Identification and characterization of fungal exposures

真菌暴露的鉴定和表征

• 批准号: 9004275
• 金额: $ 15.04万
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9004275
• 关键词: Air; Allergens; Allergic; Alternaria; Antigens; Area; Aspergillus; Asthma; Beds; Behavior; Biological Markers; Chaetomium; Child; Commercial Sectors; Connecticut; Data; Data Set; Development; Dust; Environment; Environmental Risk Factor; Exhalation; Family; Floor; Fungal Antigens; Genes; Genome; Genomics; Goals; Health; Home environment; House Dust; Housing; IgE; Immunoassay; Income; Individual; Infiltration; Location; Maine; Measures; Methodology; Methods; Modeling; Molds; Monoclonal Antibodies; Mycotoxins; Neighborhoods; New York City; Nitric Oxide; Occupational; Organism; Penicillium; Prevalence; Proteins; RNA Sequences; Reagent; Recombinant Proteins; Recombinants; Relative (related person); Ribosomal RNA; Sampling; Schools; Technology; Testing; United States; Vermont; Water; Work; airway inflammation; anthropogenesis; assay development; asthmatic; built environment; design; enolase; expression cloning; fungus; improved; indexing; insight; pullulan; remediation; transcriptome sequencing

To more completely assess the fungal species to which people are exposed, a fungal ribosomal RNA (rRNA) gene sequencing study was designed to test the hypothesis that fungal bioaerosols in the United States indoor built environments are much more diverse than previously estimated using traditional methods of analysis. In FY15, fungal diversity was evaluated in 350 dust samples derived from the bed, bedroom floor and kitchen areas from the homes of asthmatic children and/or mold remediation workers in New York City using Illumina miSeq and compared to results obtained using qPCR. The analysis of fungal diversity in samples derived from these individual studies provided new datasets that have provided greater insight into the fungal species that are frequently recovered in built environments. Analysis of the dust samples showed that Aureobasidium pullulans, Penicillium glabrum, Wallemia sebi and Alternaria alternata varied by housing type (single, multi-family or apartment) and neighborhood asthma prevalence. The preliminary results suggest that multiple environmental factors including anthropogenic behavior, housing type, and neighborhood are important variables that influence fungal diversity within middle-income homes in New York City. A. alternata measured in house dust was associated with fractional exhaled nitric oxide, specifically among children with higher combustion byproduct exposure, suggesting a possible interaction between these two exposures on airway inflammation. As an additional component of this project, NIOSH is comparing the Environmental Relative Moldiness Index (ERMI) to fungal rRNA sequencing data derived from air and dust samples collected from homes with well characterized fungal contamination. The ERMI has been increasingly used by the commercial sector and identifies fungi that typically thrive in moist and wet conditions associated with water infiltration. An expanded dataset will improve the ERMI panel so that it more fully represents the complete spectrum of fungi present in other types of environments. These studies are examining data from a number of different study locations in the United States to categorize and sequence the diversity of fungal rRNA in contaminated and non-contaminated residential and occupational environments. The study locations include homes in Atlanta, Georgia, office buildings in Hartford, Connecticut and Bennington, Vermont, and a school in Wiscasset Maine. A third aspect of this work is the identification and characterization of cross-reactive fungal biomarkers. The overall aim of this portion of the project is to identify broadly cross-reactive fungal allergens with the aim of developing improved immunological methods for detecting fungal bioaerosols and allergic sensitization. To accomplish this, NIOSH is developing species-specific monoclonal antibodies (mAbs) to recombinant fungal antigens. These mAbs will be important for the quantification of species-specific biomarkers of fungal exposure. Each mAb will be tested for reactivity against a panel of closely and distantly related occupational contaminants. MAbs that recognize the same antigen will be selected for further characterization and used in the development of either inhibition or sandwich ELISAs. Aspergillus terreus was initially used as a model fungal organism for these studies. The genome of Chaetomium globosum has now also been sequenced, allowing for the development of fungal-specific proteins using recombinant technologies. Three mAbs that react with the Chaetomium globosum enolase have been developed and characterized. A similar approach is currently being used to identify and characterize additional candidate fungal allergens from Ulocladium chartarum and to develop mAbs that can be used to track exposure and effect. Ulocladium chartarum is a recognized biomarker of moisture infiltration but there are no immunoassays available to detect this species in contaminated environments. The reagents developed in this project will be used to characterize IgE reactivity to cross-reactive allergens shared between allergenic fungal species.

为了更全面地评估人们接触的真菌种类，设计了一项真菌核糖体RNA(RRNA)基因测序研究，以检验这一假设，即美国室内建筑环境中的真菌生物气溶胶比以前使用传统分析方法估计的更多样化。在2015财年，使用Illumina MiSeq对来自纽约市哮喘儿童和/或霉菌修复工人家中的床、卧室地板和厨房区域的350个粉尘样本进行了真菌多样性评估，并与使用qPCR获得的结果进行了比较。对来自这些单独研究的样本中真菌多样性的分析提供了新的数据集，使人们能够更深入地了解经常在建筑环境中恢复的真菌物种。对粉尘样本的分析表明，不同房屋类型(单户、多户或公寓)和社区哮喘患病率不同，有出芽短梗霉、光滑青霉菌、黑斑病和交链格孢菌。初步结果表明，包括人为行为、住房类型和社区在内的多种环境因素是影响纽约市中等收入家庭真菌多样性的重要变量。在室内粉尘中测得的交链霉菌与呼出的部分一氧化氮有关，特别是在燃烧副产物暴露较高的儿童中，这表明这两种暴露之间可能存在对呼吸道炎症的相互作用。作为该项目的另一个组成部分，NIOSH正在将环境相对霉菌指数(ERMI)与真菌rRNA测序数据进行比较，这些数据来自于从真菌污染特征良好的家庭收集的空气和粉尘样本。ERMI越来越多地被商业部门使用，它识别出通常在潮湿和潮湿的条件下与水渗透相关的真菌。扩展的数据集将改进ERMI面板，使其更全面地代表其他类型环境中存在的真菌的完整光谱。这些研究正在检查美国许多不同研究地点的数据，以对污染和非污染的居住和职业环境中真菌rRNA的多样性进行分类和排序。研究地点包括佐治亚州亚特兰大的住宅，康涅狄格州哈特福德和佛蒙特州本宁顿的写字楼，以及缅因州威斯卡塞特的一所学校。这项工作的第三个方面是识别和表征交叉反应真菌生物标志物。该项目这一部分的总体目标是确定广泛交叉反应的真菌变应原，目的是开发检测真菌生物气雾剂和过敏性致敏的改进免疫学方法。为了实现这一目标，NIOSH正在开发针对重组真菌抗原的物种特异性单抗(MAbs)。这些单抗对于量化真菌暴露的物种特异性生物标志物将是重要的。每一种单抗都将接受对一组密切相关和遥远相关的职业污染物的反应性测试。识别相同抗原的单抗将被选择用于进一步鉴定，并用于抑制或夹心ELISA的开发。土曲霉最初被用作这些研究的真菌模式生物。球毛壳菌的基因组现在也已经被测序，从而可以使用重组技术开发真菌特有的蛋白质。已研制出三种与球毛壳烯醇化酶反应的单抗，并对其进行了鉴定。目前正在使用一种类似的方法来确定和表征来自轮状Ulocladium的其他候选真菌变应原，并开发可用于跟踪暴露和效果的单抗。Ulocladium chartarum是一种公认的水分渗透生物标志物，但目前还没有免疫分析方法来检测污染环境中的这种物种。在这个项目中开发的试剂将用于表征对致敏真菌物种之间共享的交叉反应变应原的IgE反应性。