Identification Of The Pathogens Of Panax Notoginseng Viral Diseases And Molecular Variation Analysis Of The Related Viruses

Panax notoginseng is a pearl in traditional Chinese herb, has very importantmedicinal and health value. However, because of its special growth and managment condition, it iseasy to cause the virus disease occurrence and result in damage. The disease seriously affects theviability of Panax notoginseng, reduces the production and quality of Panax notoginseng,and posed serious threat to production, cause heavy economic losses. But so far, few studies onPanax notoginseng virus disease and existing research only stay in Panax notoginseng viruspathogens identified level. Plant virus disease is called" plant cancer", the most important plant virusdisease prevention and control work at first is to clear the virus pathogen, then through the analysisof each pathogen transmission, host range and occurrence regularity, prevention and control theoccurrence of virus disease. For that reason, this paper aims to carry out further studies in theidentification of the Pathogens of P. notoginseng viral diseases, to explore the dominant and anothervirus species in P. notoginseng expect PnVY, clear its host range and determine the correlation ofvirus isolates/complex and Symptom types. The results woule provide a solid theoretical foundationand technical guidance for the further studies in pathogenic molecular mechanism of related virusand in the monitoring, forecasting, effective prevention and control of diseases. What’s more, thisstudy would play a role in learning and promoting at related research work in the field at home andabroad.dsRNA and viral purification were extracted from symptomatic P. notoginseng leaves. Adouble-stranded cDNA library was built based on dsRNA and negative staining electronicmicroscopy, SDS-PAGE, spectrometry technology were tested based on viral purification to obtainpossible virus species and observe the virus particle. The results showed that there were more thanone viral pathogens besides PnVY in symptomatic P. notoginseng leaves. And low concentration of dsRNA cause failed to the successful completion of the construction of cDNA libraries. Massspectrometry analysis of the results is SbDV, but SbDV was not detected in the sample bysequencing.The full-length genome sequences of TYLCCNV and TYLCCNB from one isolate, YWSh03,were obtained by PCR using virus-specific primers of TYLCCNV and degenerate primers ofgeminiviruses including DNA and betasatellite. Infectious clones of TYLCCNV DNA andbetasatellite were used to inoculate healthy P. notoginseng plants by an Agrobacterium-mediatedmethod. Viral transmission test was done by whiteflies which have been obtained virus. P.notoginseng plants inoculated showed yellow, but the plants tested by viral transmission have notobvious symptoms. However, TYLCCNV and TYLCCNB were detected in those samples. Itconfirmed TYLCCNV and TYLCCNB can infect P. notoginseng by molecular biology andbiological methods and satisfy Koch’s postulates. To our knowledge, this is the first report ofTYLCCNV infecting P. notoginseng and the family Araliaceae.Mechanical inoculation test with the symptomatic P. notoginseng leaves，dsRNA and viralpurification was carried out to12plants. Most host plants showed chlorosis after one month later.However, intermediate hosts of PnVY are N. tobacum, pepper and bean cowpea by RT-PCRdetection.Several notoginseng farms in Wenshan, Honghe, Kunming and Qujing were investigated. Thereare many symptoms of P. notoginseng viral disease and the incidence increases year by year. Thenumer of plants showed composite symptoms are large. PnVY and TYLCCNV detection The systemhas been gradually established and used for Simultaneous detecting PnVY and TYLCCNV. thesystem can accurately and quickly carry out the detection and analysis of these two kinds of virusdisease.The viruliferous percent of PnVY and TYLCCNV in different areas were detected by thesystem. The phenomenon that P. notoginseng plants were infected by the two kinds of virus is wide.It is38.7%~50.0%. On the whole, infection rate caused by PnVY was higher.The whole genome sequence of5PnVY islotes, six TYLCCNV isolates and4TYLCCNBisolates were obtained and were structure and system evolution analysised, It is supposed thatTYLCCNV and TYLCCNB may be have no relevance to notoginseng virus disease with symptomsof the complex and diverse types. The gene genetic structure of PnVY, especially PnVY P1proteinfunction may be associated with symptoms of notoginseng diversity.