Citrus yellow mottle-associated virus (CiYMaV) belonging to the subgenus Mandarivirus within the genus Potexvirus, was first identified in 2018 from Pakistan (CiYMaV-PK), where it is endemic in several regions.  Here, three full-length cDNA clones (pCiYMaV-FL-1, pCiYMaV-FL-18, and pCiYMaV-FL-22) corresponding to the genomic RNA of CiYMaV were constructed and then agroinfiltrated on Chandler pummelo (Citrus grandis) seedlings using the vacuum infiltration method.  All the inoculated plants developed severe vein yellowing, leaf mottling, and dwarfing symptoms by 40 days post-infiltration (dpi).  The results of a direct tissue blot immunoassay and reverse transcription polymerase chain reaction detection showed 94.7–100% infection rates of pCiYMaV-FL at 60 dpi.  Despite there being no observed difference among the three clones in the severity of symptom, pCiYMaV-FL-22 showed the highest accumulation levels of viral RNA and coat proteins.  Moreover, pCiYMaV-FL-22 successfully infected seven other citrus varieties and induced symptoms in five of them.  Transmission electron microscopy identified the presence of filamentous virus particles in extracts from systemic leaves of the plants infected with pCiYMaV-FL-22 at 6-months post-infiltration.  Taken together, the results indicate that Koch’s postulates were fulfilled for CiYMaV in citrus plants.  This is the first report of full-length infectious cDNA clones of CiYMaV, and thus, the data provide a basis for further study of the molecular mechanisms of virus infection and the development of a viral vector to express foreign genes in citrus plants.

Although it is usually latent on citrus, apple, and pear, apple stem grooving virus (ASGV) poses a great risk to many sensitive cultivars.  Since severe leaf yellow mottle mosaic (LYMM) symptoms have been observed on Huangjinmiyou (HJY) pummelos (Citrus grandis cv. Huangjinmiyou), a commercial variety that is widely cultivated in South China, high throughput sequencing (HTS) was used to find potential pathogens and only three divergent ASGV variants were identified.  The three ASGV variants shared 81.03–82.34% genome-wide pairwise identities with each other, and were separately closest to other ASGV variants from different hosts and/or geographical regions, as indicated by viral phylogenies.  However, these new variants may have developed from viral interstrain interactions, based on the results of recombination analysis.  A large-scale survey using reverse transcription-PCR (RT-PCR) protocols designed for the three ASGV variants revealed a high incidence (92.7–100%) of ASGV in symptomatic HJY trees from 11 major citrus-producing regions in China.  None of ASGV were detected in asymptomatic trees.  Temperature treatments applied to the symptomatic HJY plants showed that ASGV is sensitive to high temperatures (30–35°C), at which not only the plants recovered, but also the viruses were not detected by RT-PCR, while at low temperatures (20–24°C), both the symptoms and viruses remained detectable.  These data show that ASGV is associated with the LYMM disease prevalent on HJY in China, and this is the significant basis especially of taking appropriate measures timely to manage the disease.  

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.