pKpQIL-like plasmid contributes to the dissemination of blaNDM-1 and plasmid mediated quinolone resistance determinants among multi drug resistant Klebsiella pneumoniae in Assiut university hospital, Egypt.

Background and Objectives: Concomitant carriage of blaNDM-1 and plasmid mediated quinolone resistance determinants (PMQRs) by multi drug resistant (MDR) Klebsiella pneumoniae (K. pneumoniae) has increased globally, often related to their presence on transmissible plasmids. In this study, we hypothesized the presence of blaNDM-1 and PMQRs on a single conjugative plasmid that circulates among K. pneumoniae strains isolated from Assiut University Hospital. Materials and Methods: Twenty-two clinical MDR K. pneumoniae strains harboring both blaNDM-1 and PMQRs were genotyped using pulsed field gel electrophoresis. Horizontal transfer of blaNDM-1 and PMQRs was evaluated by conjugation and trans-conjugants were screened for the presence of both genes and integron by PCR. Trans-conjugant's plasmid DNA bands were purified using agarose gel electrophoresis and different DNA bands were screened for blaNDM-1 and PMQRs. Plasmids carrying blaNDM-1 and PMQRs were typed by PCR based replicon typing. Results: All MDR K. pneumoniae contained class 1 integron and belonged to 15 pulsotypes. BlaNDM-1 and PMQRs were co-transferred in each conjugation process. Multiple replicons (5-9 types) were detected in each trans-conjugant; with IncFIIK and IncFIB-KQ replicons being common among all trans-conjugants. Both blaNDM-1 and PMQRs were detected on a pKpQIL-like multi-replicon plasmid that was present in all K. pneumoniae strains. Conclusion: In view of these results, the presence of blaNDM-1 and PMQRs on pKpQIL-like plasmid that existed in multiple unrelated K. pneumoniae isolates is highly suggestive of the circulation of pKpQIL-like MDR plasmids in our hospitals. Moreover, carriage of integrons by the-circulating MDR plasmids increases the risk of dissemination of antimicrobial resistance among pathogens.

Evaluación de métodos fenotípicos para la detección de metalobetalactamasas en aislados clínicos de Pseudomonas aeruginosa / Assessment of Phenotypic Methods for Metalobetalactamase Detection in Clinical Isolates of Pseudomonas aeruginosa

Pseudomonas aeruginosa es considerado uno de los patógenos nosocomiales más importantes, siendo común su aislamiento en pacientes de unidades de cuidados intensivos, en quienes además suele presentar multirresistencia, esto limita enormemente las opciones terapéuticas para el tratamiento del paciente, lo que constituye un problema ya que no se disponen de drogas efectivas para el tratamiento. El presente trabajo tiene como objetivo evaluar cuatro métodos fenotípicos para la detección de metalo-beta-lactamasas (método de sinergia del doble disco con EDTA, método del disco combinado de imipenem y meropenem con EDTA, método de E-Test-MBL y el método de Hodge modificado), con un método de referencia (PCR), para determinar el método fenotípico o combinación de estos que sea más valido y seguro para su implementación en los laboratorios de bacteriología de rutina. La sensibilidad y especificidad de los métodos fenotípicos empleados fue buena (más de 90%), de los métodos empleados, el test de Hodge modificado fue el que proporcionó mejores resultados en la detección de aislados de P. aeruginosa productores de MBL (menor tasa de falsos positivos y negativos), al analizar los diferentes métodos combinados para determinar cuál es la asociación con mayor sensibilidad y especificidad, el método del doble disco asociado con el test de Hodge modificado reunieron estas características, estos resultados sugieren que estos dos métodos se deben emplear de rutina en un laboratorio de Bacteriología, por ser fáciles de realizar, económicos, sensibles y específicos.

Pseudomonas aeruginosa is considered one of the most important nosocomial pathogens. Its isolation is commonin intensive care unit patients who also usually have multi-resistance. This greatly limits therapeutic options, since no effective drugs are available for treatment. The study aims to evaluate four phenotypic methods for detecting metallo-beta-lactamases. The double disc synergy with EDTA method, combined imipenem and meropenem disk with EDTA method, the E-Test-MBL method and the modified Hodge method, along with a reference method (PCR), are used to determine the phenotypic method or a combination of these is used that is more valid and safe for implementation in routine bacteriology laboratories. The sensitivity and specificity of the phenotypic methods used was good (over 90%); the modified Hodge test obtained the best results in detecting MBL-producing P. aeruginosa isolates (lower rate of false positive and negative). On analyzing the different combined methods for determining the association with greater sensitivity and specificity, the double disc method associated with the modified Hodge test met these characteristics. Results suggest that these two methods should be routinely used in a bacteriology laboratory, since they are easy to perform, sensitive, specific and economical.

Detecção de metalo-beta-lactamase em Pseudomonas aeruginosa isoladas de pacientes hospitalizados em Goiânia, Estado de Goiás / Detection of metallo-beta-lactamase in Pseudomonas aeruginosa isolated from hospitalized patients in Goiânia, State of Goiás

Pseudomonas aeruginosa é uma bactéria frequentemente isolada no ambiente hospitalar. Este estudo teve como objetivo avaliar o perfil de suscetibilidade de Pseudomonas aeruginosa previamente isoladas de pacientes internados em um hospital de Goiânia (Goiás-Brasil); realizar a triagem fenotípica para a produção de metalo-beta-lactamase e detectar os genes das mesmas pela técnica de "Polimerase Chain Reaction". Foram avaliadas 75 Pseudomonas aeruginosa isoladas no período de janeiro de 2005 a janeiro de 2007. A identificação bioquímica foi realizada pelo sistema API 20E® e o antibiograma pelo método de Kirby-Bauer. Entre os 62 isolados que foram resistentes ao imipenem e à ceftazidima, 35 (56,4 por cento) apresentaram produção de metalo-beta-lactamase e em 26 (74,3 por cento) destes, foi detectado o gene blaSPM-1. A frequência de Pseudomonas aeruginosa produtoras de metalo-beta-lactamase sugere um maior controle da disseminação de resistência no ambiente hospitalar.

Pseudomonas aeruginosa is a bacterium frequently isolated from hospital environments. This study had the aims of evaluating the susceptibility profile of Pseudomonas aeruginosa previously isolated from patients in a hospital in Goiânia (Goiás, Brazil), performing phenotypic screening for metallo-beta-lactamase production and detecting its genes using the polymerase chain reaction technique. Seventy-five 75 Pseudomonas aeruginosa isolates were evaluated between January 2005 and January 2007. Biochemical identification was performed using the API 20E® system and an antibiogram was produced using the Kirby-Bauer method. Among the 62 isolates that were resistant to imipenem and ceftazidime, 35 (56.4 percent) produced metallo-beta-lactamase, while 26 (74.3 percent) showed the blaSPM-1 gene. The frequency of Pseudomonas aeruginosa that produces metallo-beta-lactamase suggests that greater control over the dissemination of resistance in hospital environments is needed.

Detecção de metalo-beta-lactamases em amostras hospitalares de Pseudomonas aeruginosa e Acinetobacter baumannii / Detection of metallo-beta-lactamases in hospital strains of Pseudomonas aeruginosa and Acinetobacter baumannii

INTRODUÇÃO E OBJETIVO: Entre os bacilos Gram-negativos não fermentadores da glicose, Pseudomonas aeruginosa e Acinetobacter baumannii são os agentes etiológicos de infecções hospitalares mais frequentes. A resistência aos carbapenemas entre esses patógenos está se tornando um problema terapêutico mundial e a produção de metalo-beta-lactamases (MBL) tem emergido como um dos mecanismos responsáveis por esta resistência. O presente estudo objetivou verificar a produção de MBL em amostras de P. aeruginosa e A. baumannii isoladas de pacientes internados no Hospital Estadual Azevedo Lima, localizado em Niterói-RJ. MATERIAL E MÉTODOS: Um total de 400 amostras (286 P. aeruginosa e 114 A. baumannii) foi identificado por meio de sistemas comerciais e sua susceptibilidade aos antimicrobianos foi testada por disco-difusão. As amostras resistentes a ceftazidima foram submetidas à avaliação da produção de MBL pelo método de disco-aproximação. RESULTADOS: A produção de MBL foi verificada em 49 amostras de P. aeruginosa (17,1 por cento) e em 20 amostras de A. baumannii (17,5 por cento). Grande parte das amostras de P. aeruginosa e de A. baumannii foi resistente a todos os antimicrobianos testados. CONCLUSÃO: A produção de MBL é um dos mecanismos de resistência aos carbapenemas e outros beta-lactâmicos entre amostras de P. aeruginosa e A. baumannii isoladas no hospital-alvo desta investigação. A rápida detecção deste fenótipo de resistência é essencial para implementar medidas rígidas de controle de infecção e permitir o início de terapia empírica adequada.

INTRODUCTION AND OBJECTIVE: P. aeruginosa and A. baumannii are the most prevalent etiological agents of hospital infections among non-fermentative gram-negative bacilli. Carbapenem resistance among these pathogens has become a therapeutic problem worldwide. Metallo-beta-lactamases (MBL) production has emerged as one of the mechanisms responsible for this resistance. The aim of the present study was to assess MBL production in clinical isolates of P. aeruginosa and A. baumannii from inpatients at Hospital Estadual Azevedo Lima, Niterói, RJ. MATERIAL AND METHODS: A total of 400 strains of P. aeruginosa (286) and A. baumannii (114) were identified through commercial systems. Antimicrobial susceptibility was tested by disk diffusion. Isolates resistant to ceftazidime were screened for MBL production by disk approximation test. RESULTS: MBL production was detected in 49 (17.1 percent) P. aeruginosa isolates and in 20 (17.5 percent) A. baumannii isolates. Most P. aeruginosa and A. baumannii samples were resistant to all antimicrobial agents. CONCLUSION: Production of MBL is one of the mechanisms of carbapenem resistance and other beta-lactams among P. aeruginosa and A. baumannii isolates in the investigated hospital. The rapid detection of this resistance phenotype is essential to implement strict infection control procedures and initiate appropriate empirical therapy.

Detecção de metalo-beta-lactamases em amostras de pseudomonas aeruginosa isoladas em Porto Alegre, Rio Grande do Sul / Detection of metallo-beta-lactamase producing pseudomonas aeruginosa isolated in Porto Alegre,Rio Grande do Sul, Brazil

As metalo-beta-lactamases (MBLs) pertencem ao grupo 3 de beta-lactamases de espectro estendido, fazendo parte de uma classefuncional de metaloenzimas classificadas com base na sua capacidade de hidrolisar os carbapenemicos como o Imipenem e em suas caracteristicas de serem inibidas por agentes quelantes como o acido etilenodiamino tetra-acetico (EDTA) e acido 2-mercaptopropionico (2-MPA). Este estudo objetivou a deteccao de MBLs em amostras nosocomiais de P. aeruginosa resistentes a Ceftazidima, provenientes de um hospital de medio porte de Porto Alegre-RS. Foram analisados 113 isolados clinicos atraves de testes fenotipicos, sendo: 80 (70,7%) do trato respiratorio inferior, 24 (21,2%) urina, 3 (2,7%) hemocultura, 3 (2,7%) ponta de cateter, 2 (1,8%) colecao de cavidadese 1 (0,9%) liquor. Com relacao a producao de MBLs, detectou-se 35 amostras (31,0%) como sendo produtoras.

Metallo-beta-lactamases belongs to the group 3 of extended spectrum beta-lactamases, making part of a common functional class of metal enzymes classified based in their ability to hydrolyze Imipenem and in your characteristics of been inhibited by ions of chelating metals, like EDTA and 2-MPA. The aim of this study was detect MBLs in samples of P. aeruginosa resistant to ceftazidime in the patients of PortoAlegre- RS Brazil. The samples was provided of different clinical specimens like: 80 (70,7%) respiratory tract, 24 (21,2%) urine, blood 3 (2,7%), catheter 3 (2,7%), 2 (1,8%) corporeal secretions and 1 (0,9%) liquor. With regard to the MBLs production, we detect 35 (31%) as being producing.

Antibacterial activities of multi drug resistant Myroides odoratimimus bacteria isolated from adult flesh flies (Diptera: Sarcophagidae) are independent of metallo beta-lactamase gene / Atividades antibacterianas de Myroides odoratimimus isolada de moscas varejeiras adultas (Diptera: Sarcophagidae) são independentes do gene metalo beta lactamase

Flesh flies (Diptera: Sarcophagidae) are well known cause of myiasis and their gut bacteria have never been studied for antimicrobial activity against bacteria. Antimicrobial studies of Myroides spp. are restricted to nosocomial strains. A Gram-negative bacterium, Myroides sp., was isolated from the gut of adult flesh flies (Sarcophaga sp.) and submitted to evaluation of nutritional parameters using Biolog GN, 16S rRNA gene sequencing, susceptibility to various antimicrobials by disc diffusion method and detection of metallo â-lactamase genes (TUS/MUS). The antagonistic effects were tested on Gram-negative and Gram-positive bacteria isolated from human clinical specimens, environmental samples and insect mid gut. Bacterial species included were Aeromonas hydrophila, A. culicicola, Morganella morganii subsp. sibonii, Ochrobactrum anthropi, Weissella confusa, Escherichia coli, Ochrobactrum sp., Serratia sp., Kestersia sp., Ignatzschineria sp., Bacillus sp. The Myroides sp. strain was resistant to penicillin-G, erythromycin, streptomycin, amikacin, kanamycin, gentamycin, ampicillin, trimethoprim and tobramycin. These strain showed antibacterial action against all bacterial strains except W. confusa, Ignatzschineria sp., A. hydrophila and M. morganii subsp. sibonii. The multidrug resistance of the strain was similar to the resistance of clinical isolates, inhibiting growth of bacteria from clinical, environmental and insect gut samples. The metallo â-lactamase (TUS/MUS) genes were absent, and resistance due to these genes was ruled out, indicating involvement of other secretion machinery.

Moscas varejeiras (Diptera: Sarcophagidae) são causa conhecida de miíase e as bactérias de seus intestinos nunca foram estudadas quanto à atividade antibacteriana. Estudos antimicrobianos de Myroides spp restringem-se à cepas hospitalares. Uma bactéria Gram negativa, Myroides sp, foi isolada do intestino de moscas varejeiras adultas (Sarcophaga sp) e submetida à avaliação de parâmetros nutricionais pelo sistema BIOLOG GN, ao sequenciamento genético 16S rRNA, à sensibilidade a vários antimicrobianos pelo método de difusão de discos e à detecção dos genes de metalo beta lactamases (TUS/MUS). Os efeitos antagonistas foram testados contra bactérias Gram negativas e Gram positivas isoladas de material clínico humano, amostras ambientais e intestino do inseto. As espécies bacterianas incluíram Aeromonas hydrophila, A. culicicola, Morganella morganii subsp sibonii, Ochrobactrum anthropi, Weissella confusa, Escherichia coli, Ochrobactrum sp, Serratia sp, Kestersia sp, Ignatzschineria sp e Bacillus sp. A cepa Myroides sp foi resistente à penicilina G, eritromicina, estreptomicina, amicacina, canamicina, gentamicina, ampicilina, trimetoprim e tobramicina. Esta cepa apresentou atividade antimicrobiana contra todas as cepas exceto W.confusa, Ignatzschineria sp, A. hydrophila e M. morgani subsp sibonii. A resistência múltipla da cepa foi semelhante à de isolados clínicos, inibindo bactérias das amostras clínicas, ambientais e do intestino do inseto. Os genes de metalo beta lactamases (TUS/MUS) estavam ausentes, excluindo-se a resistência mediada por esses genes, o que indica o envolvimento de um mecanismo alternativo de secreção.