Laboratory Studies of Abnormal Host Defense and Immunoregulatory Diseases

在20财年，我们继续研究颗粒状杆菌变性，这是慢性肉芽肿性疾病(CGD)患者的新出现病原体。根据已发表的病例，CGD患者感染这种细菌的病死率为30%。然而，先前的研究表明，这种没有临床明显疾病的生物体的长期持久性也可能发生在一些患者身上。为了更好地了解这种细菌的发病机制，我们收集了这10例报告病例中的9例，并进行了完整的基因组测序和各种实验室研究，目的是解剖这种细菌的基因/表型特征。分离株之间可以看到显著的遗传多样性，包括高达11%的开放阅读框是特定分离株所独有的，尽管&gt；99%的16S rDNA序列是相同的。对基因组岛的详细分析正在进行中，有证据表明可能影响表型的几个噬菌体整合事件。我们分离出了一种可能是致死菌株特有的荚膜多糖，并正在通过MS和核磁共振对这种物质进行分析，以确定其组成。用这种多糖免疫兔子，并用免疫印迹、免疫沉淀和其他正在进行的方法评估产生的血清，目的是确定这种物质是否有助于产生它的菌株的毒力。 COVID19大流行对该项目的影响很大，因为实验室活动减少了。然而，在过去的几个月里，通过与LCIM内部其他小组的合作，我们对175多名记录在案的SARS-CoV2感染患者的血清和血浆中重要炎症介质的浓度进行了研究，并发现了特定生物标志物与临床结果的相关性。这些结果正在准备中，准备在20财年结束前提交。 在2010财年期间，我们完成了一份手稿，目前正在印刷中，该手稿描述了血浆明胶蛋白(PGSN)的实验室研究，pGSN是一种丰富的血浆蛋白，通过隔离肌动蛋白和血小板激活因子等促炎物质来减少全身炎症。我们检测了慢性肉芽肿性疾病(CGD)患者的样本中的pGSN，发现与健康对照组相比，pGSN浓度显著降低。这一发现可能在一定程度上解释了这种疾病的过度炎症特征，并提出了提供明胶是否有助于控制这些患者的炎症反应的问题，特别是在感染期间。

During FY20, we continued our studies of Granulibacter bethesdensis, emerging pathogen in patients with chronic granulomatous disease (CGD). Based on published cases, infection of CGD patients with this organism has a case fatality rate of 30%. Previous studies have shown, however, that long-term persistence of this organism without clinically apparent disease may also occur in some patients. To better understand pathogenesis by this organism, we have collected isolates from 9 of these 10 reported cases and performed complete genome sequencing as well as a variety of laboratory studies aimed dissecting genotype/phenotype characteristics of this organism. Remarkable genetic diversity may be seen between isolates including up to 11% of open reading frames being unique to a specific isolate despite >99% identical 16S rDNA sequences. Detailed analysis of genomic islands is underway with evidence of several bacteriophage integration events that may impact phenotypes. We isolated a putative capsular polysaccharide that appears to be unique to lethal isolates and are analyzing this material by MS and NMR to identify its composition. Rabbits were immunized with this polysaccharide and resulting sera evaluated by immunoblotting, immunoprecipitation, and other ongoing approaches with an aim toward determining if this material contributes to the virulence of strains producing it. The impact of the COVID19 pandemic on this project has been significant in terms of decreased laboratory activities. Nevertheless, through collaborations with other groups within LCIM during the past several months, we have performed studies on the concentrations of important inflammatory mediators in serum and plasma from over 175 patients with documented infections by SARS-CoV2 and discovered correlations of particular biomarkers with clinical outcomes. These results are being prepared for submission before the end of FY20. During FY20 we completed a manuscript that is now in press that describes laboratory studies of plasma gelsolin (pGSN), an abundant plasma protein that decreases systemic inflammation by sequestering pro-inflammatory substances such as actin and platelet activating factor. We measured pGSN in samples from patients with Chronic Granulomatous Disease (CGD) and detected a significantly lower concentration compared to healthy controls. This finding may, in part, explain the excessive inflammation characterizing this disease and raising the question of whether or not providing gelsolin might help control inflammatory responses in these patients, particularly during infections.