Presence Of Aminoglycoside-modifying Enzymes Research Articles

Molecular Epidemiology, Antimicrobial Susceptibility, and Clinical Features of Methicillin-Resistant Staphylococcus aureus Bloodstream Infections over 30 Years in Barcelona, Spain (1990-2019).

Methicillin-resistant Staphylococcus aureus bloodstream infections (MRSA-BSI) are a significant cause of mortality. We analysed the evolution of the molecular and clinical epidemiology of MRSA-BSI (n = 784) in adult patients (Barcelona, 1990−2019). Isolates were tested for antimicrobial susceptibility and genotyped (PFGE), and a selection was sequenced (WGS) to characterise the pangenome and mechanisms underlying antimicrobial resistance. Increases in patient age (60 to 71 years), comorbidities (Charlson’s index > 2, 10% to 94%), community-onset healthcare-associated acquisition (9% to 60%), and 30-day mortality (28% to 36%) were observed during the 1990−1995 and 2014−2019 periods. The proportion of catheter-related BSIs fell from 57% to 20%. Current MRSA-BSIs are caused by CC5-IV and an upward trend of CC8-IV and CC22-IV clones. CC5 and CC8 had the lowest core genome proportions. Antimicrobial resistance rates fell, and only ciprofloxacin, tobramycin, and erythromycin remained high (>50%) due to GyrA/GrlA changes, the presence of aminoglycoside-modifying enzymes (AAC(6′)-Ie-APH(2″)-Ia and ANT(4′)-Ia), and mph(C)/msr(A) or erm (C) genes. Two CC22-IV strains showed daptomycin resistance (MprF substitutions). MRSA-BSI has become healthcare-associated, affecting elderly patients with comorbidities and causing high mortality rates. Clonal replacement with CC5-IV and CC8-IV clones resulted in lower antimicrobial resistance rates. The increased frequency of the successful CC22-IV, associated with daptomycin resistance, should be monitored.

Plazomicin Activity against 346 Extended-Spectrum-β-Lactamase/AmpC-Producing Escherichia coli Urinary Isolates in Relation to Aminoglycoside-Modifying Enzymes.

The activity of plazomicin and clinically relevant aminoglycosides was tested against 346 extended-spectrum-β-lactamase/AmpC-producing Escherichia coli urinary isolates, and the results were correlated with the presence of aminoglycoside-modifying enzymes (AMEs). Data showed that plazomicin was very active against all ESBL/AmpC-producing E. coli urinary isolates. Its activity was not related to the AME genes studied.

Frequency of Aminoglycoside-Modifying Enzymes and ArmA Among Different Sequence Groups of Acinetobacter baumannii in Iran.

We evaluated aminoglycoside resistance in 87 Acinetobacter baumannii strains isolated from four hospitals located in the North West region of Iran and typed them in sequence groups (SGs) using trilocus sequence-based scheme to compare their clonal relationships with international clones. Resistance toward aminoglycosides was assayed by minimum inhibitory concentration (MIC) and presence of aminoglycoside-modifying enzymes (AMEs), and ArmA-encoding genes were evaluated in different SGs. The majority of isolates belonged to SG1 (39%), SG2 (33.3%), and SG3 (12.6%), whereas the remaining ones were assigned to six novel variants of SGs. MIC determination revealed netilmicin as the most and kanamycin as the least active aminoglycosides against all groups. Among the varied SGs, isolates of SG2 showed more susceptibility toward all tested aminoglycosides. APH(3'')-VIa-encoding gene was predominant in SG1 (47%), SG2 (62%), and SG6-9 (100%). However, AAC(3')-Ia (100%) and ANT(2')-Ia (90.9%) were the dominant AMEs in SG3. There was significant association between harboring of aminoglycoside resistance genes and specific aminoglycosides: gene encoded by APH(3')-VIa was allied to resistance against amikacin and kanamycin, whereas ANT(2')-Ia was related to the resistance toward gentamicin and tobramycin in SG2. In SG1, tobramycin resistance was correlated with harboring of AAC(6')-Ib. Screening of armA demonstrated the presence of this gene in SG1 (58.8%), SG2 (10.3%), as well as SG3 (9%). Our results revealed definite correlation between the phenotypes and genotypes of aminoglycoside resistance in different clonal lineages of A. baumannii.

Coexistence of armA and genes encoding aminoglycoside-modifying enzymes in Acinetobacter baumannii

A total number of 104 Acinetobacter baumannii isolates obtained from a teaching hospital were studied for the genetic basis of their resistance to aminoglycosides. The rate of aminoglycoside-resistant A. baumannii was 76.9% (80/104), and 66.25% (53/80) aminoglycoside-resistant isolates were high-level resistant to either aminoglycosides including amikacin, gentamicin and tobramycin (MIC≥512μg/ml). Genes encoding aminoglycoside-modifying enzymes including ant(2″)-Ia, aac(6′)-Ib, aph(3′)-Ia, aac(3)-Iaand aph(3′)-VI, and 16S ribosomal RNA (rRNA) methylase armA, rmtA, rmtB, rmtC, rmtD,rmtE and npmA were detected by polymerase chain reaction (PCR), armA gene waspresent in 50% (52/104) of A. baumannii isolates with high-level resistance to aminoglycosides (MICs≥512 μg/ml). No rmtA, rmtB, rmtC, rmtD, rmtE or npmA genes were found. The rates of ant(2″)-Ia, aac(6′)-Ib, aph(3′)-Ia, aac(3)-Ia and aph(3′)-VI were 68.25%,20%,70%,47.5% and 1.25%, respectively. armA, ant(2″)-Ia and aph(3')-Ia genes coexisted in 62.5%(50/80) of aminoglycoside-resistant isolates, armA, ant(2″)-Ia and aac(6′)-Ib were commonly positive in same isolates. The presence of aminoglycoside-modifying enzymes was responsible for the moderate level resistance to aminoglycosides in A. baumannii, whereas armA was responsible for the high-level resistance to aminoglycosides. Aminoglycoside-resistant A. baumannii isolates were classified into genotypes by Random Amplified Polymorphic DNA PCR (RAPD-PCR), pattern A was almost throughout wards of the hospital. In summary, most A. baumannii is resistant to aminoglycosides, in the hospital coexistence of armA and genes encoding aminoglycoside-modifying enzymes are the main reasons. Key words: Acinetobacter baumannii, aminoglycoside-resistant, 16S rRNA methylase, aminoglycoside-modifying enzymes, RAPD-PCR

Correlation of Antimicrobial Resistance with β-Lactamases, the OmpA-Like Porin, and Efflux Pumps in Clinical Isolates of Acinetobacter baumannii Endemic to New York City

Acinetobacter baumannii strains resistant to all beta-lactams, aminoglycosides, and fluoroquinolones have emerged in many medical centers. Potential mechanisms contributing to antimicrobial resistance were investigated in 40 clinical isolates endemic to New York City. The isolates were examined for the presence of various beta-lactamases, aminoglycoside-modifying enzymes, and mutations in gyrA and parC. Expression of the genes encoding the beta-lactamase AmpC, the efflux systems AdeABC and AbeM, and the OmpA-like porin was also examined by real-time reverse transcription-PCR. No VIM, IMP, KPC, OXA-23-type, OXA-24-type, or OXA-58 beta-lactamases were detected, although several isolates had acquired bla(SHV-5). Most cephalosporin-resistant isolates had increased levels of expression of ampC and/or had acquired bla(SHV-5); however, isolates without these features still had reduced susceptibility to cefepime that was mediated by the AdeABC efflux system. Although most isolates with ISAba1 upstream of the bla(OXA-51)-like carbapenemase gene were resistant to meropenem, several remained susceptible to imipenem. The presence of aminoglycoside-modifying enzymes and gyrase mutations accounted for aminoglycoside and fluoroquinolone resistance, respectively. The increased expression of adeABC was not an important contributor to aminoglycoside or fluoroquinolone resistance but did correlate with reduced susceptibility to tigecycline. The expression of abeM and ompA and phenotypic changes in OmpA did not correlate with antimicrobial resistance. A. baumannii has become a well-equipped nosocomial pathogen; defining the relative contribution of these and other mechanisms of antimicrobial resistance will require further investigation.

Aminoglycoside resistance in Mycobacterium kansasii, Mycobacterium avium-M. intracellulare, and Mycobacterium fortuitum: are aminoglycoside-modifying enzymes responsible?

Aminoglycoside acetyltransferase was detected in Mycobacterium kansasii and M. fortuitum but not in M. avium-M. intracellulare when they were screened by a radioassay. Aminoglycoside phosphotransferase and nucleotidyltransferase activities were absent from all three species tested. Acetyltransferases from both M. kansasii and M. fortuitum displayed relatively high K(m)s, all at the millimolar level, for substrates including tobramycin, neomycin, and kanamycin A. The K(m) of each substrate was well above the corresponding maximum achievable level in serum. The low affinities of these enzymes for their substrates suggested that drug modification in vivo was very unlikely. Among the various substrates tested, no apparent positive correlation was found between substrate affinity and resistance level. The presence of aminoglycoside-modifying enzymes in these mycobacterial species was therefore not shown to confer resistance to aminoglycosides.

A simple method for the identification of aminoglycoside-modifying enzymes.

In order to study the correlation between the presence of aminoglycoside-modifying enzymes in bacteria and the susceptibility of these bacteria to aminoglycosides, 133 resistant strains were collected, representing the most frequently occurring modifying enzymes in clinical isolates today. Enzymes in these resistant strains were identified by the determination of substrate profiles for eight different aminoglycosides in vitro. Thirteen different enzymes or combinations of enzymes appeared to be present in this collection, whereas in seven cases the resistance appeared to be non-enzyme-mediated. The enzyme activities were not reflected in the bacterial susceptibility data for each antibiotic. Cluster analysis of the combined sensitivity data, with biochemical enzyme analysis as a guideline, proved to be unsatisfactory for the identification of aminoglycoside-modifying enzymes in clinical isolates. Much better results were obtained with a stepwise determination scheme. This method relies upon inhibition zone diameters around commercially available sensitivity discs for six aminoglycoside antibiotics. These zone diameters are compared with empirically established critical values, which are characteristic of an enzyme or group of enzymes. By this procedure proper identification of the enzyme(s) proved to be possible in 84% of the 133 resistant strains. For the evaluation of the method 100 consecutively isolated aminoglycoside resistant clinical isolates were analysed by means of the stepwise scheme and the biochemical method. Results were identical for 97 of the 100 strains.