Connecting structure and function from organisms to molecules in small-animal symbioses through chemo-histo-tomography

Our understanding of metabolic interactions between small symbiotic animals and bacteria or parasitic eukaryotes that reside within their bodies is extremely limited. This gap in knowledge originates from a methodological challenge, namely to connect histological changes in host tissues induced by beneficial and parasitic (micro)organisms to the underlying metabolites. We addressed this challenge and developed chemo-histo-tomography (CHEMHIST), a culture-independent approach to connect anatomic structure and metabolic function in millimeter-sized symbiotic animals. CHEMHIST combines chemical imaging of metabolites based on mass spectrometry imaging (MSI) and microanatomy-based micro-computed X-ray tomography (micro-CT) on the same animal. Both high-resolution MSI and micro-CT allowed us to correlate the distribution of metabolites to the same animal's three-dimensional (3D) histology down to submicrometer resolutions. Our protocol is compatible with tissue-specific DNA sequencing and fluorescence in situ hybridization for the taxonomic identification and localization of the associated micro(organisms). Building CHEMHIST upon in situ imaging, we sampled an earthworm from its natural habitat and created an interactive 3D model of its physical and chemical interactions with bacteria and parasitic nematodes in its tissues. Combining MSI and micro-CT, we present a methodological groundwork for connecting metabolic and anatomic phenotypes of small symbiotic animals that often represent keystone species for ecosystem functioning.

目前学界对于小型共生动物与其体内定植的细菌或寄生真核生物之间的代谢互作认知极为有限。这一认知空白源于一项方法学难题：即如何将有益与寄生（微）生物诱导的宿主组织的组织学变化，与其潜在的代谢产物建立关联。我们攻克了这一难题，开发出非培养依赖型的化学组织断层成像（chemo-histo-tomography, CHEMHIST）技术，用于关联毫米级小型共生动物的解剖结构与代谢功能。CHEMHIST技术可在同一样本动物上，整合基于质谱成像（mass spectrometry imaging, MSI）的代谢物化学成像，以及基于微解剖结构的微型X射线计算机断层成像（micro-computed X-ray tomography, micro-CT）。凭借高分辨率的质谱成像与微型X射线计算机断层成像技术，我们可将代谢物的分布情况，与该样本动物的三维（3D）组织学特征进行关联，分辨率可达亚微米级别。该实验方案可与组织特异性DNA测序、荧光原位杂交（fluorescence in situ hybridization）技术兼容，用于对相关微（生）物进行分类鉴定与定位。我们基于原位成像构建了CHEMHIST技术体系，从自然栖息地采集了一条蚯蚓，并构建了其体内细菌与寄生线虫的物理、化学互作的交互式三维模型。通过整合质谱成像与微型X射线计算机断层成像技术，本研究为关联小型共生动物的代谢表型与解剖表型提供了方法学基础——这类小型共生动物往往是维系生态系统功能的关键物种。