2种结核分枝杆菌抗原对潜伏感染人群鉴别诊断潜力分析

doi:10.3969/j.issn.1673-8640.2016.08.005

摘要/Abstract

摘要：

目的评价2种结核分枝杆菌（MTB）潜伏生长状态下高表达的蛋白——Rv3160c抗原、35kD抗原对MTB潜伏感染人群的血清学鉴别诊断潜力。方法纯化获得Rv3160c抗原、35kD抗原,采用C57BL/6小鼠模型评价其免疫原性。以商品化的检测抗原——早期分泌抗原靶蛋白6（ESAT-6）和38kD为阳性参照,测定Rv3160c抗原、35kD抗原在20例活动性肺结核患者（活动性肺结核组）、25例MTB潜伏感染者（MTB潜伏感染组,即与活动性肺结核患者长期密切接触者）、24名健康体检者（正常对照组）血清特异性抗体反应,即检测抗Rv3160c抗体、抗35kD抗体水平。结果 Rv3160c抗原、35kD抗原均能够在实验小鼠体内引起较高水平的体液免疫反应,血清抗体滴度达到1∶25 600,接近阳性参照抗原Ag85A的水平。MTB潜伏感染组血清抗Rv3160c抗体和抗35kD抗体水平均明显高于正常对照组（P<0.01）,且抗35kD抗体水平明显高于活动性肺结核组（P<0.05）,但MTB潜伏感染组和活动性肺结核组之间血清抗Rv3160c抗体水平差异无统计学意义（P>0.05）。MTB潜伏感染组和活动性肺结核组血清抗ESAT-6抗体水平均高于正常对照组（P<0.01）,而MTB潜伏感染组和活动性肺结核组之间差异无统计学意义（P>0.05）。MTB潜伏感染组血清抗38kD抗体水平明显高于活动性肺结核组和正常对照组（P<0.05）,而活动性肺结核组与正常对照组之间差异无统计学意义（P>0.05）。Rv3160c抗原、35kD抗原在MTB潜伏感染者的血清中产生较强的抗原-抗体反应,与正常对照者的血清反应较弱;35kD抗原与活动性肺结核患者的血清反应也较弱,其血清反应模式与38kD抗原类似;Rv3160c抗原无法区分活动性肺结核患者和潜伏感染人群,其血清反应模式与ESAT-6接近。结论 Rv3160c抗原和35kD抗原对于潜伏性结核感染具有高度特异性,将有望用于MTB潜伏感染的鉴别诊断。

关键词: 结核分枝杆菌, Rv3160c抗原, 35kD抗原, 潜伏感染, 血清诊断

Abstract:

Objective To investigate the potential of 2 antigens（Rv3160c and 35kD antigens） of Mycobacterium tuberculosis（MTB） highly expressed during latent infection for the serodiagnosis of latent tuberculosis infection（LTBI）. Methods Rv3160c and 35kD antigens were purified,and the immunogenicity was evaluated by C57BL/6 mouse model. MTB antigens,early secretory antigenic target-6（ESAT-6） and 38kD antigens which were in commercial use,were included as positive controls. Serum specificity antibody reactivities of 20 active pulmonary tuberculosis patients,25 LTBI patients（contacting closely with active pulmonary tuberculosis patients for a long time） and 24 healthy subjects were determined using Rv3160c and 35kD antigens. Serum levels of anti-Rv3160c and anti-35kD antibodies were determined. Results Rv3160c and 35kD antigens induced strong humoral immunity in mice. Serum antibody titre reached 1∶256 000,which was close to that of positive control Ag85A. Serum levels of anti-Rv3160c and anti-35kD antibodies in LTBI group were higher than those in healthy control group （P<0.01）, and serum level of anti-35kD antibody in LTBI group was higher than that in active pulmonary tuberculosis group（P<0.05）. However,serum level of anti-Rv3160c antibody had no statistical significance between LTBI and active pulmonary tuberculosis groups（P>0.05）. Serum levels of anti-ESAT-6 antibody in LTBI and active pulmonary tuberculosis groups were higher than that in healthy control group（P<0.01）,and there was no statistical significance between LTBI and active pulmonary tuberculosis groups（P>0.05）. Serum level of anti-38kD antibody in LTBI group was higher than those in active pulmonary tuberculosis and healthy control groups（P<0.05）,but there was no statistical significance between active pulmonary tuberculosis and healthy control groups（P>0.05）. Both antigens reacted more strongly with sera from LTBI group than sera from healthy control group. The 35kD antigen had a weak reactivity in active pulmonary tuberculosis group,and its pattern of reactivity was similar to that of 38kD antigen. Rv3160c antigen was unable to distinguish active pulmonary tuberculosis and LTBI,and its reactivity pattern was similar to that of ESAT-6 antigen. Conclusions Rv3160c and 35kD antigens are highly specific to LTBI. It is of potential to be used in the diagnosis of LTBI.

Key words: Mycobacterium tuberculosis, Rv3160c antigen, 35kD antigen, Latent infection, Serodiagnosis

中图分类号:

R446.1

孙睿峰, 项智灏, 陈福增, 茹欢委, 麦俊涛, 袁俐, 刘军. 2种结核分枝杆菌抗原对潜伏感染人群鉴别诊断潜力分析[J]. 检验医学, 2016, 31(8): 656-661.

SUN Ruifeng, XIANG Zhihao, CHEN Fuzeng, RU Huanwei, MAI Juntao, YUAN Li, LIU Jun. The potential of 2 antigens of Mycobacterium tuberculosis for the serodiagnosis of latent tuberculosis infection[J]. Laboratory Medicine, 2016, 31(8): 656-661.

图/表 4

参考文献 17

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引物	序列（5'~3'）
Rv3160c	F：CGTGGATCCATGCCGAGGCAGGCCG- GCCGCTG
	R：GCAAAGCTTCTAGAGCCCGCGGTCG- GGGGGTGCG
Rv2744c	F：GCGCATATGATGGCCAATCCGTTCGT- TAAAGC
	R：TATGTCGACCTACTGACCGTAGGGCT- GCTCGC

引物	序列（5'~3'）
Rv3160c	F：CGTGGATCCATGCCGAGGCAGGCCG- GCCGCTG
	R：GCAAAGCTTCTAGAGCCCGCGGTCG- GGGGGTGCG
Rv2744c	F：GCGCATATGATGGCCAATCCGTTCGT- TAAAGC
	R：TATGTCGACCTACTGACCGTAGGGCT- GCTCGC