Terminology used in the description of bennettitalean cuticle characters

Details of the bennettitalean stomatal apparatus, trichome bases and other cuticular features as seen in the light and scanning electron microscopes are described and an updated scheme of terminology is presented.     

The structure of the haemocytes of Calpodes ethlius (Lepidoptera)

The haemocytes of Calpodes ethlius are described with the light and electron microscopes. Five fine structurally distinct types are distinguishable. However only three of these, the granular haemocytes, sphaerule cells and oenocytoids can be positively identified using a series of histochemical stains on smears and on thick sections of araldite-embedded material. The classification is based entirely on the structural features even though several suggestions concerning their function can be made from their fine structure. Intermediates having features of more than one cell type suggest developmental relationships.     

Plasmolysis of Pteridium protoplasts: A study using light and scanning-electron microscopy

A study was undertaken using gametophytes of the fern Pteridium aquilinum to examine the effects of plasmolysis on the topography of protoplasts. Methods are described whereby the surfaces of non-isolated protoplasts can be observed in the plasmolysed condition using scanning electron microscopy. Plasmolysed gametophytes were also examined in the light microscope using differential interference contrast and ultra-violet fluorescence microscopy after staining with fluorescein diacetate. With scanning electron microscopy, plasmolysed protoplast surfaces appeared smooth with no evidence of wrinkling or infolding of excess membrane. The formation of irregular-shaped protoplasts, protoplasmic threads, subprotoplasts, and protoplasmic networks covering internal wall surfaces all provided evidence for strong wall adhesion of the protoplasm. The availability of membrane for uptake into folds or vesicles is therefore thought to be minimal. Transmission electron microscopy showed some protoplasmic threads to be plasmodesmata, the remainder being cell-wall contact points. Remnants of these threads were occasionally observed on isolated protoplasts in both the light and electron microscopes.     

Spermatogenesis in Bombyx mori. II. The ultrastructure of synapsed bivalents

In Bombyx mori the male is the homogametic sex, crossing over occurs only in males, and chiasmata are observed in spermatocytes, but not in oocyte nuclei. If the assembly of synaptonemal complexes is an essential prerequisite for genetic crossing over and chiasmata formation, then the nuclei of Bombyx spermatocytes should contain synaptonemal complexes. Synaptonemal complexes were found in spermatocytes from young four instar larvae. The structure of meiotic bivalents is described using micrographs taken with 100 and 1000 KV electron microscopes. These data together with that from the literature are used to construct a three-dimensional model of the synaptonemal complex and to suggest its method of origin and its function during crossing over.     

Mosquito allergy and mosquito salivary allergens

Allergic reactions to mosquito bites are caused by allergens in mosquito saliva. In this review, allergic reactions to mosquito salivary allergens, and characteristics of salivary allergens and their recombinant forms are described. The use of the recombinant allergens in the diagnosis of mosquito allergy is discussed.     

Optical and digital microscopic imaging techniques and applications in pathology

The conventional optical microscope has been the primary tool in assisting pathological examinations. The modern digital pathology combines the power of microscopy, electronic detection, and computerized analysis. It enables cellular-, molecular-, and genetic-imaging at high efficiency and accuracy to facilitate clinical screening and diagnosis. This paper first reviews the fundamental concepts of microscopic imaging and introduces the technical features and associated clinical applications of optical microscopes, electron microscopes, scanning tunnel microscopes, and fluorescence microscopes. The interface of microscopy with digital image acquisition methods is discussed. The recent developments and future perspectives of contemporary microscopic imaging techniques such as three-dimensional and in vivo imaging are analyzed for their clinical potentials.     

"Wonders unconceived": reflections on the birth of medical entomology

Prior to Patrick Manson's discovery in 1877 that the mosquito Culex fatigans was the intermediate host of filariasis, the association of insects with disease and the nature of disease transmission was almost entirely speculation. Manson's work was incomplete, however, because it showed the manner in which the mosquito acquired the infection from humans, but failed to show the way in which the mosquito passed the infection to humans. That pathogens were transmitted by the bite of an infected female mosquito was later proven experimentally with bird malaria by Manson's protégé, Ronald Ross. In 1898 Ross demonstrated that the infective stage of the malarial parasite was injected into the host when the mosquito released saliva into the wound prior to injesting blood. Insects were suspected as carriers of disease for centuries, yet it was not until the late 1870s that the uncritical acceptance of folk beliefs was supplanted by research-based scientific medicine. Why did it take so long? The answer lies in the fact that early medicine itself was imprecise and could not have pursued the subject with any hope of useful results until the last quarter of the 19th century. A better understanding of the nature of the disease process (germ theory of disease) and improved technology (microscopes and oil-immersion lenses with greater resolving power, and synthetic tissue stains) were indispensable for revealing the nexus between those partners in crime: insects and parasites.     

Morphological provision for the specialization of hares to coprophagy: The architectonics of the mucous surface of the intestine

The macrostructure of the surface of the mucous membrane of the gastrointestinal tract were studied in detail in two hare species (Lepus europaeus and L. timidus) using scanning electron and digital microscopes. The morphology of the gastrointestinal tract of hares, which is similar in the two species studied, was described in detail. The macrostructure of the inner surface of the ileocecal junction was investigated. The particularities of the architectonics of the mucous membrane of the intestine that are specific for hares were revealed. In the jejunum of both hare species, the mucous membrane is represented by villi with merged bases, which form circular plates. The mucosa of the colon forms large conical villi, the surface of which is scattered with secreting cells. The possible functional significance of the revealed morphological particularities is discussed.     

Measuring the size of biological nanostructures with spatially modulated illumination microscopy

Spatially modulated illumination fluorescence microscopy can in theory measure the sizes of objects with a diameter ranging between 10 and 200 nm and has allowed accurate size measurement of subresolution fluorescent beads ( approximately 40-100 nm). Biological structures in this size range have so far been measured by electron microscopy. Here, we have labeled sites containing the active, hyperphosphorylated form of RNA polymerase II in the nucleus of HeLa cells by using the antibody H5. The spatially modulated illumination-microscope was compared with confocal laser scanning and electron microscopes and found to be suitable for measuring the size of cellular nanostructures in a biological setting. The hyperphosphorylated form of polymerase II was found in structures with a diameter of approximately 70 nm, well below the 200-nm resolution limit of standard fluorescence microscopes.     

Resolution as a function of accelerating voltage in electron microscopy of semithick biological specimens

In the past, biological sections ranging in thickness from 0.10- to 0.50-micron have usually been examined with high-voltage (greater than 500 kV) electron microscopes (HVEM). Now investigators are increasingly using intermediate voltage (200-500 kV) electron microscopes (IVEM), which are more readily available and demand less maintenance. In a study of "typical" plastic-embedded, stained sections of mouse liver ranging from 0.10 to 1.0 micron thick, we determined the resolution obtainable at 100, 200, and 1000 kV. At all three accelerating voltages the resolution (2.7 nm) for 0.10-micron sections was limited only by the sections stain granularity. For 0.25-micron thickness the resolutions were 5.8, 3.1, and 3.1 nm at 100, 200, and 1000 kV, respectively. The maximum usable thickness at 200 kV with resolution sufficient to resolve membranes clearly was between 0.75 and 1.0 micron, depending on the magnification. Resolution at 100 kV was adequate for screening sections up to 1.0-micron thick for preparation defects prior to examination with an IVEM or HVEM.     

Phase-plate electron microscopy: a novel imaging tool to reveal close-to-life nano-structures

After slow progress in the efforts to develop phase plates for electron microscopes, functional phase plate using thin carbon film has been reported recently. It permits collecting high-contrast images of close-to-life biological structures with cryo-fixation and without staining. This report reviews the state of the art for phase plates and what is innovated with them in biological electron microscopy. The extension of thin-film phase plates to the material-less type using electrostatic field or magnetic field is also addressed.     

Simple sets for digital microphotography used and tested in the study of microorganisms

Four simple sets for digital microphotography are described that have been tested with the Carl Zeiss Jena, Meopta Prague, Lambda Prague, and LOMO Sankt Petersburg microscopes and with DSLR Nikon D 70 and Nikon D 300 cameras. They permit precise image focusing in the camera using a prism Zeiss. The sets make use of commonly available extensions Zeiss, Praktica and reductions Nikon??Praktica manufactured by ROWI (without a lens) or HAMA (with a lens). An extension has further been designed and manufactured for connecting the DSLR Nikon D 300 camera fitted with the HAMA reduction (only with a lens) and a focusing extensible prism with Zeiss Jena light measurement. It permits a precise image focusing of low light intensity objects (autofluorescence or low-contrast or moving objects when using positive or negative phase contrast). The sets are applicable to all microscopes constructed according to German DIN industrial standards.     

Soft x-ray microscopy

Soft x-ray microscopes are beginning to provide information to complement that obtained from optical and electron microscopy. Soft x-ray microscopy can deliver 30-nm resolution images of hydrated cells up to approximately 10 microns thick, and efforts towards obtaining higher resolution are under way. Although living specimens cannot be studied readily except in single exposures, fixed samples can be imaged at high resolution, and flash-frozen specimens can be studied without chemical modification and without significant radiation damage. Tomography is being developed for 3-D imaging, and spectromicroscopy offers unique capabilities for biochemical mapping of unlabelled structures beyond those of gold and fluorescent labels. Currently, most soft x-ray microscopes operate at synchrotron radiation facilities, but laboratory-scale microscopes are being developed too.     

Morphology and ultrastructure of modern and fossil spores in order Schizaeales schimp

The morphology and ultrastructure of modern and fossil spores were investigated using light, scanning, and transmission electron microscopes. We grouped spores into two groups according to ultrastructural features and sculptures of sporoderm: 1—exospore of Anemia and Klukia form the echini of sculpture; 2—exospore of Lygodium is smooth and perispore forms sculpture.     

ISOLATION OF NUCLEI FROM YEAST

A method for isolation of nuclei from Saccharomyces cervisiae in high yield is described. The DNA/protein ratio of the isolated nuclei is 10 times higher than that of whole cells. Examination of these nuclei in phase and electron microscopes has shown them to be round bodies having a double membrane, microtubules, and a dark crescent at one end. The optimum conditions for extraction and resolution of histones of these nuclei on acrylamide gels have been investigated. The nuclei have an active RNA polymerase (E.C. 2.7.7.6) and are able to synthesize RNA in vitro. They are also readily stainable with Giemsa's, Feulgen's, and acridine orange methods.     

Unconventional modes for STEM imaging of biological structures

In this paper recent developments are discussed in instrumentation and methodology associated with scanning transmission electron microscopes (STEM), which are of great potential interest for solving structural and chemical problems in biological specimens. After describing the main features of the instrument, an attempt is made to define which type of signal acquisition and processing is best suited to obtain a given type of information. Starting with a definition of cross sections of interest, a discussion follows of methods using angular selection, energy selection of the transmitted beam, and several ways of signal mixing. More specific attention is devoted to two main modes of processing signals: ratio contrast, which emphasizes slight changes in scattering factors, rather independent of thickness variations; and elemental mapping, which provides semi-quantitative information on the distribution of low Z elements of great significance in biological specimens. Data relevant to typical biological objects are presented and discussed; they allow for the definition of the capabilities and limitations of these methods. These unconventional imaging modes define a new attitude for improving the efficiency of this modern generation of electron microscopes.     

Aedes aegypti: Sensilla trichodea and stimulus-conducting structures

Much of the morphology of the olfactory sensilla on the antennae of the mosquito Aedes aegypti (L.) has been described, however little is known about the fate of odour molecules once they have been adsorbed onto the surfaces of sensilla. A stimulus-conducting system of pores, pore kettles, and pore tubules has been described for the sensilla trichodea (olfactory hairs) of several insects but not mosquitoes. Scanning electron microscopy was used to identify the s. trichodea of Ae. aegypti and to attempt visualization of their pore openings. Chemical fixation, cryopreparation, freeze drying, and negative staining, with high resolution transmission electron microscopy (TEM), were used to locate putative stimulus-conducting structures associated with the pores. TEM sections using Dalton's fixative or freeze drying showed pores and pore tubules, whereas pore tubules were poorly preserved in cryoprepared sections. The putative stimulus-conducting structures were clearly demonstrated by negative staining of whole sensilla which was quick and easy. The current hypothesis of olfactory stimulus conduction is extended to include Ae. aegypti.     

The structure of the male genitalia of Aedes diantaeus Howard, Dyar et Knab (Diptera, Culicidae)

The structure of the male genitalia of Aedes diantaeus was studied under scanning electron and optical microscopes. The tridimentional structure of the coxite is described and the structure of apical and basal warts and the arrangement of setae on the coxite are clarified. The structure of the claspette, i.e., the position of the claspette’s wing (filament) in relation to the claspette’s stem and also the shape and the structure of the wing were studied. The shape of the proctiger and the position of the paraprocts were investigated. A large membranous area forming paired lobes on the dorsal surface of the proctiger is described. Significant differences between Aedes diantaeus and A. caspius, A. cantans, and A. communis in the structure of the coxite, claspette, and proctiger, are demonstrated.