Subject(s)
Begomovirus/genetics , Solanum lycopersicum/virology , Plant Diseases/virology , Plants, Genetically Modified/virology , RNA Interference , Satellite Viruses/genetics , Begomovirus/physiology , Solanum lycopersicum/genetics , Solanum lycopersicum/immunology , Oman , Plant Diseases/genetics , Plant Diseases/immunology , Plant Diseases/prevention & control , Plants, Genetically Modified/genetics , Plants, Genetically Modified/immunology , RNA, Double-Stranded/genetics , RNA, Double-Stranded/metabolism , RNA, Viral/genetics , RNA, Viral/metabolism , Satellite Viruses/physiologyABSTRACT
Purpose: Trichomonas vaginalis, a protozoan parasite, is the causative agent of human trichomoniasis, the most common non-viral sexually transmitted disease. The infection encompasses from a complete asymptomatic presentation to severe sequelae; yet, the virulence markers have been poorly understood. It is suggested that the presence of Trichomonas vaginalis virus (TVV) in T. vaginalis may have an impact on its virulence, and its relatedness to in vitro metronidazole resistance has been reported. The aim of the study was to assess the presence of TVV in fresh and Long -Term Cultivated ( LTC) maintained T. vaginalis isolates from symptomatic (S) and asymptomatic (AS) Indian women and its relatedness, if any, with symptomatology and in vitro drug sensitivity. Materials and Methods: One thousand women (537 S and 463 AS) were screened for the presence of T. vaginalis by wet smear and culture examination of vaginal swab and urine sample. Fresh and LTC (6 months-2 years) maintained 15 isolates each from 15 S and 15 AS women were subjected to agarose gel electrophoresis following total cellular RNA extraction to evaluate the presence of double stranded (ds) RNA viral infection. The susceptibility of isolates to metronidazole was determined in vitro. Results: On agarose gel electrophoresis, three bands (5.5, 2.5 and 1.5 kb) were observed in all the 30 fresh isolates from 15 S and 15 AS women and only in 7 LTC isolates from 3 S and 4 AS women. All the fresh isolates harbouring TVV were found to be sensitive to metronidazole in vitro irrespective of the symptomatology of subjects, and out of seven LTC isolates harbouring TVV, six were sensitive to metronidazole and one showed borderline resistance. Conclusions: The results suggest that the presence of TVV alone may not be a virulence marker and loss of TVV on LTC appears to be related to drug resistance. The T. vaginalis Indian isolates are sensitive to metronidazole.
Subject(s)
Adolescent , Adult , Antiprotozoal Agents/pharmacology , Asymptomatic Diseases , Drug Resistance , Female , Humans , India , Metronidazole/pharmacology , Parasitic Sensitivity Tests , RNA Viruses/genetics , RNA Viruses/isolation & purification , RNA, Double-Stranded/genetics , Trichomonas Vaginitis/parasitology , Trichomonas vaginalis/isolation & purification , Young AdultABSTRACT
In this study, we describe Korean isolates of Trichomonas vaginalis infected with double-stranded (ds) RNA virus (TVV). One T. vaginalis isolate infected with TVV IH-2 evidenced weak pathogenicity in the mouse assay coupled with the persistent presence of a dsRNA, thereby indicating a hypovirulence effect of dsRNA in T. vaginalis. Cloning and sequence analysis results revealed that the genomic dsRNA of TVV IH-2 was 4,647 bp in length and evidenced a sequence identity of 80% with the previously-described TVV 1-1 and 1-5, but only a 42% identity with TVV 2-1 and 3 isolates. It harbored 2 overlapping open reading frames of the putative capsid protein and dsRNA-dependent RNA polymerase (RdRp). As previously observed in the TVV isolates 1-1 and 1-5, a conserved ribosomal slippage heptamer (CCUUUUU) and its surrounding sequence context within the consensus 14-nt overlap implied the gene expression of a capsid protein-RdRp fusion protein, occurring as the result of a potential ribosomal frameshift event. The phylogenetic analysis of RdRp showed that the Korean TVV IH-2 isolate formed a compact group with TVV 1-1 and 1-5 isolates, which was divergent from TVV 2-1, 3 and other viral isolates classified as members of the Giardiavirus genus.
Subject(s)
Animals , Female , Humans , Mice , Abscess/parasitology , Capsid Proteins/genetics , Cloning, Molecular , Disease Models, Animal , Frameshifting, Ribosomal , Giardiavirus/classification , Korea , Molecular Sequence Data , Open Reading Frames , Phylogeny , RNA-Dependent RNA Polymerase/genetics , RNA, Double-Stranded/genetics , RNA, Viral/genetics , Sequence Analysis, DNA , Sequence Homology , Trichomonas Infections/virology , Trichomonas vaginalis/genetics , VirulenceABSTRACT
The discovery of double-stranded RNA-mediated gene silencing has rapidly led to its use as a method of choice for blocking a gene, and has turned it into one of the most discussed topics in cell biology. Although still in its infancy, the field of RNA interference has already produced a vast array of results, mainly in Caenorhabditis elegans, but recently also in mammalian systems. Micro-RNAs are short hairpins of RNA capable of blocking translation, which are transcribed from genomic DNA and are implicated in several aspects from development to cell signaling. The present review discusses the main methods used for gene silencing in cell culture and animal models, including the selection of target sequences, delivery methods and strategies for a successful silencing. Expected developments are briefly discussed, ranging from reverse genetics to therapeutics. Thus, the development of the new paradigm of RNA-mediated gene silencing has produced two important advances: knowledge of a basic cellular mechanism present in the majority of eukaryotic cells and access to a potent and specific new method for gene silencing.
Subject(s)
Animals , Mice , Caenorhabditis elegans/genetics , RNA Interference/physiology , RNA, Small Interfering/genetics , RNA, Double-Stranded/genetics , Gene Transfer Techniques , Mice, KnockoutABSTRACT
Trichomonas vaginalis can be infected with double-stranded RNA (dsRNA) viruses designated T. vaginalis virus (TVV), which may have important implications for trichomonal virulence and disease pathogenesis. We tested for TVV in 40 fresh T. vaginalis isolates from Cuban patients by total extraction of nucleic acids (DNA and RNA). TVV was detected in 22 (55 percent) of the 40 T. vaginalis isolates. This gives an estimate of the infection rate of Cuban T. vaginalis isolates by the dsRNA virus. Future research should focus on the association between trichomonosis symptoms and the presence of TVV.