中度与重度寻常痤疮患者非皮损区毛囊菌群分析：一项单中心横断面研究

doi:10.3969/j.issn.1674-8115.2024.09.004

上海交通大学学报（医学版） ›› 2024, Vol. 44 ›› Issue (9): 1094-1103.doi: 10.3969/j.issn.1674-8115.2024.09.004

• 论著 · 基础研究 • 上一篇

中度与重度寻常痤疮患者非皮损区毛囊菌群分析：一项单中心横断面研究

梁梦晨(), 李嘉祺(), 吴心怡, 莫小辉, 鞠强()

上海交通大学医学院附属仁济医院皮肤科，上海 200127

收稿日期:2024-05-03 接受日期:2024-07-18 出版日期:2024-09-28 发布日期:2024-10-09
通讯作者: 鞠强 E-mail:2894791478@qq.com;jqedison@163.com;qiangju401@sina.com
作者简介:梁梦晨（1999—），女，博士生；电子信箱：2894791478@qq.com
李嘉祺（1996—），男，博士生；电子信箱：jqedison@163.com。第一联系人：（梁梦晨、李嘉祺并列第一作者）
基金资助:
国家自然科学基金(82373506);上海交通大学医学院附属仁济医院课题(IIT-2022-0011)

Analysis of hair follicle microbiota in non-lesional areas of patients with moderate-to-severe acne vulgaris: a single-center cross-sectional study

LIANG Mengchen(), LI Jiaqi(), WU Xinyi, MO Xiaohui, JU Qiang()

Department of Dermatology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China

Received:2024-05-03 Accepted:2024-07-18 Online:2024-09-28 Published:2024-10-09
Contact: JU Qiang E-mail:2894791478@qq.com;jqedison@163.com;qiangju401@sina.com
Supported by:
National Natural Science Foundation of China(82373506);Project of Renji Hospital, Shanghai Jiao Tong University School of Medicine(IIT?20220011)

摘要/Abstract

摘要：

目的·研究中度、重度寻常痤疮患者与健康者非皮损区毛囊菌群群落结构和载量的差异，探讨微生物与寻常痤疮及其严重程度之间的关系。方法·采用横断面研究，选取2022年8月—2023年8月在上海交通大学医学院附属仁济医院皮肤科就诊的中度、重度寻常痤疮（简称痤疮）患者和健康志愿者。取中度、重度痤疮患者及健康志愿者面部非皮损区毛囊内容物进行16S rRNA高通量测序以及实时荧光定量聚合酶链反应（quantitative real-time polymerase chain reaction，qPCR）检测，分析不同严重程度痤疮患者毛囊细菌菌群多样性、物种组成差异和菌群载量差异。结果·纳入10例中度痤疮患者、11例重度痤疮患者及11例健康志愿者，3组间年龄、性别等一般资料差异均无统计学意义。中度痤疮组和重度痤疮组相对于健康组细菌α多样性均显著下降（P=0.020，P=0.013）。主坐标分析（principal coordinates analysis，PCoA）图表明健康组人群样本分布较为集中，组内差异较小，而中度痤疮组及重度痤疮组人群样本分布有一定的趋势但较为离散，组内之间存在较大差异。3组样本趋势分布差异明显，组间微生物群落结构差异较大。相似性分析结果显示健康组与中度痤疮组（P=0.027）、健康组与重度痤疮组（P=0.017）组间β多样性有显著差异，物种组成相似度低。中度痤疮组与重度痤疮组（P=0.160）组间物种相似度较高。门水平上3组的优势菌群均为放线菌门（Actinobacteria）、厚壁菌门（Firmicutes）、变形菌门（Proteobacteria）以及拟杆菌门（Bacteroidetes）。属水平上健康组优势菌为丙酸杆菌属（Propionibacterium）、未分类的放线菌属（unclassified Actinomycetales），2组痤疮组优势菌均为葡萄球菌属（Staphylococcus）、丙酸杆菌属。相对于健康组，中度痤疮组和重度痤疮组非皮损区毛囊中的葡萄球菌属相对丰度均显著增加（P=0.010，P=0.019），毛囊菌群载量均显著增加（均P=0.001）。与中度痤疮组相比，重度痤疮组毛囊样本中细菌载量显著升高（P=0.017）。结论·中度、重度痤疮患者与健康者非皮损区毛囊微生物群落结构不同，痤疮组微生物多样性明显下降。中度痤疮组及重度痤疮组非皮损区毛囊中的葡萄球菌属相对丰度较健康组均显著增加。随着痤疮严重程度的增加，非皮损区毛囊内细菌载量显著增加。研究提示痤疮的发生以及严重程度可能与毛囊菌群的群落结构和载量有关。

关键词: 寻常痤疮, 毛囊菌群, 葡萄球菌属, 细菌载量

Abstract:

Objective ·To study the differences in the structure and load of hair follicle microbiota in non-lesional areas among patients with moderate-to-severe acne vulgaris and healthy individuals, and to explore the relationship between microorganisms and the severity of acne vulgaris. Method ·A cross-sectional study was used. Patients with moderate or severe acne vulgaris (referred to as acne) and healthy volunteers who visited the Department of Dermatology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, from August 2022 to August 2023. 16S rRNA high-throughput sequencing and quantitative real-time polymerase chain reaction (qPCR) were performed on the follicular contents from the non-lesional areas of the faces of patients with moderate or severe acne and healthy volunteers to analyze the diversity, species composition, and microbial load differences in hair follicle bacteria in patients with different severity of acne. Results ·Ten patients with moderate acne, eleven patients with severe acne, and eleven healthy volunteers were included. There were no statistically differences in general data such as age and gender ratio among the three groups. Bacterial α-diversity was significantly lower in both the moderate and severe acne groups compared to the healthy group (P=0.020, P=0.013). The principal coordinates analysis (PCoA) plot showed that the sample distribution of the healthy group was relatively concentrated, with small differences within the group, and the distribution of samples in the moderate and severe acne groups exhibited a certain trend but was relatively scattered, with differences between the groups. There were differences in the trend distribution of the three sample groups, and there were differences in the microbial community structure between the groups. The results of similarity analysis showed significant differences in β-diversity and low similarity in species composition between the healthy and moderate acne groups (P=0.027) and between the healthy and severe acne groups (P=0.017), and high species similarity between the moderate acne and severe acne groups (P=0.160). The dominant bacterial groups at the phylum level were Actinobacteria, Firmicutes, Proteobacteria, and Bacteroidetes. At the genus level, the dominant bacteria in the healthy group were Propionibacterium and unclassified Actinomycetales, and the dominant bacteria in both acne groups were Staphylococcus and Propionibacterium. Compared to the healthy group, the relative abundance of Staphylococcus species in the hair follicles in non-lesional areas of the moderate and severe acne groups was significantly increased (P=0.010, P=0.019). Compared with the healthy control group, the hair follicle microbiota load in non-lesional areas of both the moderate and severe acne groups was significantly increased (both P=0.001). Compared with the moderate acne group, the bacterial load in the hair follicle samples of the severe acne group was significantly increased (P=0.017). Conclusion ·The microbial community structure of hair follicles in non-lesional areas of patients with moderate or severe acne is different from that of healthy individuals, and the microbial diversity in the acne group is significantly reduced. The relative abundance of Staphylococcus species in the hair follicles in non-lesional areas of the moderate or severe acne groups is significantly increased compared to the healthy group. As the severity of acne increases, the bacterial load in hair follicles in non-lesional areas significantly increases. This research suggests that the occurrence and severity of acne may be related to the community structure and load of hair follicle microbiota.

Key words: acne vulgaris, hair follicle microbiota, Staphylococcus genus, bacterial load

中图分类号:

R751

梁梦晨, 李嘉祺, 吴心怡, 莫小辉, 鞠强. 中度与重度寻常痤疮患者非皮损区毛囊菌群分析：一项单中心横断面研究[J]. 上海交通大学学报（医学版）, 2024, 44(9): 1094-1103.

LIANG Mengchen, LI Jiaqi, WU Xinyi, MO Xiaohui, JU Qiang. Analysis of hair follicle microbiota in non-lesional areas of patients with moderate-to-severe acne vulgaris: a single-center cross-sectional study[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2024, 44(9): 1094-1103.

图/表 13

表1 引物序列

Tab 1 Primer sequences

Primer name	Primer sequence (5′→3′)	Amplified region
341F	CCTACGGGNGGCWGCAG	V3‒4^[9]
805R	GACTACHVGGGTATCTAATCC	V3‒4^[9]

表2 qPCR绝对定量程序

Tab 2 qPCR absolute quantification program

Step	Program
Step 1	95 ℃ 30 s
Step 2 (40 cycles)	95 ℃ 5 s; 55 ℃ 30 s; 72 ℃ 30 s
Step 3 (dissociation stage)	94 ℃ 30 s; 70 ℃ 1.5 min; 94 ℃ 10 s

表3 3组人群一般资料

Tab 3 General characteristics of the three groups

Characteristic	C group (n=11)	M group (n=10)	S group (n=11)	H/χ² value	P value
Gender/n(%)				0.182	0.913
Male	5 (45.5)	5 (50.0)	6 (54.5)
Female	6 (54.5)	5 (50.0)	5 (45.5)
Age/year	24.64±1.63	21.90±2.88	23.18±5.42	3.827	0.252
Ethnic group/n(%)				1.971	0.373
Han	10 (90.9)	10 (100.0)	11 (100.0)
Bai	1 (9.1)	0 (0)	0 (0)
Education/n(%)				2.676	0.262
Senior high school/technical secondary school	4 (36.4)	2 (20.0)	6 (54.5)
Junior college and above	7 (63.6)	8 (80.0)	5 (45.5)
Work/n(%)				1.703	0.427
In service	3 (27.3)	2 (20.0)	5 (45.5)
Not in service	8 (72.7)	8 (80.0)	6 (54.5)
Habitual residence/n(%)				2.147	0.342
Shanghai	11 (100.0)	9 (90.0)	9 (81.8)
Others	0 (0)	1 (10.0)	2 (18.2)

图1 3组人群毛囊菌群的α多样性分析Note： A. Species accumulation curve. B. Venn diagrams of the OTU numbers of the three groups. C. Box plots of Chao1 index of the three groups.

Fig 1 α diversity analysis of follicular microbiome of the three groups

图2 3组人群毛囊菌群的β多样性分析Note： PCoA scatter plot based on β diversity index between samples.

Fig 2 β-diversity analysis of hair follicle microbiome of the three groups

表4 3组样本细菌菌群在门水平上的丰度（%）

Tab 4 Abundance of bacterial flora at the phylum level in three sample groups （%）

Phylum	Abundance/%
Phylum	C group (n=11)	M group (n=10)	S group (n=11)
Actinobacteria	61.48±19.39	58.48±17.59	49.64±28.26
Firmicutes	21.94±16.10	35.42±18.37	43.90±26.42
Proteobacteria	14.84±15.23	5.27±5.79	5.51±5.41
Bacteroidetes	0.86±0.88	0.27±0.29	0.59±1.13
Others	0.88±0.55	0.56±0.57	0.36±0.49

表5 3组样本细菌菌群在属水平上的丰度（%）

Tab 5 Abundance of bacterial flora at the genus level in three sample groups （%）

Genus	Abundance/%
Genus	C group (n=11)	M group (n=10)	S group (n=11)
Staphylococcus	13.05±11.54	31.54±17.84	37.30±27.09
Propionibacterium	36.99±17.67	25.94±19.63	23.78±16.30
Unclassified Actinomycetales	18.78±19.20	23.38±15.96	19.58±17.88
Corynebacterium	2.09±2.77	8.28±11.28	5.47±4.74
Unclassified Neisseriaceae	4.36±8.64	0.06±0.08	0.86±2.48
Others	24.73±22.90	10.80±9.32	13.01±22.05

图3 3组人群毛囊菌群在门水平上相对丰度柱状图

Fig 3 Column diagram of relative abundance of hair follicle microbiome at the phylum level in the three groups

图4 3组人群毛囊菌群在属水平上相对丰度柱状图

Fig 4 Column diagram of relative abundance of hair follicle microbiome at the genus level in the three groups

图5 C组和M组人群毛囊菌群LEfSe分析进化分支图Note： The circles radiating from the inside out represent the classification levels from phylum to genus. Each circle and its shaded range at different classification levels represent the classification at that level, and the diameter of the circle is proportional to the relative abundance of the species. The species represented by the English letters next to the circles are shown in the legend. The red nodes represent microbial communities with high abundance and important roles in the red group, the green nodes represent microbial communities with important roles in the green group, and the yellow nodes represent species with no differences between groups.

Fig 5 LEfSe cladogram of analysis of follicular microbiome in C group and M group

图6 C组和S组人群毛囊菌群LEfSe分析进化分支图Note： The circles radiating from the inside out represent the classification levels from phylum to genus. Each circle and its shaded range at different classification levels represent the classifications at those levels, and the diameter of the circle is proportional to the relative abundance of the species. The species represented by the English letters next to the circles are shown in the legend. The red nodes represent microbial communities with high abundance and important roles in the red group, the green nodes represent microbial communities with important roles in the green group, and the yellow nodes represent species with no differences between groups.

Fig 6 LEfSe cladogram of analysis of hair follicle microbiome in Group C and Group S

图7 M组和S组人群毛囊菌群LEfSe分析进化分支图Note： The circles radiating from the inside out represent the classification level from phylum to genus. Each circle and its shaded range at different classification levels represent the classification at that level, and the diameter of the circle is proportional to the relative abundance of the species. The species represented by the English letters next to the circle are shown in the legend. The red nodes represent microbial communities with high abundance and important roles in the red group, the green nodes represent microbial communities with important roles in the green group, and the yellow nodes represent species with no differences between groups.

Fig 7 LEfSe cladogram of analysis of hair follicle microbiome in Group M and Group S

图8 3组人群毛囊细菌总拷贝数箱型图

Fig 8 Box plots of total number of follicular bacterial copies of the three groups

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中度与重度寻常痤疮患者非皮损区毛囊菌群分析：一项单中心横断面研究

Analysis of hair follicle microbiota in non-lesional areas of patients with moderate-to-severe acne vulgaris: a single-center cross-sectional study

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