M. Caniça,1 N. Mendonça,1 M. Costa,2 B. Coelho Baptista,2 D. Louro,1 J. Duarte-Correia,2 and C. Pomba2
Emergence of resistance to fluoroquinolones became a concerning problem since their introduction in therapy. The aim of this study was to elucidate the fluoroquinolone resistance mechanisms among uropathogenic E. coli strains isolated from dogs and cats.
A total of 182 E. coli strains were isolated from urine specimens collected from dogs (n=152) and cats (n=30) with urinary tract infection (UTI). Strains were identified using the API 20E system. Minimal Inhibitory Concentrations (MIC) were determined by broth microdilution with enrofloxacin (ENR), marbofloxacin (MAR), orbifloxacin (OBX) and ciprofloxacin (CIP), according to NCCLS guidelines. The genotype of gyrA, gyrB, parC and parE genes of thirty-six fluoroquinolone resistant strains was determined by PCR and nucleotide sequencing.
Cumulative MIC values analysis demonstrated that MIC50and MIC90 (µg/ml) values of ENR, MAR, OBX and CIP were 0.031/64, 0.031/32, 0.062/128 and 0.015/32, respectively forfeline E. coli strains. MIC50and MIC90 (µg/ml) values of ENR, MAR, OBX and CIP were 0.031/16, 0.031/16, 0.125/64 and 0.031/8, respectively for canine E. coli strains. The frequency of resistance was higher in feline strains (20% for all tested quinolones) than in canine strains (14.4% for both ENR and MAR, 15% for both OBX and CIP). Among genotyped strains 5 groups were defined: (1) 14 strains with low level resistance (Susceptible to CIP and MAR, intermediate to OBX and ENR) and only one point mutation (at nucleotide 83 of gyrA gene); (2) 2 strains with intermediate level of resistance (Susceptible to MAR, intermediate to ENR, and resistant to OBX and CIP) and two point mutations (at nucleotide 83 of gyrA gene and at nucleotide 80 of parC gene); (3) 12 strains with high level resistance (resistant to CIP, MAR, OBX and ENR) and 3 point mutations (at nucleotides 83 and 87 of gyrA gene and at nucleotide 80 of parC gene); (4) 5 strains with high level resistance (resistant to CIP, MAR, OBX and ENR) and 4 point mutations (at nucleotides 83 and 87 of gyrA gene and at nucleotides 80 and 57 or 80 and 84 of parC gene); (5) 3 strains with high level resistance (resistant to CIP, MAR, OBX and ENR) and no point mutations due to an unknown mechanism.
This study shows that high level resistant strains have identical pattern of resistance (resistance to all fluoroquinolones studied) and high number of mutations in gyrAand parC genes. Strains with low or intermediate level of resistance are also of high concern due to possible point mutations accumulation under antimicrobial selective pressure. Overall, our results provide rational explanation for the fluoroquinolone treatment failures that occur in small animals with UTI, which highlights the need of surveillance of resistance among this family of antibiotics.