| Objective: To explore the effection of chinese medicine combinated with quinolones on quinolones mutant selection to Staphylococcus Aureus and grl A、grl B、gyr A and gyr B gene mutation of bacteria, to guide clinicians using antibiotics rationally,and provide a new clinical treatment approach to Reduce bacterial drug resistance. Methods : Tissue cage infection models with staphylococcus aureus have been established in rabbits, and the animals have been given saline injection、levofloxacinonly、shuanghuanglian only、low dose Shuang Huang Lian powder injection in combination with levofloxacin、 middle dose Shuang Huang Lian powder injection combined levofloxacin and high dose Shuang Huang Lian powder injection combined levofloxacin based on MSW theory. levofloxacin concentration in tissue cage were in MSW. Bacteria recovered growth condition and different time bacterial colonies in tissue cage infection model have been studied. Different time staphylococcus aureus in tissue cage infection model have been extracted and enriched, Used the SPSS19.0 statistics software to analyze the data.Then DNA have been extracted.Bacteria grl A、grl B、gyr A and gyr B gene have been amplified and sequenced by polymerase chain reaction. Then tested grl A, grl B, gyr A and gyr B gene sequences by the dideoxy termination method of hydrogen and analyzed by DNAMAN software.Bacteria recovered growth condition,different time bacterial MIC、MPC、MSW and grl A、grl B、gyr A、gyr B gene mutation have been Compared in different treatments. Results: Bacterial colonies of the analysis results showed that the number of bacterial colonies differences between groups statistically significant(P = 0.018). Negative control group compared with other groups(P = 0.04 < 0.05) and the negative control groupnumber of bacterial colonies have a colony count rising trend, the rest of the groups have a colony count downward trend, and the differences between the and the negative control group and the rest 0f the groups were statistically significant(P < 0.05). Bacteria MIC analysis results show that: compared with negative control group, low dose Shuang Huang Lian powder injection in combination with levofloxacin, middle dose Shuang Huang Lian powder injection combined levofloxacin, high dose Shuang Huang Lian powder injection combined levofloxacin group(P = 0.047), MIC demonstrated the decline, the difference had statistical significance(P < 0.05). In the three combination drug groups,the MIC reducing trend is more obvious along with the increase of Shuang Huang Lian dose, the difference had statistical significance(P <0.05).Treated by levofloxacin only group compared with negative control group,the MIC is obviously rising trend, the difference was statistically significant(P< 0.05).Treated by Shuang Huang Lian alone group compared with negative control group, the MIC demonstrated a decline after 4 or 5 days, but there was no statistically significant difference(P = 0.422 > 0.05). Bacteria MSW analysis results show that:compared with negative control group,low dose Shuang Huang Lianpowder injection combined levofloxacin group,middledose Shuang Huang Lian powder injection combined levofloxacin group,high dose Shuang Huang Lian powder injection combined levofloxacin group,MSW demonstrated the decline, the difference had statistical significance(P < 0.05).Treaded by levofloxacin only group compared with negative control group, the MSW demonstrated the decline, the difference was statistically significant(P <0.05).In the three combination drug groups, the MSW reducing trend is more obvious along with the increase of Shuang Huang Lian dose, the difference had statistical significance(P ﹤ 0.05). Grl A gene mutation of Staphylococcus Aureusin tissue cage treated by levofloxacin mutated significantly than other experimental groups. Amino sequence of the grl A mutation of staphylococcus aureuswere:Ala(29)'Ser, Ser(32)'Val,Asn(40)'Ala,Gln(54'Asn,Phe(61)'Ser,Asp(73)'Thr,His(78)'Asn,Ile(81)'Val,Ile(90)'Val,Asn(92)'Gly,Pro(95)'Ala,Lys(104)'Arg,Lys(104)'Arg,Leu(107)'Pro,L eu(108)'Ile,Ile(116)'Leu,Ser(123)'Glu,Ile(125)'Val,Pro(126)'Ser,Tyr(128)'Phe,Thr(132)'Ser,Leu(133)'Glu,Ser(139)'Ala,Arg(140)'Ala,Asp(159)'Glu,Asn(165)'His,Tyr(175)'Arg,Asn(178)'Lys.Amino sequence of thegrl B mutation of staphylococcus aureuswas:Tyr(114)'Phe,Leu(115)'MET. Amino sequence of thegyr Amutation of staphylococcusaureuswas:Pro(61)Pro ' Thr.Gyr B gene did not mutated in all experimental groupsand staphylococcusaureus ATCC29213.Conclusions: Shuang Huang Lian combined levofloxacin group than treaded by levofloxacin only, significantly reduced staphylococcus aureus MIC 、 MSW.Within a certain range,the higher concentration of Shuang Huang Lian with levofloxacin combination, the stronger the above role. So the combination of levofloxacin and Shuang Huang Lian can reduce staphylococcus aureus resistant. The production of bacterial drug resistance caused by levofloxacin is associated with point mutations in genes grl A、grl Band gyr Agene, has nothing to do withgyr B gene. Chinese medicine Shuang Huang Lian could inhibit mutation of Staphylococcus Aureusingrl A 、grl Band gyr A genesinduced by levofloxacin. |
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