梅毒血清固定患者Tp分子分型筛选及其预测价值的评估

The prediction index for syphilis serofast reaction (or "serofast" for short) is still unclear. Our researchs have found that approximately 30% of syphilis patients could be found to have serofast state. In these serofast patients, 33% of patients were detected to have living Treponema pallidum (Tp) in their body, which will easily lead to relapse and infection of syphilis. The mechanism of serofast remians unknown, morever, the relation between serofast and neurosyphilis also aroused more attention. So far, our research data showed that 56.1% of serofast patients were finally diagnosis with neurosyphilis. It has been proved that neurosyphilis is caused by some Tp strain types. At the same time, our preliminary experiment also showed that syphilis serofast had its own predominated molecular types, which were rather different from the types among neurosyphilis. From this, we suspected that serofast may be caused by some predominated molecular types of TP strains. Firstly, in this research we will use the "CDC subtype/tp0548 sequence type" genotyping system to screenthe predominated molecular types of TP strains (maybe more than 1 type) in serofast patients. Then, complete genome sequence analysis are used to identify the different genes, which can be detected as an important prediction index for serofast. Next, based on the published genetic information about current genotyping methods, we find out some specific target genes for molecular typing of serofast. At last, we can establish a high sensitive and specific molecular typing method for serofast. This new method may provide important prediction value for the diagnosis of serofast, as well as provide scientific basis for standard diagnosis and scientific intervention of serofast. Meanwhile, it will hopefully reveal the molecular mechanism of serofast from a new perspective.

梅毒血清固定（简称血清固定）的发生机制未明，且缺乏预警指标。我们发现其发生率在30%以上，其中33 %的患者体内存有活的梅毒螺旋体（Tp），可致梅毒复发和传染；56.1%的血清固定患者最终被确诊为神经梅毒。业已证明，神经梅毒为某些基因型的Tp所致，预实验显示血清固定患者Tp有优势分子分型株，但与神经梅毒不完全一致。猜测血清固定是由某些分子分型的Tp感染所致。本项目首先采用CDC subtype/Tp0548 sequence type分型系统，初筛血清固定的优势分子分型Tp株，然后经全基因序列分析查找其差异基因，获得血清固定的一个重要标志，结合已有分型法的被检基因信息，获得特异的血清固定Tp分子分型靶基因，建立高灵敏的、特异的血清固定分子分型方法，对其诊断将具有重要预测价值，为规范其诊断，以及后续的科学干预处理提供科学依据。同时，有望从全新的角度揭示其发生的分子机制。

研究背景：梅毒的早期诊断极其重要，因为大多数类型的梅毒可以通过合理的治疗方案，得到有效的治愈或控制。临床中可见一部分梅毒患者经规则治疗 1 年后，其反应素下降到某一定的滴度后就不再下降（或反弹），持续数年甚至终身，该现象称为梅毒血清固定。遗憾的是，作为血清固定的唯一实验室检查—反应素抗体检测，是在血清固定现象出现后所作出的判断，时间是在正规治疗1年后，已经错过了治疗的时机。因此，迫切需要寻找到一种能在梅毒治疗初期就能预警血清固定存在的指标。.研究方法：我们在前期研究基础上，采用CDC subtype/Tp0548 sequencetype 分型系统，初筛血清固定的优势分子分型梅毒螺旋体株，然后经全基因序列分析查找其差异基因，获得血清固定的一个重要标志，结合已有分型法的被检基因信息，获得特异的血清固定Tp 分子分型靶基因，建立高灵敏的、特异的血清固定分子分型方法。.研究结果：项目以polA 基因作为初筛基因，其在血球和组织的阳性率分别为71.43%（50/70）和21.43%（15/70）高于血浆7.14%（15/70）、血清2.86%（15/70）、血块7.14%（15/70）和脑脊液2.86%（15/70）。polA 基因阳性的样本进一步采用CDC subtype/Tp0548 sequence type分型系统进行分子分型，梅毒血清固定患者的分子亚型分别为14d/f（70.20%），13d/f（8.20%），15d/f（8.80%）和16d/f(12.80%)；通过基因测序及CLUSTALX 软件比较，最后以tprK 基因序列分子作为诊断靶标，并构建tprK 基因测序法，盲法检测91例临床样本，阳性符合率为55.56%，阴性符合率为65.21%。.研究结论：存在梅毒血清固定基因亚型，需进一步优化tprK-DNA序列，以提高临床应用价值。