Khoshnan A, Provenzano D, Alderete J F
Department of Microbiology, University of Texas Health, Science Center, San Antonio 78284-7758.
J Virol. 1994 Nov;68(11):7108-14. doi: 10.1128/JVI.68.11.7108-7114.1994.
Most Trichomonas vaginalis isolates are carriers of the multisegmented double-stranded RNA (dsRNA) virus. In vitro polymerase assays were performed to demonstrate the RNA-dependent RNA polymerase (RDRP) activity of purified particles. Transcripts which comigrated with the dsRNAs of the virus were readily detected as synthesized products, indicating viral RDRP activity. In addition, smaller-sized dsRNA species, possibly two of approximately 700 bp (s1) and one of 500 bp (s2), were synthesized by purified virus particles of the CsCl gradient surrounding the virus peak. No cross-hybridization with either s1 or s2 and the dsRNA segments occurred, suggesting that s1 and s2 were synthesized from different templates. An RNase A protection assay revealed that the synthesized s1 and s2 polymerase products were double stranded. Furthermore, hybridization of products with strand-specific RNA of s1 generated from cDNA indicated that only one strand was synthesized in vitro. s1 and s2 were not visualized in ethidium bromide-stained agarose gels of dsRNA of infected trichomonads grown in batch cultures. However, dsRNA profiles of the same infected organisms cultivated under defined continuous-flow conditions contained readily detectable levels of s1 and s2, indicating that amplification of s1 and s2 occurred under specific environmental conditions. These newly discovered dsRNAs were not detected in all of the virus-carrying isolates. Finally, it is noteworthy that the s1 and s2 dsRNAs and the RDRP activity were not detected in trichomonal isolates without virus or in virus-negative progeny derived from virus-positive parental isolates. These data indicate the possibility of variations in the number of dsRNAs and/or of the presence of satellites in trichomonads infected with the multisegmented virus.
大多数阴道毛滴虫分离株都是多节段双链RNA（dsRNA）病毒的携带者。进行了体外聚合酶测定以证明纯化颗粒的RNA依赖性RNA聚合酶（RDRP）活性。与病毒的dsRNAs迁移一致的转录本很容易被检测为合成产物，表明存在病毒RDRP活性。此外，在围绕病毒峰的CsCl梯度的纯化病毒颗粒合成了较小尺寸的dsRNA种类，可能是两条约700 bp的（s1）和一条500 bp的（s2）。s1或s2与dsRNA片段均未发生交叉杂交，这表明s1和s2是从不同模板合成的。核糖核酸酶A保护试验表明，合成的s1和s2聚合酶产物是双链的。此外，产物与由cDNA产生的s1链特异性RNA杂交表明，体外仅合成了一条链。在分批培养的受感染滴虫的dsRNA的溴化乙锭染色琼脂糖凝胶中未观察到s1和s2。然而，在确定的连续流动条件下培养的相同受感染生物体的dsRNA图谱中含有易于检测到的s1和s2水平，这表明s1和s2在特定环境条件下发生了扩增。并非在所有携带病毒的分离株中都检测到这些新发现的dsRNAs。最后，值得注意的是，在没有病毒的滴虫分离株或来自病毒阳性亲本分离株的病毒阴性后代中未检测到s1和s2 dsRNAs以及RDRP活性。这些数据表明，感染多节段病毒的滴虫中dsRNAs数量和/或卫星存在情况可能存在差异。
