Wassermann Tina, Meinike Jørgensen Karin, Ivanyshyn Karolina, Bjarnsholt Thomas, Khademi S M Hossein, Jelsbak Lars, Høiby Niels, Ciofu Oana
Department of Clinical Microbiology, University Hospital Rigshospitalet, Copenhagen, Denmark.
Department of Immunology and Microbiology, Costerton Biofilm Center, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
Microbiology (Reading). 2016 May;162(5):865-875. doi: 10.1099/mic.0.000273. Epub 2016 Mar 7.
Ciprofloxacin is a widely used antibiotic, in the class of quinolones, for treatment of Pseudomonas aeruginosa infections. The immediate response of P. aeruginosa to subinhibitory concentrations of ciprofloxacin has been investigated previously. However, the long-term phenotypic adaptation, which identifies the fitted phenotypes that have been selected during evolution with subinhibitory concentrations of ciprofloxacin, has not been studied. We chose an experimental evolution approach to investigate how exposure to subinhibitory concentrations of ciprofloxacin changes the evolution of P. aeruginosa populations compared to unexposed populations. Three replicate populations of P. aeruginosa PAO1 and its hypermutable mutant ΔmutS were cultured aerobically for approximately 940 generations by daily passages in LB medium with and without subinhibitory concentration of ciprofloxacin and aliquots of the bacterial populations were regularly sampled and kept at - 80 °C for further investigations. We investigate here phenotypic changes between the ancestor (50 colonies) and evolved populations (120 colonies/strain). Decreased protease activity and swimming motility, higher levels of quorum-sensing signal molecules and occurrence of mutator subpopulations were observed in the ciprofloxacin-exposed populations compared to the ancestor and control populations. Transcriptomic analysis showed downregulation of the type III secretion system in evolved populations compared to the ancestor population and upregulation of denitrification genes in ciprofloxacin-evolved populations. In conclusion, the presence of antibiotics at subinhibitory concentration in the environment affects bacterial evolution and further studies are needed to obtain insight into the dynamics of the phenotypes and the mechanisms involved.
环丙沙星是一种广泛使用的抗生素，属于喹诺酮类，用于治疗铜绿假单胞菌感染。此前已研究过铜绿假单胞菌对亚抑制浓度环丙沙星的即时反应。然而，长期表型适应性，即确定在亚抑制浓度环丙沙星作用下进化过程中所选择的适应表型，尚未得到研究。我们选择了一种实验进化方法，以研究与未接触环丙沙星的群体相比，接触亚抑制浓度环丙沙星如何改变铜绿假单胞菌群体的进化。将铜绿假单胞菌PAO1及其高突变突变体ΔmutS的三个重复群体在含有和不含有亚抑制浓度环丙沙星的LB培养基中每天传代，需氧培养约940代，并定期对细菌群体进行取样，保存在−80°C以备进一步研究。我们在此研究了祖先群体（50个菌落）和进化群体（120个菌落/菌株）之间的表型变化。与祖先群体和对照群体相比，在接触环丙沙星的群体中观察到蛋白酶活性和游动能力降低、群体感应信号分子水平升高以及突变亚群的出现。转录组分析表明，与祖先群体相比，进化群体中III型分泌系统下调，而在环丙沙星进化群体中反硝化基因上调。总之，环境中亚抑制浓度抗生素的存在会影响细菌进化，需要进一步研究以深入了解表型动态及其涉及的机制。
