Stenopsyche marmorata larvae spin underwater adhesive silk for constructing nests and capture nets. The silk can be divided into fiber and adhesive regions, according to their function. The silk fiber region has a two-layer structure: a core layer situated at the center of the fiber and S. marmorata fibroin, the major component of the silk. In the anterior part of the anterior silk gland, the morphological characteristics suggest that the silk insolubilization leading to fibrillation occurs by luminal pH neutralization. The adhesive region is composed of three layers: the outermost (OM), B, and C layers. On the B layer, coated with the OM layer, numerous nano-order pillar structures (nanopillar structures) are located at regular intervals. A nanopillar structure is approximately 40 nm in diameter and 125 nm in length. The precursor materials of the nanopillar structure are electron-dense globules of approximately 25 nm in diameter that are located in the A layer of the lumen of the middle silk gland. The precursor globules autonomously connect to one another on the B layer when the liquid silk is transported to the lumen of the bulbous region. The nanopillar structures probably contribute to the strong underwater adhesion of S. marmorata silk.
The secondary tissues of woody plants consist of fragile cells and rigid cell walls. However, the structures are easily damaged during mechanical cross-sectioning for electron microscopy analysis. Broad argon ion beam (BIB) milling is commonly employed for scanning electron microscopy (SEM) of hard materials to generate a large and distortion-free cross-section. However, BIB milling has rarely been used in plant science. In the present study, SEM combined with BIB milling was validated as an accurate tool for structural observation of secondary woody tissues of two samples, living pine (Pinus densiflora) and high-density oak wood (Quercus phillyraeoides), and compared with classical microtome cross-sectioning. The BIB milling method does not require epoxy resin embedding because of prior chemical fixation and critical point drying of the sample, thus producing a three-dimensional image. The results showed that xylem structures were well-preserved in their natural state in the BIB-milled cross-section compared with the microtome cross-section. The observations using SEM combined with BIB milling were useful for wide-area imaging of both hard and soft plant tissues, which are difficult to observe with transmitted electron microscopy because it is difficult to obtain sections of such tissues, particularly those of fragile reaction woods.
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