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Date of Award

2009

Document Type

Thesis - Pacific Access Restricted

Degree Name

Master of Science (M.S.)

Department

Biological Sciences

First Advisor

Kirkwood Land

First Committee Member

Craig A. Vierra

Second Committee Member

Lisa A. Wrischnik

Abstract

Trichomonas vaginalis and Tritrichomonas foetus are the etiologic agents of human and bovine trichomoniasis, respectively. As microaerophilic protozoans; both share a wide array of clinical manifestations ranging from vaginitis to abnormal pregnancies. Human trichomoniasis receives minimal public health attention despite of its worldwide high prevalence rate. Emerging evidence of metronidazole-resistant T vaginalis strains facilitates a concern to understand this protozoan. Cysteine proteases have been implicated as important virulence factors produced by T vagina/is. This study explores the expression of one particular legumain-like cysteine protease known as Tv AE 1. Furthermore, it highlights the relationship between inhibitory effects of trichomonal cells caused by sanguinarine and chelerythrine. A system for obtaining legumains by expressing it in methylotrophic yeast, Pichia pastor is, has been described. The recombinant legumains were produced and processed by the yeast to their inactive and mature forms. Secondly, T foetus cells were transfected with TvAEl construct.

Localization and enzymatic studies on legumains will provide evidence into the pathogenicity ofT vagina/is. This study revealed the vesicularization of recombinantly unprocessed TvAEl proteins. Thirdly, plant derived compounds, sanguinarine (SA) and chelerythrine (CHE) were assessed in vitro for their inhibitory effects against T vagina/is and T. foetus. Treatment of SA and CHE for 24 h led to a significant inhibitory growth of in vitro cultures for all three trichomonal strains, G3, Tl and Dl, compared to untreated cells. For these bovine and human trichomonal strains, SA was slightly more effective inhibitor than CHE. With IC5o values between 3 - 8 micromolar for the alkaloids, CHE had less inhibitory effect compared to SA. These findings are significant considering the association between cysteine pro teases and trichomoniasis. Further elucidation of the exact anti protozoal mechanism of both compounds toward legumains may lead to the development of these potent agents against trichomonads.

Pages

139

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