Die Mehrzahl der jährlich 400.000 bis 600.000 Krankenhausinfektionen in Deutschland wird von Erregern der sog. ESCAPE-Gruppe (Enterococcus faecium, Staphylococcus aureus, Clostridium difficile, Acinetobacter baumannii, Pseudomonas aeruginosa und verschiedene Enterobacteriaceae, u.a. Klebsiella pneumoniae) verursacht. Besondere Sorge bereitet dabei die Ausbreitung von K. pneumoniae-Stämmen mit enzymvermittelter Resistenz gegenüber Carbapenem-Antibiotika (K. pneumoniae-Carbapenemase, KPC) und die Zunahme von C. difficile-Infektionen (CDI) durch hypervirulente Epidemiestämme (z.B. Ribotyp 027). Die spezifischen Erfahrungen eines prolongierten Ausbruchsgeschehens durch einen KPC-bildenden K. pneumoniae-Stamm (KPC-KP) am Leipziger Universitätsklinikum machen deutlich, dass bei diesem Erregertyp ein hohes Transmissionspotential bei enormer Tenazität (Umweltresistenz) zu berücksichtigen ist, ein Versagen von Standardhygienemaßnahmen in Betracht zu ziehen ist, und Infektionsketten oftmals unklar bleiben. Die Anwendung von Antibiotika ist bei KPC-KP-Infektionen auf einzelne Substanzen (Colistin, Tigecyclin, Gentamicin) beschränkt und vor allem bei immunsupprimierten Patienten (z.B. Lebertransplantierte) mit einem relevanten Risiko des Therapieversagens behaftet. Die Therapie von CDI wird gerade bei Immunsupprimierten durch eine steigende Zahl an Rezidiven erschwert, die teilweise antibiotisch (Vancomycin, Fidaxomicin) nicht beherrschbar sind, so dass alternative Therapieverfahren wie die fäkale Bakterientherapie („Stuhltransplantation“) zur Anwendung kommen. CDI-Rezidive, aber auch eine dauerhafte intestinale Besiedelung mit multiresistenten Enterobakterien wie KPC-KP, scheinen neben wirtsspezifischen Faktoren der Immunantwort durch eine Dysregulation der physiologischen intestinalen Standortflora mit Störung der Kolonisationsresistenz bedingt zu sein. Der Versuch einer Eradikationsbehandlung von Patienten mit persistierender intestinaler Besiedelung durch KPC-KP mittels oraler Applikation der nicht resorbierbaren Antibiotika Colistin und Gentamicin ist mit einem relevanten Risiko der Entstehung von Sekundärresistenzen behaftet. Die Zulassung neuer, besser wirksamer Antibiotika ist für die nächsten Jahre nicht in Sicht, so dass der Infektionsprävention überragende Bedeutung zukommt. Die Erfahrungen der KPC-Ausbruchsbewältigung am Leipziger Universitätsklinikum zeigen, dass nahezu lückenlose Compliance bei der Händedesinfektion, rigoros praktizierte und kontrollierte Barriere- und Isolationsmaßnahmen, Optimierung des Gebrauchs von Breitspektrum-Antibiotika (sog. „Antibiotic Stewardship“) und systematisches mikrobiologisches Erregerscreening dabei unabdingbar sind. Nachhaltige Verbesserungen hinsichtlich der globalen Ausbreitung von multiresistenten Krankenhausbakterien werden sich nur durch grundlegende Umgestaltungen in Umwelt, Landwirtschaft, Tierzucht und Gesundheitswesen mit sparsamer und möglichst gezielter Anwendung von Antibiotika erzielen lassen. Um Risikopopulationen hospitalisierter Patienten vor potentiell lebensbedrohlichen Erregertransmissionen effektiv schützen zu können, sind erweiterte Surveillance und konsequent umgesetzte krankenhaushygienische Maßnahmen erforderlich.

Ten bacteria capable of growth on the metal-cyano complex, tetracyanonickelate (II) {K2 [Ni(CN)J } (TCN), supplied as the sole nitrogen source, were isolated. Seven isolates were identified as pseudomonads while the remaining three were classified as Klebsiella species. In addition to TCN, all isolates were able to utilize KCN although it was significantly more toxic. The degradation of TCN was most complete when supplied at growth-limiting concentrations, did not occur when ammonia was present, and resulted in the formation of nickel cyanide [Ni(CN)2] as a degradation product.

Antimicrobial-resistant (AMR) bacteria are a major source of concern in modern-day nosocomial settings, leading to possible further drug resistance or spread to those who cannot fight off the infection. Previous work from our laboratory has shown that Klebsiella pneumoniae (KP) secretes an antimicrobial protein that has been shown to inhibit the growth of many species of bacteria that contain AMR properties in the Enterobacteriaceae family, a major contributor to nosocomial AMR. Klebsiella spent media is able to inhibit the growth of Citrobacter freundii (CF), Enterobacter aerogenes (EA), and Enterobacter cloacae (ECL) through an anti-microbial protein (AMP). This AMP has been shown to reduce the density and growth of CF, EA, and ECL in both biofilm and planktonic forms. To determine the genetic elements involved in AMP production, we introduced a transposon (Tn5) into the genome of Klebsiella to provide resistant selection and to create a mutant knockout to find the exact location of the gene. Upon transposon mutagenesis, the resulting genome was electroporated into Rec- E. coli. The E. coli was now able to produce the antimicrobial protein, with the zones of inhibition for CF, EA, and ECL. Upon confirmation that the plasmid mediates the AMP, the plasmid was sent for sequencing to further characterize the gene responsible for coding the AMP. This newly identified AMP may prove to be a valuable treatment for AMR bacteria once characterized.

Due to the increasing prevalence of multi-drug resistant (MDR) bacteria, it is now important to begin the search for novel means of defending against such resistant infections. Enterobacteriaceae is a clinically relevant family of bacteria that has shown extensive resistance to many antibiotics, especially after biofilm formation. Inhibitory poly-microbial interactions within this family have been observed. It is known that Citrobacter freundii (CF) growth is significantly inhibited by Klebsiella pneumoniae (KP) through a secreted protein. In this study, the potential KP bacteriocin was screened for its inhibitory effects on CF at various phases of biofilm development. The suspected KP bacteriocin was also tested for its ability to decrease the dosage of antibiotics necessary to inhibit CF growth. Using spectrophotometric analysis, it was shown that the combined treatment of streptomycin and the KP protein allowed a decrease in the minimum inhibitory concentration of streptomycin needed from 50 μM to 32 μM. The combined treatment also yielded increased inhibition at the initial attachment phase of CF infection, as well as after biofilm development. The study uses the secreted KP protein to show the use of poly-microbial interactions within clinical applications. Future projects concerning this KP molecule can pursue the use of a C. elegans model to determine its efficacy in vitro.

The purpose of this study was to examine and evaluate the K-E-S group from this community with respect to distribution in clinical materials, detailed individual biochemical characteristics, and antibiogram patterns using the Bauer-Kirby disc technique, and compare these findings with those of other investigators from other geographic locations. As far as can be determined, this is the only study of its kind on the West Coast of the United States.

Background: Bloodstream infection (BSI) caused by Klebsiella pneumoniae (KP), is a leading cause of hospitalassociated childhood mortality. There are limited data on how poor outcomes of KPBSI can be predicted in poorly resourced areas. This study aimed to assess if the profile of differential counts from full blood counts (FBC) taken at two time points in children <13 years with KPBSI could be used to predict the risk of death. Methods We conducted a retrospective study of a cohort of children admitted to hospital between 2006-2011 with KPBSI. FBC collected within 48 hours (T1) of blood culture and 5-14 days later (T2), were reviewed. Differential counts were classified as abnormal if they were higher or lower than laboratory ranges for normal results. The risk of death was assessed for each category of differential counts. Risk ratios adjusted (aRR) for potential confounders were used to estimate the effect of cell counts on risk of death using multivariable analysis. Data were stratified by HIV status. Results: Of 296 children included, median age 5 (IQR:2-13) months, 82 were HIV -infected. Ninety-five (32%) of the children with KPBSI died. Mortality in HIV-infected and uninfected children was 39/82 (48%) and 56/214 (26%), respectively (p <0.001). Independent associations with mortality were observed with leucopenia, neutropenia and thrombocytopenia. Risk of mortality in children with thrombocytopenia at T1 and T2 was aRR 2.5 (95% CI: 1.34-4.64) and 3.18 (95% CI: 1.31-7.73) respectively in the HIV-uninfected group, whereas the risk for mortality in the HIV-infected group with thrombocytopaenia at T1 and T2 was aRR 1.99 (95% CI: 0.94-4.19) and 2.01 (95% CI: 0.65-5.99) respectively. Neutropenia in the HIV-uninfected group at T1 and T2, showed aRR 2.17 (95% CI: 1.22- 3.88) and 3.70 (95% CI 1.30-10.51) respectively, while in the HIV-infected group, they were aRR 1.18 (95% CI 0.69-2.03) and 2.05 (0.87-4.85) at similar time points. Risk of mortality related to leucopenia at T2 was associated with mortality in HIV-uninfected and HIV-infected patients was aRR 3.22 (95%CI 1.22-8.51) and 2.34 (1.09-5.04) respectively. Persistently high band cell percentage at T2 in HIVinfected children indicated a risk of mortality of aRR 2.91 (95% CI 1.20-7.06). Conclusion Abnormal neutrophil counts and thrombocytopenia are independently associated with significant mortality in children with KPBSI. In resource-limited countries haematological markers have the potential to predict KPBSI mortality.

No description available.

Realizamos um estudo de corte transversal para investigar os fatores de risco para colonização de recém-nascidos por Klebsiella pneumoniae produtora de betalactamase de espectro estendido durante surto em unidade neonatal de risco intermediário. O surto se deveu à colonização crônica de profissional de saúde portadora de onicomicose. Cento e vinte recém-nascidos internados na unidade neonatal durante um período de três meses foram rastreados para colonização por Klebsiella pneumoniae produtora de ESBL através de cultura de swab retal, sendo detectados 27 colonizados. A análise multivariada mostrou que a colonização se associou de forma independente ao uso prévio de antimicrobianos e à ausência de aleitamento materno. Os antimicrobianos mais utilizados foram penicilina e amicacina. Uso prévio de antimicrobianos apresentou odds ratio (OR) igual a 12,3 [intervalo de 95% de confiança (IC): 3,66-41,2, P<0,001]. Aleitamento materno foi associado à redução do risco de colonização (OR: 0,22; IC95%: 0,05-0,99; P=0,049). Nove isolados recuperados no primeiro estágio do surto e 27 isolados de culturas de rastreamento foram posteriormente tipadas por eletroforese em gel de campo pulsado, revelando seis apresentações distintas (A a F). No primeiro estágio do surto ocorreram os clones A, C e E, enquanto entre os 27 isolados das culturas de rastreamento os seis clones foram identificados. O clone A também foi identificado nas mãos de técnica de enfermagem portadora de onicomicose. Pudemos concluir que uso prévio de antimicrobianos predispôs à colonização. O possível efeito do aleitamento materno como fator protetor deve ser mais bem investigado. A detecção de diferentes genótipos de K. pneumoniae sugere que a disseminação de elementos móveis portando o gene ESBL tenha se superposto à simples disseminação de um clone durante o surto / We describe a cross-sectional survey to identify risk factors for colonisation of neonates by extended-spectrum beta-lactamase producing Klebsiella pneumoniae. This occurred following exposure to a colonised healthcare worker during an outbreak in an intermediate-risk neonatal unit. In total, 120 neonates admitted consecutively during a three-month period were screened for ESBL-producing K. pneumoniae by rectal swabbing and 27 were identified as colonised. Multivariate analysis showed colonisation to be independently associated with use of antibiotics and absence of breastfeeding. Previous use of antibiotics presented an odds ratio (OR) of 12,3 [95% confidence interval (CI): 3,66-41,2, P<0,001]. The most commonly used antibiotics were penicillin and amikacin. Breastfeeding was associated with reduced risk for colonisation (OR: 0,22; 95% CI: 0,05-0,99; P=0,049). Nine isolates recovered during the first stage of the outbreak and 27 isolates from surveillance cultures were typed thereafter by pulsed-field gel electrophoresis, revealing six different profiles (A - F). Clones A, C, and E were implicated in the first stage of the outbreak, whereas among the 27 strains recovered from surveillance cultures, all six clones were identified. Clone A was also found on the hand of a nursing auxiliary with onychomycosis. We concluded that prior antimicrobial use predisposed to colonisation. The possible role of breastfeeding as a protective factor needs to be further elucidated. Detection of different genotypes of ESBL-producing K. pneumoniae suggests that dissemination of mobile genetic elements bearing the ESBL gene may have been superimposed on the simple dissemination of a clone during the outbreak

Submitted by Alessandra Portugal (alessandradf@ioc.fiocruz.br) on 2013-09-24T14:50:16Z No. of bitstreams: 1 Thiago Pavoni.pdf: 1955163 bytes, checksum: b15140ec10d9f1473101f075a2a57a86 (MD5) / Made available in DSpace on 2013-09-24T14:50:16Z (GMT). No. of bitstreams: 1 Thiago Pavoni.pdf: 1955163 bytes, checksum: b15140ec10d9f1473101f075a2a57a86 (MD5) Previous issue date: 2011 / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil / Drogas antimicrobianas e bactérias resistentes aos antimicrobianos estão disseminadas em grandes quantidades no ambiente, como resultado do aumento e freqüente uso indiscriminado dos antibióticos. Bactérias e seus genes de resistência têm sido detectados em diferentes ambientes, tais como esgoto hospitalar, esgoto doméstico e águas de rios contaminados. O esgoto hospitalar é um importante poluente, representando riscos para a saúde pública se chegar aos sistemas de distribuição. Ambientes fortemente seletivos, como os hospitais, permitem a geração bactérias resistentes, as quais podem ser lançadas no esgoto hospitalar. O presente trabalho tem como objetivo investigar a presença bactérias resistentes aos antimicrobianos em efluentes de uma estação de tratamento de esgoto hospitalar no Rio de Janeiro, avaliando o potencial do sistema de tratamento para a eliminação de micro-organismos. A estação de tratamento de esgoto fica localizada na região metropolitana. O sistema de lodo ativado por aeração prolongada é constituído por três partes básicas: o tanque de aeração, o decantador e o tanque de cloração. Vinte e quatro amostras de esgoto foram coletadas no período de Julho a Dezembro de 2008. Oito amostras (1000 mL) foram coletadas a partir de diferentes pontos: afluente, efluente do tanque decantador e efluente clorado. Micro-organismos indicadores também foram investigados. Os isolados bacterianos foram identificados a partir de provas bioquímicas convencionais. A sensibilidade aos antimicrobianos das bactérias isoladas foi determinada através do método fenotípico de difusão em ágar, de acordo com as orientações do Clinical and Laboratory Standards Institute (CLSI). A identificação da produção fenotípica de beta-lactamases de espectro estendido e de carbapenemases entre os isolados também seguiram as recomendações do CLSI. Ensaios de PCR foram processados para a identificação dos genes blaKPC, blaTEM, blaSHV e blaCTX-M. A genotipagem das amostras bacterianas foi realizada por eletroforese em gel de campo pulsado. Concentrações significativas de coliformes totais e fecais foram detectadas nos efluentes hospitalares. Um total de 226 isolados foi identificado, entre os quais 213 (94%) pertenciam à família Enterobacteriaceae. Outros grupos de micro-organismos, como Pseudomonas aeruginosa, Acinetobacter baumannii e Aeromonas spp., foram também observados. A maioria das cepas era sensível ao imipenem e ao meropenem; e resistente à cefalotina, à cefotaxima e ao sulfametoxazol-trimetoprim. O fenótipo de ESBL foi caracterizado em 97 (43%) isolados. Os produtores de ESBL mais comuns foram: Klebsiella pneumoniae, Enterobacter cloacae e Escherichia coli. Micro-organismos patogênicos e altas taxas de resistência ainda puderam ser observados nos efluentes clorados. Os genes blaTEM, blaSHV e blaCTX-M foram detectados em 82%, 48% e 67% dos isolados do efluente hospitalar, respectivamente. Em muitos isolados, a ocorrência de mais de um tipo de ESBL foi observada, sendo a associação dos tipos TEM e CTX-M a mais frequente. O gene blaKPC foi detectado em dois isolados do efluente. Foi possível observar isolados clínicos e do esgoto geneticamente relacionados. Concluímos que, apesar do tratamento, o esgoto hospitalar pode ser considerado um veículo ambiental de disseminação de bactérias multirresistentes. A ocorrência destes micro-organismos nos efluentes é preocupante e tem impacto sobre a saúde pública. Medidas urgentes são necessárias para enfrentar este problema. Vale ressaltar que, em muitos países em desenvolvimento, os efluentes hospitalares não recebem tratamento adequado. / Antimicrobial drugs and antimicrobial - resistant bacteria are discharged in large quantities in the environment as a result of increasing ly frequent and indiscriminate use of antibiotics . Antimicrobial - resistant bacteri a and antimicrobial - resistant genes have been detected in different environments, such as domestic sewage, hospital sewage and sewage - contaminated river waters. Hospital sewage is an important pollutant , representing risks to public health if it reaches th e distribution system. The occurrence of strongly selective environments, such as hospitals, leads to an incre ase of multiresistant bacteria, which can be released in hospital sewage. The aim of this study was to investigate the antimicrobial - resistant bac teria isolated from a hospital sewage treatment plant in Rio de Janeiro city, evaluating the treatment plant’s potential to remove these microorganisms. The sewage treatment plant serve a hospital located in the metropolitan area of the Rio de Janeiro city (RJ), Brazil. The extended aeration activated sludge plant is divided into three parts, an aeration tank, a clarifier tank and a chlorine contact tank. During the study, twenty - four sewage samples were collected in the period from July to December 2008. E ight samples (1000 ml) were collected on each day from the following: influent; clarifier tank effluent; and chlorine contact tank effluent. Total and faecal coliforms concentrations were also determined . Isolates were identified using established biochemi cal procedures. The antimicrobial susceptibilities of bacterial isolates were determined using the agar diffusion method according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Isolates were screened for the KPC - and ESBL - producing phen otype according to the CLSI. PCR experiments were used for the molecular detection of bla KPC , bla TEM , bla SHV and bla CTX - M genes. The genetic relat ionships of isolates were determined by PFGE. High concentrations of total and faecal coliforms were detected in the influent , clarifier tank and chlorine contact tank effluent. A total of 226 isolates were identified, among which 213 (94%) were Enterobacteriaceae . In addition , Pseudomonas aeruginosa , Acinetobacter baumannii and Aeromonas spp . in hospital effluent were observed . The majority of the strai ns were susceptible to imipenem and meropenem and resistant to cefalothin, cefotaxime and trimethoprim - sulphametoxazole. ESBL phenotype was characterized in 97 (43%) isolates. The most common ESBL - producing isolates were: Klebsiella pneumoniae , Enterobacter cloacae , and Escherichia coli . Pathogenic microorganisms and higher antimicrobial resistance rates were detected in chlorine contact tank effluent. The bla TEM , bla SHV and bla CTX - M genes were detected in 82%, 48% a nd 67% of isolates respectively. Many of the isolates harboured other β - lactam resistance enzymes and the a ssociation of types TEM and CTX - M was more frequent. The bla KPC was detected in isolates from effluents. PFGE analysis revealed clonal types among cl inical isolates and isolates from effluents. Despite the treatment of the wastewater, hospital effluent may be considered as a potential environmental vehicle of multiresi s tant microorganisms. The occurrence of multiresistant bacteria isolates in hospital effluents is worrisome and has a real impact on public health. Urgent measures are necessary in order to counteract this problem. It should be noted that effluents from hospitals in developing countries do not receive adequate treatment

As enterobactérias são importantes pátogenos comunitários e hospitalares e o aparecimento cada vez mais frequente de cepas multirresistentes tem sido motivo de preocupação em hospitais e instituições de saúde por todo o mundo, devido às opções terapêuticas restritas. Nas últimas décadas, o aumento global de cepas produtoras de β-lactamases plasmidiais induzíveis e β-lactamases de espectro estendido (ESBL), fizeram com que os carbapenêmicos fossem considerados a primeira opção para o tratamento de infecções graves. No entanto, a resistência aos carbapenêmicos já é considerada um problema de saúde pública em diversos países e a produção da enzima Klebsiella pneumoniae Carbapenemase (KPC) tem sido descrita como o principal mecanismo de resistência a esta classe de antibióticos na família Enterobacteriaceae. Considerando que apenas relatos esporádicos de cepas produtoras de KPC têm sido reportados no Rio Grande do Sul (RS), em contraste à situação de muitos estados brasileiros, este estudo teve por objetivo estabelecer a prevalência de KPC entre isolados com reduzida sensibilidade aos carbapenêmicos provenientes de 11 hospitais do RS, promover a caracterização molecular destes isolados, bem como avaliar as principais metodologias fenotípicas utilizadas na sua detecção. Diferenças significativas foram observadas entre os perfis de sensibilidade a imipenem (IPM) e meropenem (MEM), quando comparados ao de ertapenem (ERT), sendo que para este último menos de 10% dos isolados foram considerados sensíveis em comparação a mais de 73% para os dois primeiros. Nossos resultados também demonstraram que a redução de sensibilidade aos carbapenêmicos esteve associada à produção de β-lactamases do tipo AmpC e ESBL, em detrimento de carbapenemases. Dentre as principais carbapenemases encontradas em enterobactérias, apenas cepas produtoras de KPC foram detectadas entre os isolados estudados, cuja prevalência foi de 4%. Entre os produtores de KPC, foram identificadas espécies de E. cloacae, K. pneumoniae, S. marcescens e K. georgiana, provenientes de quatro hospitais distintos. A análise por sequenciamento revelou que todos foram produtores da enzima KPC-2. Quanto ao perfil de sensibilidade, a maioria foi altamente resistente aos β-lactâmicos e às quinolonas, enquanto que, amicacina e polimixima foram os antibióticos mais efetivos contra os isolados in vitro. Dois grupos clonais foram evidenciados entre os isolados de E. cloacae e S. marcescens e quatro entre os isolados de K. pneumoniae, na análise por PFGE. Com relação ao contexto genético que envolve o blaKPC-2, apenas uma caracterização parcial foi possível, evidenciando uma plataforma alterada em relação ao ambiente genético clássico (Tn4401). A análise plasmidial dos produtores de KPC resultou em plasmídeos de tamanhos variáveis, evidenciando a maior prevalência de plasmídeos de ~20Kb no carreamento do gene. A análise também revelou que todos foram não- tipáveis pela técnica de PBRT. Com relação às metodologias fenotípicas utilizadas na detecção de KPC, o IPM apresentou melhor desempenho que o MEM na realização do teste de discos combinados com ácido borônico (AB), resultando em 100% de sensibilidade (SN) e 96.1% de especificidade (SP). A quantificação do Teste Modificado de Hodge (MHT), proposta neste trabalho, eliminou a subjetividade na sua interpretação e evidenciou um aumento considerável na SP do teste em relação à metodologia convencional. Em conclusão, nossos resultados confirmaramm a elevada plasticidade genética e os diversos fenótipos observados na família Enterobacteriaceae; contribuíram para o conhecimento da epidemiologia local de resistência aos carbapenêmicos e dos isolados produtores de KPC; bem como reforçaram o valor das metodologias fenotípicas como ferramenta capaz de discriminar os mecanismos envolvidos na resistência aos carbapenêmicos. / The Enterobacteriaceae family includes important community and nosocomial pathogens frequently associated to multirresistance. Multidrug-resistant strains represent an important concern among hospitals and healthcare institutions around the world, due to the limited therapeutic options. In recent decades, the overall increase of strains producing inducible β-lactamases and extended spectrum β-lactamases (ESBL) has lead to the use of carbapenems as the first option for the treatment of serious infections. However, the carbapenem resistance has been considered a public health problem in many countries and the production of the enzyme Klebsiella pneumoniae carbapenemase (KPC) has been described as the major resistance mechanism to carbapenems in this family. Whereas only sporadic reports of KPC-producing strains have been reported in Rio Grande do Sul (RS), in contrast to the situation in many Brazilian states, this study aimed to establish the prevalence of KPC among isolates with reduced susceptibility to carbapenems from 11 disctinct hospitals of RS, promote the molecular characterization of these isolates, as well as evaluate the main phenotypic methods used for KPC detection. Significant differences were observed among the susceptibility profiles of imipenem (IPM) and meropenem (MEM), when compared to ertapenem (ERT): less than 10% of the isolates were classified as susceptible to ERT compared to over 73% to IPM and MEM. Our results demonstrated that the reduced susceptibility to carbapenems, was mainly due to the production of AmpC β-lactamases and ESBL, instead of true carbapenemases. Regarding the major carbapenemases found in Enterobacteriaceae, only KPC was detected among the isolates studied, at a prevalence of 4%. KPC producers included the species E. cloacae, K. pneumoniae, S. marcescens and K. georgiana, from four different hospitals. The sequencing analysis demonstrated that all of them were KPC-2 producers. According to the susceptibility profile, most were highly resistant to β-lactams and quinolones, whereas polymyxin and amikacin were the most effective drugs in vitro. Two clonal groups were detected among isolates of E. cloacae and S. marcescens and four among isolates of K. pneumoniae, by PFGE analysis. Only a partial characterization of the genetic context that involves the blaKPC-2 gene was possible for these isolates, indicating an altered platform compared to the classic genetic environment (Tn4401). The plasmid analysis indicated plasmids of variable sizes, with a higher prevalence of those of ~ 20Kb involved with the blaKPC-2 gene. The analysis also showed that all of them were non-typable by PBRT technique. With respect to the phenotypic methods used for KPC detection, IPM proved to present a better performance than MEM in the combined-disc test with boronic acid (AB), resulting in 100% of sensitivity (SN) and 96.1% of specificity (SP). The quantification of Modified Hodge Test (MHT), proposed in this study, eliminated the subjectivity of this test leading to a considerable increase in the SP of the test compared to the conventional methodology. In conclusion, our results confirmed the high genetic plasticity and the distinct phenotypes observed in the Enterobacteriaceae family; contributed to the knowledge of the local epidemiology of carbapenem resistance and for KPC-producing isolates; as well as reinforced the use of phenotypic methods as an useful tool able to discriminate the mechanisms involved in carbapenem resistance.