1. Global advances in tomato virome research : current status and the impact of high-throughput sequencingMark Paul Selda Rivarez, Ana Vučurović, Nataša Mehle, Maja Ravnikar, Denis Kutnjak, 2021, pregledni znanstveni članek Povzetek: Viruses cause a big fraction of economically important diseases in major crops, including tomato. In the past decade (2011–2020), many emerging or re-emerging tomato-infecting viruses were reported worldwide. In this period, 45 novel viral species were identified in tomato, 14 of which were discovered using high-throughput sequencing (HTS). In this review, we first discuss the role of HTS in these discoveries and its general impact on tomato virome research. We observed that the rate of tomato virus discovery is accelerating in the past few years due to the use of HTS. However, the extent of the post-discovery characterization of viruses is lagging behind and is greater for economically devastating viruses, such as the recently emerged tomato brown rugose fruit virus. Moreover, many known viruses still cause significant economic damages to tomato production. The review of databases and literature revealed at least 312 virus, satellite virus, or viroid species (in 22 families and 39 genera) associated with tomato, which is likely the highest number recorded for any plant. Among those, here, we summarize the current knowledge on the biology, global distribution, and epidemiology of the most important species. Increasing knowledge on tomato virome and employment of HTS to also study viromes of surrounding wild plants and environmental samples are bringing new insights into the understanding of epidemiology and ecology of tomato-infecting viruses and can, in the future, facilitate virus disease forecasting and prevention of virus disease outbreaks in tomato.
Objavljeno v DiRROS: 05.08.2024; Ogledov: 52; Prenosov: 62
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2. Diversity and flight activity of aphid species as potential vectors of oilseed pumpkin viruses in SerbiaAna Vučurović, Olivera Petrović-Obradović, Anđa Radonjić, Dušan Nikolić, Katarina Zečević, Ivana Stanković, Branka Krstić, 2018, izvirni znanstveni članek Povzetek: The fauna of aphids appearing on cucurbits, including oilseed pumpkin have been poorly investigated in Serbia. Yellow water traps were used to determine the diversity of aphid species visiting cucurbits in Serbia and to monitor their flight activity. During the years 2009 to 2011, a total of 1,447 specimens were collected and a total of 57 different aphid taxa were identified. In 2009, the highest total number of aphids were caught (755), followed by 203 and 489 aphids in 2010 and 2011, respectively. The most abundant species were Aphis fabae (15%), Myizus persicae (13.4%) and Acyrthosiphon pisum (11.7%), followed by Anoecia corni (6.4%)and Aphis gossypii(5.6%). All of them, except A. corni,are reported as efficient vectors of cucurbit viruses. These five species represent 52.1% of all aphid species collected within this study. Out of 57 taxa found in oilseed pumpkin crops, 17 are known as vectors of ZYMV, 18 of WMV and 15 of CMV. Generally, 25 out of a total of 57 taxa are known as vectors of at least one of three viruses present in Serbia. From total of 1,447 specimens collected during three years of investigation, 917 (63.4%) are vectors of at least one of three viruses. Individual oilseed pumpkin plants were frequently colonized by A. gossypiiduring all three years of investigation. Results of this study showed that Serbian oilseed pumpkin growing localities are grouped among medium to reach in aphidofauna, according to total Shannon-Weaver index values which varied from 1.8 to 2.8. The highest value of Shannon-Weaver (2.8) was recorded in the Bački Petrovac I locality in 2009, while the lowest Shannon-Weaver (1.8) was recorded in Kisač locality in 2010. The maximum values of Shanon-Weaver diversity index in almost all crops were recorded in the second week of investigation or in early stages of oilseed pumpkin growth when plants had 5-6 true leaves or were in flowering stage, when potential virus infection could lead to significant yield losses.
Objavljeno v DiRROS: 24.07.2024; Ogledov: 104; Prenosov: 63
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3. Detection of four new tomato viruses in Serbia using post hoc high-throughput sequencing analysis of samples from a large-scale field surveyAna Vučurović, Denis Kutnjak, Nataša Mehle, Ivana Stanković, Anja Pecman, Aleksandra Bulajić, Branka Krstić, Maja Ravnikar, 2021, izvirni znanstveni članek Povzetek: Tomato production worldwide is affected by numerous plant virus species. The early and accurate detection of viruses is a critical step for disease control. However, the simultaneous detection of the most known tomato viruses can be difficult because of the high number and diversity of tomato-infecting viruses. Here, we have identified four new viruses in Serbia by applying target-independent small RNA high-throughput sequencing (HTS). HTS was applied on pools of samples and separate samples, in total comprising 30 tomato samples that exhibited (severe) virus-like symptoms and were collected in Serbia during three annual surveys (2011 to 2013). These samples had previously tested negative for the presence of 16 tomato viruses using targeted detection methods. Three divergent complete genome sequences of Physostegia chlorotic mottled virus were obtained from different localities, indicating for the first time that this virus is widespread in Serbia and might represent an emergent viral pathogen of tomato. The tomato torrado virus was detected at one locality with devastating yield losses. The southern tomato virus was detected at two localities, and the spinach latent virus was detected at one locality. In addition, we detected the presence of one already-known virus in Serbia, the tomato spotted wilt orthotospovirus. All the HTS results were subsequently confirmed by targeted detection methods. In this study, the successful application of post hoc HTS testing of a limited number of pooled samples resulted in the discovery of new viruses. Thus, our results encourage the use of HTS in research and diagnostic laboratories, including laboratories that have limited resources to resolve disease etiology.
Ključne besede: viruses, tomato, detection, identification
Objavljeno v DiRROS: 19.07.2024; Ogledov: 116; Prenosov: 151
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4. What did we achieve with VALITEST an EU project on validation in plant pest diagnostics?Charlotte Trontin, Barbara Agstner, Denise Altenbach, Géraldine Anthoine, Hanna Baginska, Ian Brittain, A Chabirand, Anne-Marie Chappé, P. Dahlin, Tanja Dreo, C. Freye-Minks, Camilo Gianinazzi, Catherine Harrison, Glyn Jones, Marco Stefan Kaiser, Marta Luigi, Sébastien Massart, Nataša Mehle, Monica Mezzalama, Hanna Mouaziz, Maja Ravnikar, Tom Raaymakers, Jean-Philippe Renvoise, Mathieu Rolland, Marta Santos, Sam Seddas, René van der Vlugt, Ana Vučurović, Françoise Petter, 2022, izvirni znanstveni članek Ključne besede: plant pest diagnostics, validation, test performance study, high-throughput, sequencing, reference material, training
Objavljeno v DiRROS: 17.07.2024; Ogledov: 103; Prenosov: 50
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5. Biological and genetic characterization of Physostegia chlorotic mottle virus in Europe based on host range, location, and timeColine Temple, Arnaud G. Blouin, Kris De Jonghe, Yoika Foucart, Marleen Botermans, Marcel Westenberg, Ruben Schoen, Pascal Gentit, Michèle Visage, Eric Verdin, Mark Paul Selda Rivarez, Denis Kutnjak, Ana Vučurović, 2022, izvirni znanstveni članek Povzetek: Application of high throughput sequencing (HTS) technologies enabled the first identification of Physostegia chlorotic mottle virus (PhCMoV) in 2018 in Austria. Subsequently, PhCMoV was detected in Germany and Serbia on tomatoes showing severe fruit mottling and ripening anomalies. We report here how prepublication data-sharing resulted in an international collaboration across eight laboratories in five countries, enabling an in-depth characterization of PhCMoV. The independent studies converged toward its recent identification in eight additional European countries and confirmed its presence in samples collected 20 years ago (2002). The natural plant host range was expanded from two to nine species across seven families, and we confirmed the association of PhCMoV presence with severe fruit symptoms on economically important crops such as tomato, eggplant, and cucumber. Mechanical inoculations of selected isolates in the greenhouse established the causality of the symptoms on a new indexing host range. In addition, phylogenetic analysis showed a low genomic variation across the 29 near-complete genome sequences available. Furthermore, a strong selection pressure within a specific ecosystem was suggested by nearly identical sequences recovered from different host plants through time. Overall, this study describes the European distribution of PhCMoV on multiple plant hosts, including economically important crops on which the virus can cause severe fruit symptoms. This work demonstrates how to efficiently improve knowledge on an emergent pathogen by sharing HTS data and provides a solid knowledge foundation for further studies on plant rhabdoviruses.
Objavljeno v DiRROS: 17.07.2024; Ogledov: 209; Prenosov: 76
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6. Development and validation of a one-step reverse transcription real-time PCR assay for simultaneous detection and identification of tomato mottle mosaic virus and tomato brown rugose fruit virusAntonio Tiberini, Ariana Manglli, Anna Taglienti, Ana Vučurović, Jakob Brodarič, Luca Ferretti, Marta Luigi, Andrea Gentili, Nataša Mehle, 2022, izvirni znanstveni članek Povzetek: Tobamovirus species represent a threat to solanaceous crops worldwide, due to their extreme stability and because they are seed borne. In particular, recent outbreaks of tomato brown rugose fruit virus in tomato and pepper crops led to the establishment of prompt control measures, and the need for reliable diagnosis was urged. Another member of the genus, tomato mottle mosaic virus, has recently gained attention due to reports in different continents and its common features with tomato brown rugose fruit virus. In this study, a new real-time RT-PCR detection system was developed for tomato brown rugose fruit virus and tomato mottle mosaic virus on tomato leaves and seeds using TaqMan chemistry. This test was designed to detect tomato mottle mosaic virus by amplifying the movement protein gene in a duplex assay with the tomato brown rugose fruit virus target on the CP-3’NTR region, which was previously validated as a single assay. The performance of this test was evaluated, displaying analytical sensitivity 10−5–10−6-fold dilution for seeds and leaves, respectively, and good analytical specificity, repeatability, and reproducibility. Using the newly developed and validated test, tomato brown rugose fruit virus detection was 100% concordant with previously performed analyses on 106 official samples collected in 2021 from different continents.
Ključne besede: real-time PCR, tomato mottle mosaic virus, tomato brown rugose fruit virus, leaves detection, seeds detections, performance criteria
Objavljeno v DiRROS: 16.07.2024; Ogledov: 101; Prenosov: 126
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7. Tomato brown rugose fruit virus in aqueous environments : survival and significance of water-mediated transmissionNataša Mehle, Katarina Bačnik, Irena Bajde, Jakob Brodarič, Adrian Fox, Ion Gutiérrez-Aguirre, Miha Kitek, Denis Kutnjak, Yue Lin Loh, Olivera Maksimović, Maja Ravnikar, Elise Vogel, Christine Vos, Ana Vučurović, 2023, izvirni znanstveni članek Povzetek: Tomato brown rugose fruit virus (ToBRFV) has recently emerged as a major disease of tomatoes and peppers. ToBRFV is a seed- and contact-transmitted virus. In Slovenia, ToBRFV RNA was detected in samples of wastewater, river, and water used to irrigate plants. Even though the source of detected RNA could not be clearly established, this raised the question of the significance of the detection of ToBRFV in water samples and experimental studies were performed to address this question. The data presented here confirm that the release of virus particles from the roots of infected plants is a source of infectious ToBRFV particles in water and that the virus can remain infective up to four weeks in water stored at room temperature, while its RNA can be detected for much longer. These data also indicate that irrigation with ToBRFV-contaminated water can lead to plant infection. In addition, it has been shown that ToBRFV circulated in drain water in commercial tomato greenhouses from other European countries and that an outbreak of ToBRFV can be detected by regular monitoring of drain water. A simple method for concentrating ToBRFV from water samples and a comparison of the sensitivity of different methods, including the determination of the highest ToBRFV dilution still capable of infecting test plants, were also investigated. The results of our studies fill the knowledge gaps in the epidemiology and diagnosis of ToBRFV, by studying the role of water-mediated transmission, and provide a reliable risk assessment to identify critical points for monitoring and control.
Ključne besede: tomato brown rugose fruit virus, tomato, hydroponics, water-linked epidemiology, survival
Objavljeno v DiRROS: 12.07.2024; Ogledov: 117; Prenosov: 130
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8. First report of collar and root rot of lettuce caused by Plectosphaerella cucumerina in SerbiaMilica Mihajlović, Jovana Hrustić, Ana Vučurović, Mila Grahovac, Dragana Budakov, Tatjana Dudaš, Brankica Pešić, 2024, drugi znanstveni članki Povzetek: In March 2021, unusual plant stuning, collar, and wet root rot of lettuce (Lactuca sativa L.) during the rosette stage was observed in two commercial fields in Serbia (44°58'N, 20°32'E; 44°45'N, 20°43'E). Disease incidence in the fields (≈ 0.9 ha each) was approximately 15 and 20%, respectively. Initial above-ground symptoms were yellowing and wilting of leaves, while below-ground symptoms were collar, wet root rot, and lesions becoming necrotic. Eventually, whole plants wilted, collapsed, and died. A total of 35 symptomatic plants were collected from the fields, and diseased tissues were cut into small pieces, surface sterilized, and plated on potato dextrose agar (PDA). Isolation resulted in 20 morphologically uniform monoconidial isolates. The isolates formed white to creamy colonies, gradually becoming salmon pink, slimy, or moist in appearance, with sparse aerial mycelia. Numerous hyphal coils with conidiophores and hyaline, smooth-surfaced, ellipsoid to ovoid, septate or aseptate conidia were formed (4.5 to 10.1×1.2 to 3.7 μm (n = 100)). To confirm the species identity, the internal transcribed spacer (ITS) region and the D1/D2 region of a selected representative isolate 13-3-c were amplified and sequenced by using primer pairs ITS1/ITS4 (White et al. 1990) and N1/N2 (O’Donnell and Gray 1995), respectively. The sequences were deposited in GenBank (ITS: OR880564 and D1/D2: OR880567). Sequence analysis revealed 100% nucleotide identity with P. cucumerina isolates from different countries deposited in the NCBI GenBank, including isolate MH860704 (Vu et al. 2019) (ITS region) and isolate KY662256 (Su et al. 2017) (D1/D2 region). Neighbor-joining analysis was conducted based on the combined ITS and D1/D2 regions, and the tree was constructed with the substitution models (1,000 bootstrap). The combined phylogeny confirmed that the sequences shared a common clade with P. cucumerina. Hence, morphological, microscopic, and molecular characterization confirmed the pathogen as P. cucumerina (Palm et al., 1995; Carlucci et al., 2012). In a pathogenicity assay, 10 isolates were tested. Five 30-day-old lettuce plants (cv. Majska Kraljica) per isolate were root-dipped in the conidial suspensions (1×105 conidia/ml). The 10 inoculated plants were transplanted into 1 L pots containing sterile substrate (Floragard, Germany). Plants treated with sterile distilled water were used as controls. Plants were maintained in a greenhouse at 25 to 28°C under a 12-hour photoperiod (Cai et al., 2021). Four weeks after inoculation, stunting, chlorosis, and wilting of plants were observed, while collars and roots exhibited typical decaying symptoms. No symptoms were observed on the control plants. The pathogen was reisolated from symptomatic tissue as previously described. Koch's postulates were completed by confirming the identity of reisolates based on morphological features. To our knowledge, this is the first report of P. cucumerina on lettuce or any other crop in Serbia. P. cucumerina is already known as a pathogen of lettuce and other hosts grown in many countries worldwide, as well as in some European countries (Belgium, England, Italy, the Netherlands, and Switzerland) (Zhang et al. 2019). This emerging pathogen may cause significant economic losses in lettuce production in Serbia and in the entire Balkan region. Our results may help to develop effective management strategies based on accurate and timely identification and regular pathogen monitoring.
Ključne besede: detection, identification, salat, plant diseases
Objavljeno v DiRROS: 17.05.2024; Ogledov: 236; Prenosov: 75
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9. Virome analysis of irrigation water sources provides extensive insights into the diversity and distribution of plant viruses in agroecosystemsOlivera Maksimović, Katarina Bačnik, Mark Paul Selda Rivarez, Ana Vučurović, Nataša Mehle, Maja Ravnikar, Ion Gutiérrez-Aguirre, Denis Kutnjak, 2024, izvirni znanstveni članek Povzetek: Plant viruses pose a significant threat to agriculture. Several are stable outside their hosts, can enter water bodies and remain infective for prolonged periods of time. Even though the quality of irrigation water is of increasing importance in the context of plant health, the presence of plant viruses in irrigation waters is understudied. In this study, we conducted a large-scale high-throughput sequencing (HTS)-based virome analysis of irrigation and surface water sources to obtain complete information about the abundance and diversity of plant viruses in such waters. We detected nucleic acids of plant viruses from 20 families, discovered several novel plant viruses from economically important taxa, like Tobamovirus and observed the influence of the water source on the present virome. By comparing viromes of water and surrounding plants, we observed presence of plant viruses in both compartments, especially in cases of large-scale outbreaks, such as that of tomato mosaic virus. Moreover, we demonstrated that water virome data can extensively inform us about the distribution and diversity of plant viruses for which only limited information is available from plants. Overall, the results of the study provided extensive insights into the virome of irrigation waters from the perspective of plant health. It also suggested that an HTS-based water virome surveillance system could be used to detect potential plant disease outbreaks and to survey the distribution and diversity of plant viruses in the ecosystem.
Ključne besede: plant viruses, environmental water testing, high-throughput sequencing, agroecosystems, irrigation water, virome
Objavljeno v DiRROS: 29.03.2024; Ogledov: 367; Prenosov: 167
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10. Increased diversity of citrus tristeza virus in EuropeJelena Zindović, Miroslav Čizmović, Ana Vučurović, Paolo Margaria, Dijana Škorić, 2023, izvirni znanstveni članek Povzetek: This study investigated the genetic diversity of citrus tristeza virus (CTV) isolates from Montenegro and Croatia, European countries with the northernmost citrus growing regions situated on the Eastern Adriatic coast. Fifteen complete or nearly complete CTV genomes were reconstructed from high-throughput sequencing of samples collected in distinct municipalities in Montenegro and Opuzen municipality in Croatia. Phylogenetic analyses assigned some of the sequences to VT and T30 strains, previously recorded in Europe, while remarkably other isolates were placed in S1 and RB groups, which have not been reported in Europe so far. In addition, a new phylogenetic lineage including only isolates from Montenegro was delineated and tentatively proposed as the MNE cluster. Recombination analysis revealed evidence of 11 recombination events in the sequences obtained in this study, between isolates of related strains, within isolates of the same strain, and between distant strains. These findings show that CTV diversity in Europe is higher than reported before and calls for the re-evaluation of management strategies.
Ključne besede: complete genomes, genotyping, citrus tristeza virus, CTV, non-EU strain
Objavljeno v DiRROS: 29.03.2024; Ogledov: 407; Prenosov: 165
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