scholarly journals Developmental Table and Three-Dimensional Embryological Image Resource of the Ascidian Ascidiella aspersa

2021 ◽  
Vol 9 ◽  
Author(s):  
Haruka M. Funakoshi ◽  
Takumi T. Shito ◽  
Kotaro Oka ◽  
Kohji Hotta
Keyword(s):  
Laser Scanning ◽  
Developmental Stages ◽  
Model Organism ◽  
Tail Length ◽  
Developmental Time ◽  
Laser Scanning Microscopy ◽  
System Level ◽  
Confocal Laser ◽  
Cross Sectional ◽  
Ascidiella Aspersa

Ascidiella aspersa is an ascidian in the class of chordates—the closest relatives of vertebrates. A. aspersa is a potential model organism for bio-imaging studies due to its extremely transparent embryos as well as is a globally distributed cosmopolitan species. However, there is no standard developmental table for this organism. Here, as a first step to establish A. aspersa as a model organism, we report a standard developmental table as a web-based digital image resource. This resource used confocal laser scanning microscopy to scan more than 3,000 cross-sectional images and 3D-reconstructed images of A. aspersa embryos during embryogenesis. With reference to the standardized developmental table of Ciona intestinalis type A, 26 different developmental stages (Stages 1–26) from fertilized eggs to hatched larvae were redefined for A. aspersa. Cell lineages up to the cleavage period were annotated: The cleavage patterns, the embryonic morphology, and the developmental time were then compared with Ciona. We found that the cleavage patterns and developmental time up to the neurula period in A. aspersa were extremely conserved versus. Ciona. The ratio of the trunk and tail length in the tailbud period were smaller than Ciona indicating a relatively short tail. In addition, the timing of the bending of the tail is earlier than Ciona. This A. aspersa standard 3D digital resource is essential for connecting different omics data to different spatiotemporal hierarchies and is useful for a system-level understanding of chordate development and evolution.

scholarly journals Confocal Laser Scanning Microscopy and Image Analysis for Elucidating Crumb and Crust Microstructure of Bran-Enriched South African Fried Dough and Batter

Foods ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 605 ◽  
Author(s):  
Oluwatoyin O. Onipe ◽  
Daniso Beswa ◽  
Afam I. O. Jideani
Keyword(s):  
Image Analysis ◽  
Laser Scanning ◽  
Polynomial Regression ◽  
Initial Moisture Content ◽  
Soxhlet Extraction ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Cross Sectional ◽  
Linear Effect ◽  
Staining Protocol

A double staining protocol for image acquisition using confocal microscopy (CLSM) coupled with image analysis was employed to elucidate the crust and cross-sectional properties of fried dough. Penetrated oil by image analysis (POia), porosity and pore features were quantified from the cross-section micrographs. Crust surface roughness was measured using fractal metrics and fat content was determined by solvent extraction using the American Association of Cereal Chemists method. Crumb porosity ranged between 54.94%–81.84% and reduced (p < 0.05) with bran addition. Crumb pore sizes ranged from 0–475 µm with <1 circularity, indicating elliptical shape. POia values were notably higher (p < 0.05) than PO by Soxhlet extraction (POsox), except for wheat bran (WB) fried dough where the values of POia and POsox were closely ranked. The linear effect of initial moisture content and bran concentration showed a significant impact on the image properties. The mean fractal dimension (FD) decreased as initial moisture increased. The addition of WB caused a significant reduction in the FD of fried dough, while the opposite effect was noted for its oat bran counterpart. Due to non-collinearity of image properties (FD, POia and porosity), data were fitted to cubic polynomial regression with R2 values > 0.70. CLSM and image analysis were effective in measuring oil absorption and interpreting crumb properties of fried dough. The protocol used in this study can be applied to other thick deep-fried foods for qualitative observation and quantitative measurement of a specific physical or chemical property.

scholarly journals Protocol for rapid clearing and staining of fixed Arabidopsis ovules for improved imaging by confocal laser scanning microscopy

Plant Methods ◽  
2019 ◽  
Vol 15 (1) ◽  
Author(s):  
Rachele Tofanelli ◽  
Athul Vijayan ◽  
Sebastian Scholz ◽  
Kay Schneitz
Keyword(s):  
Laser Scanning ◽  
Developmental Stages ◽  
Imaging Techniques ◽  
Three Dimensional ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Imaging Data ◽  
Model Species ◽  
Cellular Models ◽  
Cellular Resolution

Abstract Background A salient topic in developmental biology relates to the molecular and genetic mechanisms that underlie tissue morphogenesis. Modern quantitative approaches to this central question frequently involve digital cellular models of the organ or tissue under study. The ovules of the model species Arabidopsis thaliana have long been established as a model system for the study of organogenesis in plants. While ovule development in Arabidopsis can be followed by a variety of different imaging techniques, no experimental strategy presently exists that enables an easy and straightforward investigation of the morphology of internal tissues of the ovule with cellular resolution. Results We developed a protocol for rapid and robust confocal microscopy of fixed Arabidopsis ovules of all stages. The method combines clearing of fixed ovules in ClearSee solution with marking the cell outline using the cell wall stain SCRI Renaissance 2200 and the nuclei with the stain TO-PRO-3 iodide. We further improved the microscopy by employing a homogenous immersion system aimed at minimizing refractive index differences. The method allows complete inspection of the cellular architecture even deep within the ovule. Using the new protocol we were able to generate digital three-dimensional models of ovules of various stages. Conclusions The protocol enables the quick and reproducible imaging of fixed Arabidopsis ovules of all developmental stages. From the imaging data three-dimensional digital ovule models with cellular resolution can be rapidly generated using image analysis software, for example MorphographX. Such digital models will provide the foundation for a future quantitative analysis of ovule morphogenesis in a model species.

scholarly journals Effect of a Bioactive Glass Ceramic on the Control of Enamel and Dentin Erosion Lesions

2017 ◽  
Vol 28 (4) ◽  
pp. 489-497 ◽  
Author(s):  
Michelle Alexandra Chinelatti ◽  
Camila Tirapelli ◽  
Silmara Aparecida Milori Corona ◽  
Renato Goulart Jasinevicius ◽  
Oscar Peitl ◽  
...  
Keyword(s):  
Bioactive Glass ◽  
Glass Ceramic ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Cross Sectional ◽  
Surface Loss ◽  
And Control ◽  
Bovine Enamel ◽  
Caries Lesions

Abstract This study evaluated the effect of a bioactive glass ceramic for the control of erosion and caries lesions. Fragments (n=10) of bovine enamel and root dentin received daily application of different treatments (Biosilicate; Acidulated Phosphate Fluoride- APF; Untreated - control) during the performance of erosive cycles. Surfaces were analyzed with 3D optical profilometry to quantify the superficial loss in four periods (1, 7, 14 and 21 days), as well as the lesion depth with confocal laser scanning microscopy. For caries progression assessment, initial Knoop microhardness was measured on enamel bovine fragments. Initial carious lesions were developed and specimens were divided into three groups (n=10), according to the daily topical application (Biosilicate; APF; no application - control), during the de-remineralization cycles for 14 days. Final microhardness was obtained to calculate the change of surface microhardness. Subsurface demineralization was analyzed using cross-sectional microhardness (depths 10, 30, 50, 70, 90, 110 and 220 µm). Data were tested using ANOVA and Tukey’s test (a=5%). Results of erosive evaluation showed that Biosilicate promoted the lowest (p<0.05) values of surface loss, regardless of time, for both enamel and dentin; APF promoted lower (p<0.05) surface loss than control; analyzing different periods of time, APF did not show difference (p>0.05) between 14 and 21 days of demineralization. Results of enamel caries assessment showed that Biosilicate resulted in higher (p<0.05) surface and subsurface microhardness than both APF and control-applications. It may be concluded that Biosilicate application showed a higher potential to reduce surface loss and development of erosion and caries lesions.

scholarly journals Ovule Development and in Planta Transformation of Paphiopedilum Maudiae by Agrobacterium-Mediated Ovary-Injection

2020 ◽  
Vol 22 (1) ◽  
pp. 84
Author(s):  
Bai-Xue Luo ◽  
Li Zhang ◽  
Feng Zheng ◽  
Kun-Lin Wu ◽  
Lin Li ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Developmental Stages ◽  
Embryo Sac ◽  
Mature Embryo ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Pcr Analysis ◽  
In Planta Transformation ◽  
In Planta ◽  
Hygromycin Selection

In this paper, the development of the Paphiopedilum Maudiae embryo sac at different developmental stages after pollination was assessed by confocal laser scanning microscopy. The mature seeds of P. Maudiae consisted of an exopleura and a spherical embryo, but without an endosperm, while the inner integument cells were absorbed by the developing embryo. The P. Maudiae embryo sac exhibited an Allium type of development. The time taken for the embryo to develop to a mature sac was 45-50 days after pollination (DAP) and most mature embryo sacs had completed fertilization and formed zygotes by about 50–54 DAP. In planta transformation was achieved by injection of the ovaries by Agrobacterium, resulting in 38 protocorms or seedlings after several rounds of hygromycin selection, corresponding to 2, 7, 5, 1, 3, 4, 9, and 7 plantlets from Agrobacterium-mediated ovary-injection at 30, 35, 42, 43, 45, 48, 50, and 53 DAP, respectively. Transformation efficiency was highest at 50 DAP (2.54%), followed by 2.48% at 53 DAP and 2.45% at 48 DAP. Four randomly selected hygromycin-resistant plants were GUS-positive after PCR analysis. Semi-quantitative PCR and quantitative real-time PCR analysis revealed the expression of the hpt gene in the leaves of eight hygromycin-resistant seedlings following Agrobacterium-mediated ovary-injection at 30, 35, 42, 43, 45, 48, 50, and 53 DAP, while hpt expression was not detected in the control. The best time to inject P. Maudiae ovaries in planta with Agrobacterium is 48-53 DAP, which corresponds to the period of fertilization. This protocol represents the first genetic transformation protocol for any Paphiopedilum species and will allow for expanded molecular breeding programs to introduce useful and interesting genes that can expand its ornamental and horticulturally important characteristics.

scholarly journals Automated Confocal Laser Scanning Microscopy and Semiautomated Image Processing for Analysis of Biofilms

1998 ◽  
Vol 64 (11) ◽  
pp. 4115-4127 ◽  
Author(s):  
Martin Kuehn ◽  
Martina Hausner ◽  
Hans-Joachim Bungartz ◽  
Michael Wagner ◽  
Peter A. Wilderer ◽  
...  
Keyword(s):  
Image Analysis ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Image Acquisition ◽  
Three Dimensional ◽  
Laser Scanning Microscopy ◽  
Three Dimensions ◽  
Confocal Laser ◽  
Cross Sectional ◽  
In Situ Conditions

ABSTRACT The purpose of this study was to develop and apply a quantitative optical method suitable for routine measurements of biofilm structures under in situ conditions. A computer program was designed to perform automated investigations of biofilms by using image acquisition and image analysis techniques. To obtain a representative profile of a growing biofilm, a nondestructive procedure was created to study and quantify undisturbed microbial populations within the physical environment of a glass flow cell. Key components of the computer-controlled processing described in this paper are the on-line collection of confocal two-dimensional (2D) cross-sectional images from a preset 3D domain of interest followed by the off-line analysis of these 2D images. With the quantitative extraction of information contained in each image, a three-dimensional reconstruction of the principal biological events can be achieved. The program is convenient to handle and was generated to determine biovolumes and thus facilitate the examination of dynamic processes within biofilms. In the present study, Pseudomonas fluorescens or a green fluorescent protein-expressing Escherichia coli strain, EC12, was inoculated into glass flow cells and the respective monoculture biofilms were analyzed in three dimensions. In this paper we describe a method for the routine measurements of biofilms by using automated image acquisition and semiautomated image analysis.

scholarly journals Extracellular excystation and development of Cryptosporidium: tracing the fate of oocysts within Pseudomonas aquatic biofilm systems

2021 ◽  
Author(s):  
Wan Koh ◽  
Andrew Thompson ◽  
Hanna Edwards ◽  
Paul Monis ◽  
Peta L Clode
Keyword(s):  
Laser Scanning ◽  
Distribution Systems ◽  
Water Distribution ◽  
Developmental Stages ◽  
Parasitophorous Vacuole ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Growth Environment ◽  
Flow Cytometric ◽  
Harsh Conditions

Background Aquatic biofilms often serve as environmental reservoirs for microorganisms and provide them with a nutrient-rich growth environment under harsh conditions. With regard to Cryptosporidium, biofilms can serve as environmental reservoirs for oocysts, but may also support the growth of additional Cryptosporidium stages. Results Here we used confocal laser scanning microscopy, scanning electron microscopy (SEM), and flow cytometry to identify and describe various Cryptosporidium developmental stages present within aquatic biofilm systems, and to directly compare these to stages produced in cell culture. We also show that Cryptosporidium has the ability to form a parasitophorous vacuole independently, in a host-free biofilm environment, potentially allowing them to complete an extracellular life cycle. Correlative data from confocal and SEM imaging of the same cells confirmed that the observed developmental stages (including trophozoites, meronts, and merozoites) were Cryptosporidium. These microscopy observations were further supported by flow cytometric analyses, where excysted oocyst populations were detected in 1, 3 and 6 day-old Cryptosporidium-exposed biofilms, but not in biofilm-free controls. Conclusions These observations not only highlight the risk that aquatic biofilms pose in regards to Cryptosporidium outbreaks from water distribution systems, but further indicate that even simple biofilms are able to stimulate oocyst excystation and support the extracellular multiplication and development of Cryptosporidium within aquatic environments.

scholarly journals From swimming towards sessility in two metamorphoses – the drastic changes in structure and function of the nervous system of the bay barnacle Amphibalanus improvisus (Crustacea, Thecostraca, Cirripedia) during development

2020 ◽  
Vol 89 (3) ◽  
pp. 324-352
Author(s):  
Paul Kalke ◽  
Thomas Frase ◽  
Stefan Richter
Keyword(s):  
Nervous System ◽  
Peripheral Nervous System ◽  
Nerve Cord ◽  
Laser Scanning ◽  
Ventral Nerve Cord ◽  
Developmental Stages ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Compound Eyes ◽  
Cephalic Shield

Knowledge about the development of the nervous system in cirripeds is limited, particularly with regard to the changes that take place during the two metamorphoses their larvae undergo. This study delivers the first detailed description of the development of the nervous system in a cirriped species, Amphibalanus improvisus by using immunohistochemical labeling against acetylated alpha-tubulin, and confocal laser scanning microscopy. The development of the nervous system in the naupliar stages corresponds largely to that in other crustaceans. As development progresses, the protocerebral sensory organs differentiate and the intersegmental nerves forming the complex peripheral nervous system appear, innervating the sensory structures of the cephalic shield. During metamorphosis into a cypris the lateral sides of the cephalic shield fold down into a bilateral carapace, which leads to a reorganization of the peripheral nervous system. The syncerebrum of the cypris exhibits the highest degree of complexity of all developmental stages, innervating the frontal filaments, nauplius eye, compound eyes and the antennules. During settlement, when the second metamorphosis occur, the closely associated frontal filaments and compound eyes are shed together with the cuticle of the carapace and the antennules. In adults, the syncerebral structures are reduced while the ventral nerve cord and the peripheral nervous system increase in complexity. The peripheral nervous system plays an important role in processing sensory input and also in settlement. In summary, through the larval development we observed a structural and thus also functional increase of complexity in favor of the peripheral nervous system and the ventral nerve cord.

scholarly journals Extracellular excystation and development of Cryptosporidium: tracing the fate of oocysts within Pseudomonas aquatic biofilm systems

2021 ◽  
Author(s):  
Wan Koh ◽  
Andrew Thompson ◽  
Hanna Edwards ◽  
Paul Monis ◽  
Peta L Clode
Keyword(s):  
Laser Scanning ◽  
Distribution Systems ◽  
Water Distribution ◽  
Developmental Stages ◽  
Parasitophorous Vacuole ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Growth Environment ◽  
Flow Cytometric ◽  
Harsh Conditions

Background Aquatic biofilms often serve as environmental reservoirs for microorganisms and provide them with a nutrient-rich growth environment under harsh conditions. With regard to Cryptosporidium, biofilms can serve as environmental reservoirs for oocysts, but may also support the growth of additional Cryptosporidium stages. Results Here we used confocal laser scanning microscopy, scanning electron microscopy (SEM), and flow cytometry to identify and describe various Cryptosporidium developmental stages present within aquatic biofilm systems, and to directly compare these to stages produced in cell culture. We also show that Cryptosporidium has the ability to form a parasitophorous vacuole independently, in a host-free biofilm environment, potentially allowing them to complete an extracellular life cycle. Correlative data from confocal and SEM imaging of the same cells confirmed that the observed developmental stages (including trophozoites, meronts, and merozoites) were Cryptosporidium. These microscopy observations were further supported by flow cytometric analyses, where excysted oocyst populations were detected in 1, 3 and 6 day-old Cryptosporidium-exposed biofilms, but not in biofilm-free controls. Conclusions These observations not only highlight the risk that aquatic biofilms pose in regards to Cryptosporidium outbreaks from water distribution systems, but further indicate that even simple biofilms are able to stimulate oocyst excystation and support the extracellular multiplication and development of Cryptosporidium within aquatic environments.

Sign in / Sign up

Export Citation Format

Share Document