The role of sex in the life cycle and transmission of Pneumocystis

肺孢子虫属是宿主专性真菌病原体，可导致致命性肺炎（PCP）， 免疫受损的宿主很少有药物对PCP有效，也没有新的治疗方法。 几十年的治疗。PCP仍然是艾滋病毒感染患者的一个问题， 新的易感人群，包括接受慢性病免疫治疗的患者。 我们最近报道了用棘白菌素治疗小鼠模型中的PCP，棘白菌素是一种抗真菌药物， 靶向β-1，3-D-葡聚糖合成（BG），耗尽了含有BG的子囊，但大量含有BG的形式 不表达BG的细胞保留在感染的肺中，显然不能增殖。值得注意的是， 小鼠在没有腹水的情况下不能传播感染。棘白菌素在给药时可预防PCP 在较高剂量下，这表明通过性周期形成asci可能是一种 生产性感染与性复制和细胞周期扰动相关的基因包括 来自阿尼芬净处理的小鼠的P. murina中最强的上调信号。这些感染缺乏 asci的结果表明，鼠疟原虫试图进行有性复制，但由于缺乏BG而不能进行。 基于这些数据，我们认为asci，因此，性复制，是需要的进展， 肺孢子虫的生命周期性周期对肺孢子虫生存的重要性将是 使用阿尼芬净治疗和预防模型进行研究。具体目标涉及两大 问题1：生命周期的完成是否需要有性复制 肺孢子虫病吗三个目标将被用来解决这个问题。首先是评估复制 通过将它们转移到新的宿主来增强用阿尼芬净处理的鼠疟原虫的能力。复制将是 通过显微镜方法、BG测量和定量PCR的组合来确定。第二 aim将使用预防性模型来允许低水平的复制，以询问复制是否通过性细胞发生。 循环，使用相同的方法。第三个目标是确定鼠肺吸虫是否能在长时间的 阿尼芬净治疗在同一宿主中进行，而不通过性周期进行，或者如果最终 通关问题2：在肺孢子虫生命周期的哪个阶段可以传播感染？的 阿尼芬净停药后性周期恢复。在戒烟后的10天到4周内，我们将跟踪 性周期的出现，并确定基因签名，生物标志物和ASCI的数量， 将这些小鼠暴露给未接受过实验的受体。利用这些数据，我们将确定在生命周期的哪个阶段 传输的发生和传输所需的阈值。基于基因选择的蛋白质 表达数据将通过蛋白质组学验证，作为与性功能恢复相关的潜在生物标志物。 循环和传输。这些研究将确定肺孢子虫生命周期中的潜在弱点， 可用于阻截，并增加我们对其生命周期和传播的了解。

Pneumocystis spp. are host obligate fungal pathogens that cause a fatal pneumonia (PCP) in immunocompromised hosts. Few drugs are effective against PCP and there have been no new therapies for its treatment in decades. PCP remains a problem in HIV-infected patients and is emerging as a clinical concern in newly susceptible populations including patients receiving immunotherapy for chronic conditions. We recently reported that treatment of PCP in murine models with the echinocandins, anti-fungal agents that target β-1,3-D-glucan synthesis (BG), depleted the asci which contain BG, but large numbers of forms that do not express BG remained in the infected lungs and were apparently unable to proliferate. Notably, the treated mice could not transmit the infection without asci. The echinocandins prevented PCP when administered prophylactically at higher doses, suggesting that formation of asci via the sexual cycle may be required for a productive infection. Genes associated with sexual replication and cell cycle perturbation comprised the strongest up-regulated signals in P. murina from anidulafungin treated mice. These infections were devoid of asci, suggesting that P. murina attempted to undergo sexual replication, but could not due to a lack of BG. Based on these data, we posit that asci, and thus sexual replication, is required for progression through the Pneumocystis life cycle. The importance of the sexual cycle for survival of Pneumocystis will be investigated using the anidulafungin treatment and prophylactic models. The specific aims address 2 major questions we propose to pursue: Question 1: Is sexual replication required for completion of the life cycle of Pneumocystis? 3 aims will be used to address this question. The first will be to assess the replication competency of P. murina treated with anidulafungin by transferring them to new hosts. Replication will be determined by a combination of microscopic methods, BG measurement, and quantitative PCR. The second aim will use the prophylactic model to allow low level replication to ask whether replication occurs via the sexual cycle, using the same methods. The third aim will determine whether P. murina can survive during prolonged anidulafungin treatment in the same hosts without proceeding through the sexual cycle or if it is eventually cleared. Question 2: At what point in the Pneumocystis life cycle can the infection be transmitted? The sexual cycle returns after withdrawal of anidulafungin. During 10 days to 4 weeks post-cessation, we will track the emergence of the sexual cycle and identify gene signatures, biomarkers and numbers of asci prior to exposing these mice to naïve recipients. Using these data, we will determine at what point in the life cycle transmission takes place and the thresholds necessary for transmission. Selected proteins based on gene expression data will be validated by proteomics as potential biomarkers associated with return of the sexual cycle and transmission. These studies will identify potential vulnerabilities in the Pneumocystis life cycle that could be exploited for interdiction and add to our understanding of its life cycle and transmission.