Comparative Structural Study of Terminal Ends of Lipoarabinomannan from Mice Infected Lung Tissues and Urine of a Tuberculosis Positive Patient
Comparative Structural Study of Terminal Ends of Lipoarabinomannan from Mice Infected Lung Tissues and Urine of a Tuberculosis Positive Patient
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DOI：
10.1021/acsinfecdis.9b00355
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发表时间：
2020-02-01
影响因子：
5.3
通讯作者：
Chatterjee, Delphi
中科院分区：
文献类型：
--
作者：
De, Prithwiraj;Shi, Libin;Chatterjee, Delphi
结核分枝杆菌脂阿拉伯甘露聚糖(LAM)是活动性结核病的生物标志物。无论HIV阳性或阴性的结核病患者尿液中LAM的存在已被气相色谱/质谱法证实，具有良好的特异性(84%)和敏感性(99%)。然而，使用免疫测定法检测尿液中结核LAM的POC方法灵敏度较低，仅限于HIV合并感染的结核病诊断。我们推测，POC方法的这些令人失望的结果可能是由于免疫分析中使用的抗体所致，因为体内LAM和体外LAM可能存在结构上的差异。为了解决这个问题，我们用北京分枝杆菌SA161感染C3HeB/FEJ小鼠，并从肺中纯化LAM。使用一组现有的单抗对LAM的这些来源进行分析，发现了表位模式的差异。传统上，LAM的非还原末端是通过它们与内源性阿拉伯纤维素酶的释放来识别的。这些表位对应于直线型四-(Ara(4))、支链六-(Ara(6))阿拉伯呋喃糖苷及其甘露糖封端形式。我们发现了两个截然不同的表位。在第一种情况下，发现结核分枝杆菌SA161株LAM的非还原末端高度琥珀酰化，特别是当从小鼠肺中分离LAM时。在第二个案例中，发现内源性纤维单胞菌对SA161和TB+HIV患者尿液中LAM的消化产生了以5个阿拉伯糖基为主要成分的表位，并且严重缺乏Ara(6)。根据结核分枝杆菌SA161的5个阿拉伯酶和人尿中的LAM的5个阿拉伯酶的表位必须是潜在的结构和表位差异的结果。这些结果建议了开发针对疑似结核病患者尿液中LAM的POC检测的特异性抗体的方法。
Mycobacterium tuberculosis lipoarabinomannan (LAM) is a biomarker for active tuberculosis (TB) disease. The presence of LAM in the urine of TB patients, whether HIV positive or negative, has been validated by a gas chromatography/mass spectral method with good specificity (84%) and sensitivity (99%). However, point -of-care (POC) methods to detect TB LAM in urine using immunoassays have poor sensitivity and are limited to only HIV coinfected TB diagnosis. We hypothesized that these disappointing results with the POC methods may be due to the antibodies used in the immunoassays as there could be structural differences between LAM in vivo and LAM in vitro. To address this issue, we infected C3HeB/FeJ mice with M.tb W. Beijing SA161 and purified LAM from the lung. Analysis of these sources of LAM using a panel of existing mAbs revealed differences in epitope patterns. Conventionally, the non-reducing termini of LAM are identified by their release with endoarabinanase. These epitopes correspond to linear tetra-(Ara(4)), branched hexa-(Ara(6)) arabinofuranosides, and their mannose-capped versions. We discovered two distinct epitopes. In the first case, it was found that the non-reducing termini of LAM from M.tb strain SA161 are highly succinylated, especially when the LAM was isolated from the mouse lungs. In the second case, it was found that Cellulomonas endoarabinanase digestion of LAM from both SA161 and LAM from a TB+ HIV- patient's urine yielded epitopes based on 5 arabinoses as major components and a profound lack of Ara(6). The epitopes based on 5 arabinoses from M.tb SA161 and from the LAM in human urine must result from underlying structural and thus epitope differences. These results suggest approaches to develop specific antibodies for POC tests for LAM in the urine of suspected TB patients.