The root system is particularly affected by unfavourable conditions because it is in direct contact with the soil environment. Casparian strips, a specialised structure deposited in anticlinal walls, are characterised by the impregnation of the primary wall pores with lignin and suberin. The Casparian strips in the endo- and exodermis of vascular plant roots appear to play an important role in preventing the non-selective apoplastic bypass of salts into the stele along the apoplast under salt stress. However, only a few investigations have examined the deposition and function of these apoplastic barriers in response to salt stress in higher plants.

Studies visualizing plant tissues and organs in three-dimension (3D) using micro–computed tomography (CT) published since approximately 2015 are reviewed. In this period, the number of publications in the field of plant sciences dealing with micro-CT has increased along with the development of high-performance lab-based micro-CT systems as well as the continuous development of cutting-edge technologies at synchrotron radiation facilities. The widespread use of commercially available lab-based micro-CT systems enabling phase-contrast imaging technique, which is suitable for the visualization of biological specimens composed of light elements, appears to have facilitated these studies. Unique features of the plant body, which are particularly utilized for the imaging of plant organs and tissues by micro-CT, are having functional air spaces and specialized cell walls, such as lignified ones. In this review, we briefly describe the basis of micro-CT technology first and then get down into details of its application in 3D visualization in plant sciences, which are categorized as follows: imaging of various organs, caryopses, seeds, other organs (reproductive organs, leaves, stems and petioles), various tissues (leaf venations, xylems, air-filled tissues, cell boundaries, cell walls), embolisms and root systems, hoping that wide users of microscopes and other imaging technologies will be interested also in micro-CT and obtain some hints for a deeper understanding of the structure of plant tissues and organs in 3D. Majority of the current morphological studies using micro-CT still appear to be at a qualitative level. Development of methodology for accurate 3D segmentation is needed for the transition of the studies from a qualitative level to a quantitative level in the future.

Although the extent of aerenchyma (interconnected gas-filled space) formed in the cortex of the roots of some species can be promoted by submergence and ethylene, such roots also form a somewhat less extensive aerenchyma under well-aerated conditions. It has been unclear whether or not ethylene is also involved in promoting this constitutive aerenchyma. To confirm the potential of ethylene to stimulate aerenchyma development and test the possibility that gas regulates constitutive aerenchyma, a novel sandwich method was employed in rice roots. This involved germinating japonica rice (Oryza sativa) caryopses sandwiched between two agar slabs with or without 1-aminocyclopropane-1-carboxylic acid (ACC) at 1 µM. The roots were then grown for 4 days in the dark in the presence or absence of gaseous 1-methylcyclopropene (1-MCP), an inhibitor of ethylene action. Examination of aerenchyma development by light microscopy demonstrated a more extensive aerenchyma in cross-section on the ACC-treated side that also commenced closer to the root tip. In the presence of 1-MCP at 0.1 or 1 ppm, aerenchyma formation was inhibited in the presence or absence of ACC. 1-Methylcyclopropene also overcame ACC-inhibited root elongation. The results indicate that ethylene signalling is involved in aerenchyma development in primary roots of rice and that this may include the regulation of constitutive aerenchyma. In addition, root elongation was slowed in control roots in the presence of 1 ppm 1-MCP, supporting previous studies demonstrating that endogenous levels of ethylene stimulate root elongation.

The effect of environmental factors on the regulation of aerenchyma formation in rice roots has been discussed for a long time, because aerenchyma is constitutively formed under aerated conditions. To elucidate this problem, a unique method has been developed that enables sensitive detection of differences in the development of aerenchyma under two different environmental conditions. The method is tested to determine whether aerenchyma development in rice roots is affected by osmotic stress. To examine aerenchyma formation both with and without mannitol treatment in the same root, germinating rice (Oryza sativa) caryopses were sandwiched between two agar slabs, one of which contained 270 mm of mannitol. The roots were grown touching both slabs and were thereby exposed unilaterally to osmotic stress. As a non-invasive approach, refraction contrast X-ray computed tomography (CT) using a third-generation synchrotron facility, SPring-8 (Super photon ring 8 GeV, Japan Synchrotron Radiation Research Institute), was used to visualize the three-dimensional (3-D) intact structure of aerenchyma and its formation in situ in rice roots. The effects of unilateral mannitol treatment on the development of aerenchyma were quantitatively examined using conventional light microscopy. Structural continuity of aerenchyma was clearly visualized in 3-D in the primary root of rice and in situ using X-ray CT. Light microscopy and X-ray CT showed that the development of aerenchyma was promoted on the mannitol-treated side of the root. Detailed light microscopic analysis of cross-sections cut along the root axis from the tip to the basal region demonstrated that aerenchyma developed significantly closer to the root tip on the mannitol-treated side of the root. Continuity of the aerenchyma along the rice root axis was morphologically demonstrated using X-ray CT. By using this ‘sandwich’ method it was shown that mannitol promoted aerenchyma formation in the primary roots of rice.

Insect-induced galls are microhabitats distinct from the outer environment that support inhabitants by providing improved nutrients, defence against enemies, and other unique features. It is intriguing as to how insects reprogram and modify plant morphogenesis. Because most of the gall systems are formed on trees, it is difficult to maintain them in laboratories and to comprehend the mechanisms operative in them through experimental manipulations. Herein, we propose a new model insect,Smicronyx madaranus, for studying the mechanisms of gall formation. This weevil forms spherical galls on the shoots ofCuscuta campestris, an obligate parasitic plant. We established a stable system for breeding and maintaining this ecologically intriguing insect in the laboratory, and succeeded in detailed analyses of the gall-forming behaviour, gall formation process, and histochemical and physiological features. ParasiticC. campestrisdepends on host plants for its nutrients, and usually shows low chlorophyll content and photosynthetic activity. We demonstrate thatS. madaranus-induced galls have significantly increased CO₂absorbance. Moreover, chloroplasts and starch accumulated in gall tissues at locations inhabited by the weevil larvae. These results suggest that the gall-inducing weevils enhance the photosynthetic activity inC. campestris, and modify the plant tissue to a nutrient-rich shelter for them.