RESUMO
OBJECTIVES: To examine the in vitro selection of aztreonam/avibactam resistance among MBL-producing Klebsiella pneumoniae and to understand the mechanism of increased resistance. METHODS: The MICs of aztreonam were determined with and without avibactam (4 mg/L) using a broth microdilution method. Single-step and multi-step mutant selection was conducted on five MBL-producing K. pneumoniae strains, including two dual carbapenemase producers. Genomic sequencing and gene cloning were performed to investigate the mechanism of increased resistance. RESULTS: We examined the MICs for 68 MBL-producing K. pneumoniae isolates, including 13 dual carbapenemase producers. Compared with aztreonam alone, the addition of avibactam (4 mg/L) reduced the MICs for all isolates by >128-fold, with MIC50 and MIC90 values of 0.25 and 1 mg/L, respectively. One NDM-1-, OXA-48-, CTX-M-15- and CMY-16-positive ST101 K. pneumoniae strain was selected to be resistant to aztreonam/avibactam, with a >16-fold increase in MIC (>128 mg/L). WGS revealed that the resistant mutants lost the blaNDM-1 gene, but acquired amino acid substitutions in CMY-16 (Tyr150Ser and Asn346His). Construction of blaCMY-16 mutants confirmed that the substitutions (Tyr150Ser and Asn346His) were primarily responsible for the decreased susceptibility to aztreonam/avibactam. In addition, transfer of blaCMY-16 mutant (Tyr150Ser and Asn346His) plasmid constructs into certain clinical carbapenemase-producing isolates demonstrated >64-fold increased MICs of aztreonam/avibactam and aztreonam/avibactam/ceftazidime. CONCLUSIONS: Aztreonam in combination with avibactam showed potent in vitro activity against MBL-producing K. pneumoniae. However, our study suggested the likelihood of aztreonam/avibactam resistance among MBL- and AmpC-co-producing strains and clinical practice should beware of the possibility of the emerging resistance.
Assuntos
Aztreonam , Klebsiella pneumoniae , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Compostos Azabicíclicos/farmacologia , Aztreonam/farmacologia , Proteínas de Bactérias , Ceftazidima , Combinação de Medicamentos , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , beta-Lactamases/genéticaRESUMO
Objectives: To examine the epidemiology of ß-lactam resistance in 'clonal group 258' (CG258), a successful KPC clonal group, over 14 years. Methods: Isolates were collected from 1999 to 2013 for a study of antibiotic resistance in Enterobacteriaceae in New York City; 515 bloodstream isolates had antibiotic susceptibility data available and 436 were available for a CG258 PCR assay. The 56 resulting CG258 isolates were characterized by MLST, capsular type and ESBL and KPC carriage. KPC-positive isolates were assessed for common KPC plasmid types, KPC subtype and Tn4401 isoform. Results: RT-PCR revealed 56 isolates were CG258. Seventeen of the 56 CG258 isolates were phenotypically susceptible to all carbapenems (all KPC negative). Five out of 17 susceptible isolates were of the cps-2 (wzi154) capsule type; none was cps-1 (wzi29). Nineteen out of 28 KPC-2 isolates were cps-1 (wzi29) and 8/10 KPC-3 isolates carried cps-2 (wzi154); however, cps-2 (wzi154) predominated among KPC-2-positive isolates in 2003 and 2004. KPC-2 was first detected in 2003 and KPC-3 was first detected in 2006. KPC-harbouring plasmids pKpQIL (all Tn4401a) and pBK30683 (all Tn4401d) were detected in 16/38 and 6/38 carbapenem-resistant isolates, respectively. Discussion: CG258-lineage Klebsiella pneumoniae isolates were completely absent in 1999, but common in 2003. Twenty-one percent of CG258 isolates were susceptible to carbapenems in addition to lacking both common ESBL and blaKPC-mediated resistance. The cps-2 (wzi154) capsule type was common in both these susceptible isolates and in early KPC-2-harbouring isolates, suggesting it was the initial capsule type in CG258. Carbapenem-resistant isolates carried common KPC-harbouring plasmids with the same KPC and Tn4401 isoforms, suggesting frequent clonal spread.
Assuntos
Enterobacteriáceas Resistentes a Carbapenêmicos/efeitos dos fármacos , Enterobacteriáceas Resistentes a Carbapenêmicos/isolamento & purificação , Epidemias , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/isolamento & purificação , Resistência beta-Lactâmica , Enterobacteriáceas Resistentes a Carbapenêmicos/classificação , Enterobacteriáceas Resistentes a Carbapenêmicos/genética , Humanos , Klebsiella pneumoniae/classificação , Klebsiella pneumoniae/genética , Epidemiologia Molecular , Tipagem de Sequências Multilocus , Cidade de Nova Iorque/epidemiologia , Plasmídeos/análise , Reação em Cadeia da Polimerase , Estudos Retrospectivos , beta-Lactamases/genéticaRESUMO
Background: Bacteremia caused by extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae (ESBL-E) is associated with inadequate empirical therapy and substantial mortality in neutropenic patients. Strategies are needed to identify neutropenic patients at high risk of these infections. Methods: From April 2014 to September 2016, we collected perianal swabs, both at admission and weekly thereafter, from patients undergoing hematopoietic stem cell transplantation (HSCT). Patients received prophylactic levofloxacin while neutropenic. Swabs were plated onto selective agar, colonies were identified and underwent antimicrobial susceptibility testing, and phenotypic ESBL testing and polymerase chain reaction for ß-lactamase genes were performed on ceftriaxone-resistant Enterobacteriaceae. We then determined the prevalence of pre-transplant ESBL-E colonization and risk of ESBL-E bacteremia. Colonizing and bloodstream isolates from patients with ESBL-E bacteremia underwent multilocus sequence typing and pulsed-field gel electrophoresis. Results: We analyzed 312 patients, including 212 allogeneic and 100 autologous HSCT recipients. Ten percent (31/312) of patients had pre-transplant ESBL-E colonization. Susceptibility rates of colonizing ESBL-E were: levofloxacin, 25%; cefepime, 9%; piperacillin-tazobactam, 84%; and meropenem, 97%. Of 31 patients colonized with ESBL-E pre-transplant, 10 (32%) developed ESBL-E bacteremia during their transplant admission, compared to 1 (0.4%) of 281 patients not colonized with ESBL-E (P < .001). All bloodstream ESBL-E were levofloxacin-resistant and colonizing and bloodstream isolates from individual patients had identical genotypic profiles. Conclusions: HSCT recipients who are colonized with levofloxacin-resistant ESBL-E pre-transplant and receive levofloxacin prophylaxis have high rates of bacteremia from their colonizing strain during neutropenia. Assessing for ESBL-E colonization in neutropenic patients could lead to optimization of empirical antibacterial therapy.
Assuntos
Antibacterianos/uso terapêutico , Infecções por Enterobacteriaceae/complicações , Enterobacteriaceae/isolamento & purificação , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Levofloxacino/uso terapêutico , Neutropenia/complicações , Adulto , Idoso , Bacteriemia/complicações , Bacteriemia/prevenção & controle , Técnicas de Tipagem Bacteriana , Farmacorresistência Bacteriana , Eletroforese em Gel de Campo Pulsado , Enterobacteriaceae/enzimologia , Infecções por Enterobacteriaceae/prevenção & controle , Feminino , Trato Gastrointestinal/microbiologia , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Tipagem de Sequências Multilocus , Neutropenia/microbiologia , Estudos Prospectivos , Fatores de Risco , beta-LactamasesRESUMO
We describe the first report of a clinical colistin-resistant ST84 Enterobacter cloacae isolate coharboring mcr-4.3 (previously named mcr-4.2) and blaNDM-1 from a patient in China. The blaNDM-1-harboring IncX3 plasmid and the novel mcr-4.3-harboring ColE plasmid were completely sequenced. Although this isolate showed a high level of resistance to colistin, mcr-4.3 plasmid transformation, gene subcloning, susceptibility testing, and lipid A matrix-assisted laser desorption ionization mass spectrometry analysis indicated that mcr-4.3 itself does not confer resistance to colistin.
Assuntos
Enterobacter cloacae/enzimologia , beta-Lactamases/metabolismo , Carbapenêmicos/farmacologia , China , Colistina/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Enterobacter cloacae/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Plasmídeos/genética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , beta-Lactamases/genéticaRESUMO
Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae have now become a global public health threat. However, the origin of this pandemic and the characterization of pre-2003 blaKPC-harboring plasmids remain unknown. Here we used next-generation sequencing to characterize two KPC-2-producing K. pneumoniae and Kmichiganensis isolates collected from a New York City hospital in 1997. Although identified in two different Klebsiella species, the blaKPC-2 gene was harbored by Tn4401b transposons on two highly similar IncN plasmids.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/enzimologia , beta-Lactamases/metabolismo , Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , Elementos de DNA Transponíveis/genética , Surtos de Doenças , Hospitais , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , Cidade de Nova Iorque , Plasmídeos/genética , beta-Lactamases/genéticaRESUMO
Carbapenemase-producing Gram-negative bacteria (CP-GNB) have increasingly spread worldwide, and different families of carbapenemases have been identified in various bacterial species. Here, we report the identification of five VIM metallo-ß-lactamase-producing Alcaligenes faecalis isolates associated with a small outbreak in a large hospital in Gaza, Palestine. Next-generation sequencing analysis showed blaVIM-2 is harbored by a chromosomal genomic island among three strains, while blaVIM-4 is carried by a novel plasmid in two strains.
Assuntos
Alcaligenes faecalis/efeitos dos fármacos , Alcaligenes faecalis/genética , Antibacterianos/farmacologia , beta-Lactamases/genética , Alcaligenes faecalis/isolamento & purificação , Farmacorresistência Bacteriana/efeitos dos fármacos , Farmacorresistência Bacteriana/genética , Feminino , Genoma Bacteriano , Ilhas Genômicas , Infecções por Bactérias Gram-Negativas/epidemiologia , Infecções por Bactérias Gram-Negativas/microbiologia , Humanos , Masculino , Testes de Sensibilidade Microbiana , Oriente Médio/epidemiologia , Filogenia , Polimorfismo de Nucleotídeo Único , beta-Lactamases/metabolismoRESUMO
Ceftazidime-avibactam is a novel ß-lactam/ß-lactamase inhibitor with activity against carbapenem-resistant Enterobacteriaceae (CRE) that produce Klebsiella pneumoniae carbapenemase (KPC). We report the first cases of ceftazidime-avibactam resistance to develop during treatment of CRE infections and identify resistance mechanisms. Ceftazidime-avibactam-resistant K. pneumoniae emerged in three patients after ceftazidime-avibactam treatment for 10 to 19 days. Whole-genome sequencing (WGS) of longitudinal ceftazidime-avibactam-susceptible and -resistant K. pneumoniae isolates was used to identify potential resistance mechanisms. WGS identified mutations in plasmid-borne blaKPC-3, which were not present in baseline isolates. blaKPC-3 mutations emerged independently in isolates of a novel sequence type 258 sublineage and resulted in variant KPC-3 enzymes. The mutations were validated as resistance determinants by measuring MICs of ceftazidime-avibactam and other agents following targeted gene disruption in K. pneumoniae, plasmid transfer, and blaKPC cloning into competent Escherichia coli In rank order, the impact of KPC-3 variants on ceftazidime-avibactam MICs was as follows: D179Y/T243M double substitution > D179Y > V240G. Remarkably, mutations reduced meropenem MICs ≥4-fold from baseline, restoring susceptibility in K. pneumoniae from two patients. Cefepime and ceftriaxone MICs were also reduced ≥4-fold against D179Y/T243M and D179Y variant isolates, but susceptibility was not restored. Reverse transcription-PCR revealed that expression of blaKPC-3 encoding D179Y/T243M and D179Y variants was diminished compared to blaKPC-3 expression in baseline isolates. In conclusion, the development of resistance-conferring blaKPC-3 mutations in K. pneumoniae within 10 to 19 days of ceftazidime-avibactam exposure is troubling, but clinical impact may be ameliorated if carbapenem susceptibility is restored in certain isolates.
Assuntos
Antibacterianos/farmacologia , Compostos Azabicíclicos/farmacologia , Proteínas de Bactérias/genética , Ceftazidima/farmacologia , Genoma Bacteriano , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/genética , Resistência beta-Lactâmica/genética , beta-Lactamases/genética , Idoso , Substituição de Aminoácidos , Proteínas de Bactérias/metabolismo , Cefepima , Ceftriaxona/farmacologia , Cefalosporinas/farmacologia , Clonagem Molecular , Combinação de Medicamentos , Escherichia coli/genética , Escherichia coli/metabolismo , Feminino , Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Infecções por Klebsiella/tratamento farmacológico , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/crescimento & desenvolvimento , Klebsiella pneumoniae/isolamento & purificação , Masculino , Meropeném , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Mutação , Plasmídeos/química , Plasmídeos/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tienamicinas/farmacologia , beta-Lactamases/metabolismoRESUMO
Infections caused by drug-resistant bacteria are a major problem worldwide. Carbapenem-resistant Klebsiella pneumoniae, most notably isolates classified as multilocus sequence type (ST) 258, have emerged as an important cause of hospital deaths. ST258 isolates are predominantly multidrug resistant, and therefore infections caused by them are difficult to treat. It is not known why the ST258 lineage is the most prevalent cause of multidrug-resistant K. pneumoniae infections in the United States and other countries. Here we tested the hypothesis that carbapenem-resistant ST258 K. pneumoniae is a single genetic clone that has disseminated worldwide. We sequenced to closure the genomes of two ST258 clinical isolates and used these genomes as references for comparative genome sequencing of 83 additional clinical isolates recovered from patients at diverse geographic locations worldwide. Phylogenetic analysis of the SNPs in the core genome of these isolates revealed that ST258 K. pneumoniae organisms are two distinct genetic clades. This unexpected finding disproves the single-clone hypothesis. Notably, genetic differentiation between the two clades results from an â¼ 215-kb region of divergence that includes genes involved in capsule polysaccharide biosynthesis. The region of divergence appears to be a hotspot for DNA recombination events, and we suggest that this region has contributed to the success of ST258 K. pneumoniae. Our findings will accelerate research on novel diagnostic, therapeutic, and vaccine strategies designed to prevent and/or treat infections caused by multidrug resistant K. pneumoniae.
Assuntos
Carbapenêmicos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Evolução Molecular , Klebsiella pneumoniae/classificação , Klebsiella pneumoniae/genética , Tipagem de Sequências Multilocus , Pareamento de Bases/genética , Sequência de Bases , Genoma Bacteriano/genética , Geografia , Humanos , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/isolamento & purificação , Dados de Sequência Molecular , Filogenia , Plasmídeos/genética , Polimorfismo de Nucleotídeo Único/genética , Análise de Sequência de DNARESUMO
BACKGROUND: Rapid molecular diagnostic (RMD) platforms may lead to better antibiotic use. Our objective was to develop analytical strategies to enhance the interpretation of RMDs for clinicians. METHODS: We compared the performance characteristics of 4 RMD platforms for detecting resistance against ß-lactams in 72 highly resistant isolates of Escherichia coli and Klebsiella pneumoniae (PRIMERS I). Subsequently, 2 platforms were used in a blinded study in which a heterogeneous collection of 196 isolates of E. coli and K. pneumoniae (PRIMERS II) were examined. We evaluated the genotypic results as predictors of resistance or susceptibility against ß-lactam antibiotics. We designed analytical strategies and graphical representations of platform performance, including discrimination summary plots and susceptibility and resistance predictive values, that are readily interpretable by practitioners to inform decision-making. RESULTS: In PRIMERS I, the 4 RMD platforms detected ß-lactamase (bla) genes and identified susceptibility or resistance in >95% of cases. In PRIMERS II, the 2 platforms identified susceptibility against extended-spectrum cephalosporins and carbapenems in >90% of cases; however, against piperacillin/tazobactam, susceptibility was identified in <80% of cases. Applying the analytical strategies to a population with 15% prevalence of ceftazidime-resistance and 5% imipenem-resistance, RMD platforms predicted susceptibility in >95% of cases, while prediction of resistance was 69%-73% for ceftazidime and 41%-50% for imipenem. CONCLUSIONS: RMD platforms can help inform empiric ß-lactam therapy in cases where bla genes are not detected and the prevalence of resistance is known. Our analysis is a first step in bridging the gap between RMDs and empiric treatment decisions.
Assuntos
Antibacterianos/uso terapêutico , Infecções por Enterobacteriaceae/tratamento farmacológico , Escherichia coli/efeitos dos fármacos , Klebsiella pneumoniae/efeitos dos fármacos , Testes de Sensibilidade Microbiana/métodos , Técnicas de Diagnóstico Molecular/métodos , Resistência beta-Lactâmica , Infecções por Enterobacteriaceae/microbiologia , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Técnicas de Genotipagem/métodos , Humanos , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/isolamento & purificação , Fatores de TempoRESUMO
The emergence and spread of Klebsiella pneumoniae carbapenemase (KPC) among Enterobacteriaceae presents a major public health threat to the world. Although not as common as in K. pneumoniae, KPC is also found in Escherichia coli strains. Here, we genetically characterized 9 carbapenem-resistant E. coli strains isolated from six hospitals in the United States and completely sequenced their blaKPC-harboring plasmids. The nine strains were isolated from different geographical locations and belonged to 8 different E. coli sequence types. Seven blaKPC-harboring plasmids belonged to four different known incompatibility groups (IncN, -FIA, -FIIK2, and -FIIK1) and ranged in size from â¼16 kb to â¼241 kb. In this analysis, we also identified two plasmids that have novel replicons: (i) pBK28610, which is similar to p34978-3 with an insertion of Tn4401b, and (ii) pBK31611, which does not have an apparent homologue in the GenBank database. Moreover, we report the emergence of a pKP048-like plasmid, pBK34397, in E. coli in the United States. Meanwhile, we also found examples of interspecies spread of blaKPC plasmids, as pBK34592 is identical to pBK30683, isolated from K. pneumoniae In addition, we discovered examples of acquisition (pBK32602 acquired an â¼46-kb fragment including a novel replication gene, along with Tn4401b and other resistance genes) and/or loss (pKpQIL-Ec has a 14.5-kb deletion compared to pKpQIL-10 and pBK33689) of DNA, demonstrating the plasticity of these plasmids and their rapid evolution in the clinic. Overall, our study shows that the spread of blaKPC-producing E. coli is largely due to horizontal transfer of blaKPC-harboring plasmids and related mobile elements into diverse genetic backgrounds.
Assuntos
Proteínas de Bactérias/metabolismo , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Plasmídeos/genética , beta-Lactamases/metabolismo , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , Estados Unidos , beta-Lactamases/genéticaRESUMO
The global spread of carbapenem-resistant Klebsiella pneumoniae (CR-Kp) has been largely associated with sequence type 258 (ST258) and its related variants (clonal group 258 [CG258]). Here we describe the molecular epidemiology of CR-Kp from five tertiary care hospitals in Medellín, the second largest city in Colombia. All CR-Kp-infected patients admitted from June 2012 to June 2014 were included (n = 193). Patients' clinical information was obtained from medical records. Carbapenemase KPC, VIM, IMP, NDM, and OXA-48 genes were detected by PCR. A CG258-tonB79 cluster-specific real-time PCR (targeting the multilocus sequence type [MLST] tonB79 allele), pulsed-field gel electrophoresis (PFGE), and MLST analysis were performed for typing. Remarkably, 62.2% (n = 120) of isolates were from STs unrelated to CG258 (non-CG258). KPC-3 predominated in CG258 isolates (86.3%), while KPC-2 prevailed in non-CG258 isolates (75.5%) (P < 0.001). Multidrug resistance (MDR) frequency was significantly higher in CG258 strains (91.4% versus 56.1%; P < 0.001). ST512 (a single-locus variant of ST258) is the main ST in CG258 (96.3%), and isolates in this group showed closely related pulsotype and similar resistance gene profiles, suggesting the clonal spread of this strain. In contrast, high heterogeneity of STs (34/54), including eight novel STs, was found in non-CG258 isolates. Among non-CG258 isolates, ST14 (13.3%; n = 16) and ST307 (14.2%; n = 17) were the most frequent, and they showed distinct molecular and clinical characteristics in comparison to CG258 isolates. Our results suggest that the dissemination of carbapenem resistance in Medellín is due to heterogeneous K. pneumoniae clones, likely the result of horizontal transmission of KPC in different unrelated lineages, further highlighting the challenge in CR-Kp infection control and the need for a multifocal intervention.
Assuntos
Proteínas de Bactérias/genética , Farmacorresistência Bacteriana Múltipla/genética , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/genética , Filogenia , beta-Lactamases/genética , Adulto , Idoso , Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Carbapenêmicos/farmacologia , Células Clonais , Colômbia/epidemiologia , Estudos Transversais , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Monitoramento Epidemiológico , Feminino , Expressão Gênica , Transferência Genética Horizontal , Humanos , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologia , Infecções por Klebsiella/transmissão , Klebsiella pneumoniae/classificação , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/enzimologia , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Epidemiologia Molecular , Tipagem de Sequências Multilocus , Plasmídeos/química , Plasmídeos/metabolismo , Centros de Atenção Terciária , beta-Lactamases/metabolismoRESUMO
Here, we report twoEnterobacter cloacaesequence type 231 isolates coproducing KPC-3 and NDM-1 that have caused lethal infections in a tertiary hospital in China. TheblaNDM-1-harboring plasmids carry IncA/C2and IncR replicons, showing a mosaic plasmid structure, and theblaNDM-1is harbored on a novel class I integron-like element.blaKPC-3is located on a Tn3-ΔblaTEM-1-blaKPC-3-ΔTn1722element, flanked by two 9-bp direct-repeat sequences and harbored on an IncX6 plasmid.
Assuntos
Enterobacter cloacae/genética , Plasmídeos/química , beta-Lactamases/genética , Antibacterianos/farmacologia , Carbapenêmicos/farmacologia , China , Enterobacter cloacae/efeitos dos fármacos , Enterobacter cloacae/crescimento & desenvolvimento , Enterobacter cloacae/patogenicidade , Infecções por Enterobacteriaceae/tratamento farmacológico , Infecções por Enterobacteriaceae/microbiologia , Infecções por Enterobacteriaceae/mortalidade , Infecções por Enterobacteriaceae/patologia , Expressão Gênica , Humanos , Integrons , Testes de Sensibilidade Microbiana , Plasmídeos/metabolismo , Replicon , Análise de Sobrevida , Centros de Atenção Terciária , Resistência beta-Lactâmica/genética , beta-Lactamases/metabolismoRESUMO
We developed and evaluated multiplexed molecular beacon probes in a real-time PCR assay to identify prominent extended-spectrum-ß-lactamase, plasmid-mediated AmpC ß-lactamase (pAmpC) and carbapenemase genes directly from perianal swab specimens within 6 h. We evaluated this assay on 158 perianal swabs collected from hematopoietic stem cell transplant recipients and found that this assay was highly sensitive and specific for detection of CTX-M-, pAmpC-, and KPC-producing Enterobacteriaceae compared to culture on chromogenic agar.
Assuntos
Canal Anal/microbiologia , Enterobacteriaceae/genética , Reação em Cadeia da Polimerase Multiplex/métodos , beta-Lactamases/genética , Proteínas de Bactérias/genética , Enterobacteriaceae/isolamento & purificação , Proteínas de Escherichia coli/genética , Transplante de Células-Tronco Hematopoéticas , Humanos , Manejo de Espécimes/métodosRESUMO
Here we completely sequenced four mcr-1-haboring plasmids, isolated from two extended-spectrum-ß-lactamase (ESBL)-producing Escherichia coli and two carbapenemase-producing Klebsiella pneumoniae clinical isolates. The mcr-1-harboring plasmids from an E. coli sequence type 2448 (ST2448) isolate and two K. pneumoniae ST25 isolates were identical (all pMCR1-IncX4), belonging to the IncX4 incompatibility group, while the plasmid from an E. coli ST2085 isolate (pMCR1-IncI2) belongs to the IncI2 group. A nearly identical 2.6-kb mcr-1-pap2 element was found to be shared by all mcr-1-carrying plasmids.
Assuntos
Proteínas de Bactérias/genética , Enterobacteriaceae/enzimologia , Enterobacteriaceae/genética , beta-Lactamases/genética , Antibacterianos/farmacologia , Enterobacteriaceae/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/enzimologia , Escherichia coli/genética , Testes de Sensibilidade Microbiana , Plasmídeos/genéticaRESUMO
The spread of the plasmid-mediated colistin resistance gene, mcr-1, into carbapenem-resistant Enterobacteriaceae (CRE) clinical isolates poses a significant threat to global health. Here we report the identification of three mcr-1-harboring carbapenem-resistant Escherichia coli strains, collected from three patients in two provinces in China. Our results show that mcr-1-harboring CRE strains have started to spread in different hospitals in China. In addition, this report presents the first description of chromosomal integration of mcr-1 into a carbapenem-resistant E. coli strain.
Assuntos
Antibacterianos/farmacologia , Carbapenêmicos/farmacologia , Colistina/farmacologia , Enterobacteriaceae/efeitos dos fármacos , China , Cromossomos Bacterianos/genética , Farmacorresistência Bacteriana/genética , Enterobacteriaceae/genética , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Hospitais , Testes de Sensibilidade MicrobianaRESUMO
Horizontal transfer of bla(KPC)-harboring plasmids contributes significantly to the inter- and intraspecies spread of Klebsiella pneumoniae carbapenemase (KPC). Here we report the complete nucleotide sequence of a bla(KPC)-harboring IncFIA plasmid, pBK32533, from Escherichia coli. pBK32533 is a cointegrate plasmid comprising of a 72-kb sequence identical to that of the nonconjugative pBK30661 plasmid plus an additional 170-kb element that harbors the genes for plasmid transfer. pBK32533 demonstrates how bla(KPC) can be spread from a nonconjugative plasmid through cointegration.
Assuntos
Escherichia coli/genética , Klebsiella pneumoniae/genética , Proteínas de Bactérias/genética , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Plasmídeos/genética , beta-Lactamases/genéticaRESUMO
We report the first multidrug-resistant Proteus mirabilis strain producing the carbapenemase OXA-48 (Pm-OXA-48) isolated at Al-Shifa hospital in Gaza, Palestine. Draft genome sequencing of Pm-OXA-48 identified 16 antimicrobial resistance genes, encoding resistance to ß-lactams, aminoglycosides, fluoroquinolones, phenicols, streptothricin, tetracycline, and trimethoprim-sulfamethoxazole. Complete sequencing of the bla(OXA-48)-harboring plasmid revealed that it is a 72 kb long IncL/M plasmid, harboring carbapenemase gene bla(OXA-48), extended spectrum ß-lactamase gene bla(CTX-M-14), and aminoglycoside resistance genes strA, strB, and aph(3')-VIb.
Assuntos
Proteínas de Bactérias/genética , Farmacorresistência Bacteriana Múltipla/genética , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/genética , beta-Lactamases/genética , Adulto , Feminino , Genoma Bacteriano/genética , Humanos , Israel , Plasmídeos/genética , Infecções por Proteus/microbiologia , Quinolonas/farmacologiaRESUMO
Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae strains have spread worldwide and become a major threat in health care facilities. Transmission of blaKPC, the plasmid-borne KPC gene, can be mediated by clonal spread and horizontal transfer. Here, we report the complete nucleotide sequences of two novel blaKPC-3-harboring IncFIA plasmids, pBK30661 and pBK30683. pBK30661 is 74 kb in length, with a mosaic plasmid structure; it exhibits homologies to several other plasmids but lacks the plasmid transfer operon (tra) and the origin of transfer (oriT) that are required for plasmid transfer. pBK30683 is a conjugative plasmid with a cointegrated plasmid structure, comprising a 72-kb element that highly resembles pBK30661 (>99.9% nucleotide identities) and an extra 68-kb element that harbors tra and oriT. A PCR scheme was designed to detect the distribution of blaKPC-harboring IncFIA (pBK30661-like and pBK30683-like) plasmids in a collection of clinical Enterobacteriaceae isolates from 10 hospitals in New Jersey and New York. KPC-harboring IncFIA plasmids were found in 20% of 491 K. pneumoniae isolates, and all carried blaKPC-3. pBK30661-like plasmids were identified mainly in the epidemic sequence type 258 (ST258) K. pneumoniae clone, while pBK30683-like plasmids were widely distributed in ST258 and other K. pneumoniae sequence types and among non-K. pneumoniae Enterobacteriaceae species. This suggests that both clonal spread and horizontal plasmid transfer contributed to the dissemination of blaKPC-harboring IncFIA plasmids in our area. Further studies are needed to understand the distribution of this plasmid group in other health care regions and to decipher the origins of pBK30661-like and pBK30683-like plasmids.
Assuntos
Proteínas de Bactérias/genética , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/genética , Plasmídeos/genética , beta-Lactamases/genética , New Jersey , New YorkRESUMO
We developed a multiplex PCR assay capable of identifying two capsular polysaccharide synthesis sequence types (sequence type 258 [ST258] cps-1 and cps-2) in epidemic Klebsiella pneumoniae ST258 strains. The assay performed with excellent sensitivity (100%) and specificity (100%) for identifying cps types in 60 ST258 K. pneumoniae sequenced isolates. The screening of 419 ST258 clonal isolates revealed a significant association between cps type and K. pneumoniae carbapenemase (KPC) variant: cps-1 is largely associated with KPC-2, while cps-2 is primarily associated with KPC-3.
Assuntos
Cápsulas Bacterianas/genética , Proteínas de Bactérias/biossíntese , Farmacorresistência Bacteriana/genética , Klebsiella pneumoniae/genética , beta-Lactamases/biossíntese , Antibacterianos/farmacologia , Cápsulas Bacterianas/classificação , Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , DNA Bacteriano/genética , Humanos , Klebsiella pneumoniae/isolamento & purificação , Klebsiella pneumoniae/patogenicidade , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Tipagem Molecular , Reação em Cadeia da Polimerase Multiplex , beta-Lactamases/genéticaRESUMO
We report here the nucleotide sequence of a novel blaKPC-2-harboring incompatibility group N (IncN) plasmid, pECN580, from a multidrug-resistant Escherichia coli sequence type 131 (ST131) isolate recovered from Beijing, China. pECN580 harbors ß-lactam resistance genes blaKPC-2, blaCTX-M-3, and blaTEM-1; aminoglycoside acetyltransferase gene aac(6')-Ib-cr; quinolone resistance gene qnrS1; rifampin resistance gene arr-3; and trimethoprim resistance gene dfrA14. The emergence of a blaKPC-2-harboring multidrug-resistant plasmid in an epidemic E. coli ST131 clone poses a significant potential threat in community and hospital settings.