Cloning of the gene encoding a protective Mycobacterium tuberculosis secreted protein detected in vivo during the initial phases of the infectious process
Cloning of the gene encoding a protective Mycobacterium tuberculosis secreted protein detected in vivo during the initial phases of the infectious process
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DOI：
10.4049/jimmunol.175.8.5298
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发表时间：
2005-10-15
影响因子：
4.4
通讯作者：
Campos-Neto, A
中科院分区：
文献类型：
--
作者：
Mukherjee, S;Kashino, SS;Campos-Neto, A
治疗剂和卡介苗的存在并未对目前结核病的流行产生重大影响。卡介苗可预防严重的儿科结核病，但不能预防成人肺结核，这是最常见和最具传染性的疾病。最近已经描述了几种候选疫苗，包括在较新佐剂中配制或在细菌质粒DNA中递送的结核分枝杆菌重组蛋白。一个有吸引力的候选疫苗来源是M.结核抗原存在于细菌体外生长的初始阶段的培养上清液中。在这项研究中，我们描述了一种Ag发现方法，以选择在感染的初始阶段在体内产生的这种Ag。我们结合反相高效液相色谱和质谱鉴定分泌或脱落的M。结核蛋白在感染后14天内从动物尿液中排出。一种含有与假设的M.鉴定了结核杆菌蛋白，并在大肠杆菌中表达了重组蛋白。该蛋白可被M.结核病和来自纯化蛋白衍生物皮肤试验呈阳性的健康捐赠者的淋巴细胞，但不是来自结核病患者的淋巴细胞。此外，该Ag诱导小鼠对M.预防结核病的水平与BCG疫苗诱导的保护水平相当。这些结果验证了M.结核蛋白在感染的早期阶段在体内分泌或脱落，并为开发潜在的候选疫苗或活性分枝杆菌增殖的标志物（因此是活性疾病）开辟了新的可能性。
The existence of therapeutic agents and the bacille Calmette-Guerin (BCG) vaccine have not significantly affected the current tuberculosis pandemic. BCG vaccine protects against serious pediatric forms of tuberculosis but not against adult pulmonary tuberculosis, the most common and contagious form of the disease. Several vaccine candidates, including Mycobacterium tuberculosis recombinant proteins formulated in newer adjuvants or delivered in bacterial plasmid DNA have recently been described. An attractive source of vaccine candidates has been M. tuberculosis Ags present in culture supernatants of the initial phases of the bacterial growth in vitro. In this study we describe an Ag discovery approach to select for such Ags produced in vivo during the initial phases of the infection. We combined RP-HPLC and mass spectrometry to identify secreted or shed M. tuberculosis proteins eliminated in animal urine within 14 days after the infection. A peptide containing sequence homology with a hypothetical M. tuberculosis protein was identified and the recombinant protein produced in Escherichia coli. The protein was recognized by Ab (IgG2a and IgG1) and T cells (Th1) of mice infected with M. tuberculosis and by lymphoid cells from healthy donors who had a positive purified protein derivative skin test but not from tuberculosis patients. Moreover, this Ag induced protection in mice against M. tuberculosis at levels comparable to protection induced by BCG vaccine. These results validate the Ag discovery approach of M. tuberculosis proteins secreted or shed in vivo during the early phases of the infection and open new possibilities for the development of potential vaccine candidates or of markers of active mycobacterial multiplication: and therefore active disease.