山东省辣椒主要病毒种类的分子检测与鉴定

doi:10.3864/j.issn.0578-1752.2017.14.009

Abstract

Abstract: 【Objective】 The objective of this study is to identify the main viruses causing pepper virus diseases in Shandong Province, China.【Method】In 2014-2015, a total of 253 susceptible pepper plant samples were collected from 10 urban areas (Linyi, Rizhao, Qingdao, Yantai, Weifang, Zibo, Jining, Heze, Liaocheng, Dezhou) in Shandong Province. Total RNA and DNA were extracted from the leaves of pepper samples, and were subjected to detect viruses with Geminivirus universal primers (PA/PB), Polerovirus universal primers (POL-F/POL-R) and specific primers of main viruses infecting pepper have been reported by PCR and reverse transcription-PCR (RT-PCR). Each amplified fragment was purified by DNA gel extraction kit, cloned into pMD18-T vector and sequenced. The obtained sequences of PMMoV, PeVYV and BWYV were subjected to BLAST search, and compared with the representative sequences of GenBank which had been registered around the world using the Megalign in DNAStar software. Clustal W method of MEGA 5.05 was used for multiple sequence alignment analysis and the phylogenetic tree were constructed by neighbor-joining (NJ) of MEGA 5.05, with the bootstrap of each branch of phylogenetic tree analyzed 1 000 times. 【Result】The PMMoV and CMV had the highest detection rate, were 61.66% and 60.08%, respectively. TMGMV, BBWV-2, BWYV and TMV were popular with the detection rate of 41.90%, 34.78%, 33.20% and 24.90%, respectively. The detection rates of PCV-2, ToMV, TYLCV, PVY, MABYV, PeVYV, PCV-1, ChiVMV and AMV were 11.86%, 9.88%, 9.09%, 6.72%, 5.53%, 3.56%, 3.16%, 0.79% and 0.40%, respectively. No CaCV, PSV, ChiRSV, TSWV and ToMMV were detected. The complex infection phenomenon showed that co-infections were up to 89.92%, three kinds of viruses mixed infection occurs the highest rate, up to 28.63%, followed by four kinds of viruses mixed infection, up to 25.00%, two kinds of viruses mixed infection, up to 21.77%, five kinds of viruses mixed infection up to 13.31%, and 6 viruses detected in one sample is the extreme case, with the mixed infection rate up to 1.21%. Phylogenetic trees were constructed with partial CP of PMMoV, partial RdRp, CP and MP genes of PeVYV and partial CP and MP genes of BWYV, respectively. PMMoV Shandong isolate SD60 and the China Guizhou isolate had the closest relationship, PeVYV-SDRZ31-1 and Italy isolate IT83 shared the closest relationship, BWYV Shandong isolate SD clustered with Korean isolate LS. 【Conclusion】 Fifteen viruses were detected, among which PMMoV and CMV were the main pathogens. The newly detected viruses are MABYV, PeVYV, BWYV, MABYV, PCV-1 and PCV-2 in pepper in Shandong Province. The occurrence of viral diseases and the primary and secondary relationship of the main types of pepper viruses in Shandong Province were clarified.

Key words: pepper virus diseases, Pepper mild mottle virus (PMMoV), Beet western yellows virus (BWYV), Melon aphid-borne yellows virus (MABYV), complex infection

WANG ShaoLi, TAN WeiPing, YANG YuanYuan, DAI HuiJie, SUN XiaoHui, QIAO Ning, ZHU XiaoPing. Molecular Detection and Identification of Main Viruses on Pepper in Shandong Province[J].Scientia Agricultura Sinica, 2017, 50(14): 2728-2738.

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