PspA-mediated aggregation protects Streptococcus pneumoniae against desiccation on fomites.
PspA-mediated aggregation protects Streptococcus pneumoniae against desiccation on fomites.
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肺炎链球菌（Spn）居住在鼻咽部，在那里它可以传播引起疾病。一个关键的Spn毒力因子是肺炎球菌表面蛋白A (PspA)，它通过阻断抗菌肽乳铁蛋白促进存活。PspA也被证明通过结合细胞表面结合的哺乳动物(m)GAPDH介导下气道死亡上皮细胞的附着。重要的是，PspA在定植过程中的作用尚不清楚。在无症状定植小鼠的鼻灌洗液中，野生型Spn的含量比其等基因pspA缺陷突变体（∆pspA）高约10倍。野生型Spn也在由脱落的上皮细胞和数百个肺炎球菌组成的粘膜分泌物中形成聚集体，而∆pspA则没有。在这些聚集体中心的Spn比单个肺炎球菌更能在螨虫上长时间的干燥中存活，并且能够感染naïve小鼠，这表明pspa介导的聚集体具有生存/传播优势。Spn在含mGAPDH的盐水中孵育也增强了对干燥的耐受性，但仅限于野生型Spn。mGAPDH足以引起野生型Spn的低水平聚集，但不能引起∆pspA的聚集。在菌株WU2中，PspA负责结合GAPDH的亚结构域（aa230-281）位于乳铁蛋白（LF）结合结构域（aa167-288）内。我们观察到LF抑制gapdh介导的聚集和干燥耐受性。利用表面等离子体共振，我们确定了Spn在其表面形成PspA-GAPDH-LF的多聚配合物，并且LF会排出GAPDH。我们的研究结果对肺炎球菌定植/传播过程和针对这种致命病原体的以pspa为重点的持续免疫工作具有重要意义。Spn是一种危险的人类病原体，能够引起肺炎和侵袭性疾病。毒力因子PspA已经被研究了近40年，在肺炎球菌逃避c反应蛋白和中和乳铁蛋白中发挥了明确的作用。在此，我们表明哺乳动物粘膜分泌物中的GAPDH以ppa依赖的方式促进肺炎球菌的聚集，而乳铁蛋白则对抗这种作用。pspa介导的gapdh依赖性细菌聚集保护鼻腔灌洗洗脱液中的Spn，并使其在体外培养时免受真菌干燥的影响。此外，在这些聚集体中存活的肺炎球菌在干燥后保留了定殖naïve宿主的能力。我们报道了Spn结合并在其表面形成由PspA、mGAPDH和乳铁蛋白组成的蛋白复合物。因此，这些蛋白水平的变化很可能对Spn的定植、在虫体上的存活和传播具有重要意义。
Streptococcus pneumoniae (Spn) resides in the nasopharynx, where it can disseminate to cause disease. One key Spn virulence factor is pneumococcal surface protein A (PspA), which promotes survival by blocking the antimicrobial peptide lactoferricin. PspA has also been shown to mediate attachment to dying epithelial cells in the lower airway due to its binding of cell surface-bound mammalian (m)GAPDH. Importantly, the role of PspA during colonization is not well understood. Wild-type Spn was present in nasal lavage elutes collected from asymptomatically colonized mice at levels ~10-fold higher than its isogenic PspA-deficient mutant (∆pspA). Wild-type Spn also formed aggregates in mucosal secretions composed of sloughed epithelial cells and hundreds of pneumococci, whereas ∆pspA did not. Spn within the center of these aggregates better survived prolonged desiccation on fomites than individual pneumococci and were capable of infecting naïve mice, indicating PspA-mediated aggregation conferred a survival/transmission advantage. Incubation of Spn in saline containing mGAPDH also enhanced tolerance to desiccation, but only for wild-type Spn. mGAPDH was sufficient to cause low-level aggregation of wild-type Spn but not ∆pspA. In strain WU2, the subdomain of PspA responsible for binding GAPDH (aa230–281) is ensconced within the lactoferrin (LF)-binding domain (aa167-288). We observed that LF inhibited GAPDH-mediated aggregation and desiccation tolerance. Using surface plasmon resonance, we determined that Spn forms multimeric complexes of PspA-GAPDH-LF on its surface and that LF dislodges GAPDH. Our findings have important implications regarding pneumococcal colonization/transmission processes and ongoing PspA-focused immunization efforts for this deadly pathogen. Spn is a dangerous human pathogen capable of causing pneumonia and invasive disease. The virulence factor PspA has been studied for nearly four decades with well-established roles in pneumococcal evasion of C-reactive protein and neutralization of lactoferricin. Herein, we show that mammalian (m)GAPDH in mucosal secretions promotes aggregation of pneumococci in a PspA-dependent fashion, whereas lactoferrin counters this effect. PspA-mediated GAPDH-dependent bacterial aggregation protected Spn in nasal lavage elutes and grown in vitro from desiccation on fomites. Furthermore, surviving pneumococci within these aggregates retained their ability to colonize naïve hosts after desiccation. We report that Spn binds to and forms protein complexes on its surface composed of PspA, mGAPDH, and lactoferrin. Changes in the levels of these proteins therefore most likely have critical implications on Spn colonization, survival on fomites, and transmission.
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