Detection of viruses of the virome affecting birches (Betula sp.) - "Case studies Europe-wide"

Opoku, Elisha Bright

• Publisher: Cuvillier Verlag
• Year: 2023
• Print ISBN: 9783736977532
• eBook ISBN: 9783736967533
• Edition: 1

Over the years, birches keep declining at various locations in Europe especially in cities, parks streets as well as forest due to biotic and abiotic stress factors.The decline of birches is characterized by die-back of twigs, branches, and can lead to theloss of tree crown, and finally to death of the tree. Virus-suspected leaf symptoms wereobserved on declining birches, which were attributed to Birch leaf rolldisease (BLRD). Investigations from this thesis give an impressionthat, the birches can be infected by more virus species than ever assumed. The birch has a complex virome than expected and the decline cannot be attributed notjust to a particular virus. Modern techniques, such as high-throughput sequencing (HTS), gave a comprehensive identification of possible viral pathogens present in the birch genome.In 2014, sequences of a previously unknown viruses could be discovered in symptomatic leafsamples of the genus Betula. Investigations of the prevalent birchviruses (BLRaV, CLRV, BiCV and ApMV) led to discovery of new viral signatures (e.g.Birch capillovirus, Birch caulimovirus, Birch benyvirus and Birch idaeovirus) in leaf samplesfrom different areas in Europe. This indicates a wide distribution of these viruses as discussedin this thesis. The sequences obtained from complex virome RNA-Seq were about 70 %similar to known plant viruses. With the aid of the high-throughput sequences, RT-PCRbaseddetection systems for newly discovered viruses have been developed. Furthermore, thegenome of these new viruses was to be further characterized and first results on thedissemination of them were presented.

Table of contents

• 1 Introduction
• 2 Materials and methods
• 2.1 Survey of selected Betula stands -Berlin- Germany
• 2.2 Betula sp. from Wildberg/Nagold seed plantation-Germany
• 2.3 Selected Betula stands -Rovaniemi-Finland
• 2.4 RNA isolation from plant materials (Boom et al., 1990)
• 2.5 Reverse transcription (cDNA synthesis)
• 2.6 Oligonucleotides (Primers)
• 2.7 Polymerase chain reaction (PCR)
• 2.8 Agarose gel electrophoresis
• 2.9 Sanger Sequencing of PCR products
• 2.10 Double Antibody Sandwich - Enzyme-linked immunosorbent assay (DAS-ELISA)
• 2.11 Bioassays
• 2.12 High-throughput sequencing (HTS) diagnostics
• 2.13 Bioinformatics including genomic database analysis - (NCBI)
• 3 Results
• 3.1 Symptomatology in birches of selected stands
• 3.2 Results of RNA extraction and RT-PCR
• 3.3 Detection of plant viruses infecting Betula sp. -Berlin
• 3.4 Detection of plant viruses infecting Betula sp -Wildberg/Nagold
• 3.5 Detection of plant viruses infecting Betula sp -Rovaniemi -Finland
• 3.6 Serological results
• 3.7 Database analysis of the Betula genome-NCBI
• 3.8 Biodiversity of plant viruses in Betula sp. -Whole genome shotgun (WGS)
• 4. Discussion
• 4.1 Selected Betula stands -Berlin -Germany
• 4.2 Single and mixed infection of plant viruses detected by RT-PCR
• 4.3 Selected leaf symptoms from Betula stands -Wildberg/Nagold -Germany
• 4.4 Selected Betula stands -Rovaniemi -Finland
• 4.5 RT-PCR detection of plant viruses
• 4.6 Bioassay
• 4.7 Serological methods (DAS- ELISA)
• 4.8 Database analysis of the Betula sequence data -NCBI
• 5 Badnaviruses state of the art
• 5.1 Symptomatology, Host Range and Mode of transmission
• 5.2 Geographical distribution of Badnaviruses
• 5.3 Genomic organization of badnaviruses
• 5.4 Current diagnostic methods for Badnaviruses detection
• 6 Summary
• 7 Zusammenfassung
• 8 References
• 9 List of figures
• 10 List of tables
• 11 Appendix
• 11.1 List of Abbreviation
• 11.2 Chemicals
• 11.3 Tables
• 11.4 Figures
• 12 Acknowledgement
• 13 Declaration on independence and resources