GH-04 Encystation of Entamoeba histolytica in Axenic Culture
Abstract
Entamoeba histolytica is a parasitic protozoan that causes amoebic dysentery through the consumption of contaminated food and water contaminated with the cyst form which is encapsulated in a hard chitin cell wall. The cysts then undergo excystation to the trophozoite form in the human host. Trophozoites can undergo encystation to convert back to the cyst form and are shed in feces to continue disease spread.Encystation has been primarily in the reptile pathogen Entamoeba invadens but recent developments now allow these studies to be conducted directly in the human pathogen.
In this project, I investigated glucose limitation and cell densityto enhance the rate of in vitro encystation. Cells were grown in standard glucose medium and transferred to medium lacking glucose at different starting cell densities for 24 to 48 hours. Initiation of encystation was assessed using reverse transcriptase PCR (RT-PCR) of two genes shown to be regulated during early encystation. The Jacob gene encodes a protein that forms part of the chitin cell wall and its gene is expressed early in encystation. The gene encoding heat shock protein 90 (Hsp90) isdownregulated during initiation of encystation. I examined expression of these two genes as a means to assess whether the conditions tested cause earlier initiation of encystation than the current standard encystation conditions.
The optimal conditions determined here will be combined with other factors to improve synchronization of encystation in E. histolytica for future studies.
GH-04 Encystation of Entamoeba histolytica in Axenic Culture
Entamoeba histolytica is a parasitic protozoan that causes amoebic dysentery through the consumption of contaminated food and water contaminated with the cyst form which is encapsulated in a hard chitin cell wall. The cysts then undergo excystation to the trophozoite form in the human host. Trophozoites can undergo encystation to convert back to the cyst form and are shed in feces to continue disease spread.Encystation has been primarily in the reptile pathogen Entamoeba invadens but recent developments now allow these studies to be conducted directly in the human pathogen.
In this project, I investigated glucose limitation and cell densityto enhance the rate of in vitro encystation. Cells were grown in standard glucose medium and transferred to medium lacking glucose at different starting cell densities for 24 to 48 hours. Initiation of encystation was assessed using reverse transcriptase PCR (RT-PCR) of two genes shown to be regulated during early encystation. The Jacob gene encodes a protein that forms part of the chitin cell wall and its gene is expressed early in encystation. The gene encoding heat shock protein 90 (Hsp90) isdownregulated during initiation of encystation. I examined expression of these two genes as a means to assess whether the conditions tested cause earlier initiation of encystation than the current standard encystation conditions.
The optimal conditions determined here will be combined with other factors to improve synchronization of encystation in E. histolytica for future studies.