Larval mosquito feeding and microbial interactions

描述(由申请人提供)：在水生幼虫栖息地的成蚊的生长和生产受到折射有机物转化为可摄取和消化的微生物生物量的限制。有机物转化为微生物生物量的速度受到颗粒基质质量的影响，表现为抑制和促进微生物和幼虫生长的双重作用。我们的长期目标是描述蚊子产生的生物学基础和限制，确定这些资源的利用效率，并制定一个现实的幼虫生长模式。我们已经确定了用于研究的树洞生态系统，其中藏有三种Ochlerotatus、Ochlerotatus Japan onicus和库蚊，这两种媒介是人类病原体的媒介，一种是外来入侵者。研究方法包括可提供包括微生物在内的个别食物来源的无菌(无菌)实验系统；追踪幼虫食物来源的稳定同位素分析(13C/12C；15N/14N)；微生物群落结构的分子分析；编码与大型生物分子降解有关的酶的微生物基因的功能基因阵列分析，包括木聚糖酶、纤维素酶和几丁质酶；以及微观世界和自然实验，其中量化微生物生物量和相对于幼虫生长的代谢活动。提出了一个生长模型，其中发育到变态(即化蛹)的时间和质量被假设为灵活的生活史特征，其最小值在营养和遗传上被限制在定义个体大小和影响成虫生存的化蛹窗口中；两者都是关键的适应度参数，都是媒介能力的重要组成部分。实验将描绘它们之间的关系，并揭示导致它们变异的环境和遗传因素。目前对幼虫生长速度的理解要求转化效率(每摄入一毫克食物所同化的镁食物)远远高于观察到的，因此我们将反复测试相对于各种质量的天然微生物食物来源的生长反应，将观察到的生长数据与模型拟合，并相应地修正转化效率估计。将对共享相同幼虫栖息地和资源的三种蚊子进行比较，以评估种内和种间激烈竞争条件下的增长率，预测竞争结果和生态置换的可能性，并量化利用幼虫食物的相对效率。将利用分子遗传工具对微生物群落中的主要成员黄杆菌进行操纵，以表达编码具有幼虫毒素和生长抑制特性的蛋白的基因，包括来自芽孢杆菌的哭泣毒素基因和几丁质酶。与公共卫生有关的蚊子是传播大量人类病原体的罪魁祸首，包括那些引起疟疾、丝虫病和虫媒病毒感染的病原体。成蚊和雌蚊的丰度和寿命(取血和病原体传播阶段)取决于幼虫环境中的营养和其他因素，而这些因素的制约是本文研究的重点。

DESCRIPTION (provided by applicant): The growth and production of adult mosquitoes from the aquatic larval habitats is constrained by conversion of refractile organic material into ingestible and digestible microbial biomass. Rates of conversion of organic matter into microbial biomass are tempered by the quality of particulate substrates that is manifested in dual roles of inhibition and stimulation of microbial and larval growth. Our long-term goal is to delineate the biological basis for and constraints on mosquito production, to identify the efficiency of utilization of these resources, and to elaborate a realistic model of larval growth. We have identified the tree hole ecosystem for study, harboring the species Ochlerotatus triseriatus, Ochlerotatus japonicus, and Culex pipiens, two vectors of human pathogens and one an exotic invader. Research approaches include an axenic (germ-free) experimental system into which individual food sources including microorganisms can be provisioned; stable isotopic analyses (13C/12C; 15N/14N) to track larval foods to their sources; molecular analyses of microbial community structure; a functional gene array analysis of microbial genes encoding enzymes involved in large biomolecule degradation, including xylanases, cellulases, and chitinases; and microcosm and naturalistic experiments in which microbial biomass and metabolic activity relative to larval growth are quantified. A growth model is proposed in which developmental time to, and mass at, metamorphosis (i.e., pupation) are postulated to be flexible life history traits whose minima are nutritionally and genetically constrained into a pupation window defining body size and affecting adult survivorship; both are key fitness parameters and both important components of vectorial capacity. Experiments will delineate relationships among them, and reveal environmental and genetic components contributing to their variation. The current understanding of larval growth rates requires conversion efficiencies (mg food assimilated per mg food ingested) far higher than observed, thus we will iteratively test growth responses relative to natural microbial food sources of various qualities, fit observed growth data to models, and revise conversion efficiency estimates accordingly. The three mosquito species sharing the same larval habitat and resources will be compared to assess growth rates under conditions of intense intra- and interspecific competition, both to make predictions of competitive outcomes and potential for ecological displacement, and to quantify comparative efficiency of utilization of larval foods. Manipulation of the Flavobacterium component of the microbial community a dominant member with molecular genetic tools to express genes encoding proteins having larval toxin and growth inhibiting properties will be conducted, including cry toxin genes from Bacillus, and chitinases. PUBLIC HEALTH RELEVANCE Mosquitoes are responsible for transmission of large number of human pathogens, including those causing malaria, filariasis, and arboviral infections. Abundance and longevity of adult, female mosquitoes (the blood feeding and pathogen transmitting stage) are dependent upon the nutritional and other factors operating in the larval environment, the constraints of which are the focus of the research here.