A study on the molecular basis of quinolone resistance mechanism in salmonella typhi

Objective To study the relationship between the gene mutations of DNA gyrase subunit A (gyrA) and quinolone resistance in Salmonella typhi. Methods The genes of gyrA DNA of Salmonella typhi S275 (a clinically isolated quinolone susceptible strain) and its spontaneous quinolone resistant mutant RG 1 were examined in this study with polymerase chain reaction (PCR), restrictive fragments length polymorphism (RFLP), single strand conformational polymorphism (SSCP) and nucleotide sequencing. Results Nudeotide sequencing of gyrA in Salmonella typhi S275 revealed that the bases of 128～426 kept highly conservative as compared with those of Escherichia coli KL 16, with only 7.49% difference in the gyrA nucleotides 128～426 between the two strains. Most of the mutations were silent mutations,which contributed to 3 amino acid substitutions in gyrase (including Thr 45→His,Arg 49→Leu and Val 56→Gly), and all these substitutions were located outside the quinolone resistance determining region (amino acids 67 106 of subunit A of gyrase). In comparison with Salmonella typhi S275, a single mutation was found at base 247 of gyrA of Salmonella typhi RG 1, with change transferred from T to G and led to a substitution of Ser 83→Ala. The mutation might be responsible for the increase of MICs of nalidixic acid, ofloxacin and ciprofloxacin against Salmonella typhi from 2,0.06 and 0.03 to 512, 2, and 1 mg/L respectively. Ser 83→Ala was also a newly discovered substitution in gyrA of Salmonella spp. The results of PCR RFLP and SSCP were in concordance with results of nucleotide sequencing. Conclusions The mutation of gyrase at the 83rd amino acid maybe play a principal role in the resistance of Salmonella typhi to quinolone.