Longitudinal TprK profiling of in vivo and in vitro-propagated Treponema pallidum subsp. pallidum reveals accumulation of antigenic variants in absence of immune pressure.
Longitudinal TprK profiling of in vivo and in vitro-propagated Treponema pallidum subsp. pallidum reveals accumulation of antigenic variants in absence of immune pressure.
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体内和体外繁殖的梅毒螺旋体亚种的纵向TprK谱梅毒显示在没有免疫压力的情况下抗原变异体积聚。
DOI：
10.1371/journal.pntd.0009753
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发表时间：
2021-09
影响因子：
3.8
通讯作者：
Greninger AL
中科院分区：
文献类型：
--
作者：
Lin MJ;Haynes AM;Addetia A;Lieberman NAP;Phung Q;Xie H;Nguyen TV;Molini BJ;Lukehart SA;Giacani L;Greninger AL
梅毒螺旋体（Treponema pallidum subspecies pallidum，T. pallidum）的致病性归因于其推定的外膜蛋白TprK的抗原变异。在TprK中，氨基酸多样性限于七个可变（V）区，并且V区内的序列多样性的产生经由非相互的节段性基因转换机制发生，其中供体盒重组到tprK表达位点中。尽管先前的研究已经显示了免疫选择在驱动TprK变体积累中的重要作用，但基线基因转换活性对变体多样性的贡献尚不清楚。在这里，将来自免疫活性和免疫抑制兔的近克隆芝加哥C的纵向tprK深度测序沿着新开发的T.苍白球，我们直接特点TprK等位基因的存在和不存在免疫选择。我们的数据证实了显着更大的序列多样性随着时间的推移，在梅毒感染期间免疫活性的兔与免疫抑制兔相比，在V6区域内，与以前的研究TprK在逃避宿主免疫反应的作用相一致。与在免疫活性兔中生长的菌株相比，在体外传代的菌株显示出与在免疫抑制兔中传代的菌株相似的TprK可变区序列的等位基因频率的低水平变化。值得注意的是，我们发现V6等位基因的产生率显着增加相对于其他可变区在体外培养的梅毒螺旋体菌株，说明这些高变区内的多样性发生在完全没有免疫选择。总之，我们的结果表明T。苍白球可以在体外进行研究，甚至在完全没有免疫压力的情况下也会发生，这使得T。苍白球种群不断逃避感染宿主的免疫系统。梅毒仍然是一种全球和公共卫生关注的疾病，尽管这种感染可以很容易地诊断并用青霉素有效治疗。虽然感染者通常对病原体产生较强的免疫力，但反复感染梅毒病原体，梅毒螺旋体梅毒亚种（T。pallidum），是可能的。一些研究指出T.苍白球TprK蛋白作为负责逃避在感染、病原体持续存在和再感染期间产生的免疫的机制。过去的研究强调了免疫清除显性变异的重要性，这反过来又允许较少代表性的变异出现。在tprK基因的变异性的免疫无关的基线生成的贡献是不太清楚的。在这里，我们使用实验室分离的T. pallidum菌株几乎与tprK等基因，其在体外随着时间的推移而繁殖，其中没有免疫压力施加在病原体上，以及在从用相同菌株感染的免疫抑制和免疫活性兔获得的样品中。我们证实，tprK积累的免疫压力下，显着更多的多样性，并证明了一个低的，但可辨别的基础率的基因转换完全没有免疫压力。
Immune evasion by Treponema pallidum subspecies pallidum (T. pallidum) has been attributed to antigenic variation of its putative outer-membrane protein TprK. In TprK, amino acid diversity is confined to seven variable (V) regions, and generation of sequence diversity within the V regions occurs via a non-reciprocal segmental gene conversion mechanism where donor cassettes recombine into the tprK expression site. Although previous studies have shown the significant role of immune selection in driving accumulation of TprK variants, the contribution of baseline gene conversion activity to variant diversity is less clear. Here, combining longitudinal tprK deep sequencing of near clonal Chicago C from immunocompetent and immunosuppressed rabbits along with the newly developed in vitro cultivation system for T. pallidum, we directly characterized TprK alleles in the presence and absence of immune selection. Our data confirm significantly greater sequence diversity over time within the V6 region during syphilis infection in immunocompetent rabbits compared to immunosuppressed rabbits, consistent with previous studies on the role of TprK in evasion of the host immune response. Compared to strains grown in immunocompetent rabbits, strains passaged in vitro displayed low level changes in allele frequencies of TprK variable region sequences similar to that of strains passaged in immunosuppressed rabbits. Notably, we found significantly increased rates of V6 allele generation relative to other variable regions in in vitro cultivated T, pallidum strains, illustrating that the diversity within these hypervariable regions occurs in the complete absence of immune selection. Together, our results demonstrate antigenic variation in T. pallidum can be studied in vitro and occurs even in the complete absence of immune pressure, allowing the T. pallidum population to continuously evade the immune system of the infected host. Syphilis continues to be a disease of global and public health concern, even though the infection can be easily diagnosed and effectively treated with penicillin. Although infected individuals often develop a strong immunity to the pathogen, repeated infection with the syphilis agent, Treponema pallidum subspecies pallidum (T. pallidum), is possible. Several studies point at antigenic variation of the T. pallidum TprK protein as the mechanism responsible for evasion of the immunity that develops during infection, pathogen persistence, and re-infection. Past studies have highlighted the importance of immune clearance of dominant variants that, in turn, allows less represented variants to emerge. The contribution of an immunity-independent baseline generation of variability in the tprK gene is less clear. Here, we used deep sequencing to profile tprK variants using a laboratory-isolated T. pallidum strain nearly isogenic for tprK that was propagated over time in vitro, where no immune pressure is exerted on the pathogen, as well as in samples obtained from immunosuppressed and immunocompetent rabbits infected with the same strain. We confirmed that tprK accumulates significantly more diversity under immune pressure, and demonstrated a low but discernible basal rate of gene conversion in complete absence of immune pressure.
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