Functional Consequence of Macrolide Resistance Mutations in T. pallidum

描述（由申请人提供）：多年来，人们一直在寻求一种单剂量口服治疗梅毒的方法。过去50年来用于治疗梅毒的苄星青霉素G有许多缺点，包括注射部位疼痛、过敏反应的可能性以及这种形式的青霉素无法在中枢神经系统（梅毒螺旋体传播的常见部位）达到密螺旋体水平。利用兔梅毒模型发表的研究表明，阿奇霉素是一种与大环内酯类相关的口服氮唑类抗生素，可有效治疗由尼科尔斯梅毒菌株引起的梅毒。在几项人体研究中，单剂量1或2克阿奇霉素对治疗早期梅毒有效。然而，最近的出版物记录了循环T. pallidum菌株23S rRNA基因中A2058G突变的高频率。在其他细菌中，这种突变与大环内酯类药物耐药性有关。梅毒T. Street 14株含有A2058G突变，在兔梅毒模型中，对大环内酯及阿奇霉素等相关药物耐药。同样，在感染含有相同突变的苍白t菌的患者中也有阿奇霉素治疗失败的记录。尽管如此，最近的一篇文章质疑了这些23S rDNA突变的临床相关性；在与疾病控制和预防中心制定新的梅毒治疗指南有关的讨论中，也表达了类似的观点。与其继续推测，更重要的是进行实验，直接确定这些和其他突变是否会影响阿奇霉素治疗的临床反应。尽管有人可能会争辩说，存在（或不存在）足够的平衡来证明阿奇霉素治疗感染生物体携带23S rDNA突变的梅毒患者的试验是合理的，但它不太可能在一个可以告知我们当前临床实践的时间框架内进行。完善的兔子梅毒模型是解决这一问题的理想方法。该项目的目标是：1)确定在广泛不同地理区域的患者样本中，苍白球绦虫23s rRNA基因中A2058G和A2059G点突变的患病率；2)在这些样本的一个子集中对完整的23s rRNA基因（两个等位基因）进行测序，以确定可能与大环内酯类药物耐药性相关的其他突变；3)在建立的兔梅毒模型中，检验鉴定出的突变与阿奇霉素治疗临床失败的相关性。为了有效地治疗我们的梅毒患者，临床医生迫切需要知道梅毒T. 23S rDNA突变是否会导致阿奇霉素治疗失败的风险。我们的工作结果将为提出合理的治疗建议提供证据基础。

DESCRIPTION (provided by applicant): For many years, a single-dose oral treatment for syphilis has been sought. Benzathine penicillin G, used to treat syphilis for the past 50 years, has many disadvantages, including pain at injection sites, the possibility of an allergic reaction, and the inability of this form of penicillin to achieve treponemacidal levels in the central nervous system (a common site for Treponema pallidum dissemination). Published studies using the rabbit syphilis model demonstrated the effectiveness of azithromycin, an oral azalide antibiotic related to macrolides, for treating syphilis caused by the Nichols strain of T. pallidum. In several human studies, a single 1 or 2 gram dose of azithromycin was effective for treatment of early syphilis. However, recent publications document high frequencies of mutation A2058G in the 23S rRNA genes of circulating T. pallidum strains. In other bacteria, this mutation is associated with macrolide resistance. The Street 14 strain of T. pallidum contains the A2058G mutation, and, in the rabbit syphilis model, is resistant to macrolide and related drugs, including azithromycin. Similarly, azithromycin treatment failure has been documented in patients infected with T. pallidum bacteria that contain the same mutation. Nonetheless, a recent publication has questioned the clinical relevance of these 23S rDNA mutations; similar views have been voiced in discussions related to the development of the new syphilis treatment guidelines by the Centers for Disease Control and Prevention. Rather than continuing to speculate, it is essential to conduct the experiments that will directly determine whether these and other mutations impact the clinical response to azithromycin therapy. Although one could argue that sufficient equipoise exists (or does not exist) to justify a trial of azithromycin for treatment of patients with syphilis whose infecting organism harbors a 23S rDNA mutation, it is unlikely to be conducted in a timeframe that can inform our current clinical practice. The well-established rabbit model of syphilis is ideal for addressing the issue. The goals of the proposed project are 1) to determine the prevalence of A2058G and A2059G point mutations in the 23s rRNA genes of T. pallidum in samples collected from patients in widely disparate geographic regions; 2) to sequence the full 23s rRNA genes (both alleles) in a subset of these samples to identify other mutations that might be associated with macrolide resistance; and 3) to test the correlation of the identified mutations with clinical failure of azithromycin treatment in the well-established rabbit model of syphilis. To effectively treat our patients with syphilis, clinicians urgently need to know whether T. pallidum 23S rDNA mutations confer risk for azithromycin treatment failure. The results of our work will provide an evidence base on which sound treatment recommendations can be made. PUBLIC HEALTH RELEVANCE: The proposed studies will 1) determine whether mutations in the bacterium that causes syphilis affect response to treatment with azithromycin and 2) determine how frequently bacteria containing these mutations are present in syphilis patients from several geographical regions. If these mutations are found to be common and are shown to cause treatment failure, physicians will be advised not to use azithromycin for treating patients with syphilis.