Kucknoor Ashwini S, Mundodi Vasanthakrishna, Alderete J F
Department of Microbiology, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Dr, San Antonio, TX 78229-3900, USA.
BMC Mol Biol. 2005 Mar 4;6:5. doi: 10.1186/1471-2199-6-5.
Trichomonosis, caused by Trichomonas vaginalis, is the number one, nonviral sexually transmitted infection that has adverse consequences for the health of women and children. The interaction of T. vaginalis with vaginal epithelial cells (VECs), a step preparatory to infection, is mediated in part by the prominent surface protein AP65. The bovine trichomonad, Tritrichomonas foetus, adheres poorly to human VECs. Thus, we established a transfection system for heterologous expression of the T. vaginalis AP65 in T. foetus, as an alternative approach to confirm adhesin function for this virulence factor.
In this study, we show stable transfection and expression of the T. vaginalis ap65 gene in T. foetus from an episomal pBS-ap65-neo plasmid. Expression of the gene and protein was confirmed by RT-PCR and immunoblots, respectively. AP65 in transformed T. foetus bound to host cells. Specific mAbs revealed episomally-expressed AP65 targeted to the parasite surface and hydrogenosome organelles. Importantly, surface-expression of AP65 in T. foetus paralleled increased levels of adherence of transfected bovine trichomonads to human VECs.
The T. vaginalis AP65 adhesin was stably expressed in T. foetus, and the data obtained using this heterologous system strongly supports the role of AP65 as a prominent adhesin for T. vaginalis. In addition, the heterologous expression in T. foetus of a T. vaginalis gene offers an important, new approach for confirming and characterizing virulence factors.
由阴道毛滴虫引起的滴虫病是首要的非病毒性性传播感染疾病，对妇女和儿童的健康会产生不良后果。阴道毛滴虫与阴道上皮细胞（VECs）的相互作用是感染的前期步骤，部分是由突出的表面蛋白AP65介导的。牛毛滴虫胎儿三毛滴虫对人VECs的黏附性较差。因此，我们建立了一个用于在胎儿三毛滴虫中异源表达阴道毛滴虫AP65的转染系统，作为确认这种毒力因子黏附素功能的另一种方法。
在本研究中，我们展示了来自游离型pBS-ap65-neo质粒的阴道毛滴虫ap65基因在胎儿三毛滴虫中的稳定转染和表达。分别通过RT-PCR和免疫印迹证实了该基因和蛋白的表达。转化后的胎儿三毛滴虫中的AP65与宿主细胞结合。特异性单克隆抗体显示游离型表达的AP65靶向寄生虫表面和氢化酶体细胞器。重要的是，胎儿三毛滴虫中AP65的表面表达与转染的牛毛滴虫对人VECs黏附水平的提高平行。
阴道毛滴虫AP65黏附素在胎儿三毛滴虫中稳定表达，使用该异源系统获得的数据有力地支持了AP65作为阴道毛滴虫主要黏附素的作用。此外，阴道毛滴虫基因在胎儿三毛滴虫中的异源表达为确认和表征毒力因子提供了一种重要的新方法。
