SUMMARY
A method is described for the in toto embedding and precipitate‐free staining of small invertebrates covered by strong protective outer surfaces.
The difficulties of tissue infiltration in undissected animals with cuticles were overcome by the use of ethanol and 1,4‐dioxane as dehydrating agents, and Spurr's medium as the embedding resin. (1) Fixation as appropriate, followed by embedment in aqueous agar; (2) dehydration: 50% ethanol, 70% ethanol, 2 times 100% 1,4‐dioxane, 15–20 min for each step; (3) infiltration: dioxane:resin 1:1 90 min, dioxane:resin 1:3 90 min, pure resin overnight, changed once for an additional 10 h, all steps with constant shaking; (4) polymerization 16 h at 343 K; (5) sectioning; (6) staining: 0·9% cacodylate buffered KMnO4 (pH 6·5) 30–60 s, washing, Reynolds' lead citrate 5 min, washing by grid floating.
The advantage of the method is discussed in relation to serial sectioning of entire invertebrates which could not be dissected prior to embedding.
The action of the chitin synthesis inhibitor complex nikkomycin on the silk and salivary glands of Tetranychus urticae (Acari, Tetranychidae) was examined with the electron microscope. Histological defects after nikkomycin treatment included a swelling of the cisterna of rough endoplasmic reticulum (ER). Subsequent defects were vacuolation, resulting from globular ER vesicles, and damage to the secretory grana of all the glands studied (the silk gland and the anterior and dorsal podocephalic glands). The consequences of the histological changes on the salivary secretions and the production of webs are discussed.
Some effects of the chitin synthesis inhibitor complex nikkomycin on oogenesis in the two‐spotted spider mite Tetranychus urticae were demonstrated by means of electron microscopy. Four main effects were evident, depending on the oogenetic stages. Nuclear defects, observed in oogonia and primary oocytes, included a widened nuclear envelope, the intermembranous spaces of which were filled with electron‐dense material, and the occurrence of small electron‐dense agglomerations and altered nucleoli inside the nuclei. Cytoplasmic defects occurred mainly in nurse cells and involved electron‐dense granular areas accompanied by mitochondria, and large cisternae and vesicles of rough endoplasmic reticulum. Yolk synthesis in oocytes could be inhibited, as shown by the lack of yolk droplets or the presence of abnormal yolk droplets. Both phases of egg shell synthesis were disturbed, resulting in an inhibition of egg deposition. Mechanisms based upon the oogenetic effects of the metabolite complex nikkomycin are discussed.
The effects of the closely related chitin synthesis inhibitors, the nucleoside peptidyl antibiotics nikkomycin X/Z, nikkomycin 2 and polyoxin D on the two-spotted spider mite, Tetranychus urricae, have been studied by electron microscopy. The application of all three compounds, nikkomycin X/Z (95% active ingredient, ratio of components unknown), nikkomycin Z (pure component) and polyoxin D (pure component) disrupted cuticle, eggshell and yolk synthesis and influenced the silk and salivary glands. Ultrahistological patterns indicated differences in the mode of action of the three compounds which were especially evident in an alteration of the nuclear ultrastructure in the mid-gut cells, which was only caused by polyoxin D. Mechanisms of action and their consequences on function, based on previously published information on the action of these compounds, are discussed.
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