EVALUATION OF A METHOD FOR DETECTION OF BOTH METALLO BETALACTAMASE (MBL) AND EFFLUX PRODUCTION USING CARBONYL CYANIDE 3-CHLOROPHENYLHYDRAZONE (CCCP)Â±COMBINATION METHOD AMONG PSEUDOMONAS AERUGINOSA ISOLATED FROM CLINICAL SPECIMENS
Objective: To standardize a method for the differential detection of Metallo Betalactamase (MBL) production and efflux pump mechanism using Carbonyl Cyanide 3-ChloroPhenylhydrazone (CCCP) among the clinical isolates of Pseudomonas aeruginosa recovered from ocular specimens.
Methods: A total of 110 clinical isolates of Pseudomonas aeruginosa were plated onto Muller-Hinton agar (MHA) plate with CCCP and without CCCP. Carbapenems-Ethylenediaminetetraacetic acid (EDTA) combination disk method was performed on CCCPÂ± plates for individual isolates and the results were compared.
Results: Out of 110 clinical isolates of P aeruginosa studied, 11(10%) were tested positive for MBL Production by carbapenem-EDTA combination test/CCCP-test. In which, six isolates showed positive for both efflux and MBL production using CCCP+plate and 5 isolates showed positive only for MBL production.
Conclusion: The current study standardized a method for detecting both MBL and efflux mechanism in a single plate consists of CCCPÂ± plate containing antibiotic disk with and without EDTA. Implication of this method in routine diagnostics will help the clinicians to treat the patient with appropriate drug choice and dosage.
2. Sy A, Srinivasan M, Mascarenhas J, Lalitha P, Rajaraman R, Ravindran M, et al. Pseudomonas aeruginosa keratitis: outcomes and response to corticosteroid treatment. Invest Ophthalmol Visual Sci 2012;53:267-72.
3. Peshattiwar PD, Peerapur BV. ESBL and MBL mediated resistance in Pseudomonas aeruginosa: an emerging threat to clinical therapeutics. J Clin Diagn Res 2011;5:1552-4.
4. Ardebili A, Talebi M, Azimi L, Lari AR. Effect of efflux pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone on the minimum inhibitory concentration of ciprofloxacin in acinetobacter baumannii clinical isolates. Jundishapur J Microbiol 2014;7(1):e8691.
5. Zechini B, Versace I. Inhibitors of multidrug resistant efflux systems in bacteria. Recent Pat Anti-Infect Drug Discovery 2009;4:37-50.
6. Murugan N, Malathi J, Therese KL, Madhavan HN. Antimicrobial susceptibility and prevalence of extended spectrum betalactamase (ESBL) and metallo betalactamase (MBL) and its co-existence among Pseudomonas aeruginosa recovered from ocular infections. Int J Pharm Pharm Sci 2015;7;147-51.
7. Murugan N, Malathi J, Umashankar V, Madhavan HNR. Comparative genomic analysis of multidrug-resistant Pseudomonas aeruginosa clinical isolates VRFPA06 and VRFPA08 with VRFPA07. Genome Announcement 2014;2(2):e00140-14.
8. Aktas Z, Kayacan Ã‡B. Investigation of metallo-beta-lactamase producing strains of Pseudomonas aeruginosa and Acinetobacter baumannii by E-test, disk synergy and PCR. Scand. J Infect Dis 2008;40:320-5.
9. Poonsuk K, Chuanchuen R. Detection of the mex efflux pumps in pseudomonas aeruginosaby using a combined resistance-phenotypic markers and multiplex RT-PCR. Open J Med Microbiol 2014;4:153-60.
10. Georgios M, Egki T, Effrosyni S. Phenotypic and molecular methods for the detection of antibiotic resistance mechanisms in gram negative nosocomial pathogens. Research Gate 2014.
11. Pournaras S, Maniati M, Spanakis N, Ikonomidis A, Tassios PT, Tsakris A, et al. Spread of efflux pump-overexpressing, non-metallo-Î²-lactamase-producing, meropenem-resistant but ceftazidime-susceptible Pseudomonas aeruginosa in a region with blaVIM endemicity. J Antimicrob Chemother 2005;56:761-4.
12. Andersen JL, He GX, Kakarla P, KC R, Kumar S, Lakra WS, et al. Multidrug efflux pumps from Enterobacteriaceae, Vibrio cholerae and Staphylococcus aureus bacterial food pathogens. Int J Environ Res Public Health 2015;12:1487-547.
13. Park S, Lee KM, Yoo YS, Yoo JS, Yoo JI, Kim HS, et al. Alterations of gyrA, gyrB, and parC and activity of efflux pump in fluoroquinolone-resistant Acinetobacter baumannii. Osong Public Health Research Perspectives 2011;2:164â€“70.
14. Rajamohan G, Srinivasan VB, Gebreyes WA. Novel role of Acinetobacter baumannii RND efflux transporters in mediating decreased susceptibility to biocides. J Antimicrob Chemother 2010;65:228â€“32.