ABSTRACT
An investigation was carried out into the genetic mechanisms responsible for multidrug resistance in nine carbapenem-resistant Pseudomonas aeruginosaisolates from different hospitals in Recife, Brazil. Susceptibility to antimicrobial agents was determined by broth microdilution. Polymerase chain reaction (PCR) was employed to detect the presence of genes encoding β-lactamases, aminoglycoside-modifying enzymes (AMEs), 16S rRNA methylases, integron-related genes and OprD. Expression of genes coding for efflux pumps and AmpC cephalosporinase were assessed by quantitative PCR. The outer membrane proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The blaSPM-1, blaKPC-2 and blaGES-1 genes were detected in P. aeruginosaisolates in addition to different AME genes. The loss of OprD in nine isolates was mainly due to frameshift mutations, premature stop codons and point mutations. An association of loss of OprD with the overexpression of MexAB-OprM and MexXY-OprM was observed in most isolates. Hyper-production of AmpC was also observed in three isolates. Clonal relationship of the isolates was determined by repetitive element palindromic-PCR and multilocus sequence typing. Our results show that the loss of OprD along with overexpression of efflux pumps and β-lactamase production were responsible for the multidrug resistance in the isolates analysed.
Subject(s)
Humans , Carbapenems/metabolism , Drug Resistance, Multiple, Bacterial/genetics , Mutation , Pseudomonas aeruginosa/drug effects , Pseudomonas aeruginosa/genetics , beta-Lactam Resistance/genetics , beta-Lactamases/metabolism , Aminoglycosides/metabolism , Amphotericin B/analogs & derivatives , Amphotericin B/metabolism , Antifungal Agents/metabolism , Brazil , Cephalosporinase/classification , Cephalosporinase/metabolism , Codon, Nonsense/metabolism , Enzyme Activation/genetics , Frameshift Mutation/genetics , Gene Expression Regulation, Bacterial/genetics , Membrane Transport Proteins/metabolism , Methyltransferases/metabolism , Nucleotidyltransferases/metabolism , Point Mutation/genetics , Porins/metabolism , Pseudomonas aeruginosa/enzymology , Pseudomonas aeruginosa/isolation & purification , Repetitive Sequences, Nucleic Acid , beta-Lactamases/geneticsABSTRACT
NDM-1 (New Delhi metallo-beta-lactamase) gene encodes a metallo-beta-lactamase (MBL) with high carbapenemase activity, which makes the host bacterial strain easily dispatch the last-resort antibiotics known as carbapenems and cause global concern. Here we present the bioinformatics data showing an unexpected similarity between NDM-1 and beta-lactamase II from Erythrobacter litoralis, a marine microbial isolate. We have further expressed these two mature proteins in E. coli cells, both of which present as a monomer with a molecular mass of 25 kDa. Antimicrobial susceptibility assay reveals that they share similar substrate specificities and are sensitive to aztreonam and tigecycline. The conformational change accompanied with the zinc binding visualized by nuclear magnetic resonance, Zn(2+)-bound NDM-1, adopts at least some stable tertiary structure in contrast to the metal-free protein. Our work implies a close evolutionary relationship between antibiotic resistance genes in environmental reservoir and in the clinic, challenging the antimicrobial resistance monitoring.
Subject(s)
Amino Acid Sequence , Anti-Bacterial Agents , Pharmacology , Aztreonam , Pharmacology , Cephalosporinase , Chemistry , Genetics , Metabolism , Computational Biology , Methods , Drug Resistance, Bacterial , Genetics , Enzyme Stability , Evolution, Molecular , Minocycline , Pharmacology , Molecular Sequence Data , Phylogeny , Protein Structure, Tertiary , Sequence Homology, Nucleic Acid , Sphingomonadaceae , Genetics , Tigecycline , Zinc , Pharmacology , beta-Lactamases , Chemistry , Genetics , MetabolismABSTRACT
Organisms producing CTX-M-beta-lactamase are emerging around the world as a source of resistance to oxyiminocephalosporins such as cefotaxime. However, the laboratory detection of these strains is not well defined. The aim of this study was to determine the presence and prevalence of known CTX-M-beta-lactamases genes in clinical isolates of Escherichia coli from hospitals of Tehran. During six months [September to February, 2006], 160 clinical isolates of Escherichia coli collected from three university hospitals of Tehran. Phenotypic screening and confirmation tests for ESBL detection was according to CLSI advised. All of the ESBL-producing isolates were examined by PCR for presence of bla CTX-M genes. Primary phenptypic tests revealed that 56.69% [n=89] of E. coli isolates produced ESBLs. In confirmatory tests by use of clavulanic acid, ESBL production were confirmed [P+C+] in 96.7% [n=86] of isolates with primary positive test. The presence of an ESBL was not confirmed [P+C-] in 3.3% [n=3] of the screen positive. Of all screen positive isolates, 34 [35.78%] were positive for bla CTX-M genes from the CTX-M-I group, indicating CTX-M-1-like beta-lactamases and Two [2.1%] were positive for bla CTX-M genes from the CTX-M-III group, indicating CTX-M-3-like beta-lactamases. The remainder 59 [62.2%] were negative for bla CTX-M genes. The levels of resistance to ceftazidim were remarkably varible among CTX-M producers. This study provides futher evidence of the global dissemination of CTX-M type ESBLs and emphasize the need for their epidemiological monitoring
Subject(s)
Escherichia coli/drug effects , Drug Resistance, Microbial , Cephalosporinase , Polymerase Chain ReactionABSTRACT
Escherichia coli is the germ most frequently isolated in espicially urinary infections. The perpetual change of its sensitivity profile to beta-lactamins is [due to several evolution factors. Some strains have acquiered resistance by gene mutation or by acquisition of movable genetic material. Also phenomenon as impermeability, excretion by efflux system or modifying penicillin liaison proteins could be seen but the production of beta-lactamases remains the most frequent mechanism of resistance that has been describe until now. The authors report, through this observation, a case of a particular strain of Escherichia coli that has been isoleted from urine of an hospitalized patiente. This strain presented an hyperproduction of cefalosporinases, which is a recently discovered mechanism of resistance. In one hand, specific bacteriologic tests as synergy test, antagonism test and the use of cloxacillin added antibiogramme on Mueller-Hinton medium were carried out and have allowed to display the phenotypic aspect of this mechanism. In the other hand, its genetic aspect was mentionned through a new literature data. The authors have also underlined the importance of the antibiogramme prior to any prescription of antibiotics which could put pressure on the germ leading to a selection of individual resistant, to this kind of difficul not controlled resistance and many potential complications ensue from it
Subject(s)
Humans , Female , Escherichia coli/drug effects , Drug Resistance, Bacterial , Microbial Sensitivity Tests , CephalosporinaseABSTRACT
Introducción. Los sistemas de vigilancia son una pieza clave para la detección y control de la resistencia bacteriana. Es indispensable recolectar constantemente la información de cada institución por la variabilidad existente entre países, ciudades y hospitales frente a los mecanismos de resistencia bacteriana y así plantear intervenciones apropiadas para cada institución. De enero 2003 a diciembre de 2005, el Centro Internacional de Entrenamiento e Investigaciones Médicas (CIDEIM) desarrolló un proyecto de vigilancia en un grupo de 10 hospitales de tercer nivel, en seis ciudades de Colombia. Objetivos. Presentar el comportamiento de Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii y Enterobacter cloacae, considerados los Gram negativos patógenos más relevantes en infección nosocomial, frente a antimicrobianos seleccionados. Materiales y métodos. Las pruebas de susceptibilidad se realizaron por métodos automatizados en 9 hospitales y por Kirby Bauer en un hospital. Se eligieron antibióticos con actividad reconocida contra Gram negativos, de acuerdo con las guías del Comité Nacional para el Control de Estándares en el Laboratorio Clínico (NCCLS). Los laboratorios realizaron control de calidad interno y externo. Mensualmente se recibió la información procedente del laboratorio de microbiología de cada institución y se centralizó en una base de datos en WHONET 5.3 en CIDEIM . Se realizó un análisis conglomerado de frecuencias y porcentajes de resistencia a antibióticos. Resultados. Los altos porcentajes de resistencia reportados en especial para A. baumannii, corroboraron la presencia de bacterias multirresistentes en las UCI en las instituciones participantes durante el periodo de estudio. Conclusiones. Es urgente crear una red nacional de vigilancia de la resistencia a antimicrobianos de los patógenos hospitalarios y de esta manera mejorar nuestra habilidad para detectar, supervisar y manejar la resistencia a antimicrob...
Introduction. Surveillance systems play a key role in the detection and control of bacterial resistance. It is necessary to constantly collect information from all institutions because the mechanisms of bacterial resistance can operate in different ways between countries, cities and even in hospitals in the same area. Therefore local information is important in order to learn about bacterial behaviour and design appropriate interventions for each institution. Between January 2003 and December 2004, the Centro Internacional de Entrenamiento e Investigaciones Médicas (CIDEIM) developed a surveillance project in 10 tertiary hospitals in 6 cities of Colombia. Objectives. Describe the trends of antibiotic resistance among the isolates of Escherichia coli, Klebsiella pneumoniae, Pseudomona aeruginosa, Acinetobacter baumannii and Enterobacter cloacae, five of the most prevalent nosocomial Gram negative pathogens. Materials and Methods: The susceptibility tests were performed by automated methods in 9 hospitals and by Kirby Bauer in 1 hospital. Antibiotics with known activity against Gram negatives, according to the Clinical Laboratory Standards Institute guidelines, were selected. The laboratories performed internal and external quality controls. During the study period, the information was downloaded monthly from the databases of each microbiology laboratory and sent to CIDEIM where it was centralized in a database using the system WHONET 5.3. Results. The high resistance rates reported especially for A. baumannii, evidenced the presence of multidrug resistant bacteria in both ICUs and wards at every studied institution. Conclusions. The creation of a national surveillance network to improve our capabilities to detect, follow up, and control the antibiotic resistance in Colombia is urgently needed.
Subject(s)
Drug Resistance, Microbial , Gram-Negative Bacterial Infections , Carbapenems , Cephalosporinase , PiperacillinSubject(s)
Humans , beta-Lactamases , Cephalosporinase , Infection Control , Epidemiological MonitoringABSTRACT
En este trabajo se évalua el problema de la resistencia bacteriana a los antibióticos betalactamicos en venezuela. Se presentan los datos de resistencia bacteriana de varios hospitales venezolanos desde 1988 hasta 1992. Se analizan los resultados de las siguientes bacterias: Staphylococcus aureus, Staphylococus epidermidis, Enterococus faecalis, Escherichia coli, Klebsiella pneumoniae, Enterobacter aerogenes, Enterobacter cloacae, Proteus mirabilis, Proteus vulgaris, Salmonella sp., Shigella sp., Pseudomonas aeruginosa, Streptococus pneumoniae, Neisseria gonorrhoeae y Neisseria meningitidis
Subject(s)
Humans , Male , Female , Cephalosporinase , Drug Resistance, Microbial , Penicillinase , Anti-Bacterial Agents , Bacteria/analysisABSTRACT
Se estudiaron 100 muestras de exudados nasales obtenidos de estudiantes de segundo año de Medicina del Instituto Superior de Ciencias Médicas de Villa Clara, de enero a julio de 1987. El objectivo es conocer el índice de portadores de estafilococos por la implicación que tiene en su futura actividad hospitalaria a partir del tercer año, y para investigar la sensibilidad y resistencia de las cepas a los diferentes antimicrobianos. Se aislaron 86 cepas de estafilococos y se les realizó la prueba coagulasa, de las que 48 resultaron positivas. Se comprobó que la novobiocina era el agente antimicrobiano más efectivo. Se verificó la alta resistencia de los estafilococos a la penicilina y se pudo comprobar que un gran por ciento de las cepas son productoras de Beta-lactamasas