A technique for preserving pigmentation in some capsalid monogeneans for taxonomic purposes

A technique is described to preserve the pigment found in the bodies and the intestine of some brightly coloured and darkly pigmented benedeniine capsalid monogeneans. Previous studies of these pigmented capsalids have proven difficult because the pigmentation usually disappears when the worms are fixed using preservatives containing concentrations of formalin over 5% and/or ethanol, acetic acid, chromic acid, picric acid and mercuric chloride. The technique developed here uses a fixative comprising glycerol, acetone and formalin (GAF). After fixation under light coverslip compression for three minutes, specimens are transferred to absolute acetone for three minutes and cleared in a mixture of nine parts cedar wood oil and one part absolute acetone before mounting in Canada balsam. Processing must be carried out quickly, as these chemicals will cause the pigments to fade if the specimens are exposed to them for too long. Pigmented benedeniines processed using this technique retain the distribution, intensity and colour observed in live worms. The colour and distribution of pigmentation in monogeneans may be of taxonomic importance and this technique aids preparation of whole-mounts suitable for registration as type-material.     

A collecting technique of eggs of fish's monogeneans in natural conditions

A device for collecting eggs produced by monogeneans parasitizing on fish is described. The device may be used in different natural conditions, i.e. lake, river, pond, sea, ocean.     

The use of geometric morphometrics in understanding shape variability of sclerotized haptoral structures of monogeneans (Platyhelminthes) with insights into biogeographic variability

The sclerotized attachment organ of monogeneans has been widely used to address fundamental questions in ecology and evolution. However, traditional morphometric techniques appear to be partially inadequate and non-optimal. Traditional linear measurements mainly provide information on the size of sclerites but provide very little information, if any, on their shape. The shape of sclerites is indeed virtually unexplored and its implication for ecological and evolutionary processes remains to be analyzed. This study aims to both introduce and illustrate the use of geometric morphometrics in order to study sclerites of monogeneans in a biogeographic context. To do this, we investigated morphological variation patterns among four populations from the Pacific Ocean and six monogenean species through traditional and geometric morphometric techniques. Unlike the traditional method, the geometric morphometric method yielded a high percentage of individuals correctly classified to the four populations, providing strong evidence for phenotypic variability, divergence and local adaptation among islands without evolutionary constraint. Moreover, the traditional method also resulted in inconsistent interpretations of shape variations. This study highlighted the limitations that may arise when using traditional morphometric techniques and emphasizes that considerable information about the shape of sclerotized haptoral parts is added by using geometric morphometrics. Given the prominent taxonomic, ecological and evolutionary role of the haptor for characterizing monogeneans, we ultimately discuss the potential broad use of geometric morphometrics in a wide variety of ecological and evolutionary contexts. This powerful approach might allow a more robust estimation of the extent to which traditional evolutionary theories based on size of sclerites are congruent with their shape.     

An SEM study of the haptoral sclerites of the genus Gyrodactylus Nordmann, 1832 (Monogenea) following extraction by digestion and sonication techniques

Techniques for the preparation of haptoral sclerites of Gyrodactylus species for scanning electron microscopy are described. Enzymatic digestion using pepsin allows fresh, frozen and alcohol-fixed material to be examined, but some sclerites, such as the dorsal and ventral bars, are lost. Sonication of fresh and frozen material retained these accessory bars, including the ventral bar membrane, the guard or sickle filament loop on the marginal hooks and even on some occasions ligaments associated with the sclerites. The sclerites of Gyrodactylus spp. from Gasterosteus aculeatus (L.) (three-spined stickleback), Oncorhynchus mykiss (Walbaum) (rainbow trout) and Salvelinus alpinus (L.) (charr) are described from SEM observations following the use of these preparation techniques, and the articulation between the hamuli and bars is commented upon.     

A technique for studying chemotaxis of leucocytes in well-defined chemotactic fields

A new technique for observing the chemotaxis of leucocytes in well-defined chemotactic fields has been developed. This technique is also applicable to other studies in cell physiology in which an efficient method of generating known chemical concentration gradients on microscope slides is needed. This study reports the range of concentrations and gradients of cyclic adenosine 3′, 5′-monophosphate which will result in chemotactic ameboid motion of human polymorphonuclear granulocytes. We have identified a threshold gradient and a range of concentration near the solubility limit in which chemotaxis is impaired.     

Sodium dodecyl sulphate as a rapid clearing agent for studying the hard parts of monogeneans and nematodes

Hard structures of helminths have often been used for taxonomic identification but are usually not clearly defined when treated with conventional methods such as ammonium picrate-glycerin for monogeneans and glycerin for nematodes. The present study reports a rapid and simple technique to better resolve the hard parts of selected monogeneans and nematodes using 5-10% alkaline sodium dodecyl sulphate (SDS). In comparison with established methods, SDS-treated worms become more transparent. In monogeneans treated with SDS, clear details of the hooks, hook filaments, anchors, bars and the sclerotized copulatory organs could be observed. In SDS-treated nematodes, spicules and ornamentations of the buccal capsules could be clearly seen.     

A simplified formaldehyde fixation and immunoprecipitation technique for studying protein-DNA interactions

Using the single cell eukaryote Tetrahymena thermophila, a simple method was developed for studying protein-DNA associations by cross-linking proteins to DNA with formaldehyde and immunoprecipitating the solubilized chromatin fragments with a specific antiserum. The protocol uses crude antiserum and involves only three steps: cross-linking, shearing to solubilize the chromatin, and immunoprecipitation. Methods for optimizing certain critical parameters, such as fixation time and NaCl concentration, are described. The method is likely to be generally useful for a variety of nuclear antigens.     

Stiffness and tanning of sclerites

Tensile studies on cuticle of different chitin fibre architecture indicate that the intimate relationship between physical stiffness and the extent of chemical tanning holds for those cuticles which are mechanically isotropic in their surface planes, but not for those which are mechanically anisotropic.     

A list of monogeneans from Chinese marine fishes

This paper contains a brief history of studies on monogeneans of Chinese marine fishes and a list of 337 monogenean species, together with their marine hosts and notes on the sea in which they were found. An additional 139 species have been added to the 198 species recorded in `Monogeneans of Chinese Marine Fishes' published by Zhang, Yang and Liu in 2001.     

Hardening of locust sclerites

The tanning process of presumptive solid cuticle was shown to be correlated with both the tensile force at fracture and the tangent modulus. Both of these physical parameters are closely related to the biochemical events associated with the tanning process.     

研究基因转录的RT-PCR技术

RT-PCR是检测和定量特异性mRNA的高度灵敏技术,被广泛应用于基因转录水平的分析研究,本文探讨RT-PCR方法的发展,应用及潜在问题。     

A continuous fermentation technique for studying the kinetics of sugar uptake by baker's yeast

1. Solutions of glucose, maltose or other sugars were pumped at controlled rates into a yeast suspension and the extracellular sugar concentration was determined. The technique was especially suitable for studying the kinetics of fermentation at low rates of sugar utilization. At high rates the fermentation system was unstable. 2. Glucose fermentation was fitted by a model in which a diffusion barrier with first-order kinetics is interposed between the environment and the site of hexokinase. 3. Aerobic conditions affected the fermentation enzyme system but not the diffusion mechanism. 4. The kinetics of maltose fermentation at low rates approximated to those of the hydrolysis of maltose by the enzyme maltase, as studied in suspensions of broken cells.     

A squash technique for studying the cytology of maize endosperm and other tissues.

A new cytological procedure specifically suited to maize endosperms is presented. It uses 8-hydroxyquinoline with sucrose and aeration to pretreat the tissues. Glusulase is used to spread the cells. The procedure makes it possible to squash endosperms into a single cell layer and to photograph as many as 70 chromosomes in the same focal plane. It also allows identification of translocation chromosomes. With a slight modification the technique has been applied successfully to root tips and other tissues.