关键词: 天疱疮, 葡萄球菌属, 皮肤菌群, 微生物组, 16S rDNA测序, 功能预测

Abstract: 【Abstract】 Objective To analyze the skin microbiota diversity in patients with pemphigus vulgaris （PV） using 16S rDNA sequencing. Methods Ten patients with PV and 10 healthy controls were collected from the Department of Dermatology, the Third People′s Hospital of Hangzhou. Skin swabs were collected from perilesional skin （PV group） and contralateral non-lesional skin （PVn group） of the patients with PV, as well as from the corresponding body sites of the healthy controls （normal control group）. The 16S rDNA amplicon sequencing technology was used for gene sequencing and classification in all microbiota samples, and Usearch software for data cluster analysis to obtain operational taxonomic units （OTUs） and assess species abundance at the phylum, class, order, family and genus levels. Observed species index, Shannon index and Simpson index were used to estimate α diversity, and principal coordinate analysis （PCoA） was performed to analyze β diversity. Linear discriminant analysis effect size （LEfSe） analysis was conducted to identify differentially abundant species in each group. PICRUSt software was used for gene function prediction. Wilcoxon rank sum test was used as nonparametric test for comparisons between 2 groups, and Kruskal Waills test as nonparametric test for comparisons among 3 groups. Results There were 2 002, 1 869, 1 751, 1 611 and 1 120 OTUs at phylum, class, order, family and genus levels respectively. Cluster analysis showed that skin microbiome in the 3 groups mainly consisted of Firmicutes, Actinobacteria, Bacteroidetes and Proteobacteria at the phylum level. At the genus level, Staphylococcus was the most abundant in the PV group and PVn group, and Corynebacterium was the most abundant in the normal control group. The observed species index, Shannon index and Simpson index all significantly differed among the 3 groups （all P < 0.05）, and the Shannon index and Simpson index were significantly lower in the PV group （3.24 ± 1.30, 0.70 ± 0.19, respectively） than in the normal control group （P < 0.05）. PCoA analysis showed no significant difference in β diversity among the 3 groups （P = 0.054）. Rank sum test showed that the abundance of 32 species significantly differed among the 3 groups （P < 0.05）. Among them, high relative abundance was observed in the class Bacilli enriched in the PV group, as well as the genera Micrococcus and Brevundimonas enriched in the normal control group. According to the disease duration, the patients with PV were divided into long-course PV group with disease duration of ≥ 3 months, and short-course PV group with disease duration of < 3 months. Clostridiales, Oscillibacter, Sphingomonas were enriched in the long-course PV group, and Gammaproteobacteria was enriched in the short-course PV group. Gene function prediction analysis showed that the genes related to infectious diseases were enriched in the pemphigus group. Conclusion The 16S rDNA-based microbiota profiling suggested differences in the diversity and composition of skin microbiota between patients with PV and healthy individuals.

Key words: Pemphigus, Staphylococcus, Skin flora, Microbiome, 16S rDNA sequencing, Function prediction