Short Communication |
Corresponding author: Attilio Rizzoli ( attilio.rizzoli@agroscope.admin.ch ) Academic editor: Oliver Martin
© 2024 Alan Oggier, Christophe Debonneville, Marco Conedera, Olivier Schumpp, Attilio Rizzoli.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Oggier A, Debonneville C, Conedera M, Schumpp O, Rizzoli A (2024) First detection of ‘Candidatus Phytoplasma ulmi’ in Switzerland and in Orientus ishidae Matsumura, 1902. Alpine Entomology 8: 29-34. https://doi.org/10.3897/alpento.8.115588
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‘Candidatus Phytoplasma ulmi’ (Ca. P. ulmi) belongs to the ribosomal subgroup 16SrV-A and is associated with dieback, shoot proliferation and yellows disease on various Ulmus spp. Other plant species, such as Carpinus betulus and Prunus spp. have also been reported infected by the same pathogen. In 2021, in the frame of research activities focused on grapevine’s Flavescence dorée (FD), one specimen of Orientus ishidae - an East Palearctic leafhopper that was identified as an alternative vector of FD phytoplasmas - was found harboring Ca. P. ulmi in southern Switzerland. No phytoplasmas were detected in plant samples taken in the same location. Orientus ishidae has already been reported to be able to acquire diverse phytoplasmas associated with other plant diseases, such as Peach X-disease. This is the first report of Ca. P. ulmi in Switzerland, as well as in O. ishidae. Ca. P. ulmi may potentially be present in the wild compartment of the Swiss Pre-alpine and Alpine range, but no dedicated survey has so far been conducted. In the case of O. ishidae, this finding highlights the broad affinity of such a species for the acquisition of several phytoplasmas. This calls for a further investigation regarding its potential role as a vector on various pathosystems of agronomic importance.
alien species, leafhopper, Neobiota, phytoplasma, vector
Phytoplasmas are cell wall-less bacteria associated with the degradation of plant tissues and the manipulation of metabolic activity often leading to important agronomic losses (
Among the diseases associated with phytoplasmas, grapevine’s Flavescence dorée (FD) is one of the most insidious and destructive ones, causing important economic losses to European viticulture (
This manuscript reports the first finding of ‘Candidatus Phytoplasma ulmi’ (Ca. P. ulmi, ribosomal subgroup 16SrV-A) in Switzerland and in O. ishidae. Ca. P. ulmi is associated with dieback, shoot proliferation and yellows disease on various Ulmus spp. (
The study area comprised Canton Ticino (southern slope of the Swiss Alps). In the frame of current research activities regarding the FD epidemics in vineyards and their surroundings, experimental plots are monitored for the presence of FDp vectors and grapevines carrying symptoms linked to Grapevine Yellows are routinely marked and sampled for molecular analysis. Leafhopper populations in vineyards are surveyed with a minimum amount of six yellow sticky traps (YST; Rebell Giallo, Andermatt Biocontrol AG, Switzerland) hanged on the highest wire of the training system. In the surrounding landscape, at least four YST mounted on wooden sticks (ca. 1.50 m off the ground) are placed in the direct proximity of spontaneous plant species, such as A. glutinosa, C. avellana, Acer spp., etc., which are known to host alternative FDp vectors (e.g., O. ishidae). In 2021, 16 new plots were added to the standard design to monitor the possible presence of FDp vectors in the surroundings of gone-wild grapevines and/or rootstock resprouts originating from incorrect or incomplete rogueing of former vineyards located in the direct proximity of currently cultivated vineyards (
Leafhoppers determination was conducted using a stereo microscope (Olympus SZX16 with SDF PLAPO 1XPF objective lenses, made in Japan). For the particular case of O. ishidae, the morphological key provided by
Each insect was individually homogenized in 900 μL of extraction buffer (3% Cetyltrimethylammonium bromide CTAB, 1.4 M NaCl, 25 mM EDTA, 1 M Tris-HCl, 2 μL β-Mercaptoethanol, pH 8.0) and shaken for 30 min at 600 rpm and 65 °C. 900 μL of Chloroform/Isoamylalcohol was added, homogenized by vortexing for 5 s and centrifuged for 5 min at 3000×g. The aqueous layer was carefully transferred to a new tube, mixed with an equal volume of cold Isopropanol, and incubated 60 min at −20 °C for DNA precipitation. Precipitated material was recovered by 2 min of centrifugation at 10000× g and washed with 1 mL of 70% Ethanol. A DNA pellet was dried overnight at room temperature and resuspended into 100 μL of PCR-grade water. For plant samples, 0.5 to 1 g of petioles and midribs from 3 to 4 different leaves per specimen were ground in 6 mL of extraction buffer using a Homex grinder (Bioreba). Subsequently, 2 mL of this homogenate was centrifuged for 10 min at 1000×g. 900 μL of the supernatant was processed as described above.
The presence of 16SrV group phytoplasma in the samples was assessed by quantitative PCR analysis according to
PCR products were controlled by electrophoresis on a 1% agarose gel and purified by ultrafiltration with NucleoFast 96 PCR plates (Macherey-Nagel). Products were sent to Fasteris (Plan-les-Ouates, Switzerland) for forward and reverse sequencing using Sanger technology. Trees were inferred by maximum likelihood method in MEGA using the General Time Reversible model and bootstrapping with 500 replicates. All trees were visualized with iTOL (https://itol.embl.de, accessed on 14 September 2023;
In the frame of the research activities associated with FD, we analyzed 16 O. ishidae specimens caught in 2021 (out of 26 YST) and 267 caught in 2022 (out of 85 YST) for a total of 283 insects. One of the three specimens of O. ishidae caught in 2021 in the additional plot of Cugnasco (WGS84 coordinates 46.18037, 8.91938) was found harboring Ca. P. ulmi. The infected insect was captured between 1 and 14 October 2021. All specimens caught the following year in the same plot resulted free of Ca. P. ulmi (N = 6, NYST = 1). No other tested insect species, such as S. titanus was found infected by Ca. P. ulmi (data not shown). The four grapevines sampled in the plot of Cugnasco in 2021 resulted free of external symptoms linked to Grapevine Yellows and phytoplasma.
The sequence of the secY-map gene obtained from the infected O. ishidae is 100.00% identical to the reference strain AM384900 Ca. P. ulmi, isolate E04-D438 found in France, Loire Atlantique, previously reported by
Phytoplasma isolate from Orientus ishidae obtained in this work and reference strains along with the relative GenBank accession number, compared genes, host, geographic origin and reference.
Sample ID or reference strain | GenBank accession no. | Gene(s) | Host | Phytoplasma strain and genotype | Geographic origin | Reference |
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Ca. P. ulmi isolate Cugnasco CH | OR594266 | map imp | Orientus ishidae | Ca. P. ulmi | Switzerland, Canton Ticino | This work |
Ca. P. ulmi isolate E04-D438 | AM384900 | secY map | Ulmus minor | Ca. P. ulmi | France, Loire Atlantique |
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Ca. P. ulmi isolate EY18_SRB | HM038478 | secY map | Ulmus laevis | Ca. P. ulmi | Serbia |
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Ca. P. ulmi isolate NK16 | KU202150 | secY map | Ulmus laevis | Ca. P. ulmi | Croatia |
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Ca. P. ulmi isolate E04-D714 | AM384901 | secY map | Ulmus glabra | Ca. P. ulmi | France, Haute-Vienne |
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Ca. P. ulmi isolate EY1 | GU004330 | map cds | Plant leaf material | Ca. P. ulmi | USA |
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map M50 isolate VF-06-27-5 | LT221945 | map | Vitis vinifera | Ca. P. vitis FD1 M50 | France, Acquitaine |
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map M38 isolate AI-031-08 | LT221933 | map | Alnus glutinosa | Ca. P. vitis FD2 M38 | Italy, Veneto |
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map M51 isolate VS-Loza37 | LT221946 | map | Vitis vinifera | Ca. P. vitis FD3 M51 | Serbia |
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map M54 isolate VF06-30-18 | LT221949 | map | Vitis vinifera | Ca. P. vitis FD2 M54 | France, Acquitaine |
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Ca. P. ziziphi isolate Jwb-nky | CP025121 | map | Ziziphus jujuba Mill | Ca. P. ziziphi | China |
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4319_Ug_SN | MT668492 | imp | Ulmus glabra | Ca. P. ulmi | Germany |
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5554_Ug_MV | MT668497 | imp | Ulmus glabra | Ca. P. ulmi | Germany |
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0865_Ug_BYs | MT668435 | imp | Ulmus glabra | Ca. P. ulmi | Germany |
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2261_Ug_HE | MT668459 | imp | Ulmus glabra | Ca. P. ulmi | Germany |
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2226_Ug_HE | MT668458 | imp | Ulmus glabra | Ca. P. ulmi | Germany |
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4167_Ul_SN | MT668491 | imp | Ulmus sp. | Ca. P. ulmi | Germany |
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ULW | MT418908 | imp | Ulmus minor | Ca. P. ulmi | Germany |
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2732_Um_BB | MT668465 | imp | Ulmus sp. | Ca. P. ulmi | Germany |
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FD70 | MT668500 | imp | Vicia faba | Ca. P. vitis FD70 | France | unpublished |
ALY1 | MT668499 | imp | Alder sp. | Alder yellows phytoplasma | Germany |
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FD-D | MK614707 | imp | Vitis vinifera | Ca. P. vitis FD-D | Italy |
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Ca. P. ziziphi isolate Jwb-nky | MG818479 | imp | Chinese jujube | Ca. P. ziziphi | China | unpublished |
Phylogenetic tree of the secY-map (A) and imp (B) genes sequences from Orientus ishidae obtained in this work and reference strains from Genbank (see Table
This communication reports the first account of Ca. P. ulmi in Switzerland, as well as the first record of O. ishidae harboring such phytoplasmas, in general. Ca. P. ulmi was identified during routine molecular analyses conducted in the frame of an ongoing research project focused on FDp in vineyards and in the adjacent wild compartment in the Swiss southern Alps.
‘Candidatus Phytoplasma ulmi’ may already be present in Swiss Ulmus spp., as well as in other host plant species both on the Alpine range and on the Swiss Plateau. However, no proper surveys have so far been conducted in order to confirm its presence and potential impact on mountainous and Alpine ecosystems. Most interestingly, O. ishidae appears to be able to acquire other phytoplasma strains in addition to genotypes belonging to the ribosomal subgroups 16SrV-C and 16SrV-D (
Alan Oggier: Conceptualization, investigation, formal analysis, data curation, writing - review and editing. Christophe Debonneville: Methodology, investigation, formal analysis, visualization, software, validation, writing - review and editing. Marco Conedera: Writing – review and editing, funding acquisition, project administration. Olivier Schumpp: Writing – review and editing, resources, validation. Attilio Rizzoli: Conceptualization, methodology, writing – original draft, writing – review and editing, validation, supervision.
This study was partially supported by the Swiss Federal Office for Agriculture FOAG (grant number 627001075).