Detection of gyrA and parC Mutations and Prevalence of Plasmid-Mediated Quinolone Resistance Genes in Klebsiella pneumoniae

Abstract

Background and Aim Recently, the extensive use of quinolones led to increased resistance to these antimicrobial agents, with different rates according to the organism and the geographical region. The aim of this study was to detect the resistance rate of Klebsiella pneumoniae Iraqi isolates toward quinolone antimicrobial agents, to determine genetic mutations in gyrA and parC , to screen for efflux-pump activity, and to screen the presence of plasmid-mediated quinolone resistance (PMQR) genes. Methods Forty-three K. pneumoniae isolates were confirmed phenotypically and genotypically by Vitek 2 system and species specific primers by PCR using the targeting rpo gene followed by sequencing. Antibiotic susceptibility test was carried out using disc diffusion method. Quinolone resistant isolates were subjected to ciprofloxacin MIC testing, and cartwheel method to screen for efflux pump activity. The presence of the plasmid mediated quinolone resistance genes qepA, qnrB, qnrS , and aac(6)Ib was tested by PCR. Sequencing of gyr A and par C was performed. Results We observed a high rate of resistance to ceftriaxone, gentamicin ciprofloxacin, and levofloxacin. Low rate of resistance was detected against amikacin and azithromycin. Ciprofloxacin MIC results revealed that 96.1% of the isolates had MICs >256 µg/mL, 83.4% had MICs >512 µg/mL while 34.6% had MIC >1024 µg/mL. Testing of isolates against ciprofloxacin mixed with EtBr at various concentrations resulted in decreased resistant. Sequencing results showed that Ser83Leu was the most common mutation in gyr A that was observed in all quinolone resistant isolates, followed by Asp87Asn. Ser80Ile mutation in par C was observed in 77.7% of the tested isolates. The prevalence of PMQR genes was 92.5% aac (6)-Ib , 51.8% qnr B, 40.7% qep A, and 37% qnr S. Conclusion Quinolone resistance is common in K. pneumoniae isolates in Baghdad. The frequent mutation in gyr A and par C, and the presence of PMQR genes is alarming.

Investigation for the Ciprofloxacin Resistance Genes Gyr A and Par C in E. Coli Isolates from Urinary Tract Infected Patients

“…Among 100 isolates four (3.8%) isolates which were showing resistance to Ciprofloxacin by MIC method were sent for molecular detection for Gyr A and Par C gene all 4 isolates were positive for both the genes. This study correlates Kareem et al, (16) the Gyr A gene was detected by PCR.…”

“…The resistance to Cotrimoxazole was 64% and resistance to Ciprofloxacin 60%. In the study of Kareem et al, (16) The antimicrobial agents tested included meropenem, imipenem, tigecycline, Ciprofloxacin, aztreonam, amikacin, levofloxacin, cefoperazone-sulbactam, cefotaxime, cefepime, and trimethoprim/sulfamethoxazole. The highest susceptibility were found to be towards the Cefoperazone-sulbactam 48% and the highest rate of resistance were found to be towards Amikiacin 82% which is least similar to our study.…”

Predictors of mortality in acute pancreatitis complicated with multidrug-resistant Klebsiella pneumoniae infection

“…The crisis of MDR bacterial infection was more frequently occurring in developing countries. K. pneumoniae , as the second most common nosocomial gram-negative bacterium, leads to significantly high all-cause mortality in worldwide [ 10 , 12 , 14 , 25 ]. Carbapenem, amikacin and tigecycline used to be effective and latest antibiotics for MDR- K. pneumoniae treatments, but recent researches revealed that drug-resistance rates of these three antibiotics were significantly increasing which may be associated with overuse during treatments [ 24 , 26 ].…”

“…Breakpoints of minimum inhibitory concentration (susceptible, resistant) for each antibiotics were according to the Clinical Laboratory Standard Institute guidelines: amoxicillin-clavulanate (≤ 8/4, ≥ 32/16 μg/ml), amikacin (≤ 16, ≥ 32 μg/ml), aztreonam (≤ 4, ≥ 16 μg/ml), ciprofloxacin (≤ 0.25, ≥ 1 μg/ml), gentamicin (≤ 4, ≥ 16 μg/ml), imipenem (≤ 2, ≥ 8 μg/ml), meropenem (≤ 2, ≥ 8 μg/ml), levofloxacin (≤ 0.5, ≥ 2 μg/ml), sulfamethoxazole (≤ 2/38, ≥ 4/76 μg/ml), piperacillin-tazobactam (≤ 16/4, ≥ 128/4 μg/ml), ceftriaxone (≤ 8, ≥ 64 μg/ml) and tigecycline (≤ 2, ≥ 8 μg/ml). MDR- K. pneumoniae was defined as the strain non-susceptible to at least one agent in three or more antimicrobial categories [ 10 ].…”

“…Recently, several studies have showed that a rising proportion of AP patients with MDR bacterial infections reached 63% [ 5 – 9 ]. MDR- K. pneumoniae has a strong drug-resistant ability through several mechanisms including target alteration, drug inactivation, decreased cell permeability, and increased efflux pump activity [ 10 – 13 ]. MDR- Klebsiella pneumoniae is found as a dangerous pathogen for hospital-acquired infections because of the limited antibiotic regimens [ 14 – 16 ].…”