ABSTRACT
En este trabajo se investigó la presencia de determinantes característicos de plásmidos de virulencia en dos aislamientos clínicos de Salmonella Infantis portadores de plásmidos de multirresistencia. Además, se estudió la capacidad de invasión y proliferación en células eucariotas no fagocíticas. Ninguno de los aislamientos de S. Infantis mostró los determinantes genéticos que caracterizan a los plásmidos de virulencia para este género (operón spv). Los ensayos de invasión sobre líneas celulares eucariotas mostraron que los aislamientos de S. Infantis presentan una capacidad de invasión disminuida pero persisten y proliferan en el citoplasma, independientemente de utilizar una línea celular permisiva (HeLa) o no permisiva (NRK) para tal fin. Finalmente, no se observaron indicios microscópicos que podrían hacer sospechar un efecto bactericida de estas líneas celulares sobre los aislamientos estudiados.
Two multidrug-resistant Salmonella Infantis isolates behave like hypo-invasive strains but have high intracellular proliferation. In this work, plasmid-encoded virulence factors in two Salmonella Infantis isolates carrying multiresistance plasmids were investigated. In addition, their invasion and proliferative ability in non-phagocytic cells was studied. None of them showed the typical determinants of virulence plasmids (spv operon). The invasion assays of S. Infantis isolates on eukaryotic cells showed a decreased ability to Invade but they remained and proliferated In the cytoplasm regardless of having used a permissive (HeLa) or non-permissive (NRK) cell line. Finally, there was no microscopic evidence suggesting a bactericidal effect of these eukaryotic cell lines on the Isolates tested.
Subject(s)
Animals , Humans , Rats , Drug Resistance, Multiple, Bacterial/genetics , Eukaryotic Cells/microbiology , R Factors/physiology , Salmonella/pathogenicity , Blood/microbiology , Cell Division , Cell Line/microbiology , Cross Infection/microbiology , Feces/microbiology , Genes, Bacterial , Genetic Markers , HeLa Cells/microbiology , Kidney/cytology , R Factors/genetics , R Factors/isolation & purification , Salmonella Infections/microbiology , Salmonella/drug effects , Salmonella/genetics , Salmonella/isolation & purification , Virulence/geneticsABSTRACT
OBJECTIVE: To determine the prevalence of resistance to ampicillin, chloramphenicol, streptomycin, sulphonamides, and tetracyclines (ACSSuT) in Salmonella serovar Typhimurium definitive [phage] type (DT) 193 strains isolated from human sources over the last four decades. METHODS: From 2008 to 2010, 553 DT193 isolates out of 810 human-origin Salmonella ser. Typhimurium phage-typed strains isolated from the 1970s through 2008 were selected and tested for ACSSuT resistance: 91 strains isolated during the 1970s, 65 from the 1980s, 70 from the 1990s, and 327 from 2000-2008. Resistance profiles were determined using the disk diffusion method. RESULTS: An antimicrobial susceptibility assay indicated 20.9 percent, or 116, of all isolates tested were ACSSuT-resistant, 52.0 percent (287) were resistant to one or more drugs in the ACSSuT profile, and 27.1 percent (150) were nonresistant (susceptible to antimicrobials). Based on the assay, overall antimicrobial resistance was extremely high in the 1970s (affecting 99.0 percent of isolates from that period) and remained high during the 1980s, when 95.4 percent of isolates had some type of antimicrobial resistance and incidence of Salmonella ser. Typhimurium DT193 R-type ACSSuT increased to 73.8 percent. R-type ACSSuT dropped to 27.1 percent (19 isolates) during the 1990s, and to 5.2 percent (17) during 2000-2008, despite a substantial increase in the number of isolates tested (397 versus 204, 111, and 98, respectively, for the previous three decades). CONCLUSIONS: Although prevalence of Salmonella ser. Typhimurium DT193 R-type ACSSuT in Brazil has decreased since the 1970s, ACSSuT resistance markers continue to circulate. Therefore, continuous surveillance should be conducted to evaluate the occurrence of Salmonella ser. Typhimurium DT193 and its antimicrobial resistance.
OBJETIVO: Determinar la prevalencia de resistencia a la ampicilina, el cloranfenicol, la estreptomicina, las sulfonamidas y las tetraciclinas (ACSSuT) en cepas de Salmonella serovariedad Typhimurium fagotipo definitivo (DT) 193 aisladas de fuentes de origen humano durante las cuatro últimas décadas. MÉTODOS: Entre el 2008 y el 2010 se seleccionaron 553 aislados de DT193 entre 810 cepas de Salmonella serovariedad Typhimurium fagotipificadas aisladas desde la década de 1970 hasta el 2008, y en ellos se analizó la resistencia a ACSSuT: se estudiaron 91 cepas aisladas durante la década de 1970, 65 aisladas durante la década de 1980, 70 aisladas durante la década de 1990, y 327 aisladas entre el 2000 y el 2008, respectivamente. Los perfiles de resistencia a los antibióticos se determinaron mediante el método de difusión en disco. RESULTADOS: El antibiograma indicó que 20,9 por ciento (116) de todos los aislados que se analizaron fueron resistentes a ACSSuT, 52,0 por ciento (287) fueron resistentes a uno o más antibióticos del grupo ACSSuT y 27,1 por ciento (150) no fueron resistentes (es decir, fueron sensibles a dichos antibióticos). Según el análisis, la resistencia general a los antibióticos fue muy alta en la década de 1970 (y comprendió a 99,0 por ciento de los aislados de ese período) y continuó siendo alta durante la década de 1980, cuando 95,4 por ciento de los aislados presentó algún tipo de resistencia a los antibióticos y la incidencia de Salmonella serovariedad Typhimurium DT193 con resistencia de tipo ACSSuT aumentó hasta 73,8 por ciento. La resistencia de tipo ACSSuT descendió a 27,1 por ciento (31 aislados) durante la década de 1990, y a 5,2 por ciento (17 aislados) entre el 2000 y el 2008, a pesar del aumento importante en el número de aislados que se evaluaron (397 frente a 204, 111 y 98 en las tres décadas anteriores, respectivamente). CONCLUSIONES: Aunque la prevalencia de Salmonella serovariedad Typhimurium DT193 con resistencia de tipo ACSSuT en el Brasil ha disminuido desde la década de 1970, los marcadores de resistencia a ACSSuT continúan en circulación. Por consiguiente, debe llevarse a cabo una vigilancia permanente para evaluar la aparición de infecciones por Salmonella serovariedad Typhimurium DT193 y su resistencia a los antibióticos.
Subject(s)
Humans , Animals , Drug Resistance, Multiple, Bacterial , R Factors/genetics , Salmonella Infections/microbiology , Salmonella typhimurium/drug effects , Anti-Bacterial Agents/pharmacology , Bacteriophage Typing , Brazil/epidemiology , Drug Resistance, Multiple, Bacterial/genetics , Food Chain , Retrospective Studies , Salmonella Infections, Animal/microbiology , Salmonella Infections/drug therapy , Salmonella Infections/epidemiology , Salmonella typhimurium/classification , Salmonella typhimurium/genetics , Salmonella typhimurium/isolation & purification , Serotyping , ZoonosesABSTRACT
The aim of this study was to investigate the presence of tetracycline and oxytetracycline resistance determinants in Bacillus cereus strains isolated from honey samples. Of a total of 77 isolates analyzed, 30 (39%) exhibited resistance to tetracyclines according to the results of a disk diffusion method. Resistant strains (n=30) were screened by PCR for the presence of the resistant determinants tetK, tetL, tetM, tetO, tetW, otrA and otrB and their MIC values for tetracycline, oxytetracycline and minocycline were assessed. According to the PCR results, 23 isolates (77%) presented at least one tetracycline or oxytetracycline resistance determinant. The tetK genotype was present in 10 isolates while the tetL, tetM, and otrA genotypes were present in 3, 2, and 5 isolates, respectively. In addition, 2 isolates of the tetK plus tetM genotype, 1 of the tetK plus tetL genotype, and 1 of the tetK plus otrA genotype were found. All isolates were tetW, tetO and otrB negatives. On the other hand, 7 isolates (23%) showed a tetracycline-resistant and/or minocyclineresistant phenotype (MIC) but did not carry any of the tet or otr determinants investigated in this study. This research has shown that B. cereus isolates from honey samples contain a variety of tetracycline and oxytetracycline resistance genes, including the tetK and tetL determinants which encode for efflux proteins, and tetM and otrA, which encode for ribosomal protection proteins. These findings indicate that strains isolated from honeys could represent a reservoir for tetracycline resistance genes. To our knowledge, this is the first report of tetracycline-resistant and oxytetracyclineresistant B. cereus strains carrying the tetK determinant, and also the first report of oxytetracycline-resistant and tetracycline- resistant Bacillus species carrying the otrA determinant.
El objetivo del presente estudio ha sido investigar la presencia de diversos determinantes de resistencia a tetraciclina y oxitetraciclina en las poblaciones de Bacillus cereus presentes en la miel. De un total de 77 aislamientos evaluados, 30 (39%) resultaron resistentes a tetraciclina y/o minociclina de acuerdo con los resultados de las pruebas de difusión en disco. Dentro del grupo que presentó un fenotipo resistente, se investigó la presencia de los determinantes tetK, tetL, tetM, tetO, tetW, otrA y otrB por PCR y se determinaron los valores de CIM para tetraciclina, oxitetraciclina y minociclina. De acuerdo con los resultados obtenidos por PCR, 23 aislamientos (77%) presentaron al menos un determinante de resistencia a tetraciclina o a oxitetraciclina; el genotipo tetK se encontró en 10 de esos aislamientos, mientras que los genotipos tetL, tetM y otrA se hallaron en 3, 2 y 5 aislamientos, respectivamente. Ningún aislamiento presentó los genotipos tetW, tetO ni otrB. Adicionalmente, se encontraron los genotipos tetK plus tetM (2 aislamientos); tetK plus tetL (1 aislamiento) y tetK plus otrA (1 aislamiento). Por otra parte, 7 cepas (23%) resultaron resistentes a tetraciclina, oxitetraciclina y/o minociclina por CIM, pero no presentaban ninguno de los determinantes tet u otr estudiados. Estos resultados indican la existencia de un alto porcentaje de cepas de B. cereus aisladas de miel con genes de resistencia a tetraciclina y oxitetraciclina, incluyendo los determinantes tetK, tetL, tetM y otrA. Este estudio constituye el primer registro de la presencia del determinante tetK de resistencia a tetraciclina en B. cereus, como así también la presencia del determinante otrA dentro del género Bacillus.
Subject(s)
Bacillus cereus/drug effects , Drug Resistance, Multiple, Bacterial/genetics , Honey/microbiology , R Factors/genetics , Tetracycline Resistance/genetics , Antiporters/genetics , Bacillus cereus/genetics , Bacillus cereus/isolation & purification , Bacterial Proteins/genetics , Genes, Bacterial , Genotype , Italy , Latin America , Microbial Sensitivity Tests , Minocycline/pharmacology , Oxytetracycline/pharmacology , Ribosomal Proteins/genetics , Sampling Studies , Tetracycline/pharmacology , United StatesABSTRACT
OBJECTIVE: In this work we report the molecular characterization of beta-lactam antibiotics resistance conferred by genes contained in plasmids from enterobacteria producing extended-spectrum beta-lactamases (ESBL). MATERIAL AND METHODS: Fourteen enterobacterial clinical isolates selected from a group of strains obtained from seven different hospitals in Mexico during 1990-1992 and 1996-1998 were analyzed at the Bacterial Resistance Laboratory (National Institute Public Health, Cuernavaca). Molecular characterization included PFGE, IEF of beta-lactamases, bacterial conjugation, PCR amplification and DNA sequencing, plasmid extraction and restriction. RESULTS: Isolates were genetically unrelated. ESBL identified were SHV-2 (5/14) and SHV-5 (9/14) type. Cephalosporin-resistance was transferable in 9 of 14 (64 percent) clinical isolates with only one conjugative plasmid, DNA finger printing showed a similar band pattern in plasmids. CONCLUSIONS: The dissemination of cephalosporin resistance was due to related plasmids carrying the ESBL genes.
OBJETIVO: En este trabajo se reporta la caracterización molecular de la resistencia a antibiótico beta-lactámicos conferida por genes contenidos en plásmidos de enterobacterias productoras de beta-lactamasas de espectro extendido (BLEEs). MATERIAL Y MÉTODOS: Catorce aislamientos clínicos de enterobacterias fueron seleccionados por conveniencia de un banco de cepas obtenidas de siete diferentes hospitales de México durante los periodos 1990-1992 y 1996-1998 y fueron procesados en el Laboratorio de Resistencia Bacteriana (Instituto Nacional de Salud Pública, Cuernavaca). En la caracterización se empleó PFGE, IEF para beta-lactamasas, conjugación bacteriana, amplificación por PCR y secuenciación de DNA, extracción y restricción de plásmidos. RESULTADOS: Las 14 cepas fueron no relacionadas genéticamente. Se identificaron BLEEs tipo SHV-2 (5/14) y SHV-5 (9/14). La resistencia a cefalosporinas fue transferida por conjugación en 9 de 14 (64 por ciento) aislamientos clínicos mediante un plásmido que mostró un patrón de restricción similar entre ellos. CONCLUSIÓN: Se sugiere que la diseminación de la resistencia a cefalosporinas fue debida a plásmidos relacionados que contienen los genes que codifican BLEEs.
Subject(s)
Humans , Bacterial Proteins/genetics , Cross Infection/microbiology , Drug Resistance, Multiple, Bacterial/genetics , Escherichia coli Infections/microbiology , Escherichia coli/enzymology , Klebsiella Infections/microbiology , Klebsiella/enzymology , R Factors/genetics , beta-Lactam Resistance/genetics , beta-Lactamases/genetics , Bacterial Proteins/classification , Bacterial Proteins/isolation & purification , Cross Infection/epidemiology , DNA Fingerprinting , DNA, Bacterial/genetics , Escherichia coli Infections/epidemiology , Escherichia coli Proteins/classification , Escherichia coli Proteins/genetics , Escherichia coli Proteins/isolation & purification , Escherichia coli/drug effects , Escherichia coli/genetics , Klebsiella Infections/epidemiology , Klebsiella/drug effects , Klebsiella/genetics , Mexico/epidemiology , beta-Lactamases/classification , beta-Lactamases/isolation & purificationABSTRACT
Investigou-se a transferência de marcadores genéticos e a presença de DNA plasmidial em 240 culturas de Escherichia coli originárias de água de esgoto (afluente e fluentes) da Estaçäo de Tratamento da Ilha do Governador, na cidade do Rio de Janeiro, RJ. Experimentos de conjugaçäo com E. coli K 12 permitiram o isolamento de transconjugantes com resistência a antibióticos (Su, Sm, Tc, Cm e Ap); a metais pesados (Cu, Hg e Zn) e fatores colicinogênicos (Col Ia, Ib e V) principalmente para os coliformes isolados nos setores terminais da estaçäo de tratamento. A distribuiçäo de plasmídeos foi prevalente nas culturas de E. coli advindas dos efluentes, com percentuais superiores a 65
Subject(s)
DNA, Bacterial/isolation & purification , Escherichia coli/genetics , R Factors/genetics , Sewage , Brazil , Genetic MarkersABSTRACT
Para analisar cepas de Salmonella ser. Typhimurium isoladas de processos entéricos e extraintestinais humanos ocorridos no período de 1970 a 2008 de diferentes regiões do país foram selecionadas, com base nos registros contidos no banco de dados do Laboratório de Enterobactérias do IOC/FIOCRUZ, RJ, amostras do fagotipo prevalente 193, visando precipuamente o reconhecimento de clones epidêmicos. Foram selecionadas 553 cepas de Salmonella ser. Typhimurium fagotipo 193 representadas por 91, 65, 70 e 327 amostras referentes as décadas de 70, 80, 90 e ao período de 2000 a 2008, respectivamente. Na análise global da sensibilidade destas cepas, 52% apresentaram um ou mais marcadores de resistência a antibióticos incluídos no perfil ACSSuT. Este perfil de resistência completo foi verificado em 20,9% dos isolados, sendo os 21,9% restantes, sensíveis a todas as drogas testadas, especialmente no período de 2000 a 2008, representadas por 121 amostras (37,0%) em relação as 327 culturas dessa época. O maior percentual de resistência foi observado nas amostras da década de 70 (99%) sendo o perfil ACSSuT detectado em 35,2% dos isolados, ressaltando-se que todas as amostras foram isoladas de processos gastroentéricos ocorridos na cidade de São Paulo. Ao longo das quatro décadas de estudo, descreve-se um ponto de ruptura entre a prevalência de resistência e a suscetibilidade na transição entre as décadas de 80 e 90. Embora o número de isolados de Salmonalla ser. Typhimurium fagotipo 193 tenha aumentado no último período considerado, o percentual de mono e multirresistência aos antimicrobianos se situou em nível elevado (63,0%). A análise do polimorfismo obtido após macrorrestrição com a enzima Xbal revelou que cepas isoladas na década de 90 apresentaram elevado percentual de similaridade (>-85%) com cepas isoladas recentemente (período de 2000-2008), sendo agrupadas nos mesmos "subclusters". Por outro lado, as cepas da década de 70 inserem-se em "subclusters" independentes...
To analyze strains of Salmonella ser. Typhimurium isolated from human cases of enteric and extraintestinal occurred during the period 1970 to 2008 of different regions of Brazil were selected, based on records in the database from Enterobacteria Laboratory of IOC/FIOCRUZ, RJ, samples prevalent phage type 193 in order to recognition of epidemic clones. We selected 553 strains of Salmonella ser. Typhimurium phage type 193 represented by 91, 65, 70 and 327 samples concerning the 1970s, 1980s, 1990s and the period from 2000 to 2008, respectively. In a global analysis of the sensitivity of these strains, 52% had one or more antibiotic resistance markers included in the profile ACSSuT. This resistance profile was found complete in 20.9% of isolates and the remaining 21.9%, sensitive to all drugs tested, especially in the period 2000 to 2008, represented by 121 samples (37.0%) compared the 327 cultures of that time. The highest percentage of resistance was observed in the samples of the 70 (99%) being the profile ACSSuT detected in 35.2% of isolates, emphasizing that all strains were isolated from gastrointestinal processes occurring in São Paulo city. Over the four decades of study, we describe a breaking point between the prevalence of resistance and susceptibility in the transition between the 1980s and 1990s. Although the number of isolates of Salmonella ser. Typhimurium phage type 193 has increased in the last period, the percentage of mono-and-multidrug resistance to antimicrobial agents stood at high level (63.0%). The analysis of polymorphism obtained after macrorestriction with the enzyme Xbal showed that isolates in the 1990s showed a high percentage of similarity (>-85%) with strains isolated recently (2000-2008) and are grouped in the same subclusters. Moreover, the strains of the 1970s fall into subclusters independent, although the percentage of similarity between such subsclusters and the other is >- 70%, the same was observed for the strains...
Subject(s)
Humans , DNA, Bacterial/analysis , Drug Resistance, Multiple, Bacterial , Electrophoresis, Gel, Pulsed-Field/methods , Salmonella Phages/isolation & purification , Salmonella Infections/microbiology , Salmonella Infections/drug therapy , Salmonella typhimurium , Salmonella typhimurium/genetics , Salmonella typhimurium/isolation & purification , R Factors/genetics , Microbial Sensitivity TestsABSTRACT
Proteus mirabilis secreta uma única protease que é uma metalo-enzima alcalina. Descrevemos aqui alguns fatores fisiológicos que afetam a secreçäo de protease em clones isolados. Mais de cem diferentes isolados clínicos foram examinados quanto a capacidade de secretar protease. Verificou-se que algumas linhagens herdam o determinante de protease como um caráter estável (säo denominadas linhagens estáveis); outras linhagens exigem um fenótipo muito instável (denominadas linhagens instáveis) com 10% das colônias-filhas perdendo o caráter de produçäo de protease. Nunca foi observada a reversäo do fenótipo protease negativa. Por outro lado, a introduçäo de fatores R em células produtoras de protease reduziu a atividade proteásica em cerca de 50%, o que sugere uma interferência na expressäo e/ou secreçäo de protease. Embora haja evidências de que um locus da protease tenha localizaçäo extracromossômica, näo foi demosntrada a presença de plasmídios nas linhagens instáveis. Um modo que pode explicar os vários dados é apresentado
Subject(s)
Peptide Hydrolases/metabolism , Proteus mirabilis/genetics , Proteus mirabilis/enzymology , R Factors/geneticsABSTRACT
Salmonella typhimurium fermentadoras de lactose säo comuns em Säo Paulo, porém, raras no Rio de Janeiro, onde descrevemos dois isolamentos. Um plasmídeo de 7,4 megadáltons, näo auto-ransferível, termorresistente e näo eliminável pelo alaranjado de acridina foi identificado em cada uma das duas linhagens isoladas no Rio de Janeiro. O fato de uma dessas linhagens ser derivada de um plasmídeo que originalmente näo expressa o caráter fermentaçäo de lactose, permite-nos especular acerca da origem deste caráter nas Salmonellas brasileiras
Subject(s)
Lactose Factors/genetics , Lactose/metabolism , Plasmids/genetics , R Factors/genetics , Salmonella typhimurium/genetics , Brazil , Conjugation, Genetic , Fermentation , Lactose Factors/physiology , R Factors/physiology , Salmonella typhimurium/metabolismABSTRACT
Se analizó el contenido plasmídico de una serie de cepas de Klebsiella pneumoniae resistentes a amikacina y otros antibióticos. Estas cepas se identificaron como causantes de epidemias intrahospitalares en unidades pediátricas situadas en distintas localidades geográficas de Argentina. En todos los casos se encontraron plásmidos que poseían determinantes genéticos para resistencia a amikacina. Mediante análisis realizados con cortes con enzimas de restricción e hibridizaciones se determinó la presencia de elementos transponibles relacionados a Tn 1331 en todos las cepas estudiadas. Estos resultados indican que la transposición de estos elementos ha jugado un papel importante en el proceso de diseminación de la resistencia a la amikacina en el género Klebsiella (y probablemente en otras bacterias gram negativas) en Argentina
Subject(s)
Amikacin/pharmacology , Klebsiella pneumoniae/genetics , R Factors/genetics , Argentina , Chromosome Mapping , DNA Transposable Elements/drug effects , Drug Resistance, Microbial/genetics , R Factors/drug effectsABSTRACT
We report a novel phenomenon of high genetic instability, related to auxotrophy, in strains of Proteus mirabilis. Among P. mirabilis strains harboring the R plasmid Kept in our laboratory collection, and some freshly isolated strians from clinical material, 54 per cent of the samples presented auxotrophy at frequencies higher than 10(-3). Prototrophic closes gave rise to auxotrophic ones at frequencies not explainable by the usual mutation mechanisms. The instability mainly affected the carbamoyl phosphate synthetase gene (car), which leads to a double requirement for arginine and uracil for growth in minimal medium. Other genes were also affected, at a lower frequency. The car mutation does not revert to prototrophy. A similar phenomenon of instability was induced in Escherichia coli strain HB 101 upon introduction of a drug-resistance plasmid from P. mirabilis. We have ruled out the hypothesis of a transposon in the generation of auxotrophy.