Modes of vector transmission of Cherry leaf roll virus (CLRV) - molecular basis and potential arthropod vector species
Final Report Abstract
Cherry leaf roll virus (CLRV) is a plant pathogen of economic and ecologic importance. It is globally distributed in a wide range of forest, fruit, and ornamental trees and shrubs. Modes and interactions responsible for the wide intergeneric host transmission as well as for the exceptional CLRV epidemic in Fennoscandia still remain unknown. The aim in this project was to investigate by systematic studies if CLRV is potentially transmittable by biological vectors. CLRV belongs to the genus Nepovirus within the family Secoviridae of the order Picornavirales. To this group of viruses genetic determinants for vector specificities are dedicated to the viral coat protein (CP). Based on structural models predicting exposed CP surface amino acids and CP sequence alignments we have identified four regions potentially indicating towards vector specificities. Two regions (R1 and R2) are unique in CLRV variants from birches and cherry trees which form the distinct phylogentic group A. These motifs may indicate specificity to a vector group occurring on both, birches and cherry. R3 and R4 constitute amino acid compositions which are shared by distinct CLRV isolates and other nepoviruses conceding potential vector specificities to the group of aphids (R3) and to soil-inhabiting nematodes of Xiphinema spp. Functional proof of these motifs should have been implemented in vector studies by the use of reverse-genetics. Therefore, we engineered wild-type full length clones of the CLRV isolate E395 and several CP-plasmid constructs of distinct CLRV isolates as templates to generate chimeric RNA2 molecules swapping regions R1-R4 by site-directed PCR-mediated mutagenesis. This however, was not conducted within the project time as the basic tool for a reverse-genetics approach, an infectious CLRV wild-type full length clone, could not have been selected from the set of engineered RNA1 and RNA2 clones so far. Moreover, vector studies with Kleidocerys resedae, Euceraphis betulae/punctipennis and Aphis sambuci were not successfully establishment in the given time due to a range of technical difficulties related to the complex interrelations of viruses infecting woody trees, the woody hosts themselves and their natural arthropod, which had to be eluded. Major problem was the provision of experimental plants (birch and black elderberry) reliably free of any virus infection as well as CLRV donor trees with virus titers and even distribution within the plantlet. Consequently, we introduced and established the tissue culture technique in our lab to consistently provide young virus-free tree plantlets with low concentrations of inhibitory substances for any experimental treatment. As vector studies with woody hosts and most of their inhabiting specialist arthropod species are complex and timeconsuming, we further need investigation to reduce the spectrum of target species. Virus-vector relations are based on protein-protein interactions between e.g. the CP and a ligand mediating the retention of the virus within the vector. The virus-overlay assay allows detection of such protein binding interaction between virus and vector. By using this method to screen diverse arthropod species occurring on birch and black elderberry, we expect to reduce the spectrum of target species for focused vector studies in the future. We introduced this method and confirmed the basic procedure by successfully detecting the positive control, however detection level was very low demonstrating the need for further improvements of several parameters related to this assay to reliably screen arthropod and nematode species occurring on CLRV hosts.
Publications
- 2012: Molecular and epidemiological characterisation of Cherry leaf roll virus (CLRV). In: Büttner, C. und Ulrichs, C. (Hrsg.): Berliner ökophysiologische und phytomedizinische Schriften, Band 25, Der Andere Verlag, Uelvesbüll, 142 S.
Langer, J.
- 2012: Variabilität Protein-kodierender Genombereiche des Cherry leaf roll virus. 58. Deutsche Pflanzenschutztagung "Pflanzenschutz alternativlos", Technische Universität Braunschweig, 10.-14. September 2012, Julius-Kühn-Archiv, 438, 399
Langer, J., Gentkow, J., von Bargen, S., Büttner, C.
- 2013: chapter 3: Forest diseases caused by viruses. In: Infectious forest diseases. Gonthier P., Nicolotti G. (eds.), CABI, p. 50-75
Büttner, C., von Bargen, S., Bandte, M., Mühlbach, H.-P.
(See online at https://dx.doi.org/10.1079/9781780640402.0050) - 2013: Vector transmission of Cherry leaf roll virus? Candidate insect species infesting Betula spp. PPPHE, IUFRO 7.02.04 – Meeting, 29.05.2013, Berlin, "Virus and phytoplasma diseases of forest and urban trees"
Langer, J., Schuster, A.-K., Bandte, M., von Bargen, S., Büttner, C.
- 2014: Biology, detection and management of plant pathogens in irrigation water; APS Press
Eds. Hong, C., Moorman, G.W., Wohanka, W., Büttner, C.
- 2014: Etablierung von Gewebekulturen Cherry leaf roll virus-infizierter Betula spp. aus Nordfinnland. 13. Forstwissenschaftliche Tagung "Wälder der Zukunft: Lebensraum, Ressourcenschutz und Rohstoffversorgung", 17.-20. September, TU Dresden/Tharandt (Fachrichtung Forstwissenschaften)
Langer, J., von Bargen, S., Aronen, T., Jalkanen, R., Büttner, C.
- 2014: Etablierung von Gewebekulturen Cherry leaf roll virus-infizierter Betula spp. aus Nordfinnland. DPG- Arbeitskreis „Viruskrankheiten der Pflanze“, 31. März-1. April 2014; Geilweilerhof, Siebeldingen; Julius Kühn-Institut (JKI)
Langer, J., von Bargen, S., Aronen, T., Jalkanen, R., Büttner, C.
- 2014: Untersuchungen zur Vektorübertragbarkeit von Cherry leaf roll virus. 59. Deutsche Pflanzenschutztagung "Forschen – Wissen – Pflanzen schützen: Ernährung sichern!", 22.-26. September, Albert-Ludwigs-Universität Freiburg, Julius-Kühn-Archiv 447, 2014, S. 254
Langer, J., von Bargen, S., Büttner, C.
- 2015: chapter 13: Phytopathogenic viruses. In: Principles of plant microbe interactions: Role of microbes in sustainable agriculture. Lugtenberg (ed.) Springer, Heidelberg, 115-122
Büttner, C., von Bargen, S., Bandte, M.
(See online at https://doi.org/10.1007/978-3-319-08575-3_13) - 2015: Cherry leaf roll virus (CLRV) - a generalist among plant viruses infecting woody hosts. XVIII. International Plant Protection Congress (IPPC), "Mission possible: food for all through appropriate plant protection"; 24-27 August 2015, Berlin (Germany), Book of abstracts, p. 370
Langer J, von Bargen S, Büttner C,
- 2016: Der Nachweis von Pflanzenviren in absterbenden Birken im Stadtgebiet Berlin Steglitz-Zehlendorf. 71. ALVA-Jahrestagung, "Eiweißpflanzen - Strategien und Chancen für Landwirtschaft und Industrie"; 30./31. Mai 2016, Klagenfurt, Österreich; Tagungsbericht
Zinnert L, Gehlsen J, Langer J, Landgraf M, Rumbou A, Bandte M, von Bargen S, Büttner C, Schreiner M, Jäckel B
