Direct Visualization of Mucus Production by the Cold-Water Coral Lophelia pertusa with Digital Holographic Microscopy

Lophelia pertusa is the dominant reef-building organism of cold-water coral reefs, and is known to produce significant amounts of mucus, which could involve an important metabolic cost. Mucus is involved in particle removal and feeding processes, yet the triggers and dynamics of mucus production are currently still poorly described because the existing tools to study these processes are not appropriate. Using a novel microscopic technique—digital holographic microscopy (DHM)–we studied the mucus release of L. pertusa under various experimental conditions. DHM technology permits μm-scale observations and allows the visualization of transparent mucoid substances in real time without staining. Fragments of L. pertusa were first maintained in flow-through chambers without stressors and imaged with DHM, then exposed to various stressors (suspended particles, particulate food and air exposure) and re-imaged. Under non-stressed conditions no release of mucus was observed, whilst mucus strings and sheaths were produced in response to suspended particles (activated charcoal and drill cuttings sediment) i.e. in a stressed condition. Mucus strings and so-called ‘string balls’ were also observed in response to exposure to particulate food (brine shrimp Artemia salina). Upon air-exposure, mucus production was clearly visible once the fragments were returned to the flow chamber. Distinct optical properties such as optical path length difference (OPD) were measured with DHM in response to the various stimuli suggesting that different mucus types are produced by L. pertusa. Mucus produced to reject particles is similar in refractive index to the surrounding seawater, suggesting that the energy content of this mucus is low. In contrast, mucus produced in response to either food particle addition or air exposure had a higher refractive index, suggesting a higher metabolic investment in the production of these mucoid substances. This paper shows for the first time the potential of DHM technology for the detection, characterization and quantification of mucus production through OPD measurements in L. pertusa.

细枝柳珊瑚（Lophelia pertusa）是冷水珊瑚礁的优势造礁生物，已知其可分泌大量黏液，而这一过程可能伴随着高昂的代谢成本。黏液参与了颗粒物清除与摄食过程，但目前学界对黏液分泌的触发因素与动态变化仍知之甚少，原因是现有研究工具无法适配此类过程的观测需求。本研究借助一种新型显微技术——数字全息显微术（digital holographic microscopy, DHM），对细枝柳珊瑚在不同实验条件下的黏液释放行为展开了观测。该技术可实现微米级尺度的观测，并能在无需染色的情况下实时可视化透明黏液类物质。研究人员首先将细枝柳珊瑚的断枝置于无应激源的流水腔室中，利用DHM进行成像观测；随后将其暴露于各类应激源（悬浮颗粒物、颗粒性食物以及空气暴露环境）中，并再次进行成像。在无应激的条件下，未观测到黏液释放；而当暴露于悬浮颗粒物（活性炭与钻井岩屑沉积物）这类应激环境时，细枝柳珊瑚会分泌黏液丝与黏液鞘。当暴露于颗粒性食物（卤虫Artemia salina）时，同样可观测到黏液丝与所谓的"丝团"结构。在空气暴露处理后，当珊瑚断枝被放回流水腔室时，其黏液分泌现象清晰可见。研究人员通过DHM测量了不同刺激下的独特光学特性，例如光程差（optical path length difference, OPD），结果表明细枝柳珊瑚可分泌不同类型的黏液。用于排斥颗粒物的黏液折射率与周围海水相近，这意味着该类黏液的能量含量较低。与之相反，针对颗粒性食物添加或空气暴露所分泌的黏液折射率更高，表明这类黏液的合成需要更高的代谢投入。本研究首次证实了DHM技术通过光程差测量，用于检测、表征并定量细枝柳珊瑚黏液分泌的潜力。