First Report of Citrus leaf blotch virus in Lemon in China

Abstract

HomePlant DiseaseVol. 101, No. 8First Report of Citrus leaf blotch virus in Lemon in China Previous DISEASE NOTES OPENOpen Access licenseFirst Report of Citrus leaf blotch virus in Lemon in ChinaM. J. Cao, Y.-Q. Yu, X. Tian, F. Y. Yang, R. H. Li, and C. Y. ZhouM. J. CaoSearch for more papers by this author, Y.-Q. YuSearch for more papers by this author, X. TianSearch for more papers by this author, F. Y. YangSearch for more papers by this author, R. H. LiSearch for more papers by this author, and C. Y. ZhouSearch for more papers by this authorAffiliationsAuthors and Affiliations M. J. Cao , National Citrus Engineering and Technology Research Center, Citrus Research Institute, Southwest University/Chinese Academy of Agricultural Sciences, Beibei, Chongqing 400712, China Y.-Q. Yu , National Citrus Engineering and Technology Research Center, Citrus Research Institute, Southwest University/Chinese Academy of Agricultural Sciences, Beibei, Chongqing 400712, China; and College of Plant Protection, Southwest University, Beibei, Chongqing 400715, China X. Tian F. Y. Yang , National Citrus Engineering and Technology Research Center, Citrus Research Institute, Southwest University/Chinese Academy of Agricultural Sciences, Beibei, Chongqing 400712, China R. H. Li , USDA-ARS, National Germplasm Resources Laboratory, Beltsville, MD 20705 C. Y. Zhou , National Citrus Engineering and Technology Research Center, Citrus Research Institute, Southwest University/Chinese Academy of Agricultural Sciences, Beibei, Chongqing 400712, China. Published Online:12 May 2017https://doi.org/10.1094/PDIS-10-16-1500-PDNAboutSections ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InRedditEmailWechat Citrus is one of most important economic fruit crops in China. Citrus leaf blotch virus (CLBV) is a virus species of genus Citrivirus in family Betaflexiviridae. This virus was first reported from Nagami kumquat in association with bud union disorder (Navarro et al. 1984). It is transmitted in citrus by grafting and seed (Guerri et al. 2004). CLBV was found recently in sweet cherry (Wang et al. 2016) and kiwifruit (Zhu et al. 2016) in China. In May 2015, one Eureka lemon sample (Citrus limon (L.) Burm.f.) displaying yellow vein clearing symptoms was found in a citrus orchard in Wanzhou in Chongqing municipality during a survey for virus-like symptoms. Leaf tissues were collected and sent to the Beijing Genomics Institute for total RNA preparation, small RNA (sRNA) library construction, and sequencing. De novo assembly of sRNA reads was performed using CLC Genomics Workbench (CLC bio v10.0, Denmark), and 623 contigs >50 nucleotides (nt) were obtained after removal of host sRNAs. BLASTn and BLASTx searches against nonredundant nucleotide and protein databases revealed eight contigs similar to Citrus yellow vein clearing virus (CYVCV) and five contigs to CLBV. The largest CLBV contig of 4,683 nt was 98% identical to CLBV isolate NZ_G78 (EU857540). From this sequence, two primer pairs, F2 (5′-AGAGGGAAAGGTCTTGGAGTG-3′)/R2 (5′-GCAATCTATCAGCAGTCAGGA-3′) and F3 (5′-AAGTCAATGGCAAAGAAGCG-3′)/R3 (5′-TCGTGGTGAGATTTCGTCTG-3′), were designed from this contig and which in RT-PCR successfully amplified 412 nt (partial replicase polyprotein and movement protein) and 597 nt (partial movement protein) fragments from the symptomatic lemon tissue. These fragments were cloned, sequenced, and found to be the appropriate CLBV genomic regions characterized from the contig. Bark grafting was performed from the lemon plant to ‘Dweet’ tangor plants and characteristic chlorotic mottle symptoms developed on leaves by 6 months after grafting; cloning and sequencing from ‘Dweet’ tissues confirmed the presence of CLBV. To further investigate the distribution of the virus, 20 citrus samples collected from different regions of China were tested by RT-PCR. Three infected tangor (C. reticulata × C. sinensis) samples were identified, one each from trees of varieties Harumi, Reikou, and Shiranuhi from Hunan, Sichuan, and Zhejiang provinces, respectively. The aforementioned 412 and 597 nt genomic regions were amplified, cloned, and sequenced from these samples. BLASTn searches of the clones from Harumi (KX924465 and KX924464), Reikou (KX924466 and KX924467), and Shiranuhi (KX924468 and KX924469) and Wanzhou (KX924470 and KX924471) revealed all sequences had at least 98% identity with CLBV isolate NZ_G78. To the best of our knowledge, this is the first report of CLBV infection on citrus in China. As CLBV has been reported to be capable of invading the apical meristem, results indicate the necessity for detection and therapy in citrus germplasm and nursery programs in China.