A study on gyrA mutant in Levofloxacin-resistant Brucella melitensis induced resistant strains in vitro / 中国地方病学杂志

ABSTRACT

Objective To study on the drug-resistance mechanism of Brucella resistance to Quinolone antibiotics to guide the selection and use of antimicrobial agents in clinical practice. Methods Six strains of Brucella melitensis(Bru1, Bru2, Bru3, Bru4, Bru5, Bru6) were selected to be induced resistance to levofloxacin in vitro respectively. The MICs of the 6 strains of Brucella melitensis and induced resistant strains were measured by agar dilution method. The sensitivity to Quinolone antibiotics (Levofloxacin, Ciprofloxacin, Lomefloxacin, Norfloxacin, Fleroxacin, Ofloxaein) of 6 strains of Brucella melitensis and induced resistant strains was measured by K-B method. The gyrA of the 6 strains of Brucella melitensis and induced resistant strains was amplified by PCR, then the nucleotide sequence of the genes were analyzed. Results The MICs of Bru1,Bru2,Bru3,Bru4, Bru6 were 0.50 μg/ml and Bru5 was 0.25 μg/ml. The strains Bin3, Bru4 were induced into drug-resistant strains by Levofloxacin, then were named LEVR3 and LEVR4 respectively. The MICs of LEVR3 and LEVR4 were 64,128 μg/ml with 128 and 256 times higher than that of the parental strains. The 6 strains of Brucella melitensis were sensitive to Quinolone antibiotics, LEVR3 and LEVR4 were resistant to Quinolone antibiotics. Neucleotide sequence analysis and comparison of the derived amino acid sequence revealed that Quinolone resistance-determing region of GyrA had a substitution at position Ala87 and Asp91 in laboratory resistant strains. Conclusion The results of in vitro experiments show that acquired resistance of Brucella melitensis strains to Levofloxacin could beinduced when exposed to high level of some antibacterial agents for short term. Two drug-resistant strains occur mutations in gyrA and have cross-resistance to other Quinolones.