| [Objective]Acne is a chronic inflammatory disease involving the sebaceous glands of hair follicles with high incidence and skin damage.Cutibacterium acnes(C.acnes)is an important causative and aggravating factor of acne,and antibiotics are one of the means of acne treatment.Because macrolide-lincosamide antibiotics have fewer side effects and are suitable for a wide range of people,they are one of the recommended drugs in acne treatment guidelines,but the frequent use of antibiotics has led to a high rate of resistance to C.acnes.This study sought to collect clinical isolates of C.acnes in early days and perform in vitro drug sensitivity tests and efflux pump inhibition tests to assess the effects of two efflux pump inhibitors on the minimum inhibitory concentration(MIC)of five macrolide-lincosamide antibacterial drugs and to understand whether efflux pump inhibitors can reduce the resistance of C.acnes to macrolide-lincosamide drugs,and whether efflux pump exists in drug-resistant bacteria,providing theoretical basis for the development of antibacterial drugs against resistant C.acnes and the treatment of acne.[Methods]The 39 strains of C.acnes collected from acne patients were frozen,followed by resuscitation,culture,and mass spectrometry identification.Then the agar dilution method was used for drug sensitivity tests to determine the MIC of five macrolide-lincosamide antibiotics(macrolide: erythromycin,azithromycin and clarithromycin;lincosamide: clindamycin and lincomycin)against the experimental strains.According to the results of drug sensitivity tests,they were divided into the multidrug resistance group,single-drug resistance group,and sensitive group.The MIC values of antimicrobial drugs combined with two efflux pump inhibitors carbonyl cyanide-3-chlorophenylhydrazone(CCCP)and phenylalanine-arginineβ-naphthylamide(PAβN)were measured in the efflux pump inhibition test.A 4-fold or greater decrease in the MIC value of the antimicrobial drug after the addition of efflux pump inhibitors was considered positive for the efflux pump phenotype.[Results]1.Results of drug sensitivity tests of 39 clinically isolated C.acnes:1)Drug resistance rate: erythromycin 69.2%(27/39),azithromycin 69.2% (27/39),clarithromycin 71.8%(28/39),clindamycin 71.8%(28/39),and Lincomycin 69.2%(27/39).2)Drug resistance phenotype: 27 multi-drug resistant strains,2 single-drug resistant strains,and 10 sensitive strains.Among them,26 multi-drug resistant strains and 2 single-drug resistant strains showed high levels of drug resistance.2.Results of efflux pump inhibition tests1)multidrug resistance group: among the 27 multi-drug resistant strains,5 strains of efflux pump phenotype-positive bacteria were screened by the combination of antimicrobial drugs and CCCP;5 strains were screened by CCCP combined with azithromycin and MIC values both decreased from >256 μg/m L to <0.0625 μg/m L;2 strains were screened by CCCP combined with clarithromycin(all in the above 5 strains),and MIC value decreased from >256 μg/m L to 128 μg/m L and 128 μg/m L to 16 μg/m L, respectively.When CCCP combined with erythromycin,clindamycin and lincomycin,the phenotype of efflux pump was negative.Antibacterial drugs combined with PAβN screened 2 strains positive for the efflux pump phenotype(one strain in the above-mentioned 5 strain),and PAβN combined with clarithromycin screened 2 strains,with MIC values reduced from 128 μg/m L to 16 μg/m L and from >256 μg/m L to 128 μg/m L,respectively.When PAβN combined with erythromycin,azithromycin,clindamycin and lincomycin,the phenotype of efflux pump was negative.Six strains with positive efflux pump phenotype were screened out by the two efflux pump inhibitors.Among them,when clarithromycin was used as efflux substrate, one strain was positive for the efflux pump phenotype when combined with two efflux pump inhibitors.2)single-drug resistance group: The efflux pump phenotype of the strain was all negative when five antibiotics were combined with two efflux pumpinhibitors.[Conclusion]1.The drug sensitivity test results of this study showed that C.acnes had a high drug resistance rate to macrolides and lincosamide,and the drug resistance pattern was bipolar,and most strains showed high drug resistance or sensitivity;The drug resistance phenotype of most drug resistant strains were multi-drug resistance.2.C.acnes multi-drug resistant strains were tested by combining macrolide and lincosamide drugs with efflux pump inhibitors,and efflux pump inhibitors could reduce drug resistance of some multi-drug resistant strains(a 4-fold or greater decrease in the MIC value of the antimicrobial drug),indicating that the generation of multi-drug resistance is related to efflux pump to some extent.3.The efflux pump inhibitors CCCP or PAβN can be used to screen the positive strains of efflux pump for C.acnes of multiple drug resistance of macrolide-lincosamide,but the effect of CCCP is better in small sample test. |
