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BACKGROUND: Certain clonal complexes (CCs) of Klebsiella pneumoniae such as CC147 (ST147 and ST392) are major drivers of blaNDM dissemination across the world. ST147 has repeatedly reported from our geographical region, but its population dynamics and evolutionary trajectories need to be further studied. METHODS: Comparative genomic analysis of 51 carbapenem-nonsusceptible strains as well as three hypervirulent K. pneumoniae (hvKp) recovered during 16-months of surveillance was performed using various bioinformatics tools. We investigated the genetic proximity of our ST147 strains with publicly available corresponding genomes deposited globally and from neighbor countries in our geographic region. RESULTS: While IncL/M plasmid harboring blaOXA-48 was distributed among divergent clones, blaNDM-1 was circulated by twenty of the 25 CC147 dominant clone and were mostly recovered from the ICU. The NDM-1 core structure was bracketed by a single isoform of mobile genetic elements (MGEs) [ΔISKpn26-NDM-TnAs3-ΔIS3000-Tn5403] and was located on Col440I plasmid in 68.7% of ST392. However, various arrangements of MGEs including MITESen1/MITESen1 composite transposon or combination of MITESen1/ISSen4/IS903B/IS5/ISEhe3 on IncFIb (pB171) were identified in ST147. It seems that ST392 circulated blaNDM-1 in 2018 before being gradually replaced by ST147 from the middle to the end of sample collection in 2019. ST147 strains possessed the highest number of resistance markers and showed high genetic similarity with four public genomes that harbored blaNDM-1 on the same replicon type. Mainly, there was a convergence between clusters and isolated neighboring countries in the minimum-spanning tree. A conserved arrangement of resistance markers/MGEs was linked to methyltransferase armA which was embedded in class 1 integron in 8 isolates of ST147/ST48 high-risk clones. CONCLUSION: Our findings highlight the dynamic nature of blaNDM-1 transmission among K. pneumoniae in Iran that occurs both clonally and horizontally via various combinations of MGEs. This is the first analysis of Iranian ST147/NDM + clone in the global context.
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Carbapenémicos , Klebsiella pneumoniae , Irán , Klebsiella pneumoniae/genética , Carbapenémicos/farmacología , Genómica , Secuencias Repetitivas Esparcidas/genéticaRESUMEN
Escherichia coli sequence type 131 (ST131) is known for its contribution to multidrug resistance and the worldwide spread of this clone has become a global problem. Understanding the trends among ST131 clades will help design strategies to prevent its rapid dissemination. In this study, 72 ST131 strains were subjected to comparative genomic analysis and 64 clade C strains were compared with clade C strains reported from other regions using publicly available whole-genome sequencing data. C1 (n = 31 [48.4%]) and C2 (n = 33 [%51.5]) strains had the same prevalence in our collection, and C1-M27 (n = 22) strains were closely related, carried a unique plasmid type (F1:A2:B20), and exhibited virotype C. Removal of 11 C2 strains with varied virotype patterns and the heterogeneous IncF type identified 22 closely related virotype E/F strains with replicon type F31/F36:A4:B1, forming what we denote as the "C2-subset." In a global context, the C2-subset constituted a distinct cluster with international virotype E strains and harbored a genomic island, GI-pheU. Association of cnf1/hlyCABD genes with 1 to 7 mobile genetic elements, mostly IS682/ISKpn37 combination within GI-pheU was identified. The C2-subset accounted for excess resistance/virulence of subclade C2 relative to C1 strains. In addition, a conserved chromosomal IS26-mediated composite transposon (IS15DIV-ISEcp1-blaCTX-M-15-WbuC cupin fold metalloprotein-Tn2-IS15DIV) was observed in the C2-subset. The local spread of the C2-subset in the hospital studied, with the carriage of higher virulence/resistance markers and a peculiar F-type plasmid, demonstrates the potential for diversification of the ST131 lineage and the emergence of subpopulations with higher survival potential to cause health care-associated outbreaks. IMPORTANCE Escherichia coli sequence type 131 (ST131) is a globally dominant multidrug-resistant clone that is commonly associated with extraintestinal infections. Specific sublineages have been shown to have emerged and spread within ST131, highlighting the complex nature of ST131 epidemiology. This study systematically compared the Iranian ST131 population to those reported from other countries and found a subpopulation harboring virotype E, a homogeneous profile of plasmid Inc-F type F31/F36:A4:B1 harboring cnf1/hemolysin genes on the genomic island GI-pheU, and up to seven mobile genetic elements (MGEs) flanking cnf1/hemolysin virulence markers. The results of this study highlight the importance of MGEs for virulence gene acquisition and the formation of new subpopulations among pandemic clones such as E. coli ST131.
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Antibacterianos/farmacología , Farmacorresistencia Bacteriana Múltiple/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Secuencias Repetitivas Esparcidas , Factores de Virulencia/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Escherichia coli/clasificación , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/microbiología , Femenino , Genoma Bacteriano , Genómica , Humanos , Masculino , Persona de Mediana Edad , Plásmidos/genética , Análisis de Secuencia de ADN , Virulencia/genética , Secuenciación Completa del Genoma , Adulto JovenRESUMEN
BACKGROUND: Sequence type 131 (ST131) of Escherichia coli is a pandemic clone that drives the increasing rates of antibiotic resistance. While the pervasiveness of ST131 clade C, especially subclades C2 and C1-M27, has been demonstrated in numerous global surveys, no report about the ST131 clades and their virotypes has been published from Iran so far. METHODS: A collection of 73 consecutive ST131 isolates from extraintestinal specimens was investigated for determination of virotypes, antibiotic susceptibility patterns, resistance/virulence determinants, and clade subsets. RESULTS: Most of the isolates belonged to subclade C2 (33/73; 45.2%), which had the highest virulence factor (VF) scores and resistance rates, followed by C1-M27 (18; 24.6%), C1-non-M27 (14; 19.1%), and A (8; 10.9%). The distinctive profiles of subclade C2 virulence genes were revealed by principle coordinates analysis testing. The distribution of the hlyA virulence gene among subclade C2 was not uniform, so that positive strains (21; 63.6%) showed significantly higher rates of resistance (bla CTX-M-15, bla OXA-1, aac(6')-Ib-cr, aac(6')-Ib, aac(3)-IIa) and virulence (hra, tia/hek, K5, cnf, papGII, papC) markers and gentamicin/tobramycin resistance. Virotype C as the most common virotype (34; 46.5%) was predominant among the subclade C1 population, while virotypes E and F (21; 28.7%) were detected among subclade C2, which had the highest VF scores and aminoglycoside resistance rates. CONCLUSIONS: The appearance of virotypes E and F among subclade C2 strains with higher rates of aminoglycoside resistance/virulence gene content shows the shifting dynamics of this pandemic clone in response to antibiotic selection pressure by establishing subsets with higher survival potential.
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BACKGROUND: Klebsiella pneumoniae is a public health concern because of its ability to develop multidrug resistance and hypervirulent genotypes, of those capsular types K1 and K2 cause community and nosocomial life-threatening infections. This study aimed to determine the antibiotic susceptibility patterns and genotypic traits of a collection of Klebsiella spp. isolates. Furthermore, the clonal relatedness of blaNDM producing strains was investigated. METHODS: During a 19-months surveillance study, 122 Klebsiella spp. isolates were cultured from extraintestinal specimens of patients admitted to the tertiary referral hospital in Semnan, Iran. Isolates were identified using biochemical tests and subjected to determination of phylogroups, capsular types and virulence/resistance genes content. Hypervirulent K. pneumoniae (hvKp) strains were detected genotypically, and Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR fingerprinting was used to determine the clonality of blaNDM producing strains. RESULTS: Multidrug resistant phenotype was detected in 75 (61.5%) isolates and amikacin was found as the most potent antibiotic with the susceptibility rate of 85.2%. The carbapenemase genes were detected in 45 (36.8%) strains, including 21 (17.2%) blaOXA-48, 7 (5.6%) blaNDM-1, 14 (11.4%) blaNDM-1/OXA-48 and 3 (2.4%) blaIMP- carrying strains, while 55 (45.08%) isolates showed carbapenem resistant phenotype. The first blaNDM-1 carrying strain was cultured from a sputum specimen on March 2015, while the last positive one was recovered from blood culture on September 2016. Most of the isolates (80.3%) belonged to phylogroup I, and blaNDM-1 was identified among all three phylogroups. The ERIC-PCR clustered the 101 blaNDM negative and 21 blaNDM-1 positive isolates into 25 and five clusters, respectively, and the latter group belonged to clonal complex 147 (CC147). One K1 and 15 K2 blaNDM-1 negative isolates were detected, of those three strains were identified as hvKp. Five K2 positive strains, including four blaOXA-48 producer and one hvKp sequence type 86 (ST86) were carbapenem resistant. Among carbapenem resistant isolates, CC147 strains harboured higher rates of siderophores iutA and ybtS. CONCLUSION: The present findings showed a hospital circulation of CC147 blaNDM-1 or blaNDM-1/OXA-48 producing strains, disseminated in different wards. The hvKp/ST86 strain expressing K2 capsular type and carbapenem resistant phenotype wasn't reported from Iran so far. So, it seems that we must be aware of the emergence and spread of new K. pneumoniae clones associated with resistant and hypermucoviscous phenotypes.
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Proteínas Bacterianas/genética , Infecciones por Klebsiella/epidemiología , Klebsiella pneumoniae/clasificación , Klebsiella pneumoniae/genética , beta-Lactamasas/genética , Cápsulas Bacterianas/genética , Carbapenémicos , Dermatoglifia del ADN , Farmacorresistencia Bacteriana Múltiple/genética , Monitoreo Epidemiológico , Humanos , Irán , Infecciones por Klebsiella/microbiología , Klebsiella pneumoniae/aislamiento & purificación , Pruebas de Sensibilidad Microbiana , Filogenia , Reacción en Cadena de la Polimerasa , Centros de Atención Terciaria , Virulencia/genéticaRESUMEN
BACKGROUND: The Escherichia coli sequence type 131 (ST131) is a well established clone causing significant extraintestinal infections worldwide. However, no studies have been reported the phenotypic and molecular traits of ST131 isolates in comparison to other clones of E. coli from Iran. So, we determined the differences between 69 ST131 strains collected during a one year surveillance study and 84 non-ST131 isolates, including 56 clinical fluoroquinolone resistant and 28 broiler colibacillosis isolates in terms of clonality and genetic background. RESULTS: ST131 isolates were associated with phylogroup B2 (68 out of 69 isolates, 98.4%), while clinical non-ST131 and fluoroquinolone resistant broiler isolates mainly belonged to phylogroup A. The highest virulence score was observed in ST131 clone, while they showed less diversity in virulence profiles than other clinical isolates. Almost all of the ST131 isolates (95.6%) were ExPEC and had the highest virulence scores, but their resistance scores were less than clinical non-ST131 isolates. Broiler isolates showed higher prevalence of ExPEC-associated virulence genes and CTX-M-G1/G9 resistance determinants as compared to clinical non-ST131 isolates. While blaOXA-48/NDM carbapenemases were mostly found in ST131 clone, resistance rate against ertapenem was higher among clinical non-ST131 strains. According to ERIC-based fingerprinting, the ST131 strains were more genetically similar, followed by non-ST131 and broiler isolates. CONCLUSIONS: ST131 isolates possess the ability to make a balance between clonality and extent of resistance/virulence genes content, so this phenomenon gives a fitness advantage over other E. coli clones. The broilers E. coli population poses a potential zoonotic risk which could be transmitted to the community through the food chain. A number of factors are involved in the dissemination of and infections due to ST131 clone.
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Infecciones por Escherichia coli/epidemiología , Escherichia coli/clasificación , Enfermedades de las Aves de Corral/epidemiología , Factores de Virulencia/genética , Animales , Proteínas Bacterianas/genética , Pollos , Farmacorresistencia Bacteriana , Ertapenem/farmacología , Escherichia coli/genética , Escherichia coli/aislamiento & purificación , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/veterinaria , Fluoroquinolonas/farmacología , Humanos , Irán/epidemiología , Fenotipo , Prevalencia , beta-Lactamasas/genéticaRESUMEN
OBJECTIVES: Escherichia coli sequence types (STs) 69, 73, 95, 127, and 131 are major STs frequently causing extraintestinal infections. The prevalence of specific clones and their virulence and resistance profiles has not been described from Iran. The aim of this study was to characterize antimicrobial-susceptibility profiles and virulence traits of five major clones of E. coli recovered from human extraintestinal infections in Semnan, Iran. We compared these traits between major ST clones and also between O25b and O16 subgroups of the ST131 clone. METHODS: We characterized the five major ST clones among 335 collected E. coli isolates obtained from extraintestinal infections, and phylogenetic groups, antimicrobial susceptibility, and virulence/resistance-gene profiles of these major STs were studied. RESULTS: The highest rates of the multidrug-resistance phenotype were detected among ST131 (85.7%) and ST69 (41.7%), and trimethoprim/sulfamethoxazole resistance was detected significantly among the latter clone. Of the 151 isolates belonging to major ST clones, bla OXA-48 was detected among all except the ST127 clone, while bla NDM genes were harbored by 14 (9.2%) isolates, which all belonged to the ST131 clone. Aggregate virulence scores (median) of ST131 isolates (11) were slightly higher than ST69 (8.50) strains, but were lower than ST73 (16), ST95 (16), and ST127 (12.50) isolates. Principal-coordinate analysis revealed distinct virulence profiles with the ST131 clone. ST73, ST95 and ST131 were enriched with "urovirulence" traits, including phylogroup B2 and group B2-associated accessory traits (chuA, iutA, yfcV, papGII, usp, kpsMTII and malX) and the derived variables extraintestinal pathogenic E. coli and uropathogenic E. coli. In contrast, ST69 was depleted of these traits, but enriched with phylogroups D and E. CONCLUSION: Our data emphasize that isolates of the ST131 clone have the ability to make a balance between resistance and virulence traits to establish a wider clone in extraintestinal pathogenic E. coli.
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Carbapenem-resistant Gram-negative bacilli (GNB) are a concern in the Middle East and worldwide. Simple screening methods have been sought to detect carbapenemase producers to determine appropriate therapeutic measures and implement infection control interventions. In this study, we evaluated the efficiency of agar disc diffusion, commercial combined disc test (Rosco), and carbapenem MIC determination in comparison to molecular detection of carbapenemase genes among 82 carbapenem non-susceptible Enterobacteriaceae (CNSE) and 37 Acinetobacter/Pseudomonas isolates. The blaOXA-48, blaNDM, blaNDM/OXA-48, and blaIMP were detected in 68 out of 82 CNSE isolates. All of the Acinetobacter baumannii isolates were positive for the blaOXA-51 (n=23), of those some were positive for blaOXA-48 (n=13) and blaNDM (n=3). Sensitivities and specificities of combined disc test for detection of blaNDM and blaOXA-48 carrying Enterobacteriaceae isolates were 92.5% and 100%, and 58.5% and 100%, respectively, while those for Acinetobacter/Pseudomonas isolates were 100%, 81.8% and 96.2%, 89%, respectively. While carbapenem MIC values had excellent concordance with phenotypic combined disc test for detection of blaOXA-48 producers (area under curve >90%), only ertapenem MIC's could precisely detect blaOXA-48 PCR-positive Enterobacteriaceae isolates (AUC 70%, sensitivity 70%, specificity 50%). The phenotypic commercial test showed excellent sensitivity for detection of blaNDM producers, but had poor sensitivity for blaOXA-48-producing Enterobacteriaceae. Ertapenem MIC values had low sensitivity and specificity for detection of the blaOXA-48-carrying Enterobacteriaceae. This is the first report of A. baumannii isolates co-harbored the blaOXA-48/blaNDM carbapenemases from Iran
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Acinetobacter baumannii/enzimología , Proteínas Bacterianas/análisis , Proteínas Bacterianas/genética , Enterobacteriaceae/enzimología , Técnicas de Genotipaje/métodos , beta-Lactamasas/análisis , Sensibilidad y Especificidad , Pruebas de Sensibilidad Microbiana/métodos , Pseudomonas/enzimología , beta-Lactamasas/genéticaRESUMEN
Carbapenem-resistant Gram-negative bacilli (GNB) are a concern in the Middle East and worldwide. Simple screening methods have been sought to detect carbapenemase producers to determine appropriate therapeutic measures and implement infection control interventions. In this study, we evaluated the efficiency of agar disc diffusion, commercial combined disc test (Rosco), and carbapenem MIC determination in comparison to molecular detection of carbapenemase genes among 82 carbapenem non-susceptible Enterobacteriaceae (CNSE) and 37 Acinetobacter/Pseudomonas isolates. The blaOXA-48, blaNDM, blaNDM/OXA-48, and blaIMP were detected in 68 out of 82 CNSE isolates. All of the Acinetobacter baumannii isolates were positive for the blaOXA-51 (n = 23), of those some were positive for blaOXA-48 (n = 13) and blaNDM (n = 3). Sensitivities and specificities of combined disc test for detection of blaNDM and blaOXA-48 carrying Enterobacteriaceae isolates were 92.5% and 100%, and 58.5% and 100%, respectively, while those for Acinetobacter/Pseudomonas isolates were 100%, 81.8% and 96.2%, 89%, respectively. While carbapenem MIC values had excellent concordance with phenotypic combined disc test for detection of blaOXA-48 producers (area under curve > 90%), only ertapenem MIC's could precisely detect blaOXA-48 PCR-positive Enterobacteriaceae isolates (AUC 70%, sensitivity 70%, specificity 50%). The phenotypic commercial test showed excellent sensitivity for detection of blaNDM producers, but had poor sensitivity for blaOXA-48-producing Enterobacteriaceae. Ertapenem MIC values had low sensitivity and specificity for detection of the blaOXA-48-carrying Enterobacteriaceae. This is the first report of A. baumannii isolates co-harbored the blaOXA-48/blaNDM carbapenemases from Iran.
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Acinetobacter baumannii/enzimología , Proteínas Bacterianas/análisis , Proteínas Bacterianas/genética , Enterobacteriaceae/enzimología , Técnicas de Genotipaje/métodos , Pruebas de Sensibilidad Microbiana/métodos , Pseudomonas/enzimología , beta-Lactamasas/análisis , beta-Lactamasas/genética , Irán , Sensibilidad y EspecificidadRESUMEN
PURPOSE OF THE RESEARCH: In order to gain a better understanding of the role of several mechanisms in antibiotic resistance in Pseudomonas aeruginosa clinical isolates obtained from CF and burn patients, we evaluated gene expression of efflux pumps MexAB-OprM and MexXY(-OprA), the natural ß-lactamase AmpC and outer membrane porin protein OprD. Also, the presence of genes encoding Ambler classes A, B ß-lactamases and aminoglycoside modifying enzymes (AMEs) was examined. PRINCIPAL RESULTS: Piperacillin-tazobactam and amikacin retained the highest in vitro activities among 21 CF and 27 burn P. aeruginosa isolates. Based on Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR, 15 distinct patterns were detected. There were 5 CF and 6 burn isolates harbored PER-1 and VEB-1, respectively. Among AMEs, involved in resistance of anti-Pseudomonas aminoglycosides, aac(6')-Ib was the most prevalent gene. Among CF isolates, mexA overexpression was the most prevalent mechanism (47.6%) followed by mexX (42.8%), ampC (9.5%) and oprD downregulation (4.7%). Among burn isolates, the prevalence of mexX, mexA, and ampC overexpression was 62.9%, 74%, and 11.1%, respectively. Downregulation of oprD was observed in 14.8% of burn isolates. MAJOR CONCLUSIONS: Among CF isolates, mexX and mexA overexpression were the major contributing factors to aminoglycoside (gentamicin) and carbapenem (meropenem) resistance, respectively while among burn isolates, AMEs in conjunction with mexX hyperexpression were identified to be responsible for aminoglycoside resistance. Also mexA overexpression was partially associated with carbapenem resistance. Moreover, cephalosporin resistance was linked to overexpression of mexA and/or mexX. The impact of interplay between different resistance mechanisms on resistant phenotypes was more complicated among burn than CF isolates.
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Quemaduras/microbiología , Fibrosis Quística/microbiología , Farmacorresistencia Bacteriana Múltiple/genética , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/metabolismo , Amicacina/uso terapéutico , Antibacterianos , Proteínas de la Membrana Bacteriana Externa/biosíntesis , Proteínas Bacterianas/biosíntesis , Cefalosporinasa , Proteínas de Escherichia coli , Regulación Bacteriana de la Expresión Génica , Humanos , Proteínas de Transporte de Membrana/biosíntesis , Pruebas de Sensibilidad Microbiana , Ácido Penicilánico/análogos & derivados , Ácido Penicilánico/uso terapéutico , Piperacilina/uso terapéutico , Porinas/biosíntesis , Infecciones por Pseudomonas/tratamiento farmacológico , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/aislamiento & purificación , Tazobactam , beta-Lactamasas/biosíntesisAsunto(s)
Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/efectos de los fármacos , Antibacterianos/farmacología , Farmacorresistencia Bacteriana Múltiple , Tetraciclinas/farmacología , Acinetobacter baumannii/clasificación , Acinetobacter baumannii/genética , Acinetobacter baumannii/aislamiento & purificación , Genes Bacterianos , Genotipo , Hospitales , Humanos , Irán , Pruebas de Sensibilidad Microbiana , Tipificación MolecularRESUMEN
OBJECTIVES: We examined the molecular epidemiology of Acinetobacter baumannii clinical isolates from two cities (Tehran and Tabriz) of Iran. METHODS: DiversiLab repetitive extragenic palindromic PCR (rep-PCR), multilocus sequence typing and sequence group multiplex PCR were performed. The presence of resistance mechanisms including metallo-ß-lactamases, extended-spectrum ß-lactamases, OXA carbapenemases, aminoglycoside-modifying enzymes and RNA methylases was also investigated. RESULTS: DiversiLab rep-PCR identified 11 clusters and 11 singleton isolates. Twelve sequence types (STs), including six novel types, were identified. Sequence groups (SGs) 1-3 as well as five additional banding patterns were detected by multiplex PCR. A local outbreak in a general hospital in Tabriz with an SG1/ST2 profile was identified. Isolates of international clone II showed the highest prevalence and the most heterogeneous combination of resistance determinants. CONCLUSIONS: Several different multiresistant strains of A. baumannii were shown to circulate in Iran. The selection and spread of the SG1/ST2 clone might have been favoured by the acquisition of resistance genes in the absence of adequate infection control measures.
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Infecciones por Acinetobacter/epidemiología , Acinetobacter baumannii/clasificación , Acinetobacter baumannii/genética , Farmacorresistencia Bacteriana Múltiple , Tipificación Molecular , Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/aislamiento & purificación , Ciudades/epidemiología , Análisis por Conglomerados , Variación Genética , Genotipo , Humanos , Irán/epidemiología , Epidemiología MolecularRESUMEN
PURPOSE: The antimicrobial activity of doripenem in comparison of imipenem, meropenem and ertapenem among Pseudomonas aeruginosa isolated from burn and Cystic Fibrosis (CF) patients were determined. METHODS: Metallo-ß-lactamase (MBL) genes in imipenem non susceptible P. aeruginosa isolates were detected using PCR method. The in vitro susceptibilities of doripenem, imipenem, meropenem and ertapenem were determined by Etests. MIC50 and MIC90 for corresponding antibiotics were determined individually in burn and CF isolates. RESULTS: Among isolates which were resistant to imipenem, 16 isolates were positive for the bla IMP gene. All isolates had no bla VIM gene. All MBL producing isolates were excluded. MIC50/MIC90 of doripenem in CF and burn isolates were 0.75/>32 and >32/>32 mg/L respectively. The corresponding values for imipenem in CF and burn isolates were 2/>32 and >32/>32 mg/L, respectively. CONCLUSION: The susceptibility rate of doripenem is higher than that of imipenem and meropenem among P.aeruginosa isolated from CF patients, whereas, there is no difference between the efficiency of doripenem and old carbapenems in non MBL producing P.aeruginosa isolates in burn patients.
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We examined the prevalence of various cephalosporins' resistance mechanisms in Acinetobacter baumannii clinical isolates. Phenotypic and molecular detection of Ambler classes A, B and D ß-lactamases was performed on 75 isolates. Clonal relatedness was defined using Repetitive Extragenic Palindromic PCR. PCR mapping was used to examine the linkage of insertion sequences and the ampC gene, and ampC expression was analyzed by TaqMan reverse transcriptase-PCR. Twenty-six (37%) isolates carried at least one of the blaPER-1 or blaTEM-1. Sixty-nine (98.5%) out of 70 cephalosporin-resistant isolates had insertions upstream of the ampC gene, of which 48 (69%) and 6 (8%) were identified as ISAba1and ISAba125, respectively. Higher level of expression was obtained in resistant isolates lacking ISAba1/ampC combination in comparison with that in positive ones. The ability to up-regulate the expression of ampC gene in association with different insertion elements has become an important factor in A. baumannii resistance to cephalosporins.
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Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/enzimología , Antibacterianos/farmacología , Resistencia a las Cefalosporinas , Cefalosporinasa/genética , Cefalosporinas/farmacología , Infecciones por Acinetobacter/tratamiento farmacológico , Infecciones por Acinetobacter/epidemiología , Acinetobacter baumannii/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Cefalosporinasa/metabolismo , Mapeo Cromosómico , Análisis Mutacional de ADN , Pruebas Antimicrobianas de Difusión por Disco , Humanos , Tipificación Molecular , Mutagénesis Insercional , Prevalencia , Reacción en Cadena en Tiempo Real de la Polimerasa , Infecciones del Sistema Respiratorio/tratamiento farmacológico , Infecciones del Sistema Respiratorio/epidemiología , Infecciones del Sistema Respiratorio/microbiología , beta-Lactamasas/genética , beta-Lactamasas/metabolismoRESUMEN
We examined the prevalence of various carbapenem resistance mechanisms in clinical isolates of Acinetobacter baumannii collected from hospitalized burn and non-burn patients. Antimicrobial susceptibility testing was performed for 43 burn and 32 non-burn isolates. Carbapenem resistance genes were identified and repetitive extragenic palindromic PCR (REP-PCR) was used to define clonal relatedness. CarO disruption was investigated by PCR and its expression analyzed by real time reverse transcription-PCR. Of the sixty-four (85%) carbapenem resistant A. baumannii isolates, 42 (66%) and 22 (34%) strains were recovered from burn and non-burn patients, respectively. Isolates were categorized into 6 major REP-PCR patterns; with the highest prevalence of non-burn and burn isolates in pattern A (63%) and B (35%), respectively. Prevalence of blaOXA-23 was 68% and isolates harbored this element belonged to all REP clusters. The blaOXA-40-like was detected in 49% of isolates, with higher prevalence among burn isolates. Three of the four isolates lacked carO gene were cultured from burn patients and level of the carO expression was decreased in carbapenem resistant isolates. These findings show that blaOXA-23 is widely distributed in carbapenem resistant A. baumannii isolates and other resistance mechanisms such as blaOXA-40-like and loss or decreased carO expression could be added in burn strains.
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Acinetobacter baumannii/efectos de los fármacos , Antibacterianos/farmacología , Quemaduras/microbiología , Carbapenémicos/farmacología , Farmacorresistencia Bacteriana Múltiple , Infecciones por Acinetobacter/epidemiología , Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/genética , Proteínas Bacterianas/genética , ADN Bacteriano/genética , Farmacorresistencia Bacteriana Múltiple/genética , Humanos , Pruebas de Sensibilidad Microbiana , Reacción en Cadena de la Polimerasa/métodos , beta-Lactamasas/genéticaRESUMEN
AIMS: The purpose of the present study was to investigate the diversity of the genes encoding aminoglycoside-modifying enzymes (AMEs) and their associations with resistance phenotypes and clonality in Acinetobacter baumannii and Pseudomonas aeruginosa isolates. METHODS: Seventy six P. aeruginosa and 75 A. baumannii isolates were collected from three University affiliated hospitals in Tehran. MIC determination of amikacin and gentamicin as well as the disk diffusion method for tobramycin, netilmicin, and kanamycin were carried out. Nine AMEs genes and three RNA methylases were investigated in all isolates using the PCR method. Clonality for A. baumannii and P. aeruginosa was investigated using repetitive extragenic palindromic and enterobacterial repetitive intergenic consensus PCR, respectively. RESULTS: aph(3')-VIa (90.6%) and aph(3')-IIb (61.8%) were the most prevalent AME genes in A. baumannii and P. aeruginosa, respectively. Eight (26%) amikacin highly resistant A. baumannii isolates were positive for armA methylase. Phenotypes and clonality did not link to the genetic determinants of resistance to aminoglycosides in our isolates. CONCLUSIONS: AMEs genes are disseminated in different clones of A. baumannii and P. aeruginosa isolates in Iran. Other than AMEs, there are more complex and multifactorial mechanisms that result in aminoglycoside-resistant phenotypes.
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Acinetobacter baumannii/genética , Proteínas Bacterianas/genética , Farmacorresistencia Bacteriana Múltiple/genética , Kanamicina Quinasa/genética , Metiltransferasas/genética , Pseudomonas aeruginosa/genética , ARN Ribosómico 16S/genética , Infecciones por Acinetobacter/tratamiento farmacológico , Infecciones por Acinetobacter/microbiología , Infecciones por Acinetobacter/transmisión , Acinetobacter baumannii/clasificación , Acinetobacter baumannii/enzimología , Acinetobacter baumannii/aislamiento & purificación , Aminoglicósidos/farmacología , Antibacterianos/farmacología , Proteínas Bacterianas/clasificación , Proteínas Bacterianas/metabolismo , Humanos , Irán , Kanamicina Quinasa/clasificación , Kanamicina Quinasa/metabolismo , Metiltransferasas/clasificación , Metiltransferasas/metabolismo , Pruebas de Sensibilidad Microbiana , Reacción en Cadena de la Polimerasa/métodos , Infecciones por Pseudomonas/tratamiento farmacológico , Infecciones por Pseudomonas/microbiología , Infecciones por Pseudomonas/transmisión , Pseudomonas aeruginosa/clasificación , Pseudomonas aeruginosa/enzimología , Pseudomonas aeruginosa/aislamiento & purificación , ARN Ribosómico 16S/clasificación , ARN Ribosómico 16S/metabolismoRESUMEN
The natural resistance-associated macrophage protein (NRAMP1), Vitamin-D receptor (VDR) and Tumor necrosis factor (TNF-¦Á) have been associated in susceptibility to tuberculosis, but the results have been inconsistent. This study aimed to determine the association of NRAMP1, VDR, and TNF-¨¢ variant with development of pulmonary tuberculosis (PTB) among Iranian patients. The single nucleotide polymorphisms (SNPs) at INT4, D543, 3'UTR of NRAMP1 gene, SNPs in restriction sites of BsmI, and FokI of the VDR gene and SNPs of TNF-¦Á at -238,-308, -244,857,-863 positions were analyzed by PCR-RFLP among two groups of individual; patients with PTB (n=117) and healthy controls (n=60). Thereafter, the frequencies of extended haplotypes and diplotypes were estimated. No statistically significant differences were observed in allele frequencies of INT4, D543, 3'UTR of NRAMPI, FokI of VDR and TNF-¦Á at -238, -244,-863 and -857 position. Although, the frequency of b allele of BsmI [ORs: 0.24 CI95 percent (0.07-0.67 (p=0.001)] and -308 A variant in TNF-¦Á promoter region [ORs:0.26 CI95 percent( 0.07-0.77) (p=0.006)] were significantly more in PTB patients than healthy controls. The frequency of extended diplotypes of NRAMP [GG TGTG++GA; 0.02(0.001-0.0035)], VDR [FFBB; 0.2(0.6-0.6] and TNF-¦Á [CCCCGGGGGG; 0.49(0.25-0.97)] were statistically different in patients and control subjects (p<0.05). This study confirmed the association of SNPs in BsmI (B/b + b/b) of VDR and SNPs in -308A (G/A +G/G) of TNF-¦Á genes with susceptibility to tuberculosis in Iranian PTB patients. Furthermore, the extended haplotypes and diplotypes analysis can be considered as an alternative way to determine the host susceptibility to TB.
Asunto(s)
Humanos , Proteínas de Transporte de Catión/genética , Predisposición Genética a la Enfermedad/genética , Receptores de Calcitriol/genética , Tuberculosis Pulmonar/genética , Factor de Necrosis Tumoral alfa/genética , Estudios de Casos y Controles , Genotipo , Irán , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , Polimorfismo de Nucleótido SimpleRESUMEN
The natural resistance-associated macrophage protein (NRAMP1), Vitamin-D receptor (VDR) and Tumor necrosis factor (TNF-alpha) have been associated in susceptibility to tuberculosis, but the results have been inconsistent. This study aimed to determine the association of NRAMP1, VDR, and TNF-á variant with development of pulmonary tuberculosis (PTB) among Iranian patients. The single nucleotide polymorphisms (SNPs) at INT4, D543, 3'UTR of NRAMP1 gene, SNPs in restriction sites of BsmI, and FokI of the VDR gene and SNPs of TNF-alpha at -238,-308, -244,857,-863 positions were analyzed by PCR-RFLP among two groups of individual; patients with PTB (n=117) and healthy controls (n=60). Thereafter, the frequencies of extended haplotypes and diplotypes were estimated. No statistically significant differences were observed in allele frequencies of INT4, D543, 3'UTR of NRAMPI, FokI of VDR and TNF-alpha at -238, -244,-863 and -857 position. Although, the frequency of b allele of BsmI [ORs: 0.24 CI95% (0.07-0.67 (p=0.001)] and -308 A variant in TNF-alpha promoter region [ORs:0.26 CI95%( 0.07-0.77) (p=0.006)] were significantly more in PTB patients than healthy controls. The frequency of extended diplotypes of NRAMP [GG TGTG++GA; 0.02(0.001-0.0035)], VDR [FFBB; 0.2(0.6-0.6] and TNF-alpha [CCCCGGGGGG; 0.49(0.25-0.97)] were statistically different in patients and control subjects (p<0.05). This study confirmed the association of SNPs in BsmI (B/b + b/b) of VDR and SNPs in -308A (G/A +G/G) of TNF-alpha genes with susceptibility to tuberculosis in Iranian PTB patients. Furthermore, the extended haplotypes and diplotypes analysis can be considered as an alternative way to determine the host susceptibility to TB.