Abstract
Amphibians are models for studying applied ecological issues such as habitat loss, pollution, disease, and global climate change due to their sensitivity and vulnerability to changes in the environment. Developmental series of amphibians are informative about their biology, and X-ray based 3D reconstruction holds promise for quantifying morphological changes during growth—some with a direct impact on the possibility of an experimental investigation on several of the ecological topics listed above. However, 3D resolution and discrimination of their soft tissues have been difficult with traditional X-ray computed tomography, without time-consuming contrast staining. Tomographic data were initially performed (pre-processing and reconstruction) using the open-source software tool SYRMEP Tomo Project. Data processing and analysis of the reconstructed tomography volumes were conducted using the segmentation semi-automatic settings of the software Avizo Fire 8, which provide information about each investigated tissues, organs or bone elements. Hence, volumetric analyses were carried out to quantify the development of structures in different tadpole developmental stages. Our work shows that synchrotron X-ray microtomography using phase-contrast mode resolves the edges of the internal tissues (as well as overall tadpole morphology), facilitating the segmentation of the investigated tissues. Reconstruction algorithms and segmentation software played an important role in the qualitative and quantitative analysis of each target structure of the Thoropa miliaris tadpole at different stages of development, providing information on volume, shape and length. The use of the synchrotron X-ray microtomography setup of the SYRMEP beamline of Elettra Synchrotron, in phase-contrast mode, allows access to volumetric data for bone formation, eye development, nervous system and notochordal changes during the development (ontogeny) of tadpoles of a cycloramphid frog Thoropa miliaris. As key elements in the normal development of these and any other frog tadpole, the application of such a comparative ontogenetic study, may hold interest to researchers in experimental and environmental disciplines.
Highlights
Amphibians are models for studying applied ecological issues such as habitat loss, pollution, disease, and global climate change due to their sensitivity and vulnerability to changes in the environment
Our goal is to investigate the ability of phase-contrast micro-CT for visualization of soft tissues and rigid internal structures at the same time, rather than to describe the anatomy in detail
Because the tadpoles are stored in formaldehyde 10%, in order to facilitate handling and avoid hazardous inhalation, specimens were previously washed in distilled water and passed in progressive 10 min baths of 20%, 50%, 70%, and absolute ethanol, in which they were stored for the tomography
Summary
Amphibians are models for studying applied ecological issues such as habitat loss, pollution, disease, and global climate change due to their sensitivity and vulnerability to changes in the environment. Data processing and analysis of the reconstructed tomography volumes were conducted using the segmentation semi-automatic settings of the software Avizo Fire 8, which provide information about each investigated tissues, organs or bone elements. Most studies of anuran development are based on whole preparations and cleared and bone-cartilage doublestained specimens[48,49,50] These studies, about bones and cartilaginous elements, have become standards for comparing sequence of skeleton development and in systematic s urveys[46]. They are d estructive[51], because the specimen’s other information (e.g. internal organs and muscles) is lost during the process of preparation. One of the main disadvantages of conventional X-ray imaging techniques for application in biological samples is that these samples generally have low density, causing the emission of hard X-rays from conventional microtomographs to have even less absorption
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
