柑橘黄化斑驳相关病毒（Citrus yellow mottle-associated virus，CiYMaV）属马铃薯X病毒属（Potexvirus）、印度柑橘病毒亚属（Mandarivirus），最早于2018年在巴基斯坦被发现，目前该病毒在巴基斯坦多地柑橘产区有发生。本研究首先借助植物双元表达载体pCass4-RZ构建了3个CiYMaV的全长cDNA克隆（pCiYMaV-FL-1、pCiYMaV-FL-18和pCiYMaV-FL-22），随后通过农杆菌真空浸润接种强德勒柚（Citrus grandis）验证其侵染活性。结果表明，在接种40-60天，3个克隆均引起严重的叶脉黄化、叶片斑驳和植株矮化的症状。直接组织点免疫（direct tissue blot immunoassay，DTBIA）和反转录聚合酶链式反应（reverse transcription polymerase chain reaction，RT-PCR）的检测结果表明3个克隆的侵染率为94.7-100%。尽管3个克隆引发的症状相似，pCiYMaV-FL-22在接种60天时，其在植株内的RNA和蛋白积累水平最高。因此，选择pCiYMaV-FL-22将其接种到7种柑橘和3种草本寄主上，结果表明pCiYMaV-FL-22能成功侵染7种柑橘寄主，并且在5个柑橘品种上引起明显症状。通过透射电镜，在接种pCiYMaV-FL-22的柑橘植株组织中观察到长线型、弯曲的病毒颗粒。综上所述，本研究构建了CiYMaV的全长感染性cDNA克隆，完成了该病毒的柯赫氏法则验证，为进一步研究该病毒侵染的分子机制和开发能应用于柑橘上的病毒载体奠定了基础。

苹果茎沟病毒 (apple stem grooving virus, ASGV) 是一种重要的潜隐类果树病毒，对柑橘、梨和苹果等多种果树的生产构成了严重的威胁。2018年，在中国南方广泛种植的黄金蜜柚 (Citrus grandis cv. Huangjinmiyou) 上观察到了严重的黄化、斑驳和花叶症状，推测其可能由病毒引起。取5株表现相关症状果树的叶片样品构建混库并送高通量测序分析，从其中鉴定到了3个ASGV变异体，通过RT-PCR和RACE技术验证了其基因组序列。序列分析显示，这3个变异体的基因组核苷酸序列一致性为81.03%–82.34%，其基因组结构与过往报道的侵染其它果树的变异体类似。基于病毒全基因组核苷酸序列和外壳蛋白氨基酸序列的系统发育分析显示，3个黄金蜜柚ASGV变异体分别与来自不同寄主和地区的ASGV变异体聚在一枝。重组分析显示，3个ASGV变异体可能来自于ASGV不同株系间的重组。在全国11个主要柑橘种植省份采集了507份黄金蜜柚样品进行RT-PCR检测发现，在每个省份表现上述相关症状的样品中，ASGV的检出率均在92.7%以上，而在40份没有症状的样品中，均未检测到ASGV。将其中6个省份的感病样品嫁接到ASGV的指示植物——Rusk枳橙上，新生的系统叶表现出典型的碎叶症状，进一步验证了黄金蜜柚中ASGV的侵染。进一步探究了病毒和症状与温度的关系，发现嫁接的感病黄金蜜柚样品在30°C–35°C条件下症状消失，同时RT-PCR也检测不到ASGV的存在。而后，再将其置于20°C–24°C的条件下一段时间后，黄金蜜柚症状恢复，且ASGV可以由RT-PCR检测到。本文揭示了黄金蜜柚黄化斑驳花叶病与ASGV侵染的相关性，并提示了该病害大面积流行的风险，为进一步的病害防控提供了相应参考。

Citrus leaf blotch virus (CLBV) is a member of the genus Citrivirus, in the family Betaflexiviridae.  It has been reported CLBV could infect kiwi, citrus and sweet cherry in China.  Of 289 citrus samples from six regions of China, 15 were detected to be infected with CLBV in this study.  The complete genome of four isolates of CLBV was obtained from Reikou in Sichuan (CLBV-LH), Yura Wase in Zhejiang (CLBV-YL), Bingtangcheng in Hunan (CLBV-BT), Fengjie 72-1 in Chongqing (CLBV-FJ), respectively.  While they all represented 8 747 nucleotides in monopartite size, excluding the poly(A) tail, each of the isolates coded three open reading frames (ORFs).  Identity of the four isolates ranged from 98.9 to 99.8% to each other and from 96.8 to 98.1% to the citrus references in GenBank by multiple alignment of genomes.  A phylogenetic tree based on the genome sequences of available CLBV isolates indicated that the four isolates were clustered together, suggesting that CLBV isolates from citrus in China did not have obvious variation.  This is the first report of the complete nucleotide sequences of CLBV isolates infecting citrus in China.

To identity the potential pathogen associated with the yellow vein clearing symptom on lemon trees, the profiles of virus-derived small interfering RNAs from citrus samples were obtained and analyzed by deep sequencing method in this study.  Twenty-seven contigs almost cover the full length genome of Citrus yellow vein clearing virus (CYVCV) isolate YN were obtained using the small RNA deep sequencing technology.  Analysis showed that this isolate CQ shared the highest nucleotide identity with isolate Y1 (JX040635) and YN (KP313242), both of which belong to the genus Mandarivirus in the family Alphaflexiviridae.  Mapping analysis of viral-derived siRNA (vsiRNA) profiles revealed an uneven distribution pattern of their generations along both positive and negative genome strands, and 22- and 21-nt vsiRNAs ranked the majority.  BLAST against viroids and other viral databases confirmed that this sample was single-infected only by CYVCV, which indicated that CYVCV was the exact causal agent for the yellow clearing symptom occurring on lemon.  This is the first CYVCV isolate detected in Chongqing and the second in China.  This result could provide a molecular basis for the investigation of citrus viral diseases to protect citrus health in this region. 

The complete nucleotide sequence of an isolate of Citrus vein enation virus (CVEV-XZG) from China has been determined for the first time. The genome consisted of 5 983 nucleotides, coding for five open reading frames (ORFs), had a similar genomic organization features with Pea enation mosaic virus (PEMV). Nucleotide and deduced amino acid sequence identity of the five ORFs compared to isolate CVEV VE-1 range from 97.1 to 99.0% and 97.4 to 100.0%, these values compared to isolate PEMV-1 range from 45.2 to 51.6% and 31.1 to 45.2%. Phylogenetic analysis based on the complete genome sequence showed that the isolate CVEV-XZG had close relationship with Pea enation mosaic virus. The results supports CVEV may be a new member of genus Enamovirus. The full sequence of CVEV-XZG presented here may serve as a basis for future study of CVEV in China.

Citrus tristeza virus (CTV) is one of the most important causal agents of citrus diseases and exists as numerous strains.CTV is replicated in phloem cells of plants within the family Rutaceae and is transmitted by a few of aphid species. CTVepidemics have caused death of millions of citrus trees in many regions all over the world, where the sour orange (Citrusaurantium) was used as rootstock. Also the production of grapefruit (C. paradisi) and sweet orange (C. sinensis) hasbeen affected by CTV strains. CTV gives uplift to three prominent syndromes, namely quick-decline (tristeza), stempittingand seedling-yellows. The disease is graft-transmissible in nature but not seed-transmitted. However, the tristezadisease in most citrus groves was a man-made problem created by the desire of horticulturists to introduce cultivars fromother citrus growing areas. The utmost importance of the disease called for review articles in numbers of plant protection,epidemiology books, citriculture and proceedings. This review collects the information with respects to disease history,distribution host range, virus isolates association, identification and detection, transmission and management; especiallyon the current status of CTV prevailing and controlling in Pakistan. It provides valuable information for CTV disease andits controlling approaches.