Ultrastructural components of trophozoites and cysts of Acanthamoeba-Hartmannella (A-H) and Naegleria amebas have been investigated. The present report describes a technique to study the morphological features of such protozoa.The A-H strains selected were cultured for 7 days in axenic amebic media, and the Naegleria strains were cultured for 7 days in axenic media with blood. In situ fixation was accomplished by adding an equal volume of 4% Osmic Acid in Cacodylate buffer at 4¶C for one hour. Following frequent agitation, the tubes were then spun at 2500 RPM for 10 minutes to obtain a pellet. The pellets were dehydrated in graded series of alcohols (from 50% to absolute) and propylene oxide and then embedded in EPON. One micron sections were stained with Paragon (PS 1301) for thirty seconds. Thin sections were cut with a diamond knife, double stained with 5% alcoholic uranyl acetate solution and lead citrate and observed in an Hitachi-HS-8F2 electron microscope.
Ultrastructural study of trophozoites and cysts in amebic encephalitides (1-4) have provided significant information pertaining to pathogenesis as well as clarification of taxonomic criteria (e.g. species, genera, etc). The use of tissues previously fixed in formalin and embedded in paraffin, then reprocessed and embedded in plastic for electron microscopic studies is useful in the evaluation of morphological features of trophozoites and cysts of these pathogenic free-living amebas. Even though the tissues disclose some post-morten autolytic alterations and distortions, trophozoite and cyst preservation is satisfactory for identification, giving excellent morphological information. Details of the overall form, which may have a role in pathogenicity, including cytoplasmic processes and projections, organelles and cyst walls, may be revealed.
Scanning electron microscopy (SEM) can serve as a valuable supplement to transmission electron microscopy (TEM) in the study of pathogenic protozoa. Details of overall form and structure of surface and organelles which may have a role in pathogenicity may be revealed. TEM studies on Naegleria have contributed much to understanding the extraordinary virulence of this ameba, but some remaining questions may be resolved by SEM. This report describes a technique which has proven useful in preparing SEM specimens. Naegleria ameba trophozoites adhere strongly to the surface on which they are growing. In culture tubes, amebae will multiply on the wall. Naegleria tends to grow from the top of the fluid downward and may multiply until a solid monolayer develops. If a strip of plastic film is introduced, the growth on the strip can be observed by direct microscope viewing through the wall of the tube.
Ultrastructural studies from tissues of Peruvian mummies ranging from 500 to 2000 years old have been carried out.Even though the tissues had undergone extensive autolytic alterations many subcellular structures were still present and it was possible to clearly identify them. The better preserved tissues were those which contained abundant collagen and connective structures (Fig. 1-3). The collagen fibrils in particular showed the characteristic cross striations. The thickness of the dark bands was 45-50 nm while the thickness of the light bands measured between 25-30 nm.
Casanova (19 74) has reported that the method of Mayor (19 61) is the best f or removal of epoxy resins. In her method, sodium methylate was made using sodium metal. We have modified this using sodium methylate powder.1. 5 gm of sodium methylate was dissolved in a beaker in 2 5 ml of methyl alcohol. As it dissolves the beaker becomes warm. When completely dissolved, 2 5 ml of benzene is added with stirring. If rings appear at the interface, mo re alcohol should be added until the rings disappear.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.