First report of Plum pox virus on plum in Finland

In July 2014, leaves showing symptoms of a viral infection were collected from a plum tree serving as a mother tree in a Finnish nursery and found to be infected by Plum pox virus (PPV). A subsequent survey revealed additional infected trees originating from the infected mother tree. This paper provides the first report of PPV, the causal agent of the most destructive viral disease of Prunus, in Finland.     

Plum pox potyvirus on sour cherry in Moldova

Plum pox potyvirus (PPV) was identified in six cultivars of sour cherry in the collection orchard of the Moldavian Horticultural Research Institute. Study of biological properties of the sour cherry isolate in herbaceous indicators showed its similarity or identity with the PPV isolate of Van Oosten and a significant difference from isolates widespread in Moldova. A purified viral preparation was used to develop antiserum with a working titre of 1:1024. Comparative serological examination of the sour cherry and conventional plum PPV isolates using ELISA, ISEM and SDS-PAGE of the protein capsid could not differentiate these isolates. The sour cherry isolate was transferred to plum resulting in weak but distinctive PPV symptoms in susceptible cv. Sopernitsa.     

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) of plum pox potyvirus Turkey (PPV-T) strain

Journal of Plant Diseases and Protection - Plum pox potyvirus (PPV) is one of the most important viral pathogens of stone fruit trees. PPV-T (Turkey) is important, unique and dominated strain in...     

Unravelling the Prunus/Plum pox virus interactions

Plum pox virus (PPV) belongs to the genus Potyvirus that contains the largest number of virus species infecting plants. Its virus genome has been extensively characterised and sequenced. However, few data are available on its interactions with woody host plants. We therefore focused, in the past 4 years, both on the cellular and molecular aspects of the compatible and incompatible Prunus /PPV interactions. GFP (Green fluorescent protein)-tagged PPV and in situ hybridisation were used to compare the localization of viral particles in stems and leaves of susceptible and resistant apricot cultivars. In parallel, molecular tools were developed through the cloning and characterization of polymorphic, homologous resistance genes and of candidate genes involved in the expression of Prunus /PPV interactions. Candidate genes are currently used to target genomic regions involved in resistance or susceptibility and to identify molecular markers indispensable for further marker assisted selection for resistance to sharka disease.     

Causal agent of sharka disease: Plum pox virus genome and function of gene products

Our understanding of the molecular biology of Plum pox virus (PPV) and of the interactions between the virus and its hosts has advanced notably in recent years. Complete genome sequences have been obtained showing up the existence of new subgroups. Functions related to replication, symptom induction, virus movement, and interference with host defense mechanisms have been assigned to the different protein products derived from the proteolytic processing of PPV polyprotein. Moreover, the use of new approaches to define protein–protein interactions between the different viral proteins and between these and the host proteins has begun to yield the first results improving our understanding of how the virus and its host interact during infection.     

Significance of the heterologous encapsidation of zucchini yellow mosaic potyvirus in transgenic plants expressing plum pox potyvirus capsid protein1

Transgenic Nicotiana benthamiana plants expressing the coat protein of an aphid-transmissible strain of plum pox potyvirus (PPV-D) were infected with an aphid non-transmissible strain of another potyvirus, zucchini yellow mosaic potyvirus (ZYMV-NAT). Non-viruliferous Myzus persicae could acquire and transmit ZYMV-NAT from these plants but not from infected N. benthamiana control plants (not transformed, or transformed by the vector alone). Immunosorbent electron microscopy experiments using the decoration technique revealed that ZYMV-NAT virus particles in the infected transgenic plants expressing the PPV coat protein could be coated not only with ZYMV antibodies but also, on segments of the particles, with PPV antibodies. This suggests that aphid transmission of ZYMV-NAT occurred through heterologous encapsidation, and reveals a potential risk of releasing genetically engineered plants expressing viral coat proteins into the environment.     

Development of an on-site plum pox virus detection kit based on immunochromatography

Sharka disease, caused by plum pox virus (PPV), is the most serious viral disease of stone fruit trees. Among the eight known strains of the virus, PPV-D is the most important due to its recent global spread. Although enzyme-linked immunosorbent assay (ELISA) is the most common approach for diagnosing sharka, it involves time-consuming steps and requires expensive equipment and trained technicians. In this study, an on-site PPV detection kit based on immunochromatography was developed using polyclonal antibodies against the coat protein (CP) of a PPV-D isolate. The immunochromatographic (IC) assay kit was as sensitive as a commercial ELISA system for detecting Japanese PPV-D isolates. Moreover, it was easy to use (a one-step procedure), and results could be obtained on-site within 15 min without special laboratory equipment. The IC assay kit detected the virus from every aerial part of symptomatic Japanese apricot trees. In a detailed study of viral localization in leaves, the most suitable plant parts for use in the IC assay were symptomatic mesophyll tissues and the region from the petiole to the main vein. A positive reaction was also observed using the CP of other major (PPV-M and PPV-Rec) and minor (PPV-EA, PPV-W, and PPV-T) strains.     

Resistance to Plum pox virus in plants expressing cytosolic and nuclear single‐chain antibodies against the viral RNA NIb replicase

The expression of engineered single‐chain variable fragments specific to the NIb RNA replicase of Plum pox virus (PPV) (scFv2A) in transgenic plants was successfully used as a strategy to interfere with viral infection. Different scFv2A fusion proteins were constructed to target those subcellular compartments, such as the cytosol, endoplasmic reticulum (ER) membrane structures and the nucleus, where NIb protein presumably accumulates. Several transgenic lines of Nicotiana benthamiana plants expressing the scFv2A targeted to the cytosol (2A lines), ER (6K2 lines) and nucleus (NLS lines) were obtained. The protective effect of scFv expression was determined by mechanical virus inoculation in five 2A, three 6K2 and four NLS transgenic lines. The strongest resistance was afforded with the 2A‐3 (six non‐infected plants out of 10), 6K2‐1 (17 out of 33) and NLS‐11 (16 out of 19) transgenic lines. The success of this interference with PPV infection opens new possibilities for the control of this RNA virus and could be exploited not only to confer resistance in transgenic plants, but also to elucidate the role of the non‐structural NIb protein in different cell compartments during viral infection.     

Detection of plum pox potyvirus using monoclonal antibodies to structural and non-structural proteins1

Eleven monoclonal antibodies specific to plum pox potyvirus (PPV) coat protein were obtained by hybridoma technology from Spanish PPV isolates. In addition, two monoclonal antibodies specific for PPV cylindrical inclusions (CIP non-structural proteins) were obtained. The monoclonal antibodies specific for PPV coat protein were assayed by DASI ELISA against 81 PPV isolates. At least nine different epitopes were found and 21 distinct serological patterns of reaction (serogroups) were established using nine selected monoclonal antibodies against the collection of PPV isolates, indicating the high variability of coat protein among PPV isolates. Changes in epitope composition were observed after aphid and mechanical transmission, indicating the occurrence of mixtures of isolates in field trees. Monoclonal antibody 5B reacted with all PPV isolates assayed, with very high affinity, using DASI ELISA. This method was compared with immunocapture-PCR on field samples in spring, and showed very good coincidence of results. The efficiency of PPV detection can be slightly increased using monoclonal antibodies specific to cylindrical inclusions mixed with monoclonal antibodies against structural proteins, and using mixtures of monoclonal antibodies against different epitopes of coat protein. ELISA-I and immunoprinting-ELISA were able to detect CIP and PPV in extracts and tissue section, respectively, of woody plants. Two monoclonal antibodies offer the possibility of distinguishing between Marcus and Dideron PPV types (M or D). These D-specific monoclonal antibodies can be used in routine tests with high affinity.     

The use of Plum pox virus as a plant expression vector

Among the biotechnological uses of plant viruses, the expression of foreign sequences through virus-based vectors represents a promising research area. The potyvirus Plum pox virus (PPV) has been used to design expression vectors which have allowed successful expression of foreign sequences in plants, either in the form of small peptides fused to the viral coat protein, or as whole independent proteins inserted on different points of the genome. The present review describes the different PPV-based vectors that have been produced, including information regarding relevant aspects of their use, such as the optimal location of peptides or the stability of inserts. Recent developments, like the expression of proteins on stone fruit trees by using PPV-based vectors capable to infect woody plants, are also described.     

Breeding for resistance: breeding for Plum pox virus resistant apricots (Prunus armeniaca L.) in the Czech Republic

Plum pox virus (PPV) causes serious damage in apricots grown in the Czech Republic and other countries where it is present. To study PPV resistance in apricot, three backcrosses between apricot cultivars or selection resistant to PPV and apricot cultivars or selections susceptible to PPV ('LE-3218' × 'Stark Early Orange', 'LE-3241' × 'Vestar' and 'LE-3246' × 'Vestar') were performed. The seedlings were repeatedly inoculated with PPV-M strain by an infected chip. The resistance of the plants was evaluated by visual inspection for symptoms and ELISA in three consecutive growth periods. The Chi-square (χ²) test was used to analyse the data. It was found that two independent dominant complementary genes conditioned PPV resistance in apricot in the case of the backcrosses. The mode of inheritance for resistance to PPV in cv. Harlayne was determined from F1 progenies. Resistance to PPV in 'Harlayne' was controlled by three independent complementary dominant genes. This knowledge will help us in effective planning of apricot breeding programmes.     

Control and monitoring: control of Plum pox virus in the United Kingdom

Plum pox virus (PPV) was first identified in the United Kingdom in 1965. Despite a rigorous eradication policy, the disease spread quickly and established itself in all the main plum-growing areas in England. In 1975, the policy was changed from a blanket eradication campaign to one of containment; retaining statutory control of PPV on propagation material but allowing the industry to control the disease in orchards. As part of the current containment campaign, annual surveys are carried out on propagation material. These surveys show that the incidence of PPV in this material is very low and that only the D-strain is present. The precise situation regarding PPV incidence in commercial orchards is unknown. Given the low incidence in propagation material, it is likely that PPV is uncommon in actively managed orchards. However, some infected orchards probably do still exist, especially older, unmanaged or abandoned ones. Overall, the history of PPV control in the UK is one of unsuccessful eradication but successful containment. The UK experience demonstrates that given the right combination of strain and host, alongside a regular testing regime, it is possible to control PPV through the establishment of a regulated certification scheme and the supply of virus-free planting material.     

Detection and characterization of plum pox potyvirus isolates from southern Germany by polymerase chain reaction (PCR)1

Plum pox potyvirus (PPV) isolates show quantitative and qualitative differences in host plants, symptomatology, aphid transmissibility and molecular biology. In order to gain an overview of the different PPV strains that exist in Germany, several PPV isolates, mainly deriving from southern Germany, were analysed by polymerase chain reaction (PCR), the reaction yielding a 243 bp product. All PPV isolates tested were amplified and the amplified fragments were analysed by restriction endonuclease digestion. An Rsa I restriction site polymorphism in the amplified fragments allowed the discrimination of two groups of isolates.     

Detection and characterization of Plum pox virus: serological methods

Tremendous progress has been made in the research and development of Plum pox virus (PPV) serological reagents and methods in recent years. Two facts have revolutionised the serological detection and characterization of the virus: the development of the ELISA method in 1977, and the later emergence of specific monoclonal antibody technology. The availability of commercial kits has popularised PPV diagnosis, now making diagnosis possible at large scale for quarantine purposes, eradication programmes and control of the disease in nurseries. The use of the universal monoclonal antibody 5B-IVIA, used in DASI-ELISA, is the most accurate system for routine PPV detection. Likewise, the use of typing monoclonal antibodies gives exact characterization of the main PPV types described: 4DG5 for PPV-D, AL for PPV-M, EA24 for PPV-EA, and TUV and AC for PPV-C. There is, in general, an excellent correlation between serological data obtained with PPV specific monoclonal antibodies and data obtained by molecular PCR based methods. ELISA using a single or a mixture of monoclonal antibodies will remain the preferred method for universal detection and routine screening of PPV for years to come. Today, other serological methods and reagents are also recommended in the EPPO Diagnostic Protocol, increasing the number of reliable tests available for PPV detection. These developments have helped to control sharka disease in recent years. International co-operation in this field has been crucial to the improvement and validation of serological tools for PPV detection and characterization.