Genes blaTEM, blaSHV y blaCTX-M en enterobacterias productoras de b-lactamasas de espectro extendido aisladas de pacientes con infección intrahospitalaria / blaTEM, blaSHV y blaCTX-M genes in extended-spectrum b-lactamases producing enterobacterias isolated from patients with hospital-acquired infections

El objetivo de este estudio fue identificar los genes blaTEM, blaSHV y blaCTX-M en aislados clínicos de enterobacterias productoras de b-lactamasas de espectro extendido (BLEE), recolectadas entre septiembre y noviembre de 2005. Además de la resistencia a las cefalosporinas de tercera generación, los aislados también mostraron resistencia a cloranfenicol (59,2%) amikacina (37,0%) y gentamicina (40,7%) y se mostraron sensibles a imipenem y meropenem. Nueve cepas lograron transferir la resistencia a las cefalosporinas de tercera generación, así como la producción de BLEE. En los aislados clínicos se detectaron los genes blaSHV, blaTEM y blaCTX-M, donde los tipos blaTEM-1, blaSHV-1, blaSHV-5 blaSHV-5-2a y blaCTX-M-1 fueron los prevalentes; mientras que en las transconjugantes sólo se detectaron blaTEM-1, blaSHV-5 y blaSHV-5-2a. Se identificaron en total siete tipos de genes, de los cuales cinco eran codificantes de enzimas tipo BLEE, lo que demuestra que en el centro hospitalario la resistencia a las cefalosporinas de tercera generación es debida a diversas enzimas.

The objective of the present investigation was to identify the blaTEM, blaSHV and blaCTX-M genes on extended-spectrum b-lactamases (ESBL) producing Enterobacteriaceae from clinical isolates, collected between September and November 2005. In addition to third-generation cephalosporin resistance, the isolates also showed resistance to chloramphenicol (59.2%), amikacin (37.0%) and gentamicin (40.7%), and demonstrated sensitivity to imipenem and meropenem. Nine strains were capable of transferring third-generation cephalosporin resistance, as well as the production of ESBL. In the clinical isolates, the genes blaSHV, blaTEM and blaCTX-M were detected, being more prevalent the types blaTEM-1, blaSHV-1, blaSHV-5 blaSHV-5-2a and blaCTX-M-1; while in the trans-conjugated only blaTEM-1, blaSHV-5 y blaSHV-5-2a were found. In total, seven types of genes were identified, five of which were codifying genes for ESBL-type enzymes. This demonstrates that in the hospital center, resistance to third-generation cephalosporin is mediated by several enzymes.

First characterisation of plasmid-mediated quinolone resistance-qnrS1 co-expressed bla CTX-M-15 and bla DHA-1 genes in clinical strain of Morganella morganii recovered from a Tunisian Intensive Care Unit.

Purpose: Aim of this study was to show the emergence of the qnr genes among fluoroquinolone-resistant, AMPC and ESBL (extended-spectrum-beta-lactamase) co-producing Morganella morganii isolate. Materials and Methods: A multi resistant Morganella morganii SM12012 isolate was recovered from pus from a patient hospitalized in the intensive care unit at the Military hospital, Tunisia. Antibiotic susceptibility was tested with the agar disk diffusion method according to Clinical and Laboratory Standards Institute guidelines. ESBLs were detected using a standard double-disk synergy test. The characterization of beta-lactamases and associated resistance genes were performed by isoelectric focusing, polymerase chain reaction and nucleotide sequencing. Results: The antimicrobial susceptibility testing showed the high resistance to penicillins, cephalosporins (MICs: 64-512 μg/ml) and fluoroquinolones (MICs: 32-512 μg/ml). But M. morganii SM12012 isolate remained susceptible to carbapenems (MICs: 4-<0.25 μg/ml). The double-disk synergy test confirmed the phenotype of extended-spectrum β-lactamases (ESBLs). Three identical β-lactamases with pI values of 6.5, 7.8 and superior to 8.6 were detected after isoelectric focusing analysis. These β-lactamases genes can be successfully transferred by the conjugative plasmid. Molecular analysis demonstrated the co-production of bla DHA-1, bla CTX-M-15 and qnrS1 genes on the same plasmid. The detection of an associated chromosomal quinolone resistance revealed the presence of a parC mutation at codon 80 (Ser80-lle80). Conclusion: This is the first report in Tunisia of nosocomial infection due to the production of CTX-M-15 and DHA-1 β-lactamases in M. morganii isolate with the association of quinolone plasmid resistance. The incidence of these strains invites continuous monitoring of such multidrug-resistant strains and the further study of their epidemiologic evolution.

Investigation of class I, II and III integrons among acinetobacter strains isolated from ventilator-associated pneumonia patients in intensive care unit of Rasoul Akram Hospital in Tehran, Iran

Multi-drug resistant strains of Acinetobacter spp. have created therapeutic problems worldwide. The objective of this study was to detect integrons in Acinetobacter spp. isolates from Ventilator-Associated Pneumonia patients using PCR method. A total 51 Bronchoalveolar lavage samples were obtained from patients in ICU and examined for Acinetobacter spp. infection by biochemical and PCR methods using blaOXA51-like primers. Antimicrobial susceptibility testing was performed using disk diffusion and MIC methods. Among 51 patients with VAP [62.7% males, 35.2% females, mean age 53 year], 50 [98%] were positive, with a high prevalence of gram-negative bacteria, mainly Acinetobacter spp. [70%], from which A. baumani was detected in 34 [68%] and A. lwoffii in 1 [2%] of isolates. More than 90% of isolates were resistant to imipenem, piperacillin+tazobactam, third generation cephalosporins and gentamicin, while the most effective antibiotic was colistin [100%]. The correlation coefficient between disk diffusion and MIC was 0.808 [p = 0.001]. Three Acinetobacter isolates [8%] harbored integrase /gene but none of isolates contained Class II or III integrons. The results showed that colistin was an effective antibiotic and can be used for treatment of patients in ICU. Due to the high number of MDR isolates lacking Integrons it can be concluded that although class I integrons are important among clinical isolates of A. baumannii, they have no significant role in dissemination of antibiotic resistance genes in Rasoul Akram Hospital in Tehran, Iran. The presence of IntI in A. Iwoffii may be related to transfer of integron to A. baumannii which can be considered as an important threat for hospitalized patients

Presencia de metalo-ß-lactamasas en Pseudomonas aeruginosa resistente a imipenem / Presence of metal lo ß-lactamases in imipenem-resistant Pseudomonas aeruginosa

Background: Metallo-ß-lactamases (MBL) confer high resistance to carbapenems in Pseudomonas aeruginosa (Psae). They are encoded in mobile elements of different genes (VIM, IMP, SMP, GIM), along with other resistance genes. Aim: To detect the presence of MBL in imipenem resistant Psae strains. Material and methods: Fifty-nine imipenem resistant Psae strains isolated from January 2004 to August 2005 in a University Clinical Hospital, were included. The presence of MBL was studied by Etest (phenotypic) and genotypic polymerase chain reaction (PCR) methods. To rule out a nosocomial outbreak, MBL positive strains, were studied by pulse field gel electrophoresis. Results: The presente of MBL was detected in eleven strains. AH were type VIM and were not clonally related. There was no concordance between phenotypic and genotypic MBL detecting methods. AH the strains were also multiresistant. Conclusions: The presence of MBL was detected in 19 percent of imipenem resistant Psae strains.

Antimicrobial Susceptibility Patterns and Macrolide Resistance Genes of beta-Hemolytic Viridans Group Streptococci in a Tertiary Korean Hospital

The aim of this study was to investigate antimicrobial susceptibilities and macrolide resistance mechanisms of beta-hemolytic viridans group streptococci (VGS) in a tertiary Korean hospital. Minimum inhibitory concentrations (MICs) of seven antimicrobials were determined for 103 beta-hemolytic VGS isolated from various specimens. The macrolide resistance mechanisms of erythromycin-resistant isolates were studied by the double disk test and polymerase chain reaction (PCR). The overall resistance rates of beta-hemolytic VGS were found to be 47.5% to tetracycline, 3.9% to chloramphenicol, 9.7% to erythromycin, and 6.8% to clindamycin, whereas all isolates were susceptible to penicillin G, ceftriaxone, and vancomycin. Among ten erythromycin-resistant isolates, six isolates expressed a constitutive MLSB (cMLSB) phenotype, and each of the two isolates expressed the M phenotype, and the inducible MLSB (iMLSB) phenotype. The resistance rates to erythromycin and clindamycin of beta-hemolytic VGS seemed to be lower than those of non-beta-hemolytic VGS in our hospital, although cMLSB phenotype carrying erm(B) was dominant in beta-hemolytic VGS.

Caracterização genotípica de cepas da família enterobacteriaceae produtoras de B-lactamases de espectro estendido, isoladas de pacientes de um hospital da rede pública da cidade de São Paulo / Genotypic characterization of extended-spectrum b-lactamase-producing enterobacteriaceae strains, isolated from patients of a public hospital in the city of Sao Paulo

Introdução - A crescente resistência antimicrobiana em bactérias responsáveis por infecções hospitalares é um grande desafio à Saúde Pública. as B-lactamases de espectro estendido (ESBL), que hidrolisam a maioria dos compostos B-lactâmicos, são reconhecidas mundialmente como um grande problema para pacientes hospitalizados, devido à localização de seus genes em elementos transferíveis, facilitando sua disseminação. Objetivo - Caracterizar geneticamente cepas de Enterobactérias produtoras de ESBL isoladas de pacientes de um hospital público da cidade de São Paulo. Material e métodos - Todas as cepas de enterobactérias produtoras de ESBL isoladas em um ano foram submetidas a análises moleculares pela PCR, com iniciadores específicos para oito genes bla, e as cepas de Klebsiella pneumoniae ESBL positivas (ESBL-Kp) identificadas nesse período foram comparadas pela técnica de PFGE.Resultados - Os genes, bla(tem), bla(shv), bla(ctx-m), bla(per-2) bla(veb) and bla(ges) foram identificados em 9 espécies: Klebsiella pneumoniae (71,5 por cento), Escherichia coli (13,5 por cento), Morganella morganii (6 por cento), Proteus mirabilis (3 por cento), Klebsiella oxytoca (1,5 por cento), Providencia rettgeri (1,5 por cento), Providencia stuartii (1,5 por cento), Enterobacter aerogenes (0,75 por cento). Os genes bla(per-1) e bla(oxa) não foram detectados. O PFGE revelou 8 perfis moleculares principais em 68,4 por cento das ESBL-Kp, e 31,6 por cento das cepas não estavam relacionadas. Conclusões - Os resultados de PCR revelaram uma grande variedade de grupos de ESBL, e aparentemente este é o primeiro relato de grupos GES e VEB em enterobactérias no Brasil

Caracterización molecular de aislamientos de enterobacter cloacae multirresistentes productores de a-lactamasas provenientes de pacientes de un hospital de tercer nivel de Bogotá / Molecular characterization of 20 isolates of resistance enterobacter cloacae from a Hospital of third level in Bogotá

Antecedentes. Las enterobacterias, antaño flora normal del tracto gastrointestinal, han cambiado su biología y emergido como agentes patógenos nosocomiales que se tornan resistentes a los antibióticos conocidos. Objetivo. Realizar la caracterización epidemiológico-molecular de 20 aislamientos de Enterobacter cloacae resistentes a cefalosporinas de tercera generación; provenientes de un hospital de tercer nivel de Bogotá-Colombia. Material y métodos. Los aislamientos fueron identificados mediante sistemas automatizados Microscan y VITEK, se utilizó el Enterobacter asbureae como control externo inter-especie. La confirmación de resistencia se hizo por técnica de difusión en agar, y una vez establecida se realizó BLEE para comprobación. La determinación de puntos isoeléctricos se hizo, mediante lisis por ultrasonido y la genotipificación mediante la metodología para bacterias Gramnegativas propuesta por Versalovic. Resultados: Los aislamientos colectados durante un año fueron causantes de 15 de infección Intrahospitalaria y dos colonizaciones. Todos los aislamientos presentaron resistencia a cefotaxima, ceftazidima, ceftriaxona, aztreonam y ciprofloxacina, 95 por ciento a amikacina, gentamicina y cloranfenicol, 75 por ciento a trimetoprim/sulfametoxazol, 20 por ciento a cefepime y todos fueron sensibles a imipenem. Dos aislamientos fueron confirmados como productores de â-lactamasas de espectro extendido (BLEE) por la técnica microbiológica de disco combinado. Por isoelectroenfoque presentaron dos â-lactamasas con puntos isoeléctricos (pI) de 5,4 y 8,2. En los 18 aislamientos no inhibidos por ácido clavulánico, se detectaron entre 2 y 4 â-lactamasas con pI de 5,4; 6,0; 7,0; 8,2 y mayor que 8,2; la resistencia a cefalosporinas de tercera generación podría ser atribuida a la hiperproducción de AmpC; los valores de pI sugieren la producción simultánea de â-lactamasas tipo SHV y TEM. La genotipificación mediante tres metodologías de rep-PCR (ERIC; REP y BOX) agrupó la población estudiada en siete clones: seis constituidos por un solo aislamiento y el clon predominante E1/B1/R1 agrupó 14 aislamientos causantes de infección en diez pacientes. Conclusión. Se identificó un clon de Enterobacter cloacae multirresistente, endémico en una institución de tercer nivel en Bogotá, causante de infección nosocomial y quirúrgica en particular

Evaluation of phenotypic and genotypic markers for clinical strains of acinetobacter baumannii

Acinetobacter baumannii es un importante patógeno oportunista. Este microorganismo adquiere con facilidad resistencia a antimicrobianos, involucrándose en infecciones nosocomiales generalmente graves. Estas características promueven el análisis epidemiológico de las infecciones provocadas por el mismo. Sin embargo, no hay aún un esquema de tipificación generalmente aceptado para este patógeno. Hemos evaluado en este trabajo diferentes procedimientos fenotípicos y genotípicos para la caracterización de aislamientos clínicos de A. baumannil aislados en un Hospital Público de Rosario (Hospital de Emergencias Clemente Alvarez, HECA), durante un período de cuatro años. Estos incluyeron PCR con oligonucleótidos degenerados (OD-PCR), PCR empleando cebadores homólogos a secuenciais palindrómicas extragénicas repetitivas (REP-PCR), electroforesis en geles de agarosa con campo pulsado (PFGE) y ensayo de susceptibilidad a antimicrobianos. OD-OCR y PFGE, entre los métodos individuales, fueron los métodos de mayor poder discriminatorio (índice discriminatorio, D, de 0.98 y 0.96; respectivamente). Por otra parte, el antibiotipo y REP-PCR presentaron menor discriminacíon (D: 0.86 y 0.77; respectivamente). La combinación de antibiotipo con cada uno de los procedimientos genotípicos mencionados originó un aumento importante en los índices discriminatorios de cada método. En particular, la combinación de OD-PCR y antibiotipo constituvó la mejor metodología para el estudio epidemiológico de A.baumannii. Así, la combinación de los procedimientos feno- y genotípicos mencionados permitó inferir las relaciones genéticas y la diseminación de clones de A.baumannii multirresistentes en el HECA en el período 1994-99. Una cepa particular, sensible a imipenem, estuvo ampliamente diseminada en el hospital durante 1994-1996. Por otra parte, un clon diferente, con resistencia adicional a carbapenemes, se diseminó rapidamente en el hospital en 1997, en coincidencia con la introducción de imipenem como terapia antibiótica.

Candidaemia in patients of a tertiary health care hospital from north India.

BACKGROUND & OBJECTIVES: Candidaemia is an important cause of mortality in hospital settings. Limited information is available from India on nosocomial candidaemia. The objective of the present study was to isolate and identify yeasts from patients suspected to have nosocomial bloodstream infection (BSI) and to determine the carriage rate of Candida species, risk factors for acquisition of infection and mortality in this group of patients. METHODS: Blood samples from 4871 patients suspected to have BSI at least 48 h after admission were cultured following standard protocol to isolate and identify the pathogens. Clinical details, possible risk factors and outcome of all candidaemic patients were recorded and analysed. Samples of hand washings and throat gargles from these patients were also cultured to determine the carriage rate. Candida albicans isolated from patients and their carriage sites were genotyped by randomly amplified polymorphic DNA (RAPD) analysis to study strain relatedness. RESULTS: Twenty one patients with candidaemia were detected with mortality of 55 per cent. Candidaemia per 1000 admissions was 1.61. Isolation of non-C. albicans Candida species was significantly higher than C. albicans (14/21 vs 7/21: P < 0.05). Use of broad-spectrum antibiotics (43%), gastrointestinal surgery (23%), immunosuppressive therapy (23%), protein calorie malnutrition with parenteral hyperalimentation (23%) and neutropaenia (14%) were identified as probable risk factors. The seven C. albicans strains isolated from patients with BSI were typed into 6 genotypes. Yeast carriage rate among the patients was 71.4 per cent. C. albicans isolated from the hand, throat and blood of two patients had identical genotype. INTERPRETATION & CONCLUSION: BSI due to non-C. albicans Candida species is more common than C. albicans in our patients and candidaemia is associated with high mortality. RAPD appears to be a simple method to study strain relatedness for C. albicans. There is a need for early diagnosis and systematic surveillance to meet the challenges of nosocomial candidaemia.