Determination of resistance to nalidixic acid - a method for the detection of salmonella with reduced susceptibility to ciprofloxacin
In recent years, in developed countries, the incidence of salmonellosis caused by non-typhoid serotypes of salmonella has increased. In the treatment of salmonellosis, fluoroquinolones have been used successfully, even with infections caused by multidrug-resistant strains. However, there is evidence that in some cases treatment has been ineffective due to the presence of salmonella (including S.typhi) which are resistant to nalidixic acid and have reduced sensitivity to fluoroquinolones. Resistance to nalidixic acid was a kind of indicator of a decrease in sensitivity to fluoroquinolones.
The purpose of this study was to establish whether it was possible to use resistance indicators to nalidixic acid to screen for salmonella with reduced susceptibility to ciprofloxacin. The study included 73 strains of Salmonella microorganisms obtained from patient feces between January 2000 and October 2001. Sensitivity was determined using the disk-to-disk method Mueller-Hinton agar and with using macro-dilutions of antibiotics in the broth. For the staging of the disc diffusion test, standard antibiotic discs (with 5 μg of ciprofloxacin and 30 μg of nalidixic acid) were used (Oxoid, UK). The MICs were determined using the macrodilution method of ciprofloxacin (Bayer, Turkey) and nalidixic acid (Ilsan & Iltas, Turkey) in broth. The strain E. coli (ATCC 25922) was used as a reference strain. The sensitivity assessment was carried out according to NCCLS standards (less than 1 and more than 4 g / l for ciprofloxacin and less than 8 and more than 32 mg / l for nalidixic acid).
The 73 isolates according to the results of macro-dilutions in the broth in accordance with the NCCLS standards were sensitive to ciprofloxacin (MPC less than 1 mg / L), and nine strains were resistant to nalidixic acid (MIC greater than 32 mg / L ).
Based on the sensitivity to nalidixic acid, all strains were divided into two groups. For the first group, including the strains sensitive to nalidixic acid, the MPC of ciprofloxacin varied from 0.25 to 8 mg / L. In the second group, which included 9 strains resistant to nalidixic acid, the MPC ciprofloxacin ranged from 4 μg / L to 32 μg / L.
When determining sensitivity to nalidixic acid (the diameter of the growth suppression zone is less than 13 mm), 8 strains out of 9 containing ciprofloxacin MPC greater than 0.008 μg / L were detected. The sensitivity of this method was 100% and the specificity 98.4%. If a GPA greater than 0.0016 μg / L was taken as the lower limit, the sensitivity and specificity of the method would then be 100% and 95.3%, respectively.
All strains (sensitive and resistant to nalidixic acid) were sensitive to ciprofloxacin in accordance with NCCLS standards (zone diameter greater than 21 mm). Eight of the twelve isolates with a growth inhibition zone of less than 33 mm had a ciprofloxacin GPA greater than 8 μg / L. The sensitivity of this method was 100% and the specificity of 93.8%. If the MPC of ciprofloxacin greater than 0.0016 μg / L was taken as the lower limit, the sensitivity and specificity of the method would be 100% and 91% respectively.
It should be noted that in salmonella resistant to nalidixic acid, the MPC of ciprofloxacin was 3 to 4 times higher than in susceptible individuals.
Thus, determining resistance to nalidixic acid using the disc diffusion method has proven to be an effective means of screening strains with reduced susceptibility to ciprofloxacin.