The prevalence of aminoglycoside-modifying enzyme and virulence genes among enterococci with high-level aminoglycoside resistance in Inner Mongolia, China

ABSTRACT This study highlights the prevalence of aminoglycoside-modifying enzyme genes and virulence determinants among clinical enterococci with high-level aminoglycoside resistance in Inner Mongolia, China. Screening for high-level aminoglycoside resistance against 117 enterococcal clinical isolates was performed using the agar-screening method. Out of the 117 enterococcal isolates, 46 were selected for further detection and determination of the distribution of aminoglycoside-modifying enzyme-encoding genes and virulence determinants using polymerase chain reaction -based methods. Enterococcus faecium and Enterococcus faecalis were identified as the species of greatest clinical importance. The aac(6')-Ie-aph(2")-Ia and ant(6')-Ia genes were found to be the most common aminoglycoside-modifying enzyme genes among high-level gentamicin resistance and high-level streptomycin resistance isolates, respectively. Moreover, gelE was the most common virulence gene among high-level aminoglycoside resistance isolates. Compared to Enterococcus faecium, Enterococcus faecalis harbored multiple virulence determinants. The results further indicated no correlation between aminoglycoside-modifying enzyme gene profiles and the distribution of virulence genes among the enterococcal isolates with high-level gentamicin resistance or high-level streptomycin resistance evaluated in our study.

Mutational and acquired carbapenem resistance mechanisms in multidrug resistant Pseudomonas aeruginosa clinical isolates from Recife, Brazil

An investigation was carried out into the genetic mechanisms responsible for multidrug resistance in nine carbapenem-resistant Pseudomonas aeruginosaisolates from different hospitals in Recife, Brazil. Susceptibility to antimicrobial agents was determined by broth microdilution. Polymerase chain reaction (PCR) was employed to detect the presence of genes encoding β-lactamases, aminoglycoside-modifying enzymes (AMEs), 16S rRNA methylases, integron-related genes and OprD. Expression of genes coding for efflux pumps and AmpC cephalosporinase were assessed by quantitative PCR. The outer membrane proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The blaSPM-1, blaKPC-2 and blaGES-1 genes were detected in P. aeruginosaisolates in addition to different AME genes. The loss of OprD in nine isolates was mainly due to frameshift mutations, premature stop codons and point mutations. An association of loss of OprD with the overexpression of MexAB-OprM and MexXY-OprM was observed in most isolates. Hyper-production of AmpC was also observed in three isolates. Clonal relationship of the isolates was determined by repetitive element palindromic-PCR and multilocus sequence typing. Our results show that the loss of OprD along with overexpression of efflux pumps and β-lactamase production were responsible for the multidrug resistance in the isolates analysed.

The prevalence of aminoglycoside-modifying enzyme genes (aac (6')-I, aac (6')-II, ant (2")-I, aph (3')-VI) in Pseudomonas aeruginosa

INTRODUCTION: Pseudomonas aeruginosa (P. aeruginosa) is one of the primary opportunistic pathogens responsible for nosocomial infections. Aminoglycosides are an import ant component of antipseudomonal chemotherapy. The inactivation of drugs by modifying enzymes is the most common mechanism of aminoglycoside resistance. OBJECTIVES: The inactivation of aminoglycosides by modifying enzymes is the primary resistance mechanism employed by P. aeruginosa. The aim of the present study was to investigate the occurrence of aminoglycoside resistance and the prevalence of four import ant modifying enzyme genes (aac (6')-I, aac (6')-II, ant (2")-I, aph (3')-VI) in P. aeruginosa in Iran. METHODS: A total of 250 clinical isolates of P. aeruginosa were collected from several hospitals in seven cities in Iran. Antimicrobial susceptibility tests (using the disk diffusion method and E-tests) were performed for all 250 isolates. In addition, all isolates were screened for the presence of modifying enzyme genes by polymerase chain reaction. RESULTS: The resistance rates, as determined by the disk diffusion method, were as follows: gentamicin 43 percent, tobramycin 38 percent, and amikacin 24 percent. Of the genes examined, aac (6')-II (36 percent) was the most frequently identified gene in phenotypic resist ant isolates, followed by ant (2")-I, aph (3')-VI, and aac (6')-I. CONCLUSIONS: Aminoglycoside resistance in P. aeruginosa remains a signific ant problem in Iran. Therefore, there is considerable local surveillance of aminoglycoside resistance.

Identificación molecular de enzimas modificantes de aminoglucósidos en cepas de Enterococcus spp. aisladas en hospitales de la Octava Región de Chile / Molecular identification of aminoglycoside-modifying enzymes among strains of Enterococcus spp. isolated in hospitals of the VIII Region of Chile

Background: Infectious diseases produced by Enterococcus spp, must be treated with a synergistic combination between a penicillin and an aminoglycoside. High level resistance to aminoglycosides is a serious therapeutic problem, since it predicts the loss of synergistic activity of this antimicrobial combination. Aim: To investigate the presence of genes encoding aminoglycoside-modifying enzymes (AMEs) among strains of Enterococcus spp with high level of resistance to aminoglycosides. Material and methods: The genes encoding some of the AMEs were investigated among 305 aminoglycoside-resistant strains of Enterococcus spp isolated in hospitals of the VIII region of Chile, by dot blot hybridization and Polymerase Chain Reaction (PCS). Results: High level resistance to some aminoglycosides was observed in 104 strains (34.1 percent) and 93 of these harbored at íeast one of the genes encoding the investigated AMEs. Three genes were detected: aac(6)Ie-aph(2")Ia (14.8 percent) encoding for the enzyme AAC(6)Ie-APH(2")Ia (resistance to all aminoglycosides, except streptomycin); aph(3)IIIa (26 percent), and ant(6)la (28.5 percent) encoding for the phosphorylating enzymes APH(3)Ilia (resistance to kanamycin, amikacin and neomycin), and ANT(6)-la (resistance only to streptomycin), respectively. None of the strains harbored the gene ant (4) which encode for the enzyme ANT (4). Conclusion: The low frequency of strains harbouring the bifunctional enzyme (<15 percent), conferring an extended resistance profile to aminoglycosides, allows us to propose the empirical use of aminoglycoside-aminocyclitols, associated to a penicillin, in the treatment of serious infections produced by species of enterococci.

Resistance due to aminoglycoside modifying enzymes in Pseudomonas aeruginosa isolates from burns patients.

BACKGROUND & OBJECTIVE: Enzymatic modifications of aminoglycosides result in high-level resistance in numerous bacterial species. However, the data on this aspect are elementary in our country. The present study was therefore designed to determine resistance rates and patterns, and to find out the prevalent aminoglycoside modifying enzymes (AMEs) in clinical isolates of Pseudomonas aeruginosa from hospitalized burn patients. METHODS: Forty two, non-repeat, clinical isolates of P. aeruginosa obtained during a period from February to July 2003, were analysed for the presence of antibiotics resistance. On the basis of aminoglycoside susceptibility patterns, resistance phenotypes and possible AMEs were inferred according to interpretative reading. Seven isolates collected during the same period and previously characterized to harbour a 48.5 kb plasmid, encoding multiple drug resistance, were also analysed for aminoglycoside susceptibility patterns, and AMEs encoded by the plasmid were inferred. RESULTS: Ninety six per cent of the isolates were multi drug-resistant and majority (71.4%) were resistant to 5 or more antibiotics. Markedly high resistance to tobramycin (83.6%) and amikacin (55.1%) was noted, whereas gentamicin resistance was present in 32.6 per cent isolates. The enzyme N-acetyl transferases (AAC) viz. AAC(6')-I was the most common isolated AME followed by AAC(3)-II in 42.8 and 20.4 per cent of isolates respectively. The plasmid harbouring isolates belonged to AAC(6') phenotype and the enzyme encoded was inferred to be AAC(6')-I. INTERPRETATION & CONCLUSION: Markedly high resistance to tobramycin and amikacin was noted in the present study. AAC(6')-I was the most common AME and was inferred to be plasmid encoded in R-plasmid harbouring isolates. This is among the premier reports regarding the aminoglycoside resistance due to AMEs especially plasmid encoded, in P. aeruginosa from India. Further studies are required from different parts of the country to findout the prevalence of aminoglycoside resistance due to AMEs in P. aeroginosa isolates.

Resistencia a gentamicina, amikacina y ciprofloxacina en cepas hospitalarias de klebsiella pneumoniae subespecie pneumoniae productoras de beta-lactamasas de espectro extendido / Resistance to gentamicin, amikacin and ciprofloxacin among nosocomial isolates of klebsiella pneumoniae subspecie pneumoniae producing extended spectrum beta-lactamases

BACKGROUND: Klebsiella pneumoniae is a pathogenic bacterium frequently isolated from nosocomial samples, specially the subspecie pneunonlae, with extensive antibiolic resistance profiles, including third generation cepbhalosporiis, aminoglycosides and quinolones. This is specially true for those strains producing extended spectrum beta lactamases (ESBL). AIM: To investigate the susceptibility to gentamicin, amikacin and ciprofloxacin and the presence of some aminogloycoside modifying enzyme (AMEs) among nosocomial strains of K pneumoniae subspecie pneumoniae producing ESBL. MATERIAL AND METHODS: The antibiotic resistant patterns and the level of resistance (minimal inhibitory concentration, MIC) of 100 strains, isoklted from sel ,eal bospitals of dcifferent Chilean cities, were deterl,in,ed. Tbe presence of some aminoglycosides modifying enzyme (AMEs) was investigated by PCR. RESULTS: Sixty five percent of strains were resistant to gentamicin, 47% were resistant to amikacin, and 29% were resistant to ciprofloxacin. The most frequent AMEs genes detected were the aac(6')-Ib gene (6'N-Acetyltransferase type Ib enzyme) in 69% of strains, conferring resistance to amikacin, kanamycin, tobramycin, and nieoniycin, and the gene aac(3)-IIa (3-Acetyltransferase type 3-IIa enzyme), in 36% of strains, conferring resistance to gentamlicin. CONCLUSIONS: Among nosocomial strains of K pneumoniae subspecie pneumoniae isolaterd from Chilean hospitals, there is an association between the production of ESBL and the resistance to others antimicrobial agents, especially aminoglycosides. Nevertheless, 71% of isolates are susceptible to ciprofloxacin.

Spontaneous Kanamycin-resistant Escherichia coli mutant with altered periplasmic oligopeptide permease protein (OPPA) and impermeability to aminoglycosides

A spontaneous kanamycin-resistant Escherichia coli mutant, showing cross resitance to five other aminoglycosides and absence of the OppA protein was isolated. [3H]- dihydrostreptomycin uptake is reduced in this mutant, implying that the oligopeptide transport system in involved in accumulation of aminoglucosides, although apparently not related with aminoglycoside permeability alteration due to bacterial adaptation to osmotic changes.