School of life science and engineering, Foshan University, Foshan, China.
Guangdong Key Laboratory for Veterinary Drug Development and Safety evaluation, South China Agricultural University, Guangzhou, China.
PLoS One. 2018 Aug 29;13(8):e0202070. doi: 10.1371/journal.pone.0202070. eCollection 2018.
Mycoplasma gallisepticum is a serious pathogen for poultry that causes chronic respiratory disease in chickens. Increased embryonic mortality, as well as reduced weight gain and egg production have been found in infected chickens, which can lead to considerable economic losses in poultry production. Increased antibiotic resistance compromises the use of tetracyclines, macrolides and quinolones in the farm environment. In the present study, danofloxacin concentrations were simulated below the MIC99, between the MIC99 and MPC (the mutant prevention concentration), and above the MPC in an in vitro dynamic model against M. gallisepticum. The relationship between the simulated danofloxacin pharmacokinetics, pharmacodynamics (PK/PD) parameters and development of resistance for M. gallisepticum was explored based on the available data obtained from various dosing regimens in the in vitro model. Danofloxacin concentration, counts of viable cell and susceptibility were determined during the experiment. The mutations in gyrA, gyrB, parC and parE as well as efflux pumps were examined. The MIC of danofloxacin against M. gallisepticum was increased when drug concentrations were between the lower and upper boundaries of the mutant selection window. The upper boundary of the selection window in vitro was estimated as a Cmax/MPC value of 1. The lower boundary was estimated as Cmax/MPC value of 0.05. Both in terms of the MIC and resistance frequency, M. gallisepticum resistance was developed when danofloxacin concentrations fell inside the mutant selection window (ratios of Cmax to MPC between 0.05 and 1). The single mutation in gyrA (Ser-83→Arg) was found in all mutants, while double mutations in gyrA and parC (Ala-64→Ser) were observed only in the mutant with the highest MIC. In addition, no change of susceptibility in the mutants was observed in the presence of reserpine and carbonyl cyanide 3-chlorophenylhydrazone (CCCP). This suggested that ATP-binding cassette superfamily (ABC transporter) and major facilitator superfamily (MFS transporter) did not play a role in danofloxacin efflux.
鸡毒支原体是一种严重的家禽病原体，可导致鸡慢性呼吸道疾病。在感染鸡中发现胚胎死亡率增加，以及体重增加和产蛋量减少，这可能导致家禽生产中巨大的经济损失。抗生素耐药性的增加使得在农场环境中使用四环素、大环内酯类和喹诺酮类药物受到限制。在本研究中，在体外动态模型中，模拟了达氟沙星浓度低于 MIC99、MIC99 与 MPC（突变预防浓度）之间以及高于 MPC 对鸡毒支原体的作用。基于从体外模型中各种给药方案获得的现有数据，探索了模拟达氟沙星药代动力学、药效学（PK/PD）参数与鸡毒支原体耐药性发展之间的关系。在实验过程中测定了达氟沙星浓度、活菌计数和药敏性。检测了 gyrA、gyrB、parC 和 parE 基因突变以及外排泵。当药物浓度在突变选择窗的下限和上限之间时，达氟沙星对鸡毒支原体的 MIC 增加。体外选择窗的上限估计为 Cmax/MPC 值为 1。下限估计为 Cmax/MPC 值为 0.05。无论是 MIC 还是耐药频率，当达氟沙星浓度落在突变选择窗内时（Cmax/MPC 比值在 0.05 到 1 之间），鸡毒支原体就会产生耐药性。所有突变体均发现 gyrA 单突变（Ser-83→Arg），而最高 MIC 突变体仅观察到 gyrA 和 parC 双突变（Ala-64→Ser）。此外，在利血平和羰基氰化物 3-氯苯腙（CCCP）存在的情况下，突变体的药敏性没有变化。这表明三磷酸腺苷结合盒超家族（ABC 转运体）和主要易化因子超家族（MFS 转运体）在达氟沙星外排中不起作用。
