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BACKGROUND: The aim of the present study was to determine the frequency of six efflux pump genes in Acinetobacter clinical isolates collected from South Korean hospitals. METHODS: In this study, we used a total of 339 Acinetobacter strains, comprising 279 Acinetobacter calcoaceticus–Acinetobacter baumannii (ACB) complex and 60 non-ACB complex strains. We performed specific PCR assays to detect adeG, adeB, adeE, adeY, abeM, and adeJ, transporter genes of the multidrug efflux pumps AdeFGH, AdeABC, AdeDE, AdeXYZ, AbeM, and AdeIJK, respectively. RESULTS: Frequencies of six efflux pump genes varied according to the species of Acinetobacter. Frequencies of adeE, abeM, and adeJ between A. baumannii group and A. nosocomialis group were found to be significantly different. Significant differences were found in the frequencies of adeB, adeE, adeY, and adeJ among the susceptible A. baumannii (SAB), multidrug-resistant A. baumannii (MDRAB), and extensively drug-resistant A. baumannii (XDRAB) groups within the 154 strains of A. baumannii. The frequencies of efflux pump genes in imipenem-susceptible and imipenem-nonsusceptible groups were significantly different for adeB, adeY, and adeJ. The frequencies of efflux pump genes in ciprofloxacin-susceptible and ciprofloxacin-nonsusceptible groups were significantly different for adeB and adeY. No significant difference was found in the frequency of efflux pump genes among groups sampled from different regions of Korea, across 86 strains of A. baumannii collected in 2012. CONCLUSIONS: The frequencies of six efflux pump genes obtained in this study demonstrate the fundamental epidemiological feature of efflux pump genes in Korean Acinetobacter clinical isolates.
Subject(s)
Acinetobacter , Gene Frequency , Genes, MDR , Korea , Polymerase Chain ReactionABSTRACT
Background & objectives: Overexpression of efflux pumps is a cause of acquired resistance to fluoroquinolones in Acinetobacter baumannii. The present study was done to investigate the presence and overexpression of AdeABC efflux system and to analyze the sequences of AdeR-AdeS regulatory system in ciprofloxacin-resistant A. baumannii isolates. Methods: Susceptibility of 50 clinical A. baumannii isolates to ciprofloxacin, imipenem, ceftazidime, cefepime and gentamicin antimicrobials was evaluated by agar dilution method. Isolates were screened for the evidence of active efflux pump. Isolates were also examined for adeR-adeS and adeB efflux genes by polymerase chain reaction (PCR). The adeR and adeS regulatory genes were sequenced to detect amino acid substitutions. Expression of adeB was evaluated by quantitative reverse-transcriptase PCR. Results: There were high rates of resistance to ciprofloxacin (88%), ceftazidime (88%), cefepime (74%) and imipenem (72%) and less resistance rate to gentamicin (64%). Phenotypic assay showed involvement of active efflux in decreased susceptibility to ciprofloxacin among 16 isolates. The 12.27-fold increase and 4.25-fold increase were found in adeB expression in ciprofloxacin-full-resistant and ciprofloxacin-intermediate-resistant isolates, respectively. Several effective mutations, including A91V, A136V, L192R, A94V, G103D and G186V, were detected in some domains of AdeR-AdeS regulators in the overexpressed ciprofloxacin-resistant isolates. Interpretation & conclusions: The results of this study indicated that overexpression of the AdeABC efflux pump was important to reduce susceptibility to ciprofloxacin and cefepime in A. baumannii that, in turn, could be triggered by alterations in the AdeR-AdeS two-component system. However, gene expression alone does not seem adequate to explain multidrug resistance phenomenon. These results could help plan improved active efflux pump inhibitors.
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Objective To detect the distribution of resistance-nodulation (RND)efflux pump system of Acineto-bacter baumannii (AB),and explore the relationship between its’expression and antimicrobial resistance.Methods Fifty-nine strains of multidrug-resistant AB isolated from clinical specimens in The First Affiliated Hospital of Nan-chang University were identified and performed antimicrobial susceptibility analysis,distribution of RND efflux sys-tem of AB was detected by polymerase chain reaction(PCR),expression of efflux pump genes in different drug-re-sistant phenotypes of AB was compared,relationship between the expression level and drug resistance was analyzed, amplified products of RND efflux system were sequenced.Results Resistance rates of AB to ampicillin/sulbactam, imipenem,gentamicin,ciprofloxacin,and levofloxacin were 93.2%,94.9%,88.1%,96.6%,and 52.5% respec-tively.PCR detection results of efflux pump and integron genes of 59 AB strains revealed that the carrying rates of adeR,adeS,adeB,adeJ,and adeG genes were 81.4%,91.5%,93.2%,100.0%,and 61.0% respectively.The expression of efflux pump genes in different strains was different,expression levels of ade B and adeJ genes among gentamicin,imipenem,ampicillin/sulbactam resistant AB group and non-resistant AB group were significantly dif- ferent (all P＜0.05).There was no mutation or insertion sequence in the base sequences of regulatory genes ade R and ade S of adeABC efflux pump.Conclusion RND efflux pump system is universally present in AB,the expres-sion upregulation of ade B and ade J genes in RND efflux pump system is related with antimicrobial resistance of bacteria to gentamycin,imipenem,and ampicillin-sulbactam.
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Background: The adeB gene in Acinetobacter baumannii regulates the bacterial internal drug effluxpump that plays a significant role in drug resistance. The aim of our study was to determine theoccurrence of adeB gene in multidrug resistant and New Delhi metallo-beta-lactamase-1 (NDM-1) gene in imipenem resistant Acinetobacter baumannii isolated from wound swab samples in atertiary care hospital of Bangladesh. Methods: A total of 345 wound swab samples were testedfor bacterial pathogens. Acinetobacter baumannii was identified by culture and biochemical tests.Antimicrobial susceptibility pattern was determined by the disc diffusion method according toCLSI standards. Extended spectrum beta-lactamases were screened using the double disc synergytechnique. Gene encoding AdeB efflux pump and NDM-1 were detected by Polymerase ChainReaction (PCR). Results: A total 22 (6.37%) Acinetobacter baumannii were identified from 345wound swab samples and 20 (91%) of them were multidrug resistant. High resistance rates to someantibiotics were seen namely, cefotaxime (95%), amoxyclavulanic acid (90%) and ceftriaxone (82%).All the identified Acinetobacter baumannii were sensitive to colistin and 82% to imipenem. Two(9%) ESBL producing Acinetobacter baumannii strains were detected. adeB gene was detectedin 16 (80%) out of 20 multidrug resistant Acinetobacter baumannii. 4 (18%) of 22 Acinetobacterbaumannii were imipenem resistant. NDM-1 gene was detected in 2 (50%) of the imipenem resistantstrains of Acinetobacter baumannii. Conclusion: The results of this study provide insight into the roleof adeB gene as a potential regulator of drug resistance in Acinetobacter baumanni in Bangladesh.NDM-1 gene also contributes in developing such resistance for Acinetobacter baumannii.
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Objetivo: Determinar las mutaciones delgen gyrA asociadas con la resistencia a fluoroquinolonas en Acinetobacter baumannii. Materiales y métodos: Entre agosto de 2005 y febrero de 2007 se recolectaron 23 aislamientos de A. baumannii de una clínica privada de tercer nivel de Montería. Se investigaron los genes gyrA, parC y adeB; este último codifica para la bomba de salida. Se realizó secuenciación del ADN y, para elanálisis de las secuencias, se usaron la base de datos GenBank y el motor de búsqueda BLASTX. Resultados: La amplificación del gen gyrA en aislamientos de A. baumannii generó unfragmento de 343 pb, el cual presentó pérdida del sitio de restricción con la enzima Hinfl en 12/23 (52,1%) de los aislamientos resistentes a fluoroquinolonas. La secuenciacióndel fragmento mostró mutación puntual con el cambio de Ser-83 a Leu, código de acceso GenBank EU886740. No se encontraron mutaciones en el gen parC, ni la presencia de la bomba de salida Ade. Conclusión: Los aislamientos de A. baumannii resistentes a fluoroquinolonas sugieren que la mutación del gen gyrA que codifica elcambio del aminoácido serina a leucina en el codón 83 de estos aislamientos, es responsable o, al menos, contribuye con la resistencia expresada a las fluoroquinolonas.
Objective: Determine mutations in thegyrA gene associated to resistance tofluoroquinolones in A. baumannii. Materials and methods: From August,2005 to February, 2007, 23 A. baumanniiisolates were collected in a third level private clinic in Monteria. Research on gyrA, parC and AdeB genes was carried out, and the latter encodes for the efflux pump. DNA sequencing was performed, and the GenBank database and BLASTX search engine were used for sequence analysis. Results: Amplification of the gyrA gene in A. baumannii isolates generated a 343pb fragment which presented loss of restriction site with the enzyme HinfI in 12/23 (52.1%) of the isolates resistant to fluoroquinolones. The fragment sequencing showed mutationcharacterized by the change of Ser-83 to Leu GenBank acces code EU886740. None of the isolates showed mutations in the parC gene or presence of the AdeB efflux pump. Conclusion: The A. baumannii isolates resistant to fluoroquinolones suggest thatmutation of the gyrA gene encoding the serine amino acid change to leucine at codon 83 of these isolates is responsible or at least contributes to the mentioned resistance to fluoroquinolones.