTIVES This study aimed isolation and identification of B. anthracis spores in soil samples of the state of Rio de Janeiro, Brazil. METHODS Phenotypic and genotypic approaches were used to identify isolates including MALDI-TOF/MS, motility test, susceptibility to gamma phage and penicillin, survey for pag and cap genes as surrogates of pXO1 and pXO2 plasmids, respectively, and sequencing of 16SrRNA-encoding gene. Physicochemical analysis of the soil samples were carried out to describe soil characteristics. FINDINGS We observed the presence of one B. anthracis pXO1+ and pXO2- isolated from clay loam soil; one B. anthracis-like strain pXO1+ and pXO2-isolated from loamy sand; and 10 Bacillus spp. strains sensitive to phage-gamma that need better characterisation to define which their species were recovered from loamy sand. MAIN CONCLUSIONS This work showed promising results and it was the first study to report results from an active surveillance for B. anthracis in Brazil.
Assuntos
Humanos , Plasmídeos/análise , Microbiologia do Solo , Esporos Bacterianos , Bacillus anthracis/isolamento & purificação , DNA Bacteriano/genética , Reação em Cadeia da Polimerase/métodos , Fatores de Virulência/genética , Plasmídeos/genética , Solo , Bacillus anthracis/genética , Bacillus anthracis/patogenicidade , Toxinas Bacterianas , Virulência , Brasil , DNA Bacteriano/análise , Análise de Sequência de DNA , Antígenos de BactériasRESUMO
En esta Parte 3 de la serie de cuatro artículos sobre micetismos se analizan los síndromes tempranos con síntomas gastrointestinales que se caracterizan por presentar un período de latencia muy corto, de menos de 6 horas después de la ingestión de los macromicetos. Los restantes síndromes tempranos con sintomatología compleja serán tratados en la Parte 4 de la serie. Actualmente se conocen más de 200 especies responsables de síndromes gastrointestinales, pero en este trabajo se abordarán solamente diez ejemplos que involucran los géneros Boletus [Boletus satanas (o Rubroboletus satanas) y Boletus venenatus (o Neoboletus venenata)], Hypholoma, Agaricus (Agaricus xanthodermus), Omphalotus, Lactarius, Russula, Entoloma, Chlorophyllum (Chlorophyllum molybdetes) y Leucoprinus (Leucoprinus birnbaumii). Las toxinas involucradas en estos casos presentan gran variedad estructural, desde proteínas hasta terpenoides, en particular sesquiterpenoides y triterpenoides, vinilglicina, fenol y azocompuestos, pero todas generan la misma sintomatología. Estas sustancias y otros componentes químicos de los hongos suelen ser indigestos, con una susceptibilidad variable entre los consumidores. El tratamiento es de apoyo y es estrictamente para esos casos con cuadros más graves de deshidratación. Normalmente, los casos evolucionan favorablemente después de 12 a 48 horas. Se analizan los síntomas, las toxinas involucradas, los mecanismos de acción, cuando se conocen y las especies causantes de los micetismos.
This part 3 of the series of four articles on mushroom poisoning refers to early-onset gastrointestinal syndromes, which are characterized by a very short latency period of less than 6 hours after mushroom ingestion. The remaining early-onset syndromes with complex symptoms will be treated in Part 4 of the series. Currently, more than 200 species responsible for gastrointestinal syndromes are known, but in this paper only ten examples will be addressed involving the genera Boletus [e.g., Boletus satanas (or Rubroboletus satanas), and Boletus venenatus (or Neoboletus venenata)], Hypholoma, Agaricus (e.g., Agaricus xanthodermus), Omphalotus, Lactarius, Russula, Entoloma, Chlorophyllum (e.g., Chlorophyllum molybdetes), and Leucoprinus (e.g., Leucoprinus birnbaumii). The toxins involved in these cases have a great structural variety, from proteins to terpenoids, in particular sesquiterpenoids and triterpenoids, vinylglycine, phenol, and azocompounds, but all show the same symptoms. These substances and other mushroom chemical constituents are usually indigestible, with varying consumer susceptibility. The treatment is supportive and is strictly for those cases with more severe dehydration. Usually, the cases progress favourably after 12 to 48 hours.The symptoms, toxins involved, mechanisms of action when known, and the species of mushrooms responsible for the mycetisms are analysed.
Nesta parte 3 da série de quatro artigos sobre intoxicação por cogumelos são analisadas as síndromes precoces com sintomas gastrointestinais que se caracterizam por apresentar um período de latência muito curto, de menos de 6 horas, após a ingestão de cogumelos. As síndromes precoces restantes com sintomatologia complexa serão tratadas na Parte 4 da série. Atualmente, são conhecidas mais de 200 espécies responsáveis por síndromes gastrointestinais, mas neste trabalho serão abordados apenas dez exemplos que envolvem os gêneros Boletus [Boletus satanas (ou Rubroboletus satanas) e Boletus venenatus (ou Neoboletus venenata)], Hypholoma, Agaricus (Agaricus xanthodermus), Omphalotus, Lactarius, Russula, Entoloma, Chlorophyllum (Chlorophyllum molybdetes) e Leucoprinus (Leucoprinus birnbaumii). As toxinas envolvidas nestes casos têm uma grande variedade estrutural, desde proteínas até terpenóides, em particular sesquiterpenóides e triterpenóides, vinilglicina, fenol e azo compostos, mas todas apresentam a mesma sintomatologia. Essas substâncias e outros constituintes químicos dos cogumelos costumam ser indigestos, com uma suscetibilidade variável entre aqueles que os consomem. O tratamento é de suporte e é rigorosamente para esses casos com quadros mais graves de desidratação. Normalmente, os casos evoluem favoravelmente após 12 a 48 horas. São analisados os sintomas, as toxinas envolvidas, os mecanismos de ação, quando conhecidos, e as espécies de cogumelos responsáveis pelas intoxicações.
Assuntos
Animais , Camundongos , Toxicologia , Agaricus/patogenicidade , Boletus satanas/toxicidade , Gastroenteropatias/complicações , Toxinas Bacterianas , Toxinas Bacterianas/análise , Latência Viral , MicotoxinasRESUMO
Resumen Introducción. Los factores de virulencia de las cepas de Vibrio cholerae no-O1, no-O139 no son claramente conocidos. La cepa de origen septicémico NN1 Vibrio cholerae no-O1, no-O139 fue secuenciada previamente mediante la plataforma Illumina, detectándose en su genoma un fragmento de la isla de patogenicidad VPaI-7 de V. parahaemolyticus. Objetivo: detectar los genes de virulencia vcsN2, vcsC2, vcsV2, vspD, toxR2 y vopF en cepas chilenas clínicas de V. cholerae no-O1, no-O139. Material y Métodos: Un total de 9 cepas chilenas de origen clínico de Vibrio cholerae no-O1, no-O139 aisladas entre 2006-2012 fueron analizadas mediante ensayos de reacción de polimerasa en cadena (RPC, en inglés PCR) convencional para los genes de secreción tipo III codificados en dicha isla: vcsN2, vcsC2, vcsV2, vspD, toxR2 y vopF. Adicionalmente se determinó la presencia de los genes de virulencia hylA y rtxA. Además, se realizaron ensayos de repetitive element palindromic PCR (REP-PCR) y Enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). Resultados: la mayoría (6/9) de las cepas chilenas de V. cholerae no-O1, no-O139 contiene todos los genes de secreción tipo III vcsN2, vcsC2, vcsV2, vspD, toxR2 y vopF, codificados en una isla de patogenicidad. Además, el total de las cepas (9/9) contiene los genes de virulencia hylA y rtxA. Conclusión: Estos resultados sugieren fuertemente la posibilidad que dichas cepas posean un potencial de virulencia importante en seres humanos.
Backgound: The virulence factors of the Vibrio cholerae non-O1, non-O139 strains are not clearly known. The strain of septicemic origin NN1 Vibrio cholerae non-O1, non-O139 was sequenced previously by the Illumina platform. A fragment of the pathogenicity island VPaI-7 of V. parahaemolyticus was detected in its genome. Aim: To detect the virulence genes vcsN2, vcsC2, vcsV2, vspD, toxR2 y vopF in Chilean strains of V. cholerae non-O1, non-O139. Methods: A total of 9 Chilean strains of clinical origin of Vibrio cholerae non-O1, non-O139 isolated between 2006-2012 were analyzed by conventional PCR assays for type III secretion genes encoded on that island: vcsN2, vcsC2, vcsV2, vspD, toxR2 and vopF. Additionally, the presence of the virulence genes hylA and rtxA was determined. In addition, REP-PCR and ERIC-PCR assays were performed. Results: most (6/9) Chilean V. cholerae non-O1, non-O139 strains contain the type III secretion genes vcsN2, vcsC2, vcsV2, vspD, toxR2 and vopF, encoded in an island of pathogenicity. In addition, all (9/9) the strains contain the virulence genes hylA and rtxA. Conclusion: These results strongly suggest the possibility that those strains possess an important virulence potential in humans.
Assuntos
Humanos , Proteínas de Bactérias/genética , Fatores de Transcrição/genética , Vibrio cholerae/genética , Fatores de Virulência/genética , Vibrio cholerae não O1/genética , Ilhas Genômicas/genética , Proteínas de Ligação a DNA/genética , Sistemas de Secreção Tipo III/genética , Toxinas Bacterianas/genética , Vibrio cholerae/isolamento & purificação , Vibrio cholerae/patogenicidade , Chile , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Vibrio cholerae não O1/isolamento & purificação , Vibrio cholerae não O1/patogenicidade , Proteínas Hemolisinas/genéticaRESUMO
Abstract INTRODUCTION: Musca domestica is resistant to many insecticides; hence, biological control is a suitable alternative. METHODS: We evaluated the lethality of strain Btk176 towards the larval and adult M. domestica and the histopathological effects in the larvae midgut. RESULTS: We observed 99% larval and 78.9% adult mortality within 48 hours of spore ingestion (dosage, 2.4×108 CFU/ml). The histopathological effects were consistent with cytotoxicity. PCR analysis showed the presence of the cry1Ba gene. Transmission electron microscopy revealed a bipyramidal parasporal body. Thurigiensin activity was not detected. CONCLUSIONS: The serovar, Btk176 might be a potential biocontrol agent for houseflies.
Assuntos
Animais , Bacillus thuringiensis , Toxinas Bacterianas/farmacologia , Moscas Domésticas/efeitos dos fármacos , Inseticidas/farmacologia , Larva/efeitos dos fármacos , Contagem de Colônia Microbiana , Controle Biológico de Vetores/métodos , Reprodutibilidade dos Testes , Análise de Variância , Microscopia Eletrônica de Transmissão , ExotoxinasRESUMO
Staphylococcus aureus es un patógeno responsable de diversos cuadros clínicos. Los marcadores moleculares son útiles para el estudio de la epidemiología microbiana. Se estudiaron 22 aislamientos de S. aureus resistentes a meticilina (SARM) y 23 sensibles a meticilina (SASM) mediante mecA, cassette SCCmec, leucocidina de Panton Valentine (LPV) y polimorfismo spa; se analizaron datos de los pacientes. SASM predominó en muestras distintas de piel y partes blandas de internados, mientras SARM en partes blandas. Predominó el SCCmec tipo IV seguido del I. Se encontró baja presencia de LPV. En SARM hubo 11 tipos de spa diferentes, t019 fue el más frecuente y en pacientes ambulatorios. En SASM se hallaron 17 tipos con prevalencia del t189. El spa t002 estuvo presente en SASM y SARM. Se hallaron 11 tipos de spa no reportados en nuestro país.
Staphylococcus aureus is a pathogen associated a different kind of infection. Molecular markers are useful tools to study microbial epidemiology. Twenty two methicillin-resistant S. aureus (MRSA) and 23 methicillin-susceptible S. aureus (MSSA) were studied by mecA gene, SCCmec cassette, Panton Valentine leucocidin (PVL) and spa polymorphism. The clinical data patients were analyzed. MSSA was prevalent in samples different from skin and soft tissue (SST) and in hospitalized patients, whereas MRSA in SST. SCCmec type IV was predominant, followed spa; by type I. Low presence of PVL was found. In MRSA 11 different types of spa were detected, SCCmec; t019 was the most frequent and associated with outpatient, 17 types were found in MSSA and Panton Valentine t189 was prevalent. spa t002 was present in MSSA and MRSA. We found 11 types of spa not leucocidin reported in our country.
Assuntos
Adulto , Humanos , Infecções Estafilocócicas , Staphylococcus aureus , Staphylococcus aureus Resistente à Meticilina , Hospitais , Argentina , Proteínas de Bactérias , Toxinas Bacterianas , Testes de Sensibilidade Microbiana , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Staphylococcus aureus Resistente à Meticilina/genética , AntibacterianosRESUMO
Clostridioides difficile es el principal agente causal de diarreas asociadas al cuidado de la salud. Esta bacteria produce toxinas y una enzima que se encuentra muy conservada en la especie: la glutamato deshidrogenasa (GDH). El diagnóstico rápido y el tratamiento efectivo permiten la pronta mejoría del paciente, con el consecuente control de la diseminación del microorganismo. Sin embargo, aún no se cuenta con un método diagnóstico óptimo y se propone la realización de diversas pruebas, cuyos resultados se interpretan en el contexto de ciertos algoritmos. En este trabajo se evaluó el desempeño de la GDH como prueba de tamizaje en el diagnóstico de la diarrea por c. difficile. Se estudiaron 615 muestras de materia fecal. Se determinó la presencia de GDH y de toxinas mediante el equipo diagnóstico de enzimoinmunoensayo de membrana C. DIFF QUIK-CHEK COMPLETE® (TECHLAB) y se realizaron cultivos para la búsqueda de C. difficile. Se calcularon los valores de sensibilidad, especificidad, VPP y VPN con un nivel de significación p < 0,05. Se detectó GDH en 266 muestras (43,25%), con una sensibilidad del 100% y una especificidad del 87,10%, IC95: 84,58-91,42. Se hallaron toxinas en 218 muestras (35,45%) y C. difficile desarrolló en 235 cultivos (38,21%). De 48 muestras GDH positivas y sin producción de toxina/s, 17 fueron positivas al cultivo de C. difficile, con 15 aislamientos toxigénicos y 2 no toxigénicos. No hubo desarrollo de C. difficile en las 31 muestras restantes. Ninguna muestra GDH negativa dio resultado positivo de toxina/s ni desarrollo en el cultivo, por lo cual el VPN de la GDH fue del 100%, mientras que el VPP fue del 81,9%. Concluimos que la determinación de GDH representa un screening adecuado para descartar casos de diarrea por C. difficile, por lo tanto de valor en los algoritmos diagnósticos de las diarreas infecciosas.
Clostridioides difficile is the main etiological agent of diarrhea associated with health care, it produces toxins and glutamate dehydrogenase (GDH), an enzyme that is highly conserved in this species. Rapid diagnosis and effective treatment produce prompt improvement of the patient and subsequent control of the microorganism spread. There are several techniques whose results are interpreted in the context of algorithms. However, the optimal diagnostic method is yet unknown. The performance of GDH as a screening test for the diagnosis of C. difficile diarrhea was assessed. Six hundred and fifteen stool samples were studied. The presence of GDH and toxins presence was determined by TECHLAB® C. DIFF QUIK-CHEK COMPLETE and the samples were cultured for the search of C. difficile. The values of sensitivity, specificity, PPV and NPV were calculated with a p value of 0.05 or less. GDH was detected in 266 samples (43.25%), with a sensitivity of 100% and specificity of 87.10%, IC95: 84.58-91.42; toxin/s were detected in 218 (35.45%) and C. difficile developed in 235 cultures (38.21%). From 48 samples with positive GDH and negative toxin/s, 15 toxigenic and 2 non-toxigenic isolates were obtained, the remaining 31 samples were negative for C. difficile. All GDH-negative samples were negative for toxins or culture, therefore, GDH NPV was 100%, while PPV was 81.9%. We conclude that GDH is a suitable screening test for the diagnostic algorithm of C. difficile diarrhea.
Assuntos
Humanos , Clostridioides difficile , Infecções por Clostridium , Glutamato Desidrogenase , Proteínas de Bactérias , Toxinas Bacterianas , Clostridioides difficile/enzimologia , Sensibilidade e Especificidade , Infecções por Clostridium/diagnóstico , Diarreia , Enterotoxinas , Fezes , Glutamato Desidrogenase/análiseRESUMO
Alfa toxina, una proteína formadora de poros con actividad citotóxica, es uno de los principales factores de virulencia secretados por la mayoría de las cepas de Staphylococcus aureus. Se ha establecido la relevancia de esta proteína en la patogenia de la neumonía asociada a infecciones por S. aureus. Por lo tanto, la inhibición de la secreción de alfa toxina puede ser una alternativa en el control de las infecciones causadas por este microorganismo. En este trabajo mostramos que quercetina, un flavonoide de origen natural, inhibe de manera dosis dependiente la actividad hemolítica y disminuye la secreción de alfa toxina en sobrenadantes de cultivos de S. aureus sensible y resistente a meticilina. Además, quercetina previene de manera significativa el daño de células alveolares humanas cuando se co-cultivan con S. aureus. Nuestros datos sugieren que quercetina puede disminuir la virulencia de S. aureus al afectar la secreción de alfa toxina.
Alpha toxin, a pore-forming protein with cytotoxic activity, is one of the major virulence factors secreted by most strains of Staphylococcus aureus. The relevance of this protein in the pathogenesis of pneumonia associated with S. aureus infections has already been esta blished. Therefore, inhibiting alpha toxin secretion can be an alternative for controlling these infections. This study shows that quercetin, a naturally occurring flavonoid, inhibits hemolytic activity in a dose-dependent manner and reduces alpha toxin secretion in culture supernatants of methicillin-sensitive and methicillin-resistant S. aureus. Furthermore, quercetin significantly prevents damage to human alveolar cells when co-cultured with S. aureus. Our results suggest that quercetin can reduce S. aureus virulence by affecting alpha-toxin secretion.
Assuntos
Humanos , Quercetina , Staphylococcus aureus , Antioxidantes , Quercetina/farmacologia , Infecções Estafilocócicas , Staphylococcus aureus/patogenicidade , Toxinas Bacterianas/metabolismo , Virulência , Fatores de Virulência , Proteínas Hemolisinas , Antioxidantes/farmacologiaRESUMO
The best laboratory diagnostic approach to detect Clostridioides --#1;Clostridium--#3; difficile infection (CDI) is a subject of ongoing debate. With the aim of evaluating four laboratory diagnostic methods, 250 unformed stools from patients with suspected CDI submitted to nine medical center laboratories from November 2010 to December 2011, were studied using: (1) an immunochromatographic rapid assay test that combines the qualitative determination of glutamate dehydrogenase (GDH) plus toxins A and B (QAB), the CDIFF QUIK CHEK COMPLETE assay; (2) an enzyme immunoassay for qualitative determination of toxins A and B, the RIDASCREENTC. difficile Toxin A/B assay (RAB); (3) a PCR for the toxin B gene assay (PCR); and (4) the toxigenic culture (TC).C. difficile isolates from direct toxin negative stools by QAB, RAB and PCR were evaluated for toxigenicity by the same direct tests, in order to assess the contribution of the TC (QAB-TC, RAB-TC, PCR-TC). A combination of the cell culture cytotoxicity neutralization assay (CCCNA) in stools, and the same assay on isolates from direct negative samples (CCCNA-TC) was considered the reference method (CCCNA/CCCNA-TC). Of the 250 stools tested, 107 (42.8%) were positive by CCCNA/CCCNA-TC. The GDH and PCR/PCR-TC assays were the most sensitive, 91.59% and 87.62%, respectively. The QAB, RAB, QAB/QAB-TC and RAB/RAB-TC had the highest specificities, ca. 95%. A negative GDH result would rule out CDI, however, its low positive likelihood ratio (PLR) of 3.97 indicates that a positive result should always be complemented with the detection of toxins. If the RAB, QAB, and PCR assays do not detect toxins from direct feces, the toxigenic culture should be performed. In view of our results, the most accurate and reliable methods to be applied in a clinical microbiology laboratory were the QAB/QAB-TC, and RAB/RAB-TC, with PLRs >10 and negative likelihood ratios <0.30.
El mejor procedimiento para realizar el diagnóstico de laboratorio de la infección causada por Clostridioides --#1;Clostridium--#3; difficile (ICD) es aún objeto de debate. Con el fin de evaluar cuatro métodos diagnósticos de laboratorio, se estudiaron 250 muestras de heces diarreicas provenientes de pacientes con sospecha de ICD remitidas a los laboratorios de nueve centros médicos entre noviembre de 2010 y diciembre de 2011. Dichas muestras se analizaron mediante los siguientes métodos:1) un ensayo rápido inmunocromatográfico que combina la detección cualitativa de la glutamato deshidrogenasa (GDH) y de las toxinas Ay B (QAB), CDIFF QUIK CHEK COMPLETE;2) un enzimoinmunoanálisis para la determinación cualitativa de las toxinas A/B, RIDASCREENTC. difficile Toxin A/B (RAB);3) un método molecular basado en PCR para la detección del gen que codifica la toxina B (PCR) y 4) el cultivo toxigénico (TC). Como método de referencia se utilizó la combinación del ensayo de citotoxicidad sobre cultivo de células con la neutralización de toxina mediante anticuerpo específico en los filtrados de las heces (CCCNA) y el mismo método en sobrenadantes de aislamientos de C. difficile (CCCNA-TC). La toxigenicidad de las cepas aisladas de muestras directas negativas con QAB, RAB y PCR se evaluó con los mismos métodos, con el propósito de detectar la contribución del TC (QAB-TC, RAB-TC, PCR-TC). De las 250 muestras estudiadas, 107 (42,8%) fueron positivas por CCCNA/CCCNA-TC. Los métodos GDH y PCR/PCR-TC fueron los más sensibles: 91,59 y 87,62%, respectivamente. Los métodos QAB, RAB, QAB/QAB-TC y RAB/RAB-TC mostraron las mayores especificidades, del 95%, aproximadamente. Un resultado negativo para GDH excluiría la ICD, pero su baja razón de verosimilitud positiva (PLR), que fue 3,97, indica que un resultado positivo debe complementarse con la detección de toxinas. Cuando no se detectan toxinas directas por RAB, QAB ni PCR, debería realizarse el TC. De acuerdo con nuestros resultados, los métodos más precisos y confiables para ser aplicados en un laboratorio de microbiología clínica son QAB/QAB-TC y RAB/RAB-TC, con una PLR> 10 y una razón de verosimilitud negativa < 0,30.
Assuntos
Humanos , Toxinas Bacterianas , Reação em Cadeia da Polimerase , Clostridioides difficile , Técnicas Imunoenzimáticas , Proteínas de Bactérias , Toxinas Bacterianas/análise , Clostridioides difficile/genética , Sensibilidade e Especificidade , Enterotoxinas , FezesRESUMO
BACKGROUND The B subunit of Escherichia coli heat-labile enterotoxin (LTB) is a potent mucosal immune adjuvant. However, there is little information about LTB's potential as a parenteral adjuvant. OBJECTIVES We aimed at evaluating and better understanding rLTB's potential as a parenteral adjuvant using the fused R1 repeat of Mycoplasma hyopneumoniae P97 adhesin as an antigen to characterise the humoral immune response induced by this construct and comparing it to that generated when aluminium hydroxide is used as adjuvant instead. METHODS BALB/c mice were immunised intraperitoneally with either rLTBR1 or recombinant R1 adsorbed onto aluminium hydroxide. The levels of systemic anti-rR1 antibodies (total Ig, IgG1, IgG2a, and IgA) were assessed by enzyme-linked immunosorbent assay (ELISA). The ratio of IgG1 and IgG2a was used to characterise a Th1, Th2, or mixed Th1/Th2 immune response. FINDINGS Western blot confirmed rR1, either alone or fused to LTB, remained antigenic; anti-cholera toxin ELISA confirmed that LTB retained its activity when expressed in a heterologous system. Mice immunised with the rLTBR1 fusion protein produced approximately twice as much anti-rR1 immunoglobulins as mice vaccinated with rR1 adsorbed onto aluminium hydroxide. Animals vaccinated with either rLTBR1 or rR1 adsorbed onto aluminium hydroxide presented a mixed Th1/Th2 immune response. We speculate this might be a result of rR1 immune modulation rather than adjuvant modulation. Mice immunised with rLTBR1 produced approximately 1.5-fold more serum IgA than animals immunised with rR1 and aluminium hydroxide. MAIN CONCLUSIONS The results suggest that rLTB is a more powerful parenteral adjuvant than aluminium hydroxide when administered intraperitoneally as it induced higher antibody titres. Therefore, we recommend that rLTB be considered an alternative adjuvant, even if different administration routes are employed.
Assuntos
Animais , Feminino , Camundongos , Toxinas Bacterianas/toxicidade , Adjuvantes Imunológicos/administração & dosagem , Adesinas Bacterianas/imunologia , Proteínas de Escherichia coli/administração & dosagem , Proteínas de Escherichia coli/imunologia , Pneumonia Suína Micoplasmática/imunologia , Pneumonia Suína Micoplasmática/prevenção & controle , Enterotoxinas/administração & dosagem , Suínos , Ensaio de Imunoadsorção Enzimática , Mycoplasma hyopneumoniae , Hidróxido de AlumínioRESUMO
Resumen Introducción. Clostridium difficile es el principal responsable de la diarrea asociada al uso de antibióticos. En Colombia y en Latinoamérica, el conocimiento sobre el comportamiento epidemiológico de la infección por C. difficile todavía es limitado. Objetivo. Describir las características de una serie de pacientes con infección por C.difficile . Materiales y métodos. Se hizo un estudio descriptivo de una serie de casos de pacientes con infección por C. difficile atendidos en la Fundación Clínica Shaio, entre enero de 2012 y noviembre de 2015. Resultados. Se estudiaron 36 pacientes con una edad promedio de 65 años. Se determinaron los siguientes factores relacionados con la infección por C. difficile: uso previo de antimicrobianos (94,4 %), hospitalización en los últimos tres meses (66,7 %) y uso de inhibidores de la bomba de protones (50 %). Las comorbilidades más comunes fueron la enfermedad renal crónica (41,7 %) y la diabetes mellitus (30,6 %). Los síntomas más frecuentes fueron más de tres deposiciones diarreicas (97,1 %) y dolor abdominal (42,9 %). En cuanto a la gravedad de los casos, 44,4 % se clasificó como leve a moderado, 38,9 % como grave, y 11,1 % como complicado o grave. El método de diagnóstico más utilizado (63,8% de los pacientes) fue la identificación de la toxina mediante reacción en cadena de la polimerasa (PCR). La mortalidad global durante la hospitalización fue de 8 %. Se identificaron cuatro cepas del serotipo NAP1/027 y nueve muestras fueron positivas para la toxina binaria. Conclusión. La infección por C. difficile debe sospecharse en pacientes con deposiciones diarreicas y factores asociados tradicionalmente a esta enfermedad. Se reportó la circulación de cepas hipervirulentas del serotipo NAP1/027 en Colombia, lo cual debe enfrentarse con la vigilancia epidemiológica y el diagnóstico temprano.
Abstract Introduction: Clostridium difficile is the main pathogen related to healthcare-associated diarrhea and it is the cause of 20 to 30% of diarrhea cases caused by antibiotics. In Colombia and Latin America, the knowledge about the epidemiological behavior of this infection is limited. Objective: To describe the characteristics of a series of patients with C. difficile infection. Materials and methods: We performed a descriptive case series study of patients with C. difficile infection hospitalized in the Fundación Clínica Shaio from January, 2012, to November, 2015. Results: We analyzed 36 patients. The average age was 65 years. The risk factors associated with the infection were: previous use of antibiotics (94.4%), prior hospitalization in the last three months (66.7%) and use of proton pump inhibitors (50%). The most common comorbidities were chronic kidney disease (41.7%) and diabetes mellitus (30.6%). The most frequent symptoms were more than three loose stools per day (97.1%) and abdominal pain (42.9%). According to the severity of the disease, 44.4% of cases were classified as mild to moderate, 38.9% as severe, and 11.1% as complicated or severe. The detection of the toxin by PCR (GeneXpert) was the most common diagnostic procedure (63.8%). Global mortality during hospitalization was 8%. We identified four strains with serotype NAP1/027 and nine samples positive for binary toxin. Conclusion: Clostridium difficile infection should be suspected in patients with diarrhea and traditional risk factors associated with this disease. We report the circulation of the hypervirulent strain serotype NAP1/027 in Colombia, which should be countered with epidemiological surveillance and a prompt diagnosis.
Assuntos
Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infecção Hospitalar/microbiologia , Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/microbiologia , Proteínas de Bactérias/análise , Toxinas Bacterianas/análise , Virulência , Sorotipagem , Dor Abdominal/etiologia , Comorbidade , Infecção Hospitalar/epidemiologia , Fatores de Risco , Clostridioides difficile/classificação , Clostridioides difficile/patogenicidade , Infecções por Clostridium/epidemiologia , Colômbia/epidemiologia , Diabetes Mellitus/epidemiologia , Diarreia/microbiologia , Diarreia/epidemiologia , Insuficiência Renal Crônica/epidemiologia , Inibidores da Bomba de Prótons/efeitos adversos , Inibidores da Bomba de Prótons/uso terapêutico , Hospitalização , Tempo de Internação/estatística & dados numéricos , Antibacterianos/efeitos adversos , Antibacterianos/uso terapêuticoRESUMO
A total of 268 Bacillus thuringiensis strains obtained from different sources of Argentina were analyzed to determine the diversity and distribution of the cryl, cry2, cry8, cry9 and vip3A genes encoding for lepidopteran-specific insecticidal proteins. Twin strains were excluded. Ten different profiles were detected among the 80 selected B. thuringiensis strains. Two of these profiles (cry1Aa, cry1Ac, cry1Ia, cry2Aa, cry2Ab and vip3Aa (35/80), and cry1Aa, cry1Ab, cry1Ac, cry1Ia, cry2Aa, cry2Ab and vip3Aa (25/80)) pooled 75% of the strains. The existence of this low diversity is rare, since in most of the studied collections a great diversity of insecticidal toxin gene profiles has been described. In addition, the most frequently detected profile was also most frequently derived from soil (70%), stored product dust (59%) and spider webs (50%). In contrast, the cry1Aa, cry1Ab, cry1Ac, cry1Ia, cry2Aa, cry2Ab and vip3Aa profiles were mainly detected in strains isolated from leaves (40%) and dead insect larvae (50%). Six of the identified insecticidal toxin gene profiles were discovered in strains isolated from stored product dust and leaves indicating higher diversity of profiles in these kinds of sources than in others. Some strains with high insecticidal activity against Epinotia aporema (Lepidoptera) larvae were identified, which is important to explore future microbial strategies for the control of this crop pest in the region.
Se analizaron 268 cepas de Bacillus thuringiensis obtenidas de diferentes fuentes de Argentina con el objeto de determinar la diversidad y distribución de genes cryl, cry2, cry8, cry9 y vip3A, que codifican proteínas insecticidas lepidóptero-específicas. Se excluyeron las cepas gemelas. Se detectaron solo diez perfiles diferentes entre los 80 B. thuringiensis seleccionados. Dos de estos perfiles, el cry1Aa, cry1Ac, crylIa, cry2Aa, cry2Ab y vip3Aa (35/80) y el cry1Aa, cry1Ab, cry1Ac, crylIa, cry2Aa, cry2Ab y vip3Aa (25/80), comprendieron el 75% de las cepas seleccionadas. La existencia de esta baja diversidad es una rareza, ya que en la mayor parte de las colecciones estudiadas se ha descrito una gran diversidad de perfiles de genes de toxinas insecticidas. El perfil detectado con mayor frecuencia se obtuvo principalmente de cepas procedentes de suelo (el 70% de los de esa fuente lo tenían), también fue mayoritario entre los procedentes de polvo de producto almacenado (59%) y en los que procedían de telas de arana (50%). En cambio, el perfil cry1Aa, cry1Ab, cry1Ac, crylIa, cry2Aa, cry2Ab y vip3Aa se detectó principalmente en las cepas aisladas de hojas (40%) y de larvas de insectos muertos (50%). Seis de los perfiles identificados fueron encontrados en cepas aisladas de polvo de producto almacenado y de hojas, lo que indica una mayor diversidad de perfiles en estas fuentes que en otras. Se identificaron algunas cepas con alta actividad insecticida contra larvas de Epinotia aporema (Lepidoptera), hallazgo importante para explorar en el futuro estrategias microbianas para el control de esta plaga en la región.
Assuntos
Animais , Bacillus thuringiensis , Toxinas Bacterianas , Genes Bacterianos , Proteínas Hemolisinas , Argentina , Bacillus thuringiensis/genética , Bacillus thuringiensis/patogenicidade , Proteínas de Bactérias , Toxinas Bacterianas/genética , Controle Biológico de Vetores , Endotoxinas , Proteínas Hemolisinas/fisiologia , Proteínas Hemolisinas/genética , Larva , LepidópterosRESUMO
Abstract The aim of this study was to determine the association between Clostridium difficile (C. difficile) and vancomycin-resistant Enterococcus (VRE) and efficacy of screening stools submitted for C. difficile toxin assay for prevalence of VRE. Between April 2012 and February 2014, 158 stool samples submitted for C. difficile toxin to the Marmara University Microbiology Laboratory, were included in the study. Stool samples were analyzed by enzyme immuno assay test; VIDAS (bioMerieux, France) for Toxin A&B. Samples were inoculated on chromID VRE (bioMerieux, France) and incubated 24 h at 37 °C. Manuel tests and API20 STREP (bioMerieux, France) test were used to identify the Enterococci species. After the species identification, vancomycin and teicoplanin MIC's were performed by E test and molecular resistance genes for vanA vs vanB were detected by polymerase chain reaction (PCR). Of the 158 stool samples, 88 were toxin positive. The prevalence of VRE was 17%(n:19) in toxin positives however, 11.4% in toxin negatives(n:70). All VRE isolates were identified as Enterococcus faecium. These results were evaluated according to Fischer's exact chi-square test and p value between VRE colonization and C. difficile toxin positivity was detected 0.047 (p < 0.05). PPV and NPV were 79% and 47% respectively. In our study, the presence of VRE in C. difficile toxin positives is statistically significant compared with toxin negatives (p < 0.05). Screening for VRE is both additional cost and work load for the laboratories. Therefore VRE screening among C. difficile toxin positive samples, will be cost effective for determination of high risk patients in the hospitals especially for developing countries.
Assuntos
Humanos , Toxinas Bacterianas/análise , Clostridioides difficile/metabolismo , Infecções por Clostridium/microbiologia , Resistência a Vancomicina , Fezes/microbiologia , Enterococos Resistentes à Vancomicina/isolamento & purificação , Toxinas Bacterianas/metabolismo , Vancomicina/farmacologia , Testes de Sensibilidade Microbiana , Clostridioides difficile/isolamento & purificação , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/genética , Infecções por Bactérias Gram-Positivas/diagnóstico , Infecções por Bactérias Gram-Positivas/microbiologia , Infecções por Clostridium/diagnóstico , Enterococos Resistentes à Vancomicina/classificação , Enterococos Resistentes à Vancomicina/efeitos dos fármacos , Enterococos Resistentes à Vancomicina/genética , Antibacterianos/farmacologiaRESUMO
Abstract The epsilon toxin, produced by Clostridium perfringens, is responsible for enterotoxemia in ruminants and is a potential bioterrorism agent. In the present study, 15 regions of the toxin were recognized by antibodies present in the serum, with different immunodominance scales, and may be antigen determinants that can be used to formulate subunit vaccines.
Assuntos
Animais , Toxinas Bacterianas/química , Clostridium perfringens/imunologia , Epitopos/química , Toxinas Bacterianas/genética , Toxinas Bacterianas/imunologia , Clostridium perfringens/química , Clostridium perfringens/genética , Enterotoxemia/microbiologia , Mapeamento de Epitopos , Epitopos/genética , Epitopos/imunologiaRESUMO
Background: Gardnerella vaginalis is a bacterial vaginosis (BV)-associated vaginal bacterium that produces the toxin vaginolysin (VLY). VLY is a pore-forming toxin that is suggested to be the main virulence factor of G. vaginalis. The high recurrence rate of BV and the emergence of antibiotic-resistant bacterial species demonstrate the need for the development of recombinant antibodies as novel therapeutic agents for disease treatment. Single-chain variable fragments (scFvs) generated against VLY exhibited reduced efficacy to neutralize VLY activity compared to the respective full-length antibodies. To improve the properties of scFvs, monospecific dimeric scFvs were generated by the genetic fusion of two anti-VLY scFv molecules connected by an alpha-helix-forming peptide linker. Results: N-terminal hexahistidine-tagged dimeric scFvs were constructed and produced in Escherichia coli and purified using metal chelate affinity chromatography. Inhibition of VLY-mediated human erythrocyte lysis by dimeric and monomeric scFvs was detected by in vitro hemolytic assay. The circulating half-life of purified scFvs in the blood plasma of mice was determined by ELISA. Dimeric anti-VLY scFvs showed higher neutralizing potency and extended circulating half-life than parental monomeric scFv. Conclusions: The protein obtained by the genetic fusion of two anti-VLY scFvs into a dimeric molecule exhibited improved properties in comparison with monomeric scFv. This new recombinant antibody might implement new possibilities for the prophylaxis and treatment of the diseases caused by the bacteria G. vaginalis.
Assuntos
Animais , Camundongos , Proteínas de Bactérias/imunologia , Toxinas Bacterianas/imunologia , Anticorpos Neutralizantes/metabolismo , Anticorpos de Cadeia Única/metabolismo , Proteínas de Bactérias/toxicidade , Toxinas Bacterianas/toxicidade , Ensaio de Imunoadsorção Enzimática , Gardnerella vaginalis , Vaginose Bacteriana , Dimerização , Fatores de Virulência , Fusão Gênica , Anticorpos Neutralizantes/imunologia , Anticorpos de Cadeia Única/imunologia , Meia-VidaRESUMO
Mannheimia haemolytica leukotoxin (LKT) is a known cause of bovine respiratory disease (BRD) which results in severe economic losses in the cattle industry (up to USD 1 billion per year in the USA). Vaccines based on LKT offer the most promising measure to contain BRD outbreaks and are already commercially available. However, insufficient LKT yields, predominantly reflecting a lack of knowledge about the LKT expression process, remain a significant engineering problem and further bioprocess optimization is required to increase process efficiency. Most previous investigations have focused on LKT activity and cell growth, but neither of these parameters defines reliable criteria for the improvement of LKT yields. In this article, we review the most important process conditions and operational parameters (temperature, pH, substrate concentration, dissolved oxygen level, medium composition and the presence of metabolites) from a bioprocess engineering perspective, in order to maximize LKT yields.