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2. Wild and globally traded ornamental aquatic plants harbor diverse plant viruses, including notable crop pathogensLana Vogrinec, Katarina Bačnik, Martina Bačič, Nataša Mehle, Aimee R. Fowkes, Živa Lengar, Valerie Harju, Ian Adams, Adrian Fox, Denis Kutnjak, 2025, original scientific article Abstract: Background Aquatic plants play key roles in ecosystems, serving as primary producers and providing habitat for other aquatic life. While many are ecologically important, some invasive species, often introduced through the ornamental plant trade, pose threats to various aquatic ecosystems. Although viral infections have been documented to some extent in aquatic crops, the viral diversity in wild and ornamental aquatic plants remains largely unexplored. Investigating the viral communities of aquatic plants is important, as their direct contact with water allows for the potential long-distance transmission of stable viruses released from infected individuals. Invasive aquatic plants exacerbate this issue by introducing novel microbes, including viruses, to new regions, increasing the potential threat to native plant populations. Results Here, we investigated the viral communities of diverse aquatic plants by mining publicly available transcriptome data of 79 wild aquatic species and sequencing the RNA from 14 plant species (some of them of different ornamental varieties), sourced from hobby aquascaping stores. Plant viruses from various families were detected in taxonomically diverse aquatic plants, ranging from algae to angiosperms. Alongside sequences of known crop pathogens, such as turnip yellows virus, cucumber mosaic virus, and lettuce chlorosis virus, we identified contigs of putative novel viral species belonging to several plant-infecting viral families. Most notably, we discovered sequences of known and novel begomoviruses, which may be causing observed ornamental phenotypes in two different aquatic plants. Further, we identified a novel potyvirus that appears to be globally present in multiple ornamental plants from the genus Sagittaria. We detected it in three plants sourced from online stores in Slovenia, as well as six plants intercepted during the import process into the UK. Conclusions Our findings expand on the so far limited knowledge of aquatic plant viruses, revealing known and putative novel plant viral species across diverse aquatic plant taxa. The detection of crop viruses, including regulated pathogens, in ornamental aquatic plants highlights the risks associated with their unregulated global trade. Further research into viruses of aquatic plants may provide insights into their role in ecosystems as well as their potential impact on agriculture.
Keywords: aquatic plants, macrophytes, viromes, high-throughput sequencing, data mining, virology
Published in DiRROS: 26.11.2025; Views: 73; Downloads: 33
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3. From vineyard to genome : optimized enrichment and sequencing of Flavescence dorée phytoplasma from grapevine samplesZala Kogej Zwitter, Denis Kutnjak, Nataša Mehle, 2025, original scientific article Abstract: Phytoplasmas are non-culturable obligate intracellular bacteria that cause considerable economic losses in agriculture. Genome sequencing provides crucial insights into their biology and vector dependence. However, genome studies on phytoplasmas are often hampered by their low abundance in naturally infected plants. Propagation in test plants is usually necessary but time-consuming and resource-intensive, especially for quarantine phytoplasmas such as the phytoplasma causing Flavescence dorée (FD), a serious threat to European viticulture. To overcome these challenges, we aimed to develop a protocol for efficient enrichment of phytoplasma DNA directly from field-collected samples, enabling genome sequencing using both Illumina and Oxford Nanopore Technologies platforms. We evaluated six sample preparation protocols that included stepwise enrichment steps to improve phytoplasma genome coverage and assembly quality. The most effective approach combined differential centrifugation, CTAB extraction and removal of CpG-methylated host DNA and resulted in a notable increase in the relative abundance of phytoplasma reads compared to other protocols. Rarefaction analysis of the dataset generated using this protocol demonstrated that the entire phytoplasma genome was covered by reads in a dataset comprising 3 billion nucleotides. We also evaluated and compared de novo phytoplasma genome assemblies generated from short Illumina reads and long nanopore sequencing reads. While Illumina sequencing yielded more accurate assemblies with longer total lengths, the assemblies derived from nanopore sequencing data contained longer individual contigs. This advantage was reflected in hybrid assemblies that combined both technologies, yielding longer phytoplasma contigs than assemblies from Illumina datasets and lower mismatch rates compared to assemblies from nanopore sequencing datasets. A hybrid de novo assembled genome of the Slovenian FD phytoplasma isolate achieved 96% reference genome coverage, with high contiguity and low error rates. This streamlined and accessible protocol enables high-quality genome sequencing of phytoplasma-infected grapevines without the need for propagation in test plants. This facilitates broader phytoplasma research and can potentially be extended to other naturally infected phytoplasma hosts or organisms infected with other non-culturable microbes.
Keywords: plant pathogen, nanopore, grapevine, enrichment, HTS, Flavescence dorée, phytoplasma
Published in DiRROS: 08.10.2025; Views: 192; Downloads: 91
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4. Validation of molecular diagnostic methods for the detection and identification of tomato mottle mosaic virus (ToMMV-detect) : final reportNataša Mehle, Ana Vučurović, Sabine Grausgruber Groeger, Amandine Lê Van, Mylène Ruh, Heiko Ziebell, 2025, final research report Keywords: validation, plant viruses, tobamovirus, ToMMV
Published in DiRROS: 03.10.2025; Views: 260; Downloads: 111
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6. Vertical transmission of tomato virusesNataša Mehle, Adrian Fox, Antonio Tiberini, Heiko Ziebell, 2025, independent scientific component part or a chapter in a monograph Abstract: The vertical transmission of tomato viruses through seeds and pollen is a significant yet often overlooked pathway for the persistence and global spread of these pathogens. This review provides a comprehensive synthesis of current knowledge on the mechanisms, epidemiological implications, and management strategies of vertically transmitted tomato viruses. While recent advances in diagnostic techniques such as high-throughput sequencing (HTS), have improved virus detection, key research gaps remain in understanding the molecular and ecological dynamics of seed and pollen transmission. The interaction between vertical and horizontal transmission modes complicates virus epidemiology, necessitating an integrated management approach that includes rigorous seed health testing, genetic resistance breeding, and biosecurity measures. Emerging threats, such as resistance-breaking virus strains and the impact of climate change on vector distribution, underscore the need for enhanced surveillance and stronger international regulatory cooperation. This review highlights the need for interdisciplinary research and collaboration to develop sustainable virus mitigation strategies. Future research priorities include optimizing detection methods, exploring next-generation breeding technologies, and strengthening international biosecurity frameworks to safeguard global tomato production against the growing threat of vertically transmitted viruses.
Keywords: Solanum lycopersicum, tomato, vertical virus transmission, high economic impact, seed-transmitted viruses, plant diseases, virology, biology
Published in DiRROS: 26.09.2025; Views: 416; Downloads: 185
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7. Tomato spotted wilt virus in tomato from Croatia, Montenegro and Slovenia: genetic diversity and evolutionDijana Škorić, Jelena Zindović, Dorotea Grbin, Patrik Pul, Vladan Božović, Paolo Margaria, Nataša Mehle, Anja Pecman, Zala Kogej Zwitter, Denis Kutnjak, Ana Vučurović, 2025, original scientific article Abstract: Tomato spotted wilt orthotospovirus (TSWV) is a major plant pathogen causing significant economic losses in tomato production worldwide. Understanding its genetic diversity and evolutionary mechanisms is crucial for effective disease management. This study analyzed TSWV isolates from symptomatic tomato plants collected across Croatia, Montenegro and Slovenia between 2020 and 2024. High-throughput sequencing (HTS) was employed to obtain whole-genome sequences, followed by phylogenetic analyses to assess genetic variability and relationships among isolates from these three countries and other isolates of worldwide geographic origin. Phylogenetic analyses placed all studied isolates within the L1-M3-S3 genotype, commonly associated with solanaceous crops in Europe. While Croatian and Slovenian isolates exhibited high genetic similarity, Montenegrin isolates clustered in a distinct subgroup, showing closer relationships to Asian and Mediterranean accessions. Despite the severe disease symptoms observed, no substitutions in the NSm protein associated with resistance-breaking (RB) phenotypes were detected. These findings suggest that additional virome components, environmental factors or so far unknown mechanism(s) may contribute to infection and disease severity in tomato and strongly support the need of continuous surveillance of TSWV genetic diversity in order to inform breeding programs and develop sustainable management strategies to mitigate future outbreaks.
Keywords: tomato, HTS, phylogeny, TSWV, plant virus
Published in DiRROS: 16.09.2025; Views: 289; Downloads: 134
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9. Digital PCR genotyping of pepino mosaic virusAmadej Jelenčič, David Dobnik, Alexandra Bogožalec Košir, Nataša Mehle, 2025, independent scientific component part or a chapter in a monograph Abstract: Pepino mosaic virus (PepMV) is a plant pathogen causing significant economic losses in tomato production. Sensitive, reliable, and robust detection methods are crucial for containing the spread of PepMV and reducing its damaging effects. Digital PCR (dPCR) presents several advantages to conventional real-time quantitative PCR (qPCR), including absolute quantification ability, robust quantitative multiplexing capabilities, and straightforward result analysis. Furthermore, dPCR is especially suitable for analysis of complex samples due to its remarkable tolerance to PCR inhibitors, which makes it a promising method for plant virus genotyping. In this chapter, we present two protocols for PepMV genotyping and quantification using one-step reverse transcription digital PCR (RT-dPCR). The first protocol outlines four simplex assays, while the second describes two duplex assays for precise and comprehensive genotyping of PepMV variants.
Keywords: plant viruses, detection, quantification, Pepino mosaic virus, PepMV, digital PCR, dPCR, RT-dPCR, duplex
Published in DiRROS: 08.09.2025; Views: 316; Downloads: 39
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10. Annual (2024) taxonomic update of RNA-directed RNA polymerase-encoding negative-sense RNA viruses (realm Riboviria: kingdom Orthornavirae: phylum Negarnaviricota)Jens H. Kuhn, Scott Adkins, Sergey V. Alkhovsky, Wenxia An, Tatjana Avšič-Županc, María A. Ayllón, Katarina Bačnik, Justin Bahl, Anne Balkema-Buschmann, Matthew J. Ballinger, Ion Gutiérrez-Aguirre, Denis Kutnjak, Nataša Mehle, Anja Pecman, Maja Ravnikar, Ana Vučurović, 2025, original scientific article Abstract: In April 2024, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was expanded by 1 new order, 1 new family, 6 new subfamilies, 34 new genera and 270 new species. One class, two orders and six species were renamed. Seven families and 12 genera were moved; ten species were renamed and moved; and nine species were abolished. This article presents the updated taxonomy of Negarnaviricota as currently accepted by the ICTV, providing an essential annual update on the classification of members of this phylum that deepen understandings of their evolution, and supports critical public health measures for virus identification and tracking.
Keywords: Aliusviridae, Arenaviridae, articulaviral, Articulavirales, Artoviridae, Aspiviridae, Bornaviridae, bunyaviral, Bunyavirales, Bunyaviricetes, bunyavirus, Crepuscuviridae, Discoviridae, Elliovirales, Filoviridae, Fimoviridae, Goujianvirales, Hantaviridae, Hareavirales, International Committee on Taxonomy of Viruses (ICTV), jingchuviral, Jingchuvirales, Konkoviridae, Lispiviridae, megaclassification, megataxonomy, mononegaviral, Mononegavirales, muviral, Muvirales, Mymonaviridae, Myriaviridae, naedreviral, Naedrevirales, Nairoviridae, Natareviridae, negarnaviricot, Negarnaviricota, Nyamiviridae, Orthomyxoviridae, Orthornavirae, Paramyxoviridae, Peribunyaviridae, Phasmaviridae, Phenuiviridae, Pneumoviridae, Rhabdoviridae, Riboviria, Sunviridae, Tenuivirus, Tosoviridae, Tospoviridae, Tulasviridae, virus classification, virus nomenclature, virus taxonomy, virology
Published in DiRROS: 24.06.2025; Views: 481; Downloads: 361
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