子宫颈微生态与宫颈鳞状上皮内病变关系的研究

Study on relationship between cervical microecology and cervical squamous intraepithelial lesions

目的:探讨子宫颈微生态与宫颈鳞状上皮内病变(squamous intraepithelial lesion,SIL)的关系。方法:选取2019年3月至5月南京医科大学附属无锡第二医院体检中心体检的和妇科就诊的12例子宫颈正常受试者(37~47岁),21例低度鳞状上皮内病变(low-grade squamous intraepithelial lesion,LSIL)患者(39~48岁),5例高度鳞状上皮内病变(high-grade squamous intraepithelial lesion,HSIL)患者(38~45岁)和3例宫颈鳞癌患者(42~43岁),填写调查问卷、行子宫颈检查,同时使用下一代测序技术(next generation sequencing,NGS)分析子宫颈分泌物的微生态,生物信息学分析NGS结果。根据NGS的人乳头状瘤病毒(human papilloma virus,HPV)检测结果将受试者分为HPV阴性组、低危HPV(low-risk HPV,lrHPV)、16/18型高危HPV(high-risk HPV,hrHPV)和其他hrHPV感染组。分析数据的韦恩图、微生态的多样性、物种的相对丰度和共现性以及受试者工作特征(receiver operating characteristic,ROC)曲线。结果:所有受试者子宫颈分泌物共获得种属水平微生态物种909种,各组之间有重叠。在不同宫颈病变受试者中,总HPV感染率、16/18型hrHPV感染率和其他hrHPV感染率差异无统计学意义(P均>0.05)。按HPV感染分组,16/18型hrHPV和其他hrHPV感染患者的子宫颈微生态多样性升高(U分别为39.00和43.00,P均<0.05),而各组之间卷曲乳酸杆菌相对丰度无差异(H=4.37,P=0.2136)。按子宫颈状况分组,子宫颈病变受试者子宫颈微生态多样性(H=14.60,P=0.0022)升高,卷曲乳酸杆菌丰度下降(H=13.98,P=0.0008)。在微生态物种中,支原体、衣原体和B族链球菌具有共现性;惰性乳酸杆菌、阴道加德纳菌、阿托波菌和二路普氏菌具有共现性。以卷曲乳酸杆菌相对丰度作为SIL诊断指标,其ROC曲线下面积(area under ROC curve,AUC)为0.874[95%置信区间(confidence interval,CI):0.732~0.957],卷曲乳酸杆菌和詹氏乳酸杆菌以及支原体联合诊断SIL的AUC为0.943[95%CI:0.822~0.991]。结论:子宫颈卷曲乳酸杆菌相对丰度下降和微生态多样性升高可能与HPV感染和子宫颈病变有关;NGS数据简化后可能有助于SIL诊断。

Objective To explore the relationship between cervical microecology and cervical squamous intraepithelial lesions(SIL).Methods All subjects were recruited from the health care center or gynecology of the Affiliated Wuxi No.2 People′s Hospital of Nanjing Medical University from March to May of 2019,including 12 subjects normal cervix with 37-47 years old,21 low-grade squamous intraepithelial lesion(LSIL)subjects with 39-48 years old,5 high-grade squamous intraepithelial lesion(HSIL)subjects with 38-45 years old and 3 cervical squamous cell carcinoma subjects with 42-43 years old.All subjects were required to fill in a questionnaire,and performed cervical examination.Meanwhile,the microecology of cervical secretions was analyzed by the next generation sequencing(NGS)and the NGS results were analyzed by bioinformatics.Subjects were divided into human papilloma virus(HPV)-negative groups,low-risk HPV(lrHPV),16/18 high-risk HPV(hrHPV)and other hrHPV infection groups based on HPV test results of NGS.The Venn diagram of data,microecology diversity,the relative abundance and co-occurrence of species,and the receiver operating characteristic(ROC)curve were analyzed.Results A total of 909 species at the species level were obtained from the cervical secretions of all the subjects,and there was overlap among the groups.There was no significant difference in total HPV infection rate,16/18 hrHPV infection rate and other hrHPV infection rates among subjects with different cervical lesions(all of P>0.05).Grouped by HPV infection,the 16/18 hrHPV-infected and other hrHPV-infected subjects had increased cervical microecology diversity(U=39.00 and 43.00,all of P<0.05),and the relative abundance of Lactobacillus crispatus(L.crispatus)had no differences among the groups(H=4.37,P=0.2136).Grouped by cervical conditions,the cervical microecology diversity of the subjects with cervical lesions increased(H=14.60,P=0.0022),while the L.crispatus relative abundance decreased(H=13.98,P=0.0008).Among all the detected species,Mycoplasma,Chlamydia and Streptococcus B had a co-occurrence,while Lactobacillus iners,Gardnerella vaginalis,Atopobium vaginae,and Prevotella bivia had a co-occurrence.As the SIL diagnostic index,the area under the ROC curve(AUC)of the relative L.crispatus relative abundance was 0.874[95%confidence interval(CI):0.732-0.957].L.crispatus combined with Lactobacillus jensenii(L.jensenii)and Mycoplasma had an AUC of 0.943[95%CI:0.822-0.991]in the SIL diagnosis.Conclusions The decreased L.crispatus relative abundance and the increased cervical microecology diversity may be related to HPV infection and cervical lesions;simplified NGS data may be helpful to the SIL diagnosis.