Detection, Inhibition and Molecular Analysis of Multidrug Resistant Aerobic Gram-negative Clinical Isolates from a Tertiary Hospital in Nigeria

Abstract

The challenge of combating the ever emerging multi-drug resistant (MDR) clinical isolates in the face of a slow rate of discovery 
of new classes of antibiotics is a problem in antibiotic chemotherapy. This study was aimed at (i) linking phenotypic antibiotic 
drug-resistance characteristics detected in randomly-sampled clinical isolates with detectable genetic markers. (ii) screening a 
suspected efflux pump inhibitor (EPI) [1-(3-(trifluoromethyl)benzyl]-piperazine (TFMBP)], which could be helpful in 
combating this challenge. Fifty-one isolates; 28 Klebsiella pneumonia, 3 E. Coli, 1 Enterobacter cloacae, 1 E. aerogenes, 5
Proteus mirabilis, 4 Providencia rettgeri, 1 P. stuartii, 1 Serratia liquefaciens, 6 S. odorifera, and 1 Acinetobacter baumannii
obtained from infections of urinary tract, upper respiratory tract, gastrointestinal tract, ear swab, eye swab, and blood culture 
were screened for (i) antibiotic-susceptibility over a range of 11 classes of antibiotics, (ii) β-lactamase production, (iii) ESBL 
production and (iv) Efflux pump activity (EPA) in the presence and absence of 1-[3-(trifluoromethyl) benzyl]-piperazine 
(TFMBP). Molecular analysis was done using DNA extraction by boiling and the randomly-amplified polymorphic DNA 
(RAPD) polymerase chain reaction (PCR) procedure with 2% agarose gel electrophoresis stained with ethidium bromide at 10 
µg/ml and visualized by UV trans-illumination. AmpC β-lactamase (4%) and K1 β-lactamase (5.8%) were detected with no 
carbapenemase producers. AcrA and AcrB marker genes were detected in 12% of the isolates while blaCTX-M (8%) and blaTEM 
(4%) were also detected. Antibiotic resistance due to EPA can be combated with a suitable EPI as demonstrated by TFMBP 
when combined with specific antibiotics.