RESUMO
IncQ-type plasmids have become important vectors in the dissemination of blaGES among different bacterial genera and species from different environments around the world, and studies estimating the occurrence of Guiana extended-spectrum (GES)-type ß-lactamases are gaining prominence. We analyzed the genetic aspects of two IncQ1 plasmids harboring different blaGES variants from human and environmental sources. The blaGES variants were identified using polymerase chain reaction (PCR) in Aeromonas veronii isolated from hospital effluent and Klebsiella variicola isolated from a rectal swab of a patient admitted to the cardiovascular intensive care unit in a different hospital. Antimicrobial-susceptibility testing and transformation experiments were performed for phenotypic analysis. Whole-genome sequencing was performed using Illumina and Oxford Nanopore platforms. The comparative analysis of plasmids was performed using BLASTn, and the IncQ1 plasmids showed a high identity and similar size. A. veronii harbored blaGES-7 in a class 1 integron (In2061), recently described by our group, and K. variicola carried blaGES-5 in the known class 1 integron. Both integrons showed a fused gene cassette that encodes resistance to aminoglycosides and fluoroquinolones, with an IS6100 truncating the 3'-conserved segment. The fused genes are transcribed together, although the attC site is disrupted. These gene cassettes can no longer be mobilized. This study revealed a mobilome that may contribute to the dissemination of GES-type ß-lactamases in Brazil. Class 1 integrons are hot spots for bacterial evolution, and their insertion into small IncQ-like plasmids displayed successful recombination, allowing the spread of blaGES variants in various environments. Therefore, they can become prevalent across clinically relevant pathogens.
Assuntos
Plasmídeos , beta-Lactamases , Plasmídeos/genética , Brasil , beta-Lactamases/genética , Humanos , Genômica/métodos , Antibacterianos/farmacologia , Klebsiella/genética , Klebsiella/efeitos dos fármacos , Aeromonas/genética , Aeromonas/efeitos dos fármacos , Aeromonas/isolamento & purificação , Testes de Sensibilidade Microbiana , Sequenciamento Completo do Genoma , Genoma Bacteriano , Integrons/genéticaRESUMO
Aquatic environments are recognized as one of the main reservoirs for the emergence and dissemination of high-risk lineages of multidrug-resistant (MDR) bacteria of public health concern. However, the genomic characteristics of antibiotic-resistant Escherichia coli isolates from aquatic origins remain limited. Herein, we examined the antibiotic resistance and virulence genomic profiles of three E. coli recovered from surface water in northwest Mexico. Antimicrobial susceptibility testing, whole-genome sequencing (WGS), and in-depth in silico analysis were performed. Two E. coli exhibited MDR phenotypes. WGS-based typing revealed genetic diversity, and phylogenetic analysis corroborated a notable divergent relationship among the studied E. coli. One E. coli strain, harboring enterotoxigenic and extraintestinal pathogenic-associated virulence genes, was assigned to the ST4 lineage. MDR E. coli, belonging to the international high-risk clones ST410 and ST617, carried genes and mutations conferring resistance to aminoglycosides, ß-lactams, quinolones, sulfonamides, tetracyclines, and trimethoprim. This study describes, for the first time, the detection and genomic profiling of high-risk lineages of E. coli ST410 and ST617 from surface water in Mexico. Additionally, our results underscore the role of surface water as a reservoir for critical pathogenic and MDR E. coli clones and the need for the surveillance and monitoring of aquatic environments via WGS from the One Health perspective.
RESUMO
OBJECTIVES: The main objective of this study was the genetic characterization of clinically relevant class 1 integrons carried by multidrug resistant bacteria isolated from the intestinal microbiota of aquaculture salmon treated with high concentrations of antibiotics. METHODS: In 82 multidrug resistant bacterial isolates, the prevalence of both the conserved elements of the integrons, qacEΔ1 and sul1 genes, and the variable region (VR) was determined. Further, whole genome sequencing and complete genetic analysis was performed in VR-positive isolates. RESULTS: Despite the fact that 100% of the bacterial isolates presented the intI1 gene, only 12.3% carried the qacEΔ1 and sul1 genes and only two (2.4%) presented a VR with gene cassettes. In the Pseudomonas baetica 25P2F9 isolate, a VR carrying aac(6')31, qacH, and blaOXA-2 gene cassettes was described, whereas the VR of Aeromonas salmonicida 30PB8 isolate showed a dfrA14 gene cassette. The array of gene cassettes found in the Pseudomonas isolate appears with high frequency in clinically relevant pathogens such as Pseudomonas aeruginosa or Escherichia coli. Additionally, it was possible to determine that these integrons are contained in plasmids and coul be easily transferred. Resistome analysis demonstrated that both isolates carried a great diversity of antibiotic resistance genes, including many ß-lactamases. Even in the Aeromonas isolate a new oxacillin-hydrolyzing beta-lactamase gene was described (blaOXA-956). CONCLUSION: The presence of multidrug resistant bacteria and clinically relevant genetic elements in the salmon intestinal microbiota make the aquaculture a hotspot in the phenomenon of antibiotic resistance; therefore, the control of antibiotics used in this activity is a key point to avoid its escalation.
Assuntos
Microbioma Gastrointestinal , Salmo salar , Animais , Antibacterianos/farmacologia , Escherichia coli/genética , Integrons/genética , Testes de Sensibilidade Microbiana , Pseudomonas aeruginosa/genéticaRESUMO
This study aimed to identify and characterize integrons among multidrug-resistant (MDR) uropathogenic Escherichia coli (UPEC) from outpatients in Mexico City, Mexico. PCR assays were used to screen for the presence of class 1, 2 and 3 integrons, whose PCR products were sequenced to identify the inserted gene cassettes within the variable regions. Out of 83 tested strains, 53 (63.9%) were positive for the presence of class 1 integrons, whereas no integrons were detected in the remaining strains, regardless of their classes. Most of the strains carrying the intI1 gene belonged to the extraintestinal B2 (41.5%) and commensal A (32.1%) phylogroups, and to a lesser extent, the extraintestinal D (20.8%) and commensal B1 (5.7%) phylogroups. Moreover, 8 different gene cassette arrangements were detected, with dfrA17 and aadA5 being the most common (32.1% of the class 1 integron-positive strains), which confer resistance to trimethoprim/sulfamethoxazole and aminoglycosides, respectively. Our results suggest that class 1 integrons are widely distributed among MDR-UPEC strains in Mexico, which may directly or indirectly contribute to the selection of MDR strains. These findings are important for a better understanding of the factors and mechanisms that promote multidrug resistance among UPEC strains.
Assuntos
Infecções por Escherichia coli , Escherichia coli Uropatogênica , Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Humanos , Integrons/genética , México , Escherichia coli Uropatogênica/genéticaRESUMO
INTRODUCTION: Infections acquired in hospitals are the cause of high morbidity and mortality and with the emergence of resistant bacteria, the problem is greater. The aim of this work was to determine the genetic characteristics and timeline of Klebsiella pneumoniae blaNDM-1 carrying a class 1 integron involved in an intrahospital outbreak. METHODOLOGY: Investigation was made from the first detection of K. pneumoniae blaNDM-1, strain "466", and the last clone "423". 16S rRNA gene analysis showed that 466 strain and clones were related to K. pneumoniae. Extended-spectrum ß-lactamases (ESBL) was detected according to the Clinical and Laboratory Standards Institute (CLSI) and real time-PCR. Typing of K. pneumoniae blaNDM-1 strains was carried by ERIC-PCR and sequencing the variable region of the integrons were performed. RESULTS: A cluster of six resistant isolates of K. pneumoniae blaNDM-1 was detected in intensive care unit (ICU), internal medicine (IM) and orthopedics (OT). Timeline revealed that the first bacterial identification was in ICU and the last clone in OT service. The array genetic of variable region was "IntI/aadA5-drfA17/qacEΔ1-Sul1". CONCLUSIONS: The evidences highlight the importance of the epidemiological surveillance of Extended-spectrum ß-lactamases (ESBL) strains, as well as the need for molecular epidemiological studies to identify the routes of transmission and the contamination sources within health personnel.
Assuntos
Infecção Hospitalar/epidemiologia , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/isolamento & purificação , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Infecção Hospitalar/tratamento farmacológico , Infecção Hospitalar/microbiologia , Surtos de Doenças , Farmacorresistência Bacteriana Múltipla , Feminino , Hospitais , Humanos , Integrons , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/metabolismo , Masculino , México/epidemiologia , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , beta-Lactamases/metabolismoRESUMO
The dissemination of antimicrobial resistance genes and the bacterium that harbor them have increasingly become a public concern, especially in low- and middle-income countries. The present study used whole-genome sequencing to analyze 10 KPC-2-producing Klebsiella pneumoniae isolates obtained from clinical specimens originated from Brazilian hospitals. The study documents a relevant "snapshot" of the presence of class 1 integrons in 90% of the strains presenting different gene cassettes (dfrA30, dfrA15, dfrA12, dfrA14, aadA1, aadA2, and aac(6')Iq), associated or not with transposons. Two strains presented nonclassical integron (lacking the normal 3'conserved segment). In general, most strains showed a complex resistome, characterizing them as highly resistant. Integrons, a genetically stable and efficient system, confer to bacteria as highly adaptive and low cost evolution potential to bacteria, even more serious when associated with high-risk clones, indicating an urgent need for control and prevention strategies to avoid the spread of resistance determinants in Brazil. Despite this, although the class 1 integron identified in the KPC-2-producing K. pneumoniae clones is important, our findings suggest that other elements probably have a greater impact on the spread of antimicrobial resistance, since many of these important genes were not related to this cassette.
Assuntos
Klebsiella pneumoniae/genética , beta-Lactamases/genética , Brasil , Elementos de DNA Transponíveis/genética , Farmacorresistência Bacteriana Múltipla/genética , Humanos , Integrons , Sequenciamento Completo do Genoma/métodosRESUMO
The aim of this study was to investigate the presence of class 1 integrons in a collection of Shiga toxin-producing Escherichia coli (STEC) from different origins and to characterize pheno- and genotypically the antimicrobial resistance associated to them. A collection of 649 isolates were screened for the class 1 integrase gene (intI1) by Polymerase chain reaction The variable region of class 1 integrons was amplified and sequenced. Positive strains were evaluated for the presence of antimicrobial resistance genes with microarray and for antimicrobial susceptibility by the disk diffusion method. Seven out of 649 STEC strains some to serogroups, O26, O103 and O130 isolated from cattle, chicken burger, farm environment and pigs were identified as positive for intl1. Different arrangements of gene cassettes were detected in the variable region of class 1 integron: dfrA16, aadA23 and dfrA1-aadA1. In almost all strains, phenotypic resistance to streptomycin, tetracycline, trimethoprim/sulfamethoxazole, and sulfisoxazole was observed. Microarray analyses showed that most of the isolates carried four or more antimicrobial resistance markers and STEC strains were categorized as Multridrug-resistant. Although antimicrobials are not usually used in the treatment of STEC infections, the presence of Multridrug-resistant in isolates collected from farm and food represents a risk for animal and human health.
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This study aimed to characterize multidrug-resistant Proteus mirabilis clones carrying a novel class 1 integron-borne blaIMP-1 In1359 was inserted into a large conjugative plasmid that also carried blaCTX-M-2 The production of carbapenemases in Enterobacteriaceae that are intrinsically resistant to polymyxins and tigecycline is very worrisome, representing a serious challenge to clinicians and infection control teams.
Assuntos
Regulação Bacteriana da Expressão Gênica , Integrons , Plasmídeos/química , Proteus mirabilis/genética , beta-Lactamases/genética , Antibacterianos/farmacologia , Brasil/epidemiologia , Carbapenêmicos/farmacologia , Células Clonais , Farmacorresistência Bacteriana Múltipla/genética , Humanos , Testes de Sensibilidade Microbiana , Plasmídeos/metabolismo , Polimixinas/farmacologia , Infecções por Proteus/tratamento farmacológico , Infecções por Proteus/epidemiologia , Infecções por Proteus/microbiologia , Infecções por Proteus/transmissão , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/enzimologia , Proteus mirabilis/isolamento & purificação , Centros de Atenção Terciária , Tigeciclina/farmacologia , beta-Lactamases/metabolismoRESUMO
Abstract Fecal bacteria are considered to be a potential reservoir of antimicrobial resistance genes in the aquatic environment and could horizontally transfer these genes to autochthonous bacteria when carried on transferable and/or mobile genetic elements. Such circulation of resistance genes constitutes a latent public health hazard. The aim of this study was to characterize the variable region of the class 1 integron and relate its genetic content to resistance patterns observed in antimicrobial-resistant Escherichia coli isolated from the surface waters of Patos Lagoon, Southern Brazil. Genetic diversity of the isolates and presence of the qacEΔ1 gene, which confers resistance to quaternary ammonium compounds, were also investigated. A total of 27 isolates were analyzed. The variable region harbored dfrA17, dfrA1 and dfrA12 genes, which confer resistance to trimethoprim, and aadA1, aadA5 and aadA22 genes that encode resistance to streptomycin/spectinomycin. Most of the isolates were considered resistant to quaternary ammonium compounds and all of them carried the qacE Δ1 gene at the 3′ conserved segment of the integron. ERIC-PCR analyses of E. coli isolates that presented the integrons showed great genetic diversity, indicating diverse sources of contamination in this environment. These results suggest that fecal bacteria with class 1 integrons in aquatic environments are potentially important reservoirs of antibiotic-resistance genes and may transfer these elements to other bacteria that are capable of infecting humans.
Assuntos
Integrons , Escherichia coli/isolamento & purificação , Escherichia coli/genética , Água Doce/microbiologia , Antibacterianos/farmacologia , Filogenia , Variação Genética , Brasil , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismoRESUMO
Forty-five multi-resistant Salmonella enterica subsp. enterica serovar (S.) Typhimurium isolates obtained at five pig abattoirs in Southern Brazil were characterized. Their relatedness was determined by XbaI-macrorestriction analysis. Resistance genes, integrons and plasmid-mediated quinolone resistance genes (PMQR) were investigated by PCR. Amplicons for the variable part of class 1 integrons and the quinolone resistance-determining regions (QRDR) were sequenced. Plasmids were characterized by conjugation assays and replicon typing. Eighteen XbaI-macrorestriction patterns and 19 plasmid profiles were seen. Resistance to ampicillin (blaTEM), chloramphenicol (catA1 and floR), streptomycin (strA-strB), streptomycin/spectinomycin (aadA variants), sulphonamides (sul1, sul2, sul3) and tetracyclines [tet(A) and tet(B)] were commonly found. A trimethoprim resistance gene, dfrA8, was identified on a 100-kb plasmid. Single substitutions in the QRDR of GyrA but no PMQR genes were found. Twenty-five isolates carried class 1 integrons with an aadA23 gene cassette or unusual class 1 integrons with a dfrA12-orfF-aadA27 gene cassette array. Both integrons were found on large conjugative plasmids. Salmonella plasmid-located virulence genes spvR, spvA, spvB, rck and pefA were found on an IncFIB resistance plasmid. Hybrid virulence-resistance plasmids or plasmids harbouring class 1 integrons may play a role in the maintenance and dissemination of antimicrobial resistance among S. Typhimurium in this pig production system.
Assuntos
Matadouros , Farmacorresistência Bacteriana/genética , Genes Bacterianos/genética , Integrons/genética , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/genética , Animais , Antibacterianos/farmacologia , Brasil , Salmonella typhimurium/patogenicidade , Suínos , Virulência/genéticaRESUMO
Fecal bacteria are considered to be a potential reservoir of antimicrobial resistance genes in the aquatic environment and could horizontally transfer these genes to autochthonous bacteria when carried on transferable and/or mobile genetic elements. Such circulation of resistance genes constitutes a latent public health hazard. The aim of this study was to characterize the variable region of the class 1 integron and relate its genetic content to resistance patterns observed in antimicrobial-resistant Escherichia coli isolated from the surface waters of Patos Lagoon, Southern Brazil. Genetic diversity of the isolates and presence of the qacE1 gene, which confers resistance to quaternary ammonium compounds, were also investigated. A total of 27 isolates were analyzed. The variable region harbored dfrA17, dfrA1 and dfrA12 genes, which confer resistance to trimethoprim, and aadA1, aadA5 and aadA22 genes that encode resistance to streptomycin/spectinomycin. Most of the isolates were considered resistant to quaternary ammonium compounds and all of them carried the qacE 1 gene at the 3 conserved segment of the integron. ERIC-PCR analyses of E. coli isolates that presented the integrons showed great genetic diversity, indicating diverse sources of contamination in this environment. These results suggest that fecal bacteria with class 1 integrons in aquatic environments are potentially important reservoirs of antibiotic-resistance genes and may transfer these elements to other bacteria that are capable of infecting humans.(AU)
Assuntos
Escherichia coli/imunologia , Anti-Infecciosos , Águas SuperficiaisRESUMO
The integron-gene cassette system has typically been associated with antibiotic-resistant pathogens. However, the diversity of gene cassettes and the abundance of class 1 integrons outside of the clinical context are not fully explored. Primers targeting the conserved segments of attC recombination sites were used to amplify gene cassettes from the sediment of the Mina stream, which exhibited a higher degree of stress to metal pollution in the dry season than the rainy season. Of the 143 total analyzed sequences, 101 had no matches to proteins in the database, where cassette open reading frames could be identified by homology with database entries. There was a predominance of sequences encoding essential cellular functions. Each season that was sampled yielded a specific pool of gene cassettes. Real-time PCR revealed that 8.5 and 41.6 % of bacterial cells potentially harbored a class 1 integron in the rainy and dry seasons, respectively. In summary, our findings demonstrate that most of the gene cassettes have no ascribable function and, apparently, historically metal-contaminated sediment favors the maintenance of bacteria containing the intI1 gene. Thus, the diversity of gene cassettes is far from being fully explored deserving further attention.
Assuntos
Poluição Ambiental , Genes Bacterianos , Variação Genética , Sedimentos Geológicos/microbiologia , Integrases/genética , Microbiota , Arsênio/análise , Bacteroidetes/genética , Bacteroidetes/isolamento & purificação , Sedimentos Geológicos/química , Ferro/análise , Proteobactérias/genética , Proteobactérias/isolamento & purificaçãoRESUMO
Fecal bacteria are considered to be a potential reservoir of antimicrobial resistance genes in the aquatic environment and could horizontally transfer these genes to autochthonous bacteria when carried on transferable and/or mobile genetic elements. Such circulation of resistance genes constitutes a latent public health hazard. The aim of this study was to characterize the variable region of the class 1 integron and relate its genetic content to resistance patterns observed in antimicrobial-resistant Escherichia coli isolated from the surface waters of Patos Lagoon, Southern Brazil. Genetic diversity of the isolates and presence of the qacEΔ1 gene, which confers resistance to quaternary ammonium compounds, were also investigated. A total of 27 isolates were analyzed. The variable region harbored dfrA17, dfrA1 and dfrA12 genes, which confer resistance to trimethoprim, and aadA1, aadA5 and aadA22 genes that encode resistance to streptomycin/spectinomycin. Most of the isolates were considered resistant to quaternary ammonium compounds and all of them carried the qacEΔ1 gene at the 3' conserved segment of the integron. ERIC-PCR analyses of E. coli isolates that presented the integrons showed great genetic diversity, indicating diverse sources of contamination in this environment. These results suggest that fecal bacteria with class 1 integrons in aquatic environments are potentially important reservoirs of antibiotic-resistance genes and may transfer these elements to other bacteria that are capable of infecting humans.
Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Água Doce/microbiologia , Integrons , Brasil , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Variação Genética , FilogeniaRESUMO
Se han descrito aislados de Vibrio cholerae resistentes a una amplia variedad de antibióticos. En Venezuela, durante el brote de cólera ocurrido entre noviembre de 1998 y enero 2000 fueron reportados por primera vez aislados de V. cholerae O1 resistentes a ampicilina, trimetoprim-sulfametoxazole. Usando experimentos de conjugación se determinó la capacidad de transferir los determinantes de resistencia a ampicilina y trimetoprim-sulfametoxazole en 11 aislados. La visualización de plásmidos se realizó utilizando la digestión con nucleasa S1 y electroforesis en campo pulsante. La presencia de integrones de clase 1 fue establecida por PCR y se obtuvo la secuencia de la región variable del integrón. Los determinantes de resistencia fueron transferidos en un plásmido conjugativo de aproximadamente 170 kbp, común a todos los aislados. La resistencia a trimetoprim esta codificada en el gen dfra15, el cual se encuentra en un integrón clase 1 presente en el plásmido. En este estudio, se caracterizó la localización genética de los determinantes que codifican la resistencia a los antibióticos, y al conocer el mecanismo probable de dispersión de los determinantes de resistencia se podrán implementar medidas de control más adecuadas.
Vibrio cholerae has been reported to be resistant to a wide range of antibiotics. V. Cholerae O1 strains resistant to ampicillin, trimethoprim-sulfamethoxazole were isolated for the first time in Venezuela during a cholera outbreak that occurred between November 1998 and January 2000. Using conjugation experiments, the capacity of transfer of the resistance determinants in 11 strains resistant to ampicillin and trimethoprim-sulfamethoxazole was investigated. Plasmid analysis was done by S1 nuclease digestion and pulsed field gel electrophoresis. The presence of class 1 integrons was determined by PCR and the sequence of the gene harbored in the variable region of the integron was obtained. The antibiotic resistance determinants were transferred by a conjugative plasmid of approximately 170 kbp, common to all the isolates. Resistance to trimethoprim is encoded by the dfra15 gene that is harbored by a class 1 integron present in the plasmid. In this study, the genetic location of the determinants that code for resistance to antibiotics was characterized, and knowing the probable mechanism of dispersion of the determinants of resistance, control measures can be implemented most appropriate
Assuntos
Humanos , Masculino , Feminino , Criança , Adolescente , Idoso , Trimetoprima , Vibrio cholerae , Resistência Microbiana a Medicamentos , Cólera , Antibacterianos , Plasmídeos , Anti-InfecciososRESUMO
Drug resistant Pseudomonas aeruginosa represents a therapeutic challenge. To assess the diversity of P. aeruginosa antibiotic resistant variants, isolates were recovered from hospital patients in Colombia. Thirty of 60 isolates contained class 1 integrons and five were of Sequence Type ST235 having appeared in a single intensive care unit. All five possessed an unusual integron but showed differences in gene cassette content and the presence/absence of insertion sequence IS26. This showed that differences can arise rapidly, even within a single ICU. Also, the emergence of IS26 in P. aeruginosa is contributing to the evolution of resistance in this bacterium.
Assuntos
Infecção Hospitalar/microbiologia , Farmacorresistência Bacteriana Múltipla , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/isolamento & purificação , Antibacterianos/farmacologia , Colômbia/epidemiologia , Infecção Hospitalar/epidemiologia , Humanos , Unidades de Terapia Intensiva/estatística & dados numéricos , Testes de Sensibilidade Microbiana , Infecções por Pseudomonas/epidemiologia , Pseudomonas aeruginosa/classificação , Pseudomonas aeruginosa/genéticaRESUMO
Introduction The high prevalence of Klebsiella pneumoniae infections is related to the ability of K. pneumoniae to acquire and disseminate exogenous genes associated with mobile elements, such as R plasmids, transposons and integrons. This study investigated the presence of class 1 integrons in clinical and microbiota isolates of K. pneumoniae belonging to different phylogenetic groups and correlated these results with the antimicrobial resistance profiles of the studied isolates. Methods Of the 51 isolates of K. pneumoniae selected for this study, 29 were from multidrug-resistant clinical isolates, and 22 were from children's microbiota. The susceptibility profile was determined using the disk diffusion method, and class 1 integrons were detected through polymerase chain reaction (PCR). Results The results showed that none of the 22 microbiota isolates carried class 1 integrons. Among the 29 clinical isolates, 19 (65.5%) contained class 1 integrons, and resistance to sulfamethoxazole/trimethoprim was identified in 18 of these isolates (94.7%). Among the K. pneumoniae isolates with class 1 integrons, 47% belonged to the KpI phylogenetic group, and one isolate (14.3%) carrying these genetic elements belonged to the KpIII group. Conclusions The wide variety of detected class 1 integrons supports the presence of high rates of antimicrobial resistance, genetic variability, and rapid dissemination of beta-lactamase genes among K. pneumoniae clinical isolates in recent years in hospitals in Recife-PE, Brazil. The findings of this study indicate that the surveillance of K. pneumoniae integrons in clinical isolates could be useful for monitoring the spread of antibiotic resistance genes in the hospital environment. .
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Humanos , Antibacterianos/farmacologia , Integrons/genética , Klebsiella pneumoniae/genética , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , DNA Bacteriano/genética , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/isolamento & purificação , Microbiota/genética , Filogenia , Reação em Cadeia da PolimeraseRESUMO
BACKGROUND: The occurrence and prevalence of integrons in clinical microorganisms and their role played in antimicrobial resistance have been well studied recently. As screening and detection of integrons are concerned, current diagnostic methodologies are restricted by significant drawbacks and novel methods are required for integrons detection. RESULTS: In this study, three loop-mediated isothermal amplification (LAMP) assays targeting on class 1, 2 and 3 integrons were implemented and evaluated. Optimization of these detection assays were performed, including studing on the reaction temperature, volume, time, sensitivity and specificity (both primers and targets). Application of the established LAMP assays were further verified on a total of 1082 isolates (previously identified to be 397 integron-positive and 685 integron-negative strains). According to the results, the indispensability of each primer had been confirmed and the optimal reaction temperature, volume and time were found to be 65°C, 45 min and 25 µL, respectively. As application was concerned, 361, 28 and 8 isolates carrying intI1, intI2 and intI3 yielded positive amplicons, respectively. Other 685 integron-negative bacteria were negative for the integron-screening LAMP assays, totaling the detection rate and specificity to be 100%. CONCLUSIONS: The intI1-, intI2- and intI3-LAMP assays established in this study were demonstrated to be the valid and rapid detection methodologies for the screening of bacterial integrons.