Mutant Prevention Concentrations of Some Aminoglycoside Antibiotics for Fecal Isolates of Escherichia coli under different Growth Temperatures

Abstract

For optimal efficacy, an antibiotic must achieve and sustain at the site of infection, a concentration that can inhibit growth of the bacteria. However, a bacterial infection may contain subpopulations of mutant variants with reduced susceptibility to the antimicrobial agent. There is a great need to periodically evaluate the mutant prevention concentration (MPC) of antibiotic to provide a basis for altering dosing regimens such that the growth of resistant organisms could be curtailed. To evaluate the mutant prevention concentrations (MPCs) of streptomycin, gentamicin and amikacin for fecal Escherichia coli isolates under different growth temperatures and determine the extent of recovery of resistant mutants at such temperatures. Fifty (50) isolates of E. coli were isolated from stools of patients attending Nasarawa State University Keffi Health Centre in Keffi, Nigeria and identified using standard protocol. Antibiotic minimum inhibitory concentrations (MICs) were determined using macro-broth dilution method of the Clinical and Laboratory Standards Institute (CLSI) with incubation for 24 h at 37°C and 41°C. MIC for 50% (MIC₅₀) and 90% (MIC₉₀) of isolates were then generated from the plot of cumulative frequency curve. MPCs were measured by spreading a series of agar plates containing known aminoglycoside concentrations with approximately 10¹⁰ CFU of E. coli culture and incubated for 48 h at 37°C and 41°C. The lowest aminoglycoside concentration that prevented the growth of resistant colonies was taken as the MPC. MPCs for 50% (MPC₅₀) and 90% (MPC₉₀) of isolates were then generated from the plot of cumulative frequency of the MPCs obtained. MPC/MIC ratios for 50% (MPC₅₀/MIC₅₀) and 90% (MPC₉₀/MIC₉₀) of isolates were also determined. Bacteria surviving (persisting) at MPC were isolated and quantified after 48 h. Statistical analyses of data were done by one-way Analysis of Variance (ANOVA). For each of the drugs, MPC₅₀ and MPC₅₀/MIC₅₀ were the same at both 37°C and 41°C. MPC₅₀ values were: streptomycin (44.2 ?g/ml [?32.0 ?g/ml]); gentamicin (44.2 ?g/ml [?32.0 ?g/ml]); and amikacin (37.4 ?g/ml [?32.0 ?g/ml]); and MPC₅₀/MIC₅₀ ratios for each drug at both temperatures were ? 3. MPC₉₀ and MPC₉₀/MIC₉₀ were the same for each drug at both 37°C and 41°C. MPC₉₀ values were: streptomycin (253.2 ?g/ml [?256.0 ?g/ml]), gentamicin (209.0 ?g/ml [?256.0 ?g/ml]), and amikacin (128.0 ?g/ml); and MPC₉₀/MIC₉₀ ratios for each drug at both temperatures were ? 4. Mutant recoveries at the MPCs of the drugs for 50% of the isolates were significantly (P < 0.05) different both at 37°C (P = 0.0089) and 41°C (P = 0.0011). However, mutant recoveries at the MPCs of the drugs for 90% the isolates were insignificantly (P >0.05) different at 37°C (P = 0.0055) but significantly (P > 0.005) different at 41°C (P = 0.0080). Whether at normal body temperature or at a higher body temperature usually obtained during fever, E. coli selects and enrich for resistant mutants less easily against streptomycin than gentamicin or amikacin. The extent of recovery of mutants however, is higher at the higher temperature, justifying the common practice of administering high dosage of antimicrobial agent at high body temperature during therapy of bacterial disease.

Keywords: Mutant Prevention Concentration, Escherichia coli, Aminoglycoside