| Sweet potato is the third largest tuber and stem crop in the world,and it is also an important food,feed,industrial raw materials and energy crop in China.In recent years,sweet potato virus disease has become increasingly serious in Hunan Province,affecting the stable and healthy development of sweet potato industry.In order to understand the occurrence of viruses in sweet potato germplasm resources in Hunan Province,the small RNA deep sequencing was used to analyze the types of sweet potato DNA viruses in sweet potato germplasm resources.The full sequences of sweet potato leaf curl virus and sweet potato badnavirus genes were identified by PCR,and the infectious clone library of sweet potato leaf curl virus were constructed for pathogenicity analysis.The prevalence of DNA virus in sweet potato germplasm was analyzed,and the main conclusion are as follows.1,Seven RNA viruses and two DNA viruses were identified in sweet potato by small RNA deep sequencing.The RNA viruses included sweet potato chlorotic fleck virus,sweet potato chlorotic stunt virus,sweet potato feather mottle virus,sweet potato latent virus,sweet potato virus 2,sweet potato virus C,and sweet potato virus G,and the DNA viruses included Sweepoviruses and SPBV.The assembled contigs of DNA viruses showed that sweet potato badnavirus B and sweet potato badnavirus A accounted for 63.23 % and 30.4 % of the total DNA genome sequences,respectively,sweet potato golden vein virus accounted for 24.40 % of the whole genome,and the13 sweet potato leaf curl virus strains accounted for 14.39-36.83 % of the each total genome sequences.2,The full sequences of two sweet potato badnavirus B genes were identified by PCR.Two full sequences of sweet potato badnavirus B genes,MK052980 and MK052981,were identified from sweet potato samples with fan-shaped leaves,short internodes and deformity.The phylogenetic tree analysis showed that the similarities between the two isolates(MK052980 and MK052981)and SPBV-B(FJ560944)were86% and 94%,respectively,the similarities between the two isolates(MK052980 and MK052981)and SPBV(FJ560943)were 81 % and 83 %,respectively.The amino acid analysis indicated at the 414 th and 560 th amino acid of the MK052980 ORF3 a were missing,and 6 new amino acids were detected at the 459–461aa,509 aa and540–541aa,respectively.3,A total of 12 sweet potato leaf curl virus whole genome sequences were identified by PCR for pathogenicity analysis.The 12 whole genome sequences of the sweet potato leaf curl virus genes were identified from sweet potato samples with symptoms of leaf curl,shrinkage and chlorotic spots,which were MK052982-MK052988 and MW021472-MW021476.Phylogenetic tree analysis showed that 12 isolates could be clustered into 4 groups,and SPLCV(41-56)had a relatively distant relationship with isolates KJ013578 and KJ013574.The genetic distance between the isolates SPLCV(6-1)and SPLCV(6-A5)was the nearest,0.001.The genetic distance between the clustered SPLCV(8-76)and the isolate MK931311 from Shandong was 0.007.Recombination analysis showed that SPLCV(3-3),SPLCV(5-1),SPLCV(6-1),SPLCV(6-A5),SPLCV(8-47),SPLCV(6-68),SPLCV(194),SPLCV(77),and SPLCV(41-56)were recombinant strains,of which seven SPLCV isolates were concentrated in the IR region and AV1,AV2 region,two isolates were in the AC1 and AC2 region,and three isolates were in the AC3 and AC4 region.Further,the p BA002-myc-SPLCV(21-5)infectious clone library was constructed to infect the sweet potato variety ’Xiangcaishu No.3 ’ and Sanshengyan,resulting in shrinkage,rolled leaves,chlorosis and other symptoms with two receptor plants.The infected leaves were used to inoculate virus on the same receptor plants by in-touch friction,resulting shrinkage and leaf curl on leaves that containing sweet potato leaf curl virus.4,Epidemic analysis of sweet potato DNA virus was conducted using PCR method with sweet potato germplasm resources in Hunan Province.Twenty two of the 180 sweet potato germplasm samples,were detected to be SPLCV positive,accounting for12.2 %,32 were SPBV positive accounting for 17.78 %,8 with both SPLCV and SPBV positive,accounting for 3.9%.The samples with SPLCV were collected from Changsha,Yongzhou,Huaihua,Shaoyang,Chenzhou,Changde,Loudi,Yiyang,Yueyang and Zhuzhou.SPBV was detected in samples collected from Changsha,Yongzhou,Huaihua,Hengyang and Shaoyang.Samples with both SPLCV and SPBV positive were sampled from Huaihua,Changsha,Yongzhou and Shaoyang.The results indicated the two viruses were widely distributed in Hunan,particularly in southwest region of Hunan Province.In this study,two DNA viruses,SPBV and Sweepoviruses,were detected from sweet potato plants,and the full sequence of SPBV-B gene was identified.A total of12 whole sequences of SPLCV gene and 9 recombinant sequences were identified.The recombinant sites were located in IR region,AV1,AV2 and AC1-AC4 regions.The p BA002-myc-SPLCV(21-5)infectious clones were constructed.The symptoms of shrinkage and chlorosis were observed in ’ Xiangcaishu 3 ’ and Sanshengyan.It indicates that SPLCV had pathogenicity.Further,the epidemic analysis showed the epidemiological characteristics of sweet potato DNA virus mainly existed in southwest region of Hunan.This study provides strong theoretical support for the prevention and control of sweet potato DNA virus and the healthy development of sweet potato industry. |
