Genotyping of the parasite Trichomonas vaginalis

项目描述（由申请人提供）：本项目的总体目标是建立第一个成功的综合基因分型系统，在遗传水平上明确区分鞭毛原虫阴道毛滴虫的个体、耐药和毒力分离株，为未来的发病机制、毒力和耐药性的流行病学和生化研究提供依据。基因分型系统还将应用于连锁研究、绘制基因组图谱和确定基因组之间的变异性，包括检测耐药、致病性和毒性菌株。滴虫病是最常见的性传播感染。症状包括处女和生殖器粘膜急性炎症性疾病，感染与早产、出生体重低和婴儿死亡率增加以及易患艾滋病毒/艾滋病和宫颈癌有关。滴虫病是所有性传播感染中患病率和发病率最高的，根除滴虫病很可能是降低艾滋病毒发病率的最具成本效益的单一步骤。该方案利用毛滴虫基因组的限制性内切酶切割的非常大的染色体DNA片段的脉冲场凝胶电泳，结合基因杂交，产生一个基因分型系统，以区分单个分离物。由于广泛的内源性核酸酶活性，这种分析以前没有实现。基因分型系统将与全染色体原位杂交和电泳核型相结合，用于基因连锁研究，绘制基因组组织和结构，确定分离株之间的基因组变异性，并定义毒力和耐药性标记。

DESCRIPTION (provided by the applicant): The overall goal of this project is to establish the first successful comprehensive genotyping system to clearly distinguish at the genetic level individual, drug resistant and virulent isolates of the flagellated protozoan parasite Trichomonas vaginalis for future epidemiological and biochemical studies of pathogenesis, virulence and drug resistance. The genotyping system will also be applied to linkage studies, mapping the genome, and determining variability among genomes including the detection of drug resistant, pathogenic and virulent strains. Trichomoniasis is the most common, sexually transmitted infection. Symptoms include virginities and acute inflammatory disease of the genital mucosa, and infections have been associated with pre-term delivery, low birth weight and increased infant mortality, as well as predisposing to HIV/AIDS and cervical cancer. Trichomoniasis has the highest prevalence and incidence of any STI, and its eradication may well be the single most cost-effective step in HIV incidence reduction. This proposal utilises pulsed field gel electrophoresis of very large chromosomal DNA segments cleaved by restriction endonucleases of the Trichomonas genome, combined with gene hybridization, to generate a genotyping system, which distinguishes individual isolates. Such an analysis has not previously been achieved due to extensive endogenous nuclease activity. The genotyping system will be coupled to in situ hybridization of whole chromosomes and electrophoretic karyotyping for gene linkage studies, mapping genome organization and structure, determining genome variability among isolates, and defining markers for virulence and drug resistance.