Research On Detection And Molecular Identification And Genetic Diversity Of Viruses Infected Melon And Watermel In Guangxi Province

Melon and watermelon are important cash crops in Guangxi province.for better control virus diseases in melon and watermelon,detection and identification of virus were conducted by using serological and molecular biological methods,and analyses of genetic diversity were performed to main viruses genomic sequences,which were cloned by reverse transcription polymerase chain reaction(RT-PCR).A rapid diagnosis method was developed to detect the three RNA viruses successf-ully.Research showed that Cucumber mosaic virius(CMV),Tobacco mosaic virus(TMV),Watermelon mosaic virus(WMV),Squash mosaic virus(SqMV),Cucumber green mottle mosaic virus(CGMMV),Zucchini yellow mosaic virus(ZYMV),and Papaya ringspot virus(PRSV)were detected from watermelon samples,the dominant viruses were ZYMV and PRSV,which account for 41.9%,22.8%among detected viruses respectively.There were 9 viruses in melon samples,they are CMV,TMV,WMV,CGMMV,ZYMV,PRSV and Cucurbit chlorotic yellows virus(CCYV),Melon yellow spot virus(MYSV)and Melon necrotic spot virus(MNSV).ZYMV account for 18.3%in melon samples most collected in fields,whilst the detection rate of CCYV and MYSV were 43.1%,40.5%rspectively in melon samples most collected in greenhouse.MNSV was first detected in melon from Guangxi.The results of statistic about mixed infection were showed that 7 types of co-infection were detected in watermelon,whilst 9 types detected in melon.The results of nearly full-length nucleotide sequence analysis showed that the whole genome sequence(RNA1 and RNA2)of CCYV Guangxi isolates had 99.5%-100%and 99.6%-100%identity with the nucleotide sequences of RNA1 and RNA2 registered on GenBank respectively.Phylogenetic analyses based on the CP gene sequence and complete genomic sequence placed Guangxi isolates and the other isolates of the CCYV in a major cluster.S RNA full-length sequence of MYSV Guangxi isolates had 94.9%-99.8%identity with other sequence registered on GenBank respectively.The identity of amino acid sequence encoded from nucleocapsid(N)and non structural protein(NSS)gene aligned with other isolates were 98.9%-100%and 95.3%-99.8%respectively.Phylogenetic analyses based on the S RNA full-length sequence and N gene sequence indicated that MYSV isolates from Guangxi is phylogenetically more closely related with isolates from china and thailand Physalis than others.The genomic nucleotide sequence identity of MNSV Guangxi isolate with sequence registered on GenBank range from 74.4%to 93.8%,whilst the amino acid sequences from each opening reading frame were between 93.0%and 100%identical to those of the other MNSV isolates(except two isolates infected watermelon from Japan).3'UTR of MNSV Guangxi isolate has a segment on the 5'terminal which has 90%identity with the counterparts of Cucurbit aphid-borne yellow mosaic virus(CABYV),whether its function is consistent with the isolate of MNSV-N remains to be studied.Phylogenetic analysis indicated that MNSV isolate from Guangxi was placed in a cluster alone,phylogenetically more closely related with some isolates from europe and America.In order to improve the efficiency of detection,a multiplex RT-PCR detection method was established to detect the three RNA viruses(CCYV,MYSV and MNSV)successfully.