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Phenotypic Detection of High-Level Aminoglycoside Resistance (HLAR) in Enterococcus Species in a Tertiary Care Centre

Phenotypic Detection of High-Level Aminoglycoside Resistance (HLAR) in Enterococcus Species in a Tertiary Care Centre

14. Swenson, J.M., Ferraro, M.J., National Committee for Clinical Laboratory Standard Working Group on Enterococci. et al. Multilaboratory evaluation of screening methods for detection of high-level aminoglycoside resistance in enterococci. J Clin Microbiol, 33: 3008-18 (1995). Facklam 15. David Paterson , Janet botman and Mee Len Thong, Royal Brisbane Hospital, Herston Road, Hersten, Queensland 4006. High level aminoglycoside resistance in Enterococcal blood culture isolates. Comm Dis Intell., 20: 532-535 (1996).

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Detection of high-level aminoglycoside resistance by disc diffusion and e-test amongst the enterococcus species isolated from various clinical samples in a tertiary care hospital

Detection of high-level aminoglycoside resistance by disc diffusion and e-test amongst the enterococcus species isolated from various clinical samples in a tertiary care hospital

The study was conducted in a tertiary care hospital, on various clinical samples of IPD and OPD patients attending NIMS hospital Jaipur Study Period: January 2015 to June 2016. Study Population: Includes patients of all age group and gender both from outpatients and in patient departments. Inclusion Criteria are including the patient of all age group and all samples (like urine, blood, pus, stool, wound swab, sputum, body fluids, etc) received in our lab and were collected as per standard guidelines only. Exclusion Criteria is Nil. Methodology Followed: Collected samples received in lab and then processed in microbiology lab and isolates were identified and confirmed as Enterococcus species by various conventional methods (Gram staining, catalase test, bile esculin test, PYR test, growth at 45˚C, salt tolerance test 6.5%, growth at alkaline pH 9.6, arginine dihydrolase test, hippurate hydrolysis test, potassium tellurite reduction test, sugar fermentation test). These isolates were subjected to Antibiotic susceptibility testing using Kirby-Baeur disc diffusion method as per CLSI guidelines 2016. The antibiotics disc used are ampicillin 10µg, nitrofurantoin 300µg, gentamicin (HLG) 120µg, and streptomycin (HLS) 300µg, ciprofloxacin 5µg, vancomycin 30µg, linezolid 30µg, teicoplanin 30µg, quinpristin / dalfopristin 15µg with E. faecalis ATCC 29212 as quality control. E-test was done for all high level aminoglycoside resistance Enterococcus species isolated by disc diffusion test. Statistical Analysis: Statistical analysis is done by using SPSS software version 17.
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<p>High Level Aminoglycoside Resistance And Distribution Of The Resistance Genes In <em>Enterococcus faecalis</em> And <em>Enterococcus faecium</em> From Teaching Hospital In Malaysia</p>

<p>High Level Aminoglycoside Resistance And Distribution Of The Resistance Genes In <em>Enterococcus faecalis</em> And <em>Enterococcus faecium</em> From Teaching Hospital In Malaysia</p>

The most common clinical manifestations of entero- coccal infections were urinary tract and wound infections, septicemia, and endocarditis. The recommended manage- ment of enterococcal infections involves a synergistic combination of aminoglycosides such as streptomycin and gentamicin with cell wall active inhibitors such as glycopeptides or beta-lactams. The emergence of high- level aminoglycoside resistant (HLAR) enterococci has posed signi fi cant challenges for infection management due to production of aminoglycosides modifying enzymes (AME) that inactivate aminoglycosides. 5 Acquisition of AME genes such as aac(6 ′ )-Ie-aph(2 ″ )-Ia, aph(2 ″ )-Ib, aph(2 ″ )-Ic, aph(2 ″ )-Id, and aph(3 ′ )IIIa by enterococci has seriously affected the treatment for enterococcal infections. 6 High-level aminoglycosides resistance (HLAR) in enterococci was fi rst detected in 1980s and is de fi ned by MIC ≥ 512 μ g/mL. 7
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Detection of high level aminoglycoside resistance genes among clinical isolates of Enterococcus species

Detection of high level aminoglycoside resistance genes among clinical isolates of Enterococcus species

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.
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Multilaboratory evaluation of screening methods for detection of high level aminoglycoside resistance in enterococci  National Committee for Clinical Laboratory Standards Study Group on Enterococci

Multilaboratory evaluation of screening methods for detection of high level aminoglycoside resistance in enterococci National Committee for Clinical Laboratory Standards Study Group on Enterococci

For gentamicin, sensitivity and specificity of the broth mi- crodilution test were excellent at all concentrations, but they were best when 500 m g/ml was used and reading was done at 24 h (Table 5). For streptomycin (Table 6), the most effective parameters were not as obvious. Sensitivity was very good with streptomycin at 500 m g/ml, but specificity was poor, especially when Difco BHIB was used, whereas the reverse was true for testing with streptomycin at 2,000 m g/ml. The best combination of sensitivity and specificity was achieved with 1,000 m g/ml, although with the medium from Becton Dickinson Microbiol- ogy Systems, the sensitivity was not quite as good as that with the medium from Difco. Since the results from all seven lab- oratories were combined for this phase, each of the 48 strains included in the study was used seven times for evaluation. The most significant errors, then, would be with the resistant or- ganisms that were missed in more than one or two laboratories. More than half of the resistant organisms included in phase 3 were characterized as being borderline resistant to streptomy- cin, and as such, they would be the most difficult to detect. Only two strains were missed in more than two laboratories: one strain characterized as being borderline resistant and strain 211, for which the MICs were consistently less than 1,000 m g/ml (see above). The specificity errors occurred among seven isolates, with only one error (involving a strain [strain 566] for TABLE 11. Results of streptomycin high-content (300 m g) disk diffusion screen determined in a single laboratory with 146 strains of
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Detection of various virulence factors in high level aminoglycoside resistance and vancomycin resistant enterococci isolates of uropathogenic Enterococci

Detection of various virulence factors in high level aminoglycoside resistance and vancomycin resistant enterococci isolates of uropathogenic Enterococci

By observing the various parameters of present study, it can be concluded that enterococci are now emerging as a potential pathogen, particularly among hospitalized patients. E. faecalis and E. faecium found to be the most prevalent species which confer resistance to various groups of antibiotics. In our institution Enterococcus isolates was more resistant to fluoroquinolones, aminoglycosides and β-lactams agents like Ampicillin & Piperacillin. This may be due to selection pressure of these antibiotics in our set up. Early detection of enterococcal species and resistance to Aminoglycoside and vancomycin can be helpful in limiting the morbidity in hospital set up. A continuous monitoring is needed particularly for resistance to Aminoglycoside and Vancomycin to stop their institutional spread. Judicial use
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Effects of medium and inoculum variations on screening for high level aminoglycoside resistance in Enterococcus faecalis

Effects of medium and inoculum variations on screening for high level aminoglycoside resistance in Enterococcus faecalis

Because agar screen results showed that inocula of 104 to 106 CFU generally resulted in high specificities and sensitivities Tables 2 and 3, respectively, and because 105 CFU/ml is the f[r]

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Factors influencing determination of high level aminoglycoside resistance in Enterococcus faecalis

Factors influencing determination of high level aminoglycoside resistance in Enterococcus faecalis

There was complete agreement between UC and UN when these criteria were used for interpreting standard agar screen results, but difficulties in interpreting light growth may account for [r]

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Vancomycin Resistant Enterococci in Preterm Neonates: A Rising Thunder in Intensive Care Units

Vancomycin Resistant Enterococci in Preterm Neonates: A Rising Thunder in Intensive Care Units

The study shows upsurge of Vancomycin resistant Enterococci (11 %) and High level Aminoglycoside resistance (78 %) in Enterococcal neonatal septicemia (16 %), considered as only colonizers previously. With this increase in mass spread, increasing resistance to other glycopeptides, macrolides and lipopeptides leads to development of multi drug resistant Enterococci. It is a burning issue as after these drugs, no other effective treatment is available. So judicious use of Vancomycin and other higher antibiotics like Linezolid, Daptomycin, Teicoplanin should be done. Further, VRE can be transmitted from one neonate to another, so Active surveillance culture programs with more such studies should be done to identify colonizers, understand resistance pattern of the organisms and to curtail morbidity and mortality in neonates. In addition to that, periodic training and surveillance are needed to control and increase awareness amongst the health care workers about the rising multi drug resistant infectious incidence of Enterococci.
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Amplification of Aminoglycoside Resistance Gene aphA1 in Acinetobacter baumannii Results in Tobramycin Therapy Failure

Amplification of Aminoglycoside Resistance Gene aphA1 in Acinetobacter baumannii Results in Tobramycin Therapy Failure

The mechanism by which Aph(3=)-I confers tobramycin resis- tance is of interest, as tobramycin lacks the 3= hydroxyl group which is the target of this phosphotransferase (27). Tobramycin resistance in Escherichia coli by overproduction of Aph(3=)-I has been reported (28). In that study, high-level expression of aphA1 was due to the presence of a strong promoter upstream of the structural gene for the enzyme, and resistance resulted from the formation of a complex between the protein and tobramycin in a stoichiometric manner (28). Thus, trapping of tobramycin by the enzyme could account for the reproducible correlation between increasing tobramycin MICs and higher aphA1 copy number (Fig. 5). As in E. coli, the resistance achieved, although of clinical relevance, was of intermediate level and reached a plateau at 16 ␮ g/ml (Fig. 5). The saturable intracellular sequestration of the drug suggests that the binding between tobramycin and Aph(3=)-I is not very tight. Our observations are compatible with the notion that a tobramycin concentration of 4 ␮g/ml represents a crude threshold: at higher concentrations, the aphA1 copy number ap- peared to coalesce between ca. 30 and 55 in isolate MRSN 57 and between 60 to 120 copies in MRSN 58. At lower drug concentra- tions, the copy number was maintained at approximately 8 and 15 copies in MRSN 57 and MRSN 58, respectively (Fig. 3).
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Laboratory detection of high level aminoglycoside aminocyclitol resistance in Enterococcus spp

Laboratory detection of high level aminoglycoside aminocyclitol resistance in Enterococcus spp

The resistant strain detected by microtiter only was susceptible by both methods on initial testing, kanamycin resistant on repeat testing, and resistant to synergistic killing by penici[r]

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Prevalence of Multidrug Resistant Enterococci in a Tertiary Care Hospital in India: A Growing Threat

Prevalence of Multidrug Resistant Enterococci in a Tertiary Care Hospital in India: A Growing Threat

Introduction: Enterococci are members of the healthy human intestinal flora, but are also leading causes of highly antibiotic-resistant infections. Serious enterococcal infections are often difficult to treat since the organisms have a tremendous capacity to acquire resistance to penicillin, high concentration of aminoglycoside & vancomycin. Careful review of in vitro susceptibility data is required to treat infections caused by MDR Enterococci. Therefore we conducted the study to find out prevalence of MDR Enterococci. Aims & Objectives: To study the prevalence of Vancomycin re- sistance, High Level Streptomycin Resistance (HLSR) & High Level Gentamicin Resistance (HLGR) in different enterococcal isolates. Materials & Methods: Total 180 enterococcal isolates were stu- died. Identification was done by conventional biochemical methods. Antibiotic susceptibility test- ing was done by Kirby-Bauer disc diffusion method on Mueller–Hinton agar and results were in- terpreted as per CLSI guidelines. HLSR & HLGR was determined by disc diffusion method using high level Gentamicin disc (120 µg) & Streptomycin (300 µg) discs. Minimum inhibitory concen- tration (MIC) determination for Vancomycin was done by vancomycin E test strips. Results: Total 180 entetococcal isolates were studied. E. faecalis was 60%, E. faecium was 32.2%, E. durans and E. raffinosus were 4.4% & 3.3% respectively. Enterococcus fecium showed resistance in high percen- tage as compared to E. faecalis. 15 isolates were found to be vancomycin resistant. Conclusion: Re- sistance to aminoglycoside is of great concern. Regular screening of enterococcal isolates for van- comycin resistance detection should be implemented. It is very important to implement infection control measures, screening of health care workers, surveillance cultures in intensive care units which can control spread of multidrug resistant enterococci.
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A solution to combat aminoglycoside and quinolone resistant  gram negative organisms

A solution to combat aminoglycoside and quinolone resistant gram negative organisms

A very high level of resistance to monotherapies were observed where cefepime was 55%-74%, tobramycin 20.8%- 70.2%, gentamycin 40%-65.6%, levofloxacin 32.9%-62.4%, moxifloxacin 40.1%-47.7%, ofloxacin 29.9%-48.6%, imipenem plus cilastatin 30%-60%, ciprofloxacin 52.3%- 75.7%, ceftazidime 50.3%-60%, azithromycin 40.5%-86%, and amikacin 40.4%-79.7% among all isolated Gram negative bacterial pathogens. Potentox (cefepime+amikacin) showed very low resistance (8.2%-13.4%) among all Gram-negative pathogens (Figure 2). Khalili et al. (2012) observed the resistance rate of Gram- negative bacilli to cefepime 60, 67.9, 37.9 and 50% in 2007, 2008, 2009 and 2010, respectively in Iran. Other studies observed 51 to 84 % resistance to cefepime in Gram-negative organisms (Jazani et al., 2010; De Macedo and Santos, 2005). A similar study performed by Aliakbarzade et al. (2014) reported that Acinetobacter spp. showed differential resistance pattern to aminoglycosides i.e. gentamicin (86%), tobramycin (63%) and amikacin (81%). Resistance to gentamicin and ciprofloxacin against E. coli observed in this study was comparable to a study by Yismav et al. (2010). Rajat et al. (2012) reported that P. aeruginosa isolated from various samples are resistant to tobramycin (68%) followed by gentamycin (63%), ciprofloxacin (49%) and ceftazidime (43%). Mandal et al. (2010) also showed that 44.5% of Pseudomonas spp. isolates were found to be resistant to ceftazidime which is similar to that reported in the current study where 50.3% P. aeruginosa isolates were resistant to ceftazidime. In our study when the activity of cefepime/amikacin (Potentox) was compared with that of eleven other antibiotics including cefepime and amikacin alone, the susceptibility for all pathogens were significantly higher for Potentox.
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Rapid Aminoglycoside NP Test for Rapid Detection of Multiple Aminoglycoside Resistance in Enterobacteriaceae

Rapid Aminoglycoside NP Test for Rapid Detection of Multiple Aminoglycoside Resistance in Enterobacteriaceae

However, plasmid-mediated 16S rRNA methyltransferases conferring a high level of resistance to multiple AG is also reported (4), and they are identified at a high frequency, especially among producers of NDM-like carbapenemases (5). Enzymes encoded by these genes methylate the aminoglycoside tRNA recognition site (A-site) of the 16S rRNA, which is the intracellular target of the AG. The 16S rRNA methylases identified in Enterobacteriaceae are ArmA, RmtB to RmtF, and NpmA, with ArmA being the most frequently identified methylase (4). Those methylases exhibit a significant amino acid heterogeneity, sharing similar amino acid motifs in their active sites. They confer resistance to almost all AG (amikacin, gentamicin, tobramycin, and kanamycin,
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High incidence of virulence determinants, aminoglycoside and vancomycin resistance in enterococci isolated from hospitalized patients in Northwest Iran

High incidence of virulence determinants, aminoglycoside and vancomycin resistance in enterococci isolated from hospitalized patients in Northwest Iran

Aminoglycosides alone are considered inactive in the treatment of enterococcal infections and are usually com- bined with inhibitors of cell wall synthesis such as vanco- mycin or ampicillin [6]. High-level aminoglycoside-resistant (HLAR) and vancomycin-resistant enterococci (VRE) have created serious problems for antibiotic therapy [6]. Vanco- mycin-resistant enterococci are more common in North America, Europe, and Asia. Eight genotypes (vanA, vanB, vanC, vanD, vanE, vanG, vanM and vanL) have been de- scribed, of which, vanA (Tn1546) genotype with acquired inducible resistance to vancomycin and teicoplanin and vanB (Tn1549/Tn5382) genotype with variable resistance to vancomycin and susceptibility to teicoplanin are the most common [7].
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Identification of clinically antibiotic resistant genes Aac(3)-IIa and Aac(6’)-Ib in wastewater samples by multiplex PCR

Identification of clinically antibiotic resistant genes Aac(3)-IIa and Aac(6’)-Ib in wastewater samples by multiplex PCR

Background: Aminoglycoside antibiotics are widely used in medical centers, particularly to treat infections. The resistance developed against these agents is a huge concern in health care. A number of researchers have reported that hospital and municipal wastewaters are among the most important dissemination sources of these agent into the environment. Some, however, do not agree with this opinion. In the present study, the prevalence of aminoglycoside resistance genes was investigated in raw and effluent wastewater from hospital and municipal wastewater treatment plants.
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Mechanism of High-Level Daptomycin Resistance in Corynebacterium striatum

Mechanism of High-Level Daptomycin Resistance in Corynebacterium striatum

ABSTRACT Daptomycin, a last-line-of-defense antibiotic for treating Gram-positive infections, is experiencing clinical failure against important infectious agents, includ- ing Corynebacterium striatum. The recent transition of daptomycin to generic status is projected to dramatically increase availability, use, and clinical failure. Here we confirm the genetic mechanism of high-level daptomycin resistance (HLDR; MIC ⫽ ⬎ 256 ␮ g/ml) in C. striatum, which evolved within a patient during daptomycin ther- apy, a phenotype recapitulated in vitro. In all 8 independent cases tested, loss-of- function mutations in phosphatidylglycerol synthase (pgsA2) were necessary and suf- ficient for high-level daptomycin resistance. Through lipidomic and biochemical analysis, we demonstrate that daptomycin’s activity is dependent on the membrane phosphatidylglycerol (PG) concentration. Until now, the verification of PG as the in vivo target of daptomycin has proven difficult since tested cell model systems were not viable without membrane PG. C. striatum becomes daptomycin resistant at a high level by removing PG from the membrane and changing the membrane com- position to maintain viability. This work demonstrates that loss-of-function mutation in pgsA2 and the loss of membrane PG are necessary and sufficient to produce high- level resistance to daptomycin in C. striatum.
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Staphylococcus aureus adaptation to aerobic low-redox-potential environments: implications for an intracellular lifestyle

Staphylococcus aureus adaptation to aerobic low-redox-potential environments: implications for an intracellular lifestyle

aureus aerobic respiration was impaired, the intracellular 26 NADH:NAD+ ratio increased, lactate dehydrogenase was induced, resistance to the 27 aminoglycoside antibiotic gentamicin was [r]

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A Novel aadA Aminoglycoside Resistance Gene in Bovine and Porcine Pathogens

A Novel aadA Aminoglycoside Resistance Gene in Bovine and Porcine Pathogens

I n the last decade, multidrug-resistant (MDR) bacterial isolates of the bovine respira- tory disease (BRD) complex have emerged that harbor mobile genetic elements (MGEs) containing genes that confer resistance to many veterinary antimicrobials (1). Widespread resistance in BRD bacterial pathogens—primarily the gammaproteobacte- rial Pasteurellaceae family members Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni (2)—would be devastating to the beef and dairy industries (3). Discovered in the 1940s, aminoglycosides are broad-spectrum antimicrobials; veterinary formulations may include spectinomycin, dihydrostreptomycin, genta- micin, hygromycin B, and neomycin (4, 5). Collectively, they are the fifth-most-used drug class, accounting for 2%, by weight (344,120 kg), of the veterinary antimicro- bials sold (5). Notwithstanding off-label usage, spectinomycin has been withdrawn from therapeutic BRD usage in Canada but continues to be used in the United States and is approved in both countries for in-feed prophylaxis for chickens and swine, often in combination with lincomycin (FDA Center for Veterinary Medicine, Health Canada Drug Product Database). Aminoglycoside resistance may involve 16S rRNA target modification or methylation, efflux, decreased permeability, and enzymatic inactivation by aminoglycoside-modifying enzymes (AMEs) (4, 6). The AMEs include
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Tuberculosis: an overview of current literature on types, diagnosis and drug therapy

Tuberculosis: an overview of current literature on types, diagnosis and drug therapy

Pyrazinamide (PZA), a pro-drug, is highly effective against semi-dormant bacilli in an acidic environment (such as the phagosome). PZAis activated by pyrazinamidase (PncA) to form pyrazinoic acid, which inactivates a vital stepin fatty acid synthesis. PZA also inhibits protein translation and the ribosome-sparing process of translation by binding to ribosomal protein S1 (RpsA) in M. tuberculosis. Most (68-95%) PZA-resistant M. tuberculosis strains contain mutations in pncA or rpsA. Ethambutol (EMB) inhibits the synthesis and poly- merization of cell wall arabinan, which leads tothe accumulation of free mycolic acids and incom-plete cell wall assembly. EMB mainly interacts with membrane- associated arabinosyltransferases encoded by three contiguous genes (embCABoperon) as an arabinose analog.7 The molecular basis of resistance to EMB is not completelydefined. Resistance-conferring mutations in three emb genes have been identified in M. tuberculosis strains.
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