Study Of Pathogenicity And Erythromycin Resistance Of Cutibacterium Acnes Based On The Pan-genome Level

Object:C.acnes is the main symbiotic bacteria on the surface of human skin.It has the function of maintaining skin homeostasis and regulating the immune system of skin.C.acnes is considered to be the main pathogen of acne vulgaris,and ectopic colonization of C.acnes may cause serious diseases.At present,the research on C.acnes genome is still blank in China.This study carried out genome-wide sequencing,assembly and annotation of multiple strains of C.acnes strains from Chinese people,and obtained the genome-wide information of C.acnes from Chinese people.Through the pan-genome and other bioinformatics analysis,the evolutionary relationship and genomic characteristics of C.acnes strains from Chinese population were revealed,and the pathogenicity,drug resistance and genotypes of C.acnes isolated from Chinese population were explored.The research is intended to provide a theoretical basis for maintaining the skin health of the Chinese population and the prevention and treatment of infection caused by C.acnes.Method:2-6 C.acnes strains were isolated from the cultures of each patient with acne and each healthy individual in anaerobic condition,and the whole genome of all trains were sequenced by NGS;the quality control of the sequenced data was analyzed using the Fast QC and NGS QC Toolkit;Use SOAPdenovo,velvet and spades to assemble the C.acnes genome;use prokka to annotate the gene information of C.acnes;use BPGA to complete pan-genome analysis,including power-law regression analysis,phylogenetic tree,COG analysis,KEGG analysis;using MLST method to type C.acnes;genomic characteristics and differences of subtype strains were evaluated according to the heat map of the accessory genes of 50 C.acnes strains;pathogenic subtypes were screened according to the distribution of C.acnes strains in two groups;screening for pathogenic factors according to the difference of C.acnes genome from patients and healthy individuals.The erythromycin resistance of all monoclonal was verified using the paper method,the reason of erythromycin resistance was analyzed according to the monoclonal gene differences;Themic value of erythromycin resistance and non-erythromycin resistance of monoclonal was determined by dilution method.Result:50 strains of C.acnes were isolated from 10 samples of facial skin cotton swabs from Chinese population.For the first time,50 strains of C.acnes strains from the Chinese population were sequenced.The sequencing depth was 100×,and the quality of sequencing was good.Three assembling software were tested and the monoclonal genomic reads was performed using the optimal software and optimized parameters.Using the prokka to complete the monoclonal gene annotation.Quantitative regression analysis of 50 monoclones showed that the C.acnes genome was conserved and that the number of core genes was reduced and the rate of non-essential gene was slowed down when more than 12 monoclones were analyzed,the results were verified by 50 strains of C.acnes from the NCBI database.COG analysis showed that the main function of the core gene and the subsidiary gene of C.acnes was focused on maintaining the transcriptional transcription and material metabolism.KEGG analysis showed that most of the genetic functions of C.acnes were related to the metabolism of three major nutrients,the transport and transcription of the substance;the C.acnes of patient has more genes involved in carbohydrate metabolism and fewer genes participated in the vitamin synthesis.The rec A gene is not suitable for C.acnes evolution tree construction.The phylogenetic tree of the core gene shows that the same source of C.acnes has a more recent evolutionary relationship.The pan-genome evolutionary tree showed that six type ? C.acnes strains may be associated with acne.Based on the cluster analysis of the matrix,it is found that each subtype of C.acnes has a large difference in the group of accessory genes.90 genes that may be associated with acne?p<0.05?were found based on the comparison of genomes of 50 strains.The two genes b₀2220 and b₀0131 may be the virulence factor genes of subtype ?.The acquirement of erythromycin resistance of C.acnes may be the horizontal transferation of erm X mediated by transposon Tn5432.Conclusion:1.The genome of C.acnes strains is highly conserved;SNPs of C.acnes core genes can be used to analyze the source of C.acnes strains;pan-genome evolution tree of C.acnes can better reflect the difference between the genomes of different strain.2.C.acnes strains derived from healthy individuals and patients with acne have different percentage of genes involved in carbohydrate metabolism and vitamin synthesis.3.It was found that each subtype of C.acnes strains has unique genes,and these genes are significantly different in function based on the presence or absence of C.acnes accessory genes for the first time.4.The subtype ? strains of C.acnes is associated with acne vulgaris in chinese patients.5.Tt is found that the two genes,b₀2220 and b₀0131,may be potential virulence factors of the C subtype ? strain of C.acnes.6.The acquirement of erythromycin resistance of C.acnes strains may be the horizontal transferation of erm X mediated by transposon Tn5432,and there is another unknown resistance mechanism need to be studied.