c.i.-fluorescent-brightening-agent-28 and Amebiasis

Acanthamoeba cysts are resistant to unfavorable physiological conditions and various disinfectants. Acanthamoeba cysts have 2 walls containing various sugar moieties, and in particular, one third of the inner wall is composed of cellulose. In this study, it has been shown that down-regulation of cellulose synthase by small interfering RNA (siRNA) significantly inhibits the formation of mature Acanthamoeba castellanii cysts. Calcofluor white staining and transmission electron microscopy revealed that siRNA transfected amoeba failed to form an inner wall during encystation and thus are likely to be more vulnerable. In addition, the expression of xylose isomerase, which is involved in cyst wall formation, was not altered in cellulose synthase down-regulated amoeba, indicating that cellulose synthase is a crucial factor for inner wall formation by Acanthamoeba during encystation.

Topics: Acanthamoeba castellanii; Aldose-Ketose Isomerases; Amebiasis; Benzenesulfonates; Cell Wall; Cellulose; Down-Regulation; Encephalitis; Glucosyltransferases; Keratitis; Microscopy, Electron, Transmission; RNA Interference; RNA, Small Interfering

Topics: Acanthamoeba; Amebiasis; Animals; Benzenesulfonates; Humans; Staining and Labeling

In search of an easy and quick method for the diagnosis of fungal and Acanthamoeba infections of the eye, the authors have used calcofluor white (CFW), a fluorescent brightener with marked affinity for chitin and cellulose which are present in the cell walls of fungi and cysts of Acanthamoeba. Paraffin-embedded tissues containing mycotic organisms were stained with CFW and examined under a fluorescent microscope. Several fungi such as Candida sp, Aspergillus sp, Phycomycetes, Fusarium solani, Histoplasma capsulatum, Cryptococcus neoformans, Rhinosporidium seeberi, and others were studied. Trephine corneal biopsy specimens (2 mm) and methanol-fixed corneal scrapings containing Acanthamoeba sp were also examined. The walls of fungi and Acanthamoeba cysts stained bright greenish white in contrast to the reddish-orange background of the supporting tissues. The results demonstrate that the CFW method is a highly reliable and rapid technique for identification of mycotic organisms as well as cysts of Acanthamoeba. This method may also be applied in surgical pathology when performing frozen sections, as well as cytologic examination of corneal smears, vitrectomy specimens, and others.

Topics: Amebiasis; Benzenesulfonates; Eye; Eye Diseases; Humans; Microscopy, Fluorescence; Mycoses; Orbit; Orbital Diseases; Staining and Labeling

Calcofluor white (CFW) is a chemofluorescent dye with an affinity for the polysaccharide polymers of amebic cysts. Using CFW staining with fluorescent microscopy, we demonstrated amebic cysts in corneal scrapings and keratectomy specimens from four patients with culture-proved Acanthamoeba keratitis and from one in whom CFW was the only positive laboratory test. Calcofluor white staining is simple, rapid, and highly reliable in the diagnosis of Acanthamoeba keratitis.

Topics: Adolescent; Adult; Amebiasis; Amoeba; Benzenesulfonates; Cornea; Female; Fluorescent Dyes; Humans; Keratitis; Male; Middle Aged; Staining and Labeling