黏菌素压力下mcr-1阳性大肠杆菌适应性演化及其质粒传播的分子机制

Colistin is a last-line drug for treatment of infections caused by multidrug resistant Gram-negative bacteria. The issue of colistin resistance has attracted much attention after the mobile colistin resistance gene mcr-1 was discovered by Chinese scientists in 2015. mcr-1 is mainly located in three plasmids of Escherichia coli including IncI2, IncX4 and IncHI2, and it has been prevalent over 50 countries in six continents so far. Several mcr-1-positive Escherichia coli (MCRPEC) could decrease fitness cost against acquiring mcr-1-positive plasmids (MCRPP), which can facilitate stability and spread of MCRPP. However, the mechanism of compensatory mutation is unknown. The colistin has been banned for using as a growth promotor and can only be used for treatment, which significantly decreased the colistin pressure, however the effect on fitness of MCRPEC and spread of MCRPP under low concentration of colistin needs further analyses. Thus, this project aims to reveal the compensatory mutations of evolved MCRPEC via approaches of bioinformatic, molecular biology and so on. This project also aims to ascertain the effective threshold value of colistin that could impact the fitness of MCRPEC and spread of MCRPP, and clarify the corresponding molecular mechanism. All findings will extend the theory on evolution of MCRPEC and spread of MCRPP.

黏菌素是治疗多重耐药革兰阴性菌感染的“最后一道防线”药物。自2015年我国科学家发现可转移的黏菌素耐药基因mcr-1后，黏菌素耐药性问题备受关注。mcr-1基因主要存在于大肠杆菌中的IncI2、IncX4和IncHI2型质粒上，目前已在全球6大洲超过50个国家中广泛流行。部分mcr-1阳性大肠杆菌（MCRPEC）降低了获得mcr-1阳性质粒（MCRPP）带来的适应性代价，促进了MCRPP的稳定性和播散，但其中的补偿突变机制不明。我国于2017年禁止黏菌素用于动物促生长仅保留治疗用途后黏菌素压力显著下降，然而低浓度的黏菌素对MCRPEC适应性和MCRPP传播的影响还有待解析。因此，本项目拟通过生物信息学和分子生物学等手段揭示MCRPEC演化的补偿突变机制，并明确黏菌素影响MCRPEC适应性和MCRPP传播的作用阈值和相应的分子机制。研究成果可丰富MCRPEC演化和MCRPP传播的理论。

黏菌素是治疗多重耐药革兰阴性菌感染的“最后一道防线”药物。自2015年我国科学家发现可转移的黏菌素耐药基因mcr-1后，黏菌素耐药性问题备受关注。部分mcr-1阳性大肠杆菌（MCRPEC）降低了获得mcr-1阳性质粒（MCRPP）带来的适应性代价，其中的补偿突变机制不明。我国于2017年禁止黏菌素用于动物促生长仅保留治疗用途后黏菌素压力显著下降，然而低浓度的黏菌素对MCRPP传播的影响还有待解析。本项目研究发现X基因缺失后可显著降低mcr-1基因所在质粒的拷贝数进而降低mcr-1基因的转录，减少其适应性代价，并可增强宿主菌的竞争、生物膜形成、抗氧化及细胞黏附功能。研究证明了杀菌浓度的黏菌素（4 µg/mL）通过促进携带mcr-1的IncI2型质粒接合转移系统活性的提升、细菌菌毛产生及细胞膜通透性的增加，从而促进该质粒的接合转移，同时ROS和SOS系统对该过程无显著影响。研究揭示了人肠道源MCRPEC菌株在黏菌素禁止作为饲料促生长剂使用前后的三个重要变化：（1）mcr-1通过丢失ISApl1来稳定存在于适应性代价更低的质粒如IncI2上；（2）mcr-1与其他抗生素抗性基因共同传播来实现共同选择效应；（3）MCRPEC中毒力基因的数量显著增多，ExPECs的比例显著增加。这些变化可能会增强MCRPEC的定植能力，占据共生大肠杆菌的生态位，对公众健康的风险更高。以上成果丰富了MCRPEC演化和MCRPP传播的理论，为遏制黏菌素耐药性传播扩散提供了科学依据和思路。