Breaking Biomphalaria black box by in situ revelation of fluorescent Schistosoma mansoni parasites.

Tissue clearing is an old-fashioned method developed in the 1900's and used to turn an opaque biological object into a 3D visualizable transparent structure. Developed and diversified over the last decade, this method is most of the time applied to mammals' tissues, and especially mouse and human tissues for cytological, histological and pathophysiological studies. Through autofluorescence, immunofluorescence, in situ hybridization, intercalating agents, fluorescent transfection markers or fluorescent particle uptake, optically cleared samples can be monitored to discover new biological structures and cellular interactions through 3D-visualization, which can be more challenging in some extend through classical histological methods. Most of the tissue clearing procedures have been developed for specific applications like endogenous fluorescence visualization, immunolabeling or for revealing specific organs. Thus, choosing the adapted protocol may be empirical for non-model species, especially for mollusks for which very little related literature is available. Herein, we suggest an effective optical tissue clearing procedure for the freshwater snail Biomphalaria glabrata, known as the intermediate host of the human parasite Schistosoma mansoni. This clearing procedure involves solvents with a minimal toxicity, preserves the endogenous fluorescence of labeled parasites inside snail tissues and is compatible with an immunolabeling procedure.

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Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.