The antiviral effect of the acyclic nucleoside phosphonate tenofovir (R)-PMPA on double-stranded DNA Cauliflower mosaic virus (CaMV) in Brassica pekinensis plants grown in vitro on liquid medium was evaluated. Double antibody sandwich ELISA and PCR were used for relative quantification of viral protein and detecting nucleic acid in plants. (R)-PMPA at concentrations of 25 and 50 mg/l significantly reduced CaMV titers in plants within 6-9 weeks to levels detectable neither by ELISA nor by PCR. Virus-free plants were obtained after 3-month cultivation of meristem tips on semisolid medium containing 50 mg/l (R)-PMPA and their regeneration to whole plants in the greenhouse. Studying the metabolism of (R)-PMPA in B. pekinensis revealed that mono- and diphosphate, structural analogs of NDP and/or NTP, are the only metabolites formed. The data indicate very low substrate activity of the enzymes toward (R)-PMPA as substrate. The extent of phosphorylation in the plant's leaves represents only 4.5% of applied labeled (R)-PMPA. In roots, we detected no radioactive peaks of phosphorylated metabolites of (R)-PMPAp or (R)-PMPApp.

• MeSH
• Adenine analogs & derivatives   metabolism   pharmacology   MeSH
• Antiviral Agents metabolism   pharmacology   MeSH
• Biotransformation MeSH
• Brassica metabolism   virology   MeSH
• Caulimovirus drug effects   growth & development   MeSH
• DNA, Viral analysis   MeSH
• Enzyme-Linked Immunosorbent Assay methods   MeSH
• Phosphorous Acids metabolism   pharmacology   MeSH
• Polymerase Chain Reaction methods   MeSH
• Viral Load MeSH
• Viral Proteins analysis   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

A new method was developed for testing antiviral compounds against plant viruses based on rapidly growing brassicas in vitro on liquid medium. This method enables exchange of media containing tested chemicals in various concentrations and simultaneous evaluation of their phytotoxicity and antiviral activity. While using ribavirin as a standard for comparison, phytotoxicity and ability of the acyclic nucleotide analogues (R)-PMPA, PMEA, PMEDAP, and (S)-HPMPC to eliminate ssRNA Turnip yellow mosaic virus (TYMV) were evaluated by this method. Double antibody sandwich ELISA and real-time PCR were used for relative quantification of viral protein and nucleic acid in plants. Ribavirin had the most powerful antiviral effect against TYMV. On the other hand, (R)-PMPA and PMEA had no antiviral effect and almost no phytotoxicity compared to the control. (S)-HPMPC and PMEDAP showed moderate antiviral effect, accompanied by higher phytotoxicity. The tested compounds can be screened within 6-9 weeks in contrast to the 6 months for traditionally used explants on solid medium. The method enables large-scale screening of potential antivirals for in vitro elimination of viruses from vegetatively propagated crops and ornamentals.

• MeSH
• Hydrocarbons, Acyclic pharmacology   therapeutic use   MeSH
• Antiviral Agents pharmacology   therapeutic use   MeSH
• Brassica drug effects   growth & development   virology   MeSH
• Hydroponics methods   MeSH
• Culture Media MeSH
• Microbial Sensitivity Tests methods   MeSH
• Plant Diseases therapy   virology   MeSH
• Nucleosides pharmacology   therapeutic use   MeSH
• Virus Replication drug effects   MeSH
• Ribavirin pharmacology   therapeutic use   MeSH
• Tymovirus drug effects   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH