Determination of Plasmid Profile Multiple Antibiotics Resistant Pseudomonas aeruginosa Isolated from Clinical Specimens in Healthcare Facilities across Anambra and Imo States Nigeria

Abstract

The emergence and dissemination of antibiotic resistant plasmids of P. aeruginosa is posing a major public threat and huge concern in hospital facilities. This study was done across Anambra and Imo States with a total number of 100 P. aeruginosa isolates, 50 from each State to determine the plasmid profile of multidrug resistant isolates. Methods of re-identifying P. aeruginosa were based upon cultural methods coupled with biochemical tests. To study the susceptibility of these isolates using disk diffusion method, seven (7) antibiotics were used namely: Piperacillin–tazobactam (100/10 µg), Ceftazidime (30 µg), Amikacin (30 µg), Ciprofloxacin (5 µg), Cefuroxime (30 µg), Ceftriazone (30 µg) and Gentamicin (10 µg). Plasmid extraction was done using alkaline lysis method after growing isolates resistant to more than four antibiotics on a nutrient broth. Agarose gel electrophoresis of plasmid DNA was carried out on 2% agarose gel slab in 1X TAE buffer. The results show that, the nutrient agar at 37˚C aerobically, P. aeruginosa isolates were recovered, which produced greenish-yellow pigment colonies, oxidase was positive and negative for gram stain. In Anambra, the result showed 100% resistance to Cefuroxime, Ceftazidime and Ciprofloxacin, 90% and 86% for Ceftriaxone and Piperacillin-tazobactem, 48% and 40% to Gentamicin and Amikacin respectively. Whereas in Imo state, the result showed 100% resistance to Ceftazidime and Ciprofloxacin, 80% to Ceftriaxone, Cefuroxime and Piperacillin-tazobactem while the least resistance was seen in Amikacin, and Gentamicin. Plasmid size ranging from 100bp to >1000bp was detected from most of the multidrug resistant isolates. Not all the isolates with multidrug resistance were found to possess plasmids. It can be seen from this study that multidrug resistance in P. aeruginosa is not strictly plasmid-dependent (mediated).