Molecular and phenotypic characterization of diarrheagenic Escherichia coli isolated from groundwater in rural areas in southern Brazil.

Water-borne diseases like diarrheagenic Escherichia coli (DEC)-induced gastroenteritis are major public health problems in developing countries. In this study, the microbiological quality of water from mines and shallow wells was analyzed for human consumption. Genotypic and phenotypic characterization of DEC strains was performed. A total of 210 water samples was analyzed, of which 153 (72.9%) contained total coliforms and 96 (45.7%) E. coli. Of the E. coli isolates, 27 (28.1%) contained DEC genes. The DEC isolates included 48.1% Shiga toxin-producing E. coli (STEC), 29.6% enteroaggregative E. coli (EAEC), 14.9% enteropathogenic E. coli (EPEC), 3.7% enterotoxigenic E. coli (ETEC), and 3.7% enteroinvasive E. coli (EIEC). All the STECs had cytotoxic effects on Vero cells and 14.8% of the DEC isolates were resistant to at least one of the antibiotics tested. All DEC formed biofilms and 92.6% adhered to HEp-2 cells with a prevalence of aggregative adhesion (74%). We identified 25 different serotypes. One EPEC isolate was serotype O44037:H7, reported for the first time in Brazil. Phylogenetically, 63% of the strains belonged to group B1. The analyzed waters were potential reservoirs for DEC and could act as a source for infection of humans. Preventive measures are needed to avoid such contamination.

Antimicrobial activity evaluation and comparison of methods of susceptibility for Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacter spp. isolates.

The production of KPC (Klebsiella pneumoniae carbapenemase) is the major mechanism of resistance to carbapenem agents in enterobacterias. In this context, forty KPC-producing Enterobacter spp. clinical isolates were studied. It was evaluated the activity of antimicrobial agents: polymyxin B, tigecycline, ertapenem, imipenem and meropenem, and was performed a comparison of the methodologies used to determine the susceptibility: broth microdilution, Etest® (bioMérieux), Vitek 2® automated system (bioMérieux) and disc diffusion. It was calculated the minimum inhibitory concentration (MIC) for each antimicrobial and polymyxin B showed the lowest concentrations for broth microdilution. Errors also were calculated among the techniques, tigecycline and ertapenem were the antibiotics with the largest and the lower number of discrepancies, respectively. Moreover, Vitek 2® automated system was the method most similar compared to the broth microdilution. Therefore, is important to evaluate the performance of new methods in comparison to the reference method, broth microdilution.

Caracterização de Escherichia coli diarreiogênica isolada de água subterrânea para consumo humano em um assentamento rural / Characterization of diarrheagenic Escherichia coli isolated from underground water for human consumption in a rural settlement

Escherichia coli diarreiogênica (DEC) é um importante agente de infecções gastrointestinais transmitidas pela água. O trabalho teve como objetivo avaliar a qualidade microbiológica e físicoquímica de 58 amostras de água subterrânea in natura para consumo de um assentamento rural e caracterizar os isolados de E. coli, genotipicamente dentro dos patotipos de DEC, pela técnica da PCR, e fenotipicamente. Todas as amostras apresentaram contaminação por coliformes totais e 36 (62,1%) por E. coli. Em células HEp-2, dos 170 isolados de E. coli, 106 (62,36%) apresentaram AA, 15 (8,82%) AD, 17 (10%) adesão não caracterizado e 32 (18,82%) foram não aderentes. Quanto à formação de biofilme, 126 (74,12%) cepas são formadoras, e 44 (25,88%) não formaram biofilme. Foram identificadas DEC em 6,89% das amostras de água: três (5,17%) ETEC e uma (1,72%) EAEC. EAEC apresentou AA e as três cepas de ETEC apresentaram AD, AA e não definida. Todas as DEC foram formadoras de biofilme. Dois isolados de ETEC apresentaram resistência à ampicilina e tetraciclina, uma ETEC à aminoglicosídeos, e EAEC foi sensível a todos os antibióticos testados. As ETEC foram classificadas no filogrupo B1 e a EAEC no E. Os sorotipos encontrados foram: O24:H21, OR:H21, O17:H46 e O6:H12. Todas as amostras estavam dentro dos parâmetros normais de flúor e 13 (22,4%) apresentaram resultados acima do padrão permitido de turbidez. A presença de E. coli e DEC nas amostras de água indica a necessidade de adoção de medidas que evitem a contaminação da água fornecida à população, evitando, assim, a transmissão de doenças.(AU)

Diarrheogenic Escherichia coli (DEC) is an important agent of waterborne gastrointestinal infections. The objective of this work was to evaluate the microbiological and physico-chemical quality of 58 freshwater groundwater samples for the consumption of a rural settlement and to characterize the E. coli isolates, genotyped within the DEC pathophyses, by the PCR technique, and phenotypically. All water samples presented contamination with total coliforms, and 36 (62.1%) with E. coli. In HEp-2 cells, of 170 E. coli isolates, 106 (62.36%) showed AA, 15 (8.82%) AD, 17 (10%) noncharacterized adhesion and 32 (18.82%) were non-adherent. As for biofilm formation, 126 (74.12%) strains are forming, and 44 (25.88%) did not form biofilm. DEC was identified in 6.89% of the water samples with E. coli: three (5.17%) with enterotoxigenic E. coli (ETEC), and one (1.72%) with enteroaggregative E. coli (EAEC). EAEC showed aggregative adherence, whereas various ETEC strains presented diffuse, aggregative, and non-defined adherence. All DEC strains were biofilm forming. Two isolates of ETEC showed resistance to ampicillin and tetracycline, and one isolate showed resistance to aminoglycosides. The EAEC isolate was sensitive to all antibiotics tested. All ETECs were classified into phylogenetic B1 and EAEC in E. The serotypes identified in our study were: O24:H21, ONT:H21, O17:H46, and O6:H12. All water samples were within normal fluorine parameters; however, measured turbidity was above the permitted standard in 13 (22.4%) samples. The presence of E. coli and DEC in water samples indicates the need to take action to prevent contamination of water resources accessed by people to prevent the transmission of diseases.(AU)

Antimicrobial activity evaluation and comparison of methods of susceptibility for Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacter spp. isolates

Abstract The production of KPC (Klebsiella pneumoniae carbapenemase) is the major mechanism of resistance to carbapenem agents in enterobacterias. In this context, forty KPC-producing Enterobacter spp. clinical isolates were studied. It was evaluated the activity of antimicrobial agents: polymyxin B, tigecycline, ertapenem, imipenem and meropenem, and was performed a comparison of the methodologies used to determine the susceptibility: broth microdilution, Etest® (bioMérieux), Vitek 2® automated system (bioMérieux) and disc diffusion. It was calculated the minimum inhibitory concentration (MIC) for each antimicrobial and polymyxin B showed the lowest concentrations for broth microdilution. Errors also were calculated among the techniques, tigecycline and ertapenem were the antibiotics with the largest and the lower number of discrepancies, respectively. Moreover, Vitek 2® automated system was the method most similar compared to the broth microdilution. Therefore, is important to evaluate the performance of new methods in comparison to the reference method, broth microdilution.

Evolução da resistência de Klebsiella pneumoniae no Hospital Universitário de Londrina no período de 2000 a 2011 / Evolution of resistance of Klebsiella pneumoniae in Londrina University Hospital from 2000 to 2011

O presente estudo teve como objetivo avaliar a evolução da resistência a antimicrobianos em isolados clínicos de Klebsiella pneumoniae, no período de 2000 a 2011, no Hospital Universitário da Universidade Estadual de Londrina (HU-UEL). Foi realizada uma análise retrospectiva de 2.318 testes de sensibilidade aos antimicrobianos de K. pneumoniae, a partir de um banco de dados do setor de Microbiologia do Laboratório Clínico do HU. No período de 2000 a 2009, o principal mecanismo de resistência aos antimicrobianos β-lactâmicos observado foi a produção de β-lactamases do tipo ESBL (β-lactamase de espectro ampliado),que pode ser verificado pelo aumento da resistência de K. pneumoniae às cefalosporinas de 3a. e 4a. geração. No entanto, a partir de 2009 apareceram as primeiras cepas de K. pneumoniae produtoras de carbapenemase, comprometendo a eficácia dos carbapenêmicos. Os índices de resistência ao ertapenem variaram de 16%,em 2005, para 40%, em 2011. Outra classe de antimicrobianos comprometida foi a das fluoroquinolonas; para ciprofloxacina, os índices de resistência variaram de 13% a 62%, em 2001 e 2011, respectivamente.Os aminoglicosídeos tiveram oscilações de resistência durante o período estudado, chegando, em 2011,a valores de 56% e 30% para gentamicina e amicacina, respectivamente. Enquanto isso, sulfametoxazol/trimetoprim e piperacilina/tazobactam alcançaram índices de resistência de 60%, nesse mesmo período. O aumento de resistência em K. pneumoniae neste hospital evidencia a necessidade de adequação do tratamento de infecções por este agente e de adoção o de medidas apropriadas que visem ao controle de infecções, bem como ao uso adequado dessas drogas.

The present study aimed to evaluate the evolution of antibiotic resistance in clinical isolates of Klebsiella pneumoniae in the period of 2000 to 2011, at the University Hospital of Londrina (HU-UEL). A retrospective analysis of 2,318 antimicrobial susceptibility tests of Klebsiella pneumoniae was performed from a database of the Clinical Laboratory of Microbiology of the University Hospital. In the period of 2000 to 2009, the main mechanism of resistance observed to β-lactam antimicrobials was due to the production of ESBL β-lactamase type (β-lactamase wide spectrum), which can be verified by the increased resistance of Klebsiella pneumoniae to 3rd generation cephalosporins and cefepime. However, the first strains of Klebsiella pneumoniae carbapenemase-producing appeared in 2009, compromising the efficacy of carbapenems. The rates of resistance to ertapenem ranged from 16%, in 2005, to 40% in 2011. Another class of committed antibiotics was the fluoroquinolones; for ciprofloxacin, resistance rates ranged from 13% to 62%, in 2001 and 2011, respectively. Aminoglycosides exhibited oscillations of resistance during the period studied, reaching,in 2011, values of 56% and 30% for gentamicin and amikacin, respectively. Meanwhile, trimethoprim/sulfamethoxazole and piperacillin/tazobactam reached about 60%, in the same period. Therefore, knowing the antimicrobial resistance of Klebsiella pneumoniae strains is essential for proper treatment of patients and adoption of appropriate measures that aims infection control, and proper use of these drugs.