The Mechinism Of Colistin Resistance And Heteroresistance In Carbapenem-resistant Enterobacteriaceae

Currently,multidrug-resistant gram-negative bacteria,especially carbapenem-resistant Enterobacteriaceae,have become a major challenge in the treatment of anti-infection.Polymyxins are playing a more and more important role in the treatment of multidrug-resistant Enterobacteriaceae.However,with the increasing application of polymyxin in clinical,the polymyxin-resistant Enterobacteriaceae have been isolated word-wide.Currently,the existing resistance mechanisms cannot fully explain the cause of the clinical strains of polymyxin resistance,this study will explore gradually the mechanisms of colistin resistance and heteroresistance in Enterobacteriaceae.The first part is mainly about the detection of colistin resistance and colistin heteroresistance in KPC-producing K.pneumoniae.We collected clinical KPC-producing K.pneumoniae strains to perform colistin susceptibility test and performed population analysis profiling(PAP)to measure heteroresistance to colistin.The results showed that the rate of colistin resistance in clinical KPC-producing K.pneumoniae strains was still very low(1%),but they showed a high rate of colistin heteroresistance(100%).All the 22 colistin sensitive KPC-producing K.pneumoniae isolates showed heteroresistance to colistin(100%).Most of the subpopulations selected in the PAP experiments contained an inactivated mgrB gene(20/22).The mutations in two-component systems also caused heteroresistance of colistin(2/22).The second part we mainly studied the plasmid-mediated colistin resistance gene mcr-l in carbapene-resistant K.pneumoniae and E.coli.Here we studied the impact of mcr-l on the development of high-level colistin resistance(HLCR)mutations in them.We detected the high-level colistin resistance mutation rates of three clinical carbapenem-resistant strains(XH209.KP10 and Q3)and their combinations with mcr-l gene.The HLCR mutation rates of two K.pneumoniae strains(XH209,KP10)showed no statistically significant differences with their derived strains containing mcr-l.With or without mcr-l gene.mutation of gene was the most important mechanism for the development of high-level colistin resitance in K.pneumoniae.But mcr-l result in higher mutation rates of HLCR mutants in the E.coli strain Q3.Chromosomal mutations in pmrA/pmrB or phoQ showed synergistic effect with mcr-l in colistin resistance.Our results show that the mcr-l gene has greater impact on the E.coli.It facilitates development of HLCR mutants in E.coli.but does not affect the K.pneumoniae.In the third part of this study,we detected the impact of mcr-l gene in K.pneumoniae.The results showed that the effect of mcr-l gene on the growth rate of K.pneumoniae was strain specific.and the mcr-1 gene did not cause the change of virulence of K.pneumoniae.That is,the expression of the mcr-l gene does not lead to the apparent fitness cost of K.pneumoniae.