Aminoglycoside resistance genes in Enterococcus faecium: mismatch with phenotype (original) (raw)
Related papers
2018
Enterococcus faecium and Enterococcus faecalis are among the predominant species causing hospital-acquired infections, with high mortality and morbidity rates. Enterococci which exhibit high level aminoglycoside (HLAR) and vancomycin resistance (VRE) possess a significant problem in therapeutic treatment. These resistance strains are becoming more widespread in Europe, USA and Asia. Studies have shown that the different distributions of aminoglycoside modifying enzymes (AMEs) among HLAR are based on the geographical regions. In Malaysia, however, data on the prevalence of HLAR and the distribution of AMEs are still limited. Hence, the aims of this study were to determine the antibiotic susceptibility patterns of the clinical isolates E. faecalis and E. faecium and the distribution of HLAR genes among the resistant isolates. Seventy-five clinical enterococci isolates used in this study were originally obtained from a tertiary centre, in the year 2009 and 2010. These isolates were isolated from different sources including pus (50%), blood (32%), urine (11%) and other sources such as CSF and HVS (7%). Reidentification of these isolates was carried out using several methods including sub-culturing on selective medium and Gram staining, followed by confirmatory tests using a commercial biochemical profiling (API20 strep) and 16s rRNA PCR amplification. Antimicrobial susceptibility tests were performed using disc diffusion, E-test and broth microdilution methods. The antibiotics used were aminoglycosides (gentamicin, streptomycin, kanamycin, amikacin, tobramycin) and other antibiotics which are commonly © C O P U P M ii used in the hospital to treat enterococcal infections such as ampicillin, vancomycin, erythromycin, tetracycline, chloramphenicol and linezolid. Detection of HLAR genes was performed on resistance isolates using single PCR amplifications. Out of 25 (33.3%) isolates of E. faecium, 84% and 68% showed high level gentamicin resistance (HLGR) and high level streptomycin resistance (HLSR), respectively. Resistance against erythromycin and tetracycline were 100%, while ampicillin and chloramphenicol showed 84% and 32% resistance rates, respectively. None of the E. faecium isolates showed resistance towards vancomycin and linezolid. On the other hand, all 50 (66.7%) E. faecalis isolates were resistance to all aminoglycosides tested with the following percentages; amikacin (100%), kanamycin (90%), tobramycin (70%), HLGR (36%) and HLSR (46%). These isolates were also resistance to tetracycline (98%), erythromycin (96%), chloramphenicol (46%), ampicillin (24%), linezolid (8%), and vancomycin (4%). Interestingly, the E. faecalis isolates that were resistant to vancomycin were previously reported as susceptible. In this study, E. faecium exhibited higher resistance rates to all antibiotics except vancomycin and linezolid compared to E. faecalis. Additionally, 46% of E. faecalis showed MIC of streptomycin up to 1042 μg/mL while another 36% showed MIC of gentamicin equal to 512 μg/mL. The MIC of aminoglycosides determined in this study showed similar level as observed in other countries, despite the variation of the methods used. The presence of aminoglycoside modifying enzyme (AME) genes [aac(6)-Ie-aph(2)-Ia, aph(2)-Ib, aph(2)-Ic, aph(2)-Id, aph(3)-IIIa] were detected using PCR amplification. The results demonstrated that the HLGR gene, aac(6)-Ie-aph(2)-Ia and the HLSR gene, aph(3)-IIIa were detected in 32% and 40% of E. faecalis and E. faecium resistance isolates, respectively. The spread of these genes were responsible for high level resistance to gentamicin and streptomycin among enterococci isolated in this study. As HLAR genes are highly transferable to not only among the enterococci species but also among various bacterial species, continuous antibiotic surveillance in Malaysian hospitals is warranted in future and preventive measures can be implemented accordingly.
Jundishapur Journal of Microbiology, 2016
Background: Enterococci are important pathogens in nosocomial infections. Various types of antibiotics, such as aminoglycosides, are used for treatment of these infections. Enterococci can acquire resistant traits, which can lead to therapeutic problems with aminoglycosides. Objectives: This study was designed to identify the prevalence of, and to compare, the aac(6')-aph(2") and aph(3)-IIIa genes and their antimicrobial resistance patterns among Enterococcus faecalis and E. faecium isolates from patients at Imam Reza hospital in Kermanshah in 2011-2012. Patients and Methods: One hundred thirty-eight clinical specimens collected from different wards of Imam Reza hospital were identified to the species level by biochemical tests. Antimicrobial susceptibility tests against kanamycin, teicoplanin, streptomycin, imipenem, ciprofloxacin, and ampicillin were performed by the disk diffusion method. The minimum inhibitory concentrations of gentamicin, streptomycin, kanamycin, and amikacin were evaluated with the microbroth dilution method. The aminoglycoside resistance genes aac(6')-aph(2") and aph(3")-IIIa were analyzed with multiplex PCR. Results: The prevalence of isolates was 33 (24.1%) for E. faecium and 63 (46%) for E. faecalis. Eighty-nine percent of the isolates were high-level gentamicin resistant (HLGR), and 32.8% of E. faecium isolates and 67.2% of E. faecalis isolates carried aac(6')-aph(2"). The prevalence of aph(3")-IIIa among the E. faecalis and E. faecium isolates was 22.7% and 77.3%, respectively. Conclusions: Remarkably increased incidence of aac(6')-aph(2") among HLGR isolates explains the relationship between this gene and the high level of resistance to aminoglycosides. As the resistant gene among enterococci can be transferred, the use of new-generation antibiotics is necessary.
Characterization of High-Level Aminoglycoside-Resistant Enterococci in Kuwait Hospitals
Microbial Drug Resistance, 2004
This study investigated the distribution of genes for aminoglycoside-modifying enzymes (AME) and the genetic relatedness of high-level aminoglycoside-resistant enterococci isolated in Kuwait hospitals. A total of 117 enterococci, consisting of 109 Enterococcus faecalis, seven Enterococcus faecium, and one Enterococcus casseliflavus were studied. The MICs of gentamicin, kanamycin, amikacin, tobramycin, and streptomycin were determined by agar dilution and the genes encoding the AAC(6)-APH(2؆), ANT(4), APH(3), APH (2؆)-Ib, APH (2؆)-Ic, APH (2؆)-Id, and ANT(6) enzymes were amplified by PCR. They were typed by pulsed-field gel electrophoresis (PFGE). Filter mating was used to transfer gentamicin resistance determinants. They were all resistant to kanamycin (MIC Ͼ2000 mg/L). Fifty-five isolates were resistant to gentamicin (MIC Ͼ500 mg/L), 72 were resistant to tobramycin (MIC Ͼ64 mg/L), 115 were resistant to amikacin (MIC Ͼ 64 mg/L), and 97 were resistant to streptomycin (MIC Ͼ 1000 mg/L). The aac(6)-Ie-aph(2؆)-Ia was detected in all isolates with gentamicin MIC Ͼ 500 mg/L and in 15 isolates with gentamicin MIC 256 mg/L. The aph(3)-IIIa gene was detected in 101 isolates, whereas the ant(6)-Ia gene was detected in 85 of the 97 streptomycin-resistant isolates with MIC Ն 1000 mg/L. The aac(6)-Ii gene was detected only in the seven E. faecium isolates. None of them contained ant(4)-Ia, aph(2؆)-Ib, aph(2؆)-Ic and aph(2؆)-Id. PFGE revealed heterogeneous patterns with no dominant clone. The results demonstrated that AME are common in aminoglycoside-resistant enterococci isolated in Kuwait. However, the absence of a dominant clone suggests that they acquired high-level aminoglycoside independently.
Egyptian Journal of Medical Human Genetics, 2019
Background: Enterococci are intrinsically resistant to clinically achievable concentrations of aminoglycosides. However, high-level resistance to aminoglycosides (HLAR) is primarily due to the acquisition of genes encoding aminoglycoside-modifying enzymes (AMEs). Aminoglycosides along with cell wall inhibitors are given clinically for treating enterococcal infections. The current study was conducted to investigate the rate of HLAR and to determine aminoglycoside resistance encoding genes profile in enterococcal isolates from different clinical specimens. Results: From 120 Enterococcus species, 50 (41.7%) enterococcal isolates were proven to have HLAR, 78% (39/50) have high-level gentamicin resistance (HLGR), and 74% (37/50) were high-level streptomycin-resistant (HLSR). HLGR isolates carried aminoglycoside modifying gene aac (6′)-Ie-aph (2′)-Ia in 26/39 (66.7%) of isolates, whereas 32/37 (86.5%) of HLSR carried aph (3′)-IIIa gene and were observed in E. faecalis, E. faecium, E. gallinarum, and E. casseliflavus. The aph (2′)-Ib, aph (2′)-Ic, and aph (2′)-Id that encode HLGR could not be detected. Conclusions: The high detection rate of HLAR among the studied Enterococcus species and the coexistence of HLGR and HLSR strains provide crucial insights to the necessity of routine testing for HLAR in the microbiology lab. The main AME genes among HLGR and HLSR enterococci were aac (6′)-Ie-aph (2″)-Ia and aph (3′)-IIIa, respectively.
Microbial Drug Resistance, 2006
Disks containing 120 g of gentamicin were used to detect high-level gentamicin-resistant phenotype (HLGR) among isolates of Enterococcus faecalis (n ؍ 79) and E. faecium (n ؍ 35). These isolates were collected from three hospitals in Tehran during 2002-2004. The macrobroth dilution assay was then used to determine the minimum inhibitory concentration (MIC) of gentamicin. The susceptibility of isolates against amikacin, netilmicin, tobramycin, and kanamycin were also determined by Kirby-Bauer method. All isolates were subjected to polymerase chain reaction (PCR) assays targeting aminoglycoside modifying enzyme (AMEs) genes including aac(6)-aph(2؆), aph(2؆)-Ib, aph(2؆)-Ic, aph(2؆)-Ia, aph(2؆)-Id, aph(3)-IIIa, and ant(4)-Ia. Fifty-nine isolates (52%) showed HLGR phenotype. All isolates with HLGR phenotype and those showing 64 Ͻ MIC Ͻ 500 g/ml contained aac(6)-aph(2؆). The aph(3)-IIIa was found in 61% of the isolates with HLGR phenotypes and in 65% of isolates with MIC Ͻ 500. Coexistence of aac(6)-aph(2؆) and aph(3)-IIIa gene among HLGR isolates of E. faecalis and E. faecium were 60% and 65%, respectively. The gene aph(2؆)-Ic was amplified in two isolates of E. faecium. The results of PCR for aph(2؆)-Id, ant(4)-Ia and aph(2؆)-Ib genes were negative. The aac(6)-aph(2؆) was the most frequent gene encoding resistance to gentamicin and other aminoglycosides followed by aph(3)-IIIa. Isolates lacking these genes were susceptible to all aminoglyocosides tested in this study.
Infection and Drug Resistance, 2019
Background: Enterococcus faecium and Enterococcus faecalis are among the predominant species causing hospital-acquired infections. Currently, enterococcal infections are treated using combination therapy of an aminoglycoside with cell-wall active agents, which led to high level aminoglycoside resistance (HLAR) and vancomycin resistance (VRE) among enterococci. The aim of this study was to determine the prevalence of HLAR and the distribution of the resistance genes among clinical E. faecalis and E. faecium isolates in Malaysia. Materials and methods: Seventy-five enterococci isolates recovered from different clinical sources were re-identified by subculturing on selective medium, Gram staining, biochemical profiling (API 20 Strep), and 16s rRNA sequencing. Antimicrobial susceptibility testing (AST) was performed using Kirby-Bauer disc diffusion, E-test, and broth microdilution methods. PCR amplification was used to detect the presence of aminoglycoside modifying enzyme (AME) genes [aac(6')-Ie-aph(2")-Ia, aph(2")-Ib, aph(2")-Ic, aph(2")-Id, aph (3')-IIIa]. Descriptive data analysis was used to analyze the antibiotic susceptibility profiles and the distribution of HLAR genes. Results: The majority of the isolates recovered from the clinical samples are E. faecalis (66.7%), with the highest recovery from the pus. The prevalence of HLGR (51%) is higher when compared to HLSR (45-49%). Analysis of the resistance genes showed that bifunctional genes aac(6')-Ie-aph(2")-Ia and aph(3')-IIIa contributed to the HLAR E. faecalis and E. faecium. The other AME genes [aph(2")-Ib, aph(2")-Ic, aph(2")-Id] were not detected in this study. Conclusion: This study provides the first prevalence data on HLAR and the distribution of the AME genes among E. faecalis and E. faecium isolates from Malaysia. These highlight the need for continued antibiotic surveillance to minimize its emergence and further dissemination.