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Resume La ciguatera est une intoxication frequente en zone tropicale. Depuis 1993, le Centre Antipoison de Marseille travaille sur le sujet de la Toxinologie marine et des intoxications par produits de la mer. Les auteurs rapportent l’experience de ce service sur la ciguatera de 1993 a 2017. Quarante-neuf intoxications ont ete etudiees concernant 175 patients (25 cas avec un seul patient et 24 cas collectifs avec 2 a 30 patients). Cent douze patients etaient des touristes de metropole intoxiques durant leurs vacances et 62 patients etaient des habitants d’Outremer intoxiques chez eux. Un seul patient, un marin civil, a ete intoxique lors de son activite professionnelle. L’espece de poisson incriminee a ete identifiee dans 70 % des cas et appartenait alors majoritairement a trois familles : 10 merous (Serranidae), 5 barracudas (Sphyraenidae) et 5 vivaneaux (Lutjanidae). Les intoxications ont eu lieu en Atlantique dans 23 cas (98 patients), dans l’ocean Indien dans 12 cas (53 patients) et dans l’ocean Pacifique dans 14 cas (24 patients). Au total, 61 % des cas de ciguatera (30/49 dossiers) ont ete pris en charge au cours de la derniere decennie. Les symptomes sont apparus entre 2 a 12 heures apres le repas contaminant, et le tableau clinique de la phase initiale etait variable selon l’origine geographique. Tous les cas en Atlantique ont debute par des signes digestifs, alors que plus de 50 % des cas dans la region Pacifique n’ont jamais presente de signe gastro-intestinal, avec alors un tableau initial domine par des signes neurosensoriels d’emblee comme des paresthesies et/ou des dysesthesies. Un cas mortel a ete collecte en 1999 : un homme de 73 ans est decede d’un arret respiratoire brutal apres ingestion d’un barracuda dans un restaurant local a Cuba. L’evolution des 174 autres patients a ete classique avec des symptomes persistant plusieurs semaines (63 % des patients), voire plus d’un trimestre (18 % des patients). Trente-six pour cent des patients de cette serie ont rapporte des episodes de rechute des signes neurologiques (paresthesies, dysesthesies, allodynie, prurit) plusieurs mois apres l’intoxication, et ce apres consommation de boissons alcoolisees ou de chair de poisson. De tels episodes de resurgence sont rapportes dans la litterature mais leur mecanisme physiopathologique reste mysterieux.

On the French island of Corsica, a 57-y-old woman without significant medical history was stung on the left thumb while she was taking care of an ornamental Ficus benjamina plant. Immediately, she felt intense pain in her hand. She saw a strange caterpillar, later identified by the local poison center as Acharia stimulea. The pain in her hand was evaluated as 8 of 10 using the numerical pain rating scale; only a slight erythema was visible on her skin. Symptoms disappeared within 2 h with use of local anti-inflammatory ointment and oral painkillers. Three other caterpillars emerged out of the soil of the potted plant during the following week. This sting by a saddleback caterpillar is exceptional in Corsica. French garden store owners and healthcare professionals should be informed that caterpillars can be imported across the oceans to Europe on different plants.

Virus infection of plants can result in various degrees of detrimental impacts and disparate symptom types and severities. Although great strides have been made in our understanding of the virus–host interactions in herbaceous model plants, the mechanisms underlying symptom development are poorly understood in perennial fruit crops. Grapevine fanleaf virus (GFLV) causes variable symptoms in most vineyards worldwide. To better understand GFLV-grapevine interactions in relation to symptom development, field and greenhouse trials were conducted with a grapevine genotype that exhibits distinct symptoms in response to a severe and a mild strain of GFLV. After validation of the infection status of the experimental vines by high-throughput sequencing, the transcriptomic and metabolomic profiles in plants infected with the two viral strains were tested and compared by RNA-Seq and LC-MS, respectively, in the differentiating grapevine genotype. In vines infected with the severe GFLV strain, 1023 genes, among which some are implicated in the regulation of the hypersensitive-type response, were specifically deregulated, and a higher accumulation of resveratrol and phytohormones was observed. Interestingly, some experimental vines restricted the virus to the rootstock and remained symptomless. Our results suggest that GFLV induces a strain- and cultivar-specific defense reaction similar to a hypersensitive reaction. This type of defense leads to a severe stunting phenotype in some grapevines, whereas others are resistant. This work is the first evidence of a hypersensitive-like reaction in grapevine during virus infection.

Objectifs La pervenche francaise (Vinca minor) est une plante grimpante de la famille des apocynacees. Les tiges sont fines, les feuilles sont opposees, ovales ou oblongues, et les fleurs, tres souvent violettes, ont cinq petales. La floraison a lieu pendant les mois de mars a juin et les fruits, qui se trouvent au milieu de la fleur contiennent 7-8 graines en juin et juillet. Toute la plante est toxique et contient plus de 50 alcaloides. Un de ces alcaloides, la vincamine, est aussi utilise dans le traitement des troubles de la perfusion cerebrale. Nous decrivons un rare cas d’intoxication collige en France metropolitaine apres ingestion de fleurs et de feuilles de cette espece dans une salade. Methode Une dame de 71 ans s’est fait pour son repas du soir une salade avec 8 fleurs et feuilles de Vinca minor avec des asperges et de la vinaigrette. Resultats Quelques minutes apres, une vasodilatation persistante avec une forte rougeur apparait au niveau du visage de la dame, ce qui l’amene a appeler le centre antipoison de Marseille 2h30 apres son repas. La dame precise par ailleurs qu’elle presente comme antecedent une HTA traitee par irbesartan et qu’elle s’est deja auto-traitee avec du charbon. Il lui est conseille de se reposer et de rappeler s’il y a de moindres soucis. Lorsque le CAP rappelle pour prendre des nouvelles une demi-heure plus tard, la dame presente toujours une rougeur du visage et une tension systolique a 170 (ce qui serait habituel les derniers jours). Au troisieme appel (3h30 de l’ingestion) la dame se sent bien, mais son visage est toujours un peu rouge. Vu le temps passe et la non-aggravation des symptomes pendant les dernieres heures, il est decide de laisser la dame a la maison. Au suivi le lendemain la dame indique que pendant la nuit elle aurait fait une petite malaise, qui serait passe apres l’ingestion d’un biscuit. Depuis elle se sentait bien et est consideree comme guerie. Conclusion Le cas clinique illustre la dangerosite potentielle des intoxications involontaires avec des plantes du jardin non comestibles mais attirantes en insistant sur la necessite d’identifier l’espece avant toute ingestion.

International audience; Introduction: Ciguatera fish poisoning (CFP) is a common Poisoning in the tropical countries. France is directly concerned with French tourists in endemic area and with French citizens living in the French overseas territories. Method: Retrospective, descriptive study of CFP cases handled by the French Poison Control Centre Network from 2012 through 2019. Results: Fifty-two events were studied concerning 130 patients. The fish species was identified for 41 events, mainly belonging to five fish families: 14 groupers, 11 snappers, 5 jacks, 4 parrotfishes, 4 barracudas. The origin of the fish was the Atlantic Ocean (23 events), the Indian Ocean (17 events) and the Pacific Ocean (12 events). 91% of the poisonings occurring in the Atlantic Ocean began with gastrointestinal effects while in 44% of events occurring in the Pacific Ocean, the patients had no gastrointestinal effects (onset with neurological symptoms: paraesthesia and dysesthesia). The evolution of the 130 patients has been classic for CFP with persistent symptoms during 1 to 45 weeks. Numerous patients reported exacerbation of neurological signs several months after poisoning following consumption of alcoholic beverages (23 patients) or seafood (19 patients). Discussion: Medical practitioners in Europe must be trained to manage CFP as cases are reported with tourists returning from endemic areas but also with poisoned patients far from tropical areas after consumption of imported fish.

In the world, the impact of environmental conditions on the number of scorpion events was evaluated in North Africa,Middle East, and the Amazonian region but not in Europe. In mainland France, scorpion species described are Buthus occitanus (Amoreux, 1789), Belisarius xambeui (Simon, 1879) and 4 Euscorpiidae: Euscorpius concinnus (Koch, 1837), Euscorpius italicus (Herbst, 1800), Euscorpius tergestinus (Koch, 1837), and Tetratrichobothrius flavicaudis (De Geer, 1778). We aimed to describe the impact of environmental conduction on the number of scorpion events. For this, a retrospective multi-center study was conducted with data from the French poison control centers files about scorpion events between 1 January 2011 and 31 December 2020. During the study period, 975 incoming calls for scorpion events were recorded and 574 were related to scorpions native to mainland France and Corsica: B. occitanus (n = 86), Euscorpiidae species (n = 222), B. xambeui (n = 1), and undetermined species (n = 265). Cases were mostly reported along the Mediterranean coast, along rivers, and in cities with a trading port. The number of scorpion events was linked to the rivers' water level, rivers' flow, temperature, sunshine, and pluviometry (P < 0.05 for all variables). B. occitanus need warmest and driest environment than Euscorpiidae spp. A link between the severity of the envenoming and climatic condition or seasonality was not demonstrated.

International audience; Concentrated ammonia solutions are strong mineral bases that can induce severe lesions of the upper gastrointestinal tract when ingested. Little is known about the circumstances of poisoning with these solutions. The aim of this study was to compare ammonia exposure between patients who were drug users or abusers with non-user patients. A retrospective study was conducted on the Poison Control Center in Marseille (PCCM) data over a period of 11 years (2009–2019). A total of 614 patients were registered with 564 non-user drugs (NUD) and 50 patients with drug use or addiction (UD). Results founds that UD patients contacted the PCCM mostly for oral exposure (86%), whereas NUD patients were exposed to inhaled ammonia (64%). Repackaging accidents were the major circumstance of poisoning in UD patients (68%) with predominant digestive symptoms (86%). Severity scores evaluated according to the PSS were higher in UD patients even if only the oral route is considered (72% PSS ≥ 2, P < 0.001). Only one death was recorded, in a 65-year-old patient with a history of alcoholism. The annual number of poisoning cases in UD patients was increasing with 56% patients reported during the last two years. Medical teams in contact with patients at risk need to be made aware of this type of accident, which may be rare but can have potentially catastrophic consequences for the user and their close family and friends.

International audience; In European countries, vitamin A toxicity is most often the result of an excessive intake of vitamin supplements and rarely the consequence of the ingestion of a large carnivorous fish liver. We report 3 cases of vitamin A poisoning after fish liver ingestion in mainland and overseas France. The patients were a 12-y-old girl, a 36-y-old pregnant woman, and a 62-y-old man. They experienced headache, nausea, emesis, and desquamation. Laboratory examination showed a high serum retinol level in the girl. The woman's pregnancy progressed to a miscarriage. This case series shows that this kind of poisoning is not restricted to the polar regions. In patients presenting with flushing combined with signs of intracranial hypertension, accurate questioning of the patient's diet is crucial to avoid misdiagnosis and unnecessary examinations. Pregnant women or women of child-bearing age should be informed of the risk to pregnancy in the case of excessive fish liver ingestion.

The purification of plant cell walls is challenging because they constitute an open compartment which is not limited by a membrane like the cell organelles. Different strategies have been established to limit the contamination by proteins of other compartments in cell wall proteomics studies. Non-destructive methods rely on washing intact cells with various types of solutions without disrupting the plasma membrane in order to elute cell wall proteins. In contrast, destructive protocols involve the purification of cell walls prior to the extraction of proteins with salt solutions. In both cases, proteins known to be intracellular have been identified by mass spectrometry in cell wall proteomes. The aim of this chapter is to provide tools to assess the subcellular localization of the proteins identified in cell wall proteomics studies, including: (1) bioinformatic predictions, (2) immunocytolocalization of proteins of interest on tissue sections and (3) in muro observation of proteins of interest fused to reporter fluorescent proteins by confocal microscopy. Finally, a qualitative assessment of the work can be performed and the strategy used to prepare the samples can be optimized if necessary.

The purification of plant cell walls is challenging because they constitute an open compartment which is not limited by a membrane like the cell organelles. Different strategies have been established to limit the contamination by proteins of other compartments in cell wall proteomics studies. Non-destructive methods rely on washing intact cells with various types of solutions without disrupting the plasma membrane in order to elute cell wall proteins. In contrast, destructive protocols involve the purification of cell walls prior to the extraction of proteins with salt solutions. In both cases, proteins known to be intracellular have been identified by mass spectrometry in cell wall proteomes. The aim of this chapter is to provide tools to assess the subcellular localization of the proteins identified in cell wall proteomics studies, including: (1) bioinformatic predictions, (2) immunocytolocalization of proteins of interest on tissue sections and (3) in muro observation of proteins of interest fused to reporter fluorescent proteins by confocal microscopy. Finally, a qualitative assessment of the work can be performed and the strategy used to prepare the samples can be optimized if necessary.

International audience; Grapevine fanleaf virus (GFLV) is a picorna-like plant virus transmitted by nematodes that affects vineyards worldwide. Nanobody (Nb)-mediated resistance against GFLV has been created recently, and shown to be highly effective in plants, including grapevine, but the underlying mechanism is unknown. Here we present the high-resolution cryo electron microscopy structure of the GFLV-Nb23 complex, which provides the basis for molecular recognition by the Nb. The structure reveals a composite binding site bridging over three domains of one capsid protein (CP) monomer. The structure provides a precise mapping of the Nb23 epitope on the GFLV capsid in which the antigen loop is accommodated through an induced-fit mechanism. Moreover, we uncover and characterize several resistance-breaking GFLV isolates with amino acids mapping within this epitope, including C-terminal extensions of the CP, which would sterically interfere with Nb binding. Escape variants with such extended CP fail to be transmitted by nematodes linking Nb-mediated resistance to vector transmission. Together, these data provide insights into the molecular mechanism of Nb23-mediated recognition of GFLV and of virus resistance loss.

International audience; Introduction: For many years, applications for baclofen have widened in the treatment of substance abuse disorder (SUD), mainly alcohol use disorder, with a growing rate of off-label prescriptions in Europe. Clinical effects seem to be both a decrease of craving and anxiety, leading to a decrease of drug or alcohol consumption. We described baclofen poisoning circumstances, therapeutic options and outcomes when used in substance use disorders. Areas covered: This review summarizes the toxicological considerations where baclofen was prescribed in humans for substance use or abuse disorder in randomized clinical trials, case series, case reports and observational studies between 1990 and 2020 according to the Preferred Reporting Items for Systemic reviews and Meta-Analysis. Expert opinion: The most frequent cause of severe intoxication is self-poisoning. A dose above 180 mg are expected to cause severe toxicity and death. The treatment is only symptomatic as no antidote is available. Off-label prescription remains unsafe because the optimal dose is not known and varies greatly between patients. As SUD are frequently associated with psychiatric disorders and such patients may have suicidal thoughts, the risk of self-poisoning is high. Potential co-ingestants should also be considered, especially CNS depressants, and they need to be closely monitored.

ZusammenfassungPalytoxine, die in Korallen vorkommen und unter anderem durch Dinoflagellaten synthetisiert werden, wirken bei Exponierten über einen Eingriff in die Natrium-Kalium-Pumpe und führen somit zu einer Änderung des Konzentrationsgradienten zwischen intra- und extrazellulärem Raum. Die Symptomatik kann aus Atemnot, Übelkeit und Erbrechen, Kopfschmerzen, Tachykardie und EKG-Veränderungen, aber auch aus Ulzerationen der Hornhaut bestehen. Die symptomatische Therapie setzt sich zusammen aus inhalativen Bronchodilatatoren, evtl. Infusion und Schmerztherapie, Antibiotikatherapie bei Infizierung der Schnittwunden, Vorstellung beim Augenarzt und Behandlung mit Vitamin-A-haltigen sowie antibiotischen Augentropfen. Um eine Exposition mit Palytoxinen zu vermeiden, gilt es, entweder auf Korallen im Aquarium zu verzichten oder bereits vorhandene Korallen nicht aus dem Aquarium zu entfernen.

Objectif La solution d’ammoniaque a de nombreux usages tant domestiques que professionnels. Les usagers de cocaine connaissent egalement ses proprietes basiques qui permettent la synthese de crack (ou free base) [1] . Les cas d’ingestion accidentels ou volontaires de ce produit caustique sont rares mais potentiellement graves. L’objectif de cette etude est de decrire une serie de cas d’intoxication a l’ammoniaque rapportes au centre Antipoison de Marseille (CAPM). Methode Tous les cas d’intoxication a l’ammoniaque rapportes au CAPM sur 10 ans (1er janvier 2009 au 31 decembre 2018) ont ete analyses retrospectivement. Pour chaque cas, les antecedents, les symptomes, les circonstances d’intoxication et la severite ont ete recueillis. Resultats 543 cas d’intoxication par ammoniaque ont ete analyses avec 321 (59 %) par voie inhalee, 176 (32,5 %) par voie orale, 21 (3,8 %) par voie oculaire, 23 (4,2 %) par voie cutanee et 2 (0,5 %) par voie IV. Il y avait 37 patients aux antecedents de toxicomanie, essentiellement des hommes consommateurs de cocaine. Ces patients etaient plus nombreux au cours des annees avec 59 % des cas signales lors des 2 dernieres annees (p Discussion En utilisant de l’ammoniaque deconditionnee, les usagers de crack sont plus exposes au risque d’accident de confusion avec un soda ou de la biere. Ces cas d’ingestions accidentelles surviennent principalement lors de soirees ou tot le matin, en lendemain de soiree, lorsque le patient a soif. Nos resultats sont probablement sous-estimes du fait que les consommateurs de substances illicites sont mefiants et refusent souvent de preciser leurs antecedents medicaux ou de decrire les circonstances d’intoxication. L’entourage direct est egalement a risque comme l’illustre la seule publication d’un cas d’intoxication a l’ammoniaque en lien avec le milieu toxicomane. Il s’agissait d’un enfant de 2 ans dont les parents etaient usagers [1] . Il aurait ete interessant de savoir si l’entourage des deux enfants de notre etude etait usager de substances illicites ou non. Conclusion Ces premiers resultats montrent que les intoxications accidentelles par ammoniaque chez les usagers de cocaine ne sont pas anecdotiques. Une etude nationale serait necessaire pour confirmer ces donnees. En attendant, les equipes medicales en lien avec les patients a risque doivent etre sensibilisees a ce type d’accident, certes rare mais aux consequences potentiellement catastrophiques pour l’usager comme pour son entourage direct.

Objectifs Les violets de roche du genre Microcosmus sont des fruits de mer consommes crus en periodes festives en Provence. Ce sont des animaux marins sessiles de la classe des ascidies. A partir de 2011, le centre antipoison de Marseille (CAPM) a ete consulte pour plusieurs intoxications apres consommation de violets avec un tableau clinique n’evoquant aucune intoxication marine connue. Deux objectifs ont ete definis : decrire le tableau clinique de cette pathologie emergente et obtenir des restes de repas pour identifier la toxine responsable. Methode Du point de vue clinique, une etude retrospective nationale a ete effectuee pour verifier l’existence d’intoxications similaires plus anciennes du 01/01/2002 au 01/01/2011. Une etude prospective a Marseille a ete mise en place du 01/01/2011 au 31/03/2018 pour realiser un suivi de toxicovigilance pour tout nouveau dossier collecte. Du point de vue analytique, les echantillons de chair de violet ont ete analyses par chromatographie liquide d’interaction hydrophile couplee a la spectrometrie de masse basse resolution (HILIC-LRMS) pour quantification puis haute resolution (HILIC-HRMS) pour confirmation. Resultats Resultats cliniques : l’etude retrospective a permis de retrouver une seule intoxication similaire en decembre 2004 (dossier du CAPM avec ingestion de 6 violets par un homme de 50 ans). Les 18 premiers cas colliges par l’enquete prospective de janvier 2011 a mars 2018 (au total 26 patients), ont ete publies [1] permettant de definir un tableau clinique homogene. Ainsi, 10 a 90 minutes apres le repas, un syndrome cerebelleux se developpe pendant 1 a 24 heures : par ordre decroissant de frequence, ataxie, vertiges, vision trouble, diplopie, troubles de l’accommodation, myalgies, crampes, cephalees, paresthesies, tremulations, hypersudation. Des troubles digestifs sont possibles, mais restent secondaires : vomissements, diarrhee, douleurs abdominales pour moins de la moitie des patients. Depuis cette synthese publiee, d’autres observations ont ete colligees soit de facon retrospective par l’Anses (une observation dans le Morbihan en 2013 avec des violets achetes a Marseille), soit de facon prospective par le CAPM avec recuperation d’echantillons de violets pour deux cas (decembre 2019 dans le Var, janvier 2020 dans le Gard). Resultats analytiques : des restes de violets recuperes lors de 3 cas ont ete analyses. L’absence de toxines marines lipophiles a ete confirmee par LC-HRMS. L’analyse par HILIC-LRMS (confirmation par HILIC-HRMS) a permis de retrouver des concentrations variables en anatoxine-a dans tous les echantillons impliques dans des syndromes cerebelleux (190 a 1240 μg/kg de chair de violet), mais aussi dans de moindres concentrations dans des violets « temoins » achetes a Paris (22,5 μg/kg de chair de violet, pas de toxine dans la chair de moule temoin) [2] . Conclusion L’anatoxine-a est une cyanotoxine hydrophile redoutable impliquee dans des deces de canides, de betail ou d’oiseaux sur tous les continents. En France, il s’agit de la principale toxine responsable de la mort de chiens et chevreuils en ete, au bord de cours d’eau lents ou des pullulations de cyanobacteries sont possibles. Il n’existe pas d’intoxication humaine clairement attribuee a cette toxine, ce qui ferait de cette serie la premiere connue impliquant l’anatoxine-a comme probable responsable de cas humains. Tous les echantillons de chair de violet analyses, y compris certains qui servaient de temoin, contenaient cette cyanotoxine des eaux douces. Enfin, il semble que les violets surexploites soient une ressource quasi epuisee en France. L’origine geographique precise des lots consommes est difficile a obtenir mais plusieurs enquetes du CAPM tendent a montrer que les violets impliques dans des intoxications proviennent de mer Adriatique, plus particulierement de Croatie qui est le premier fournisseur en violets du marche francais. Enquete a suivre…

International audience; Since their discovery, single-domain antigen-binding fragments of camelid-derived heavy-chainonly antibodies, also known as nanobodies (Nbs), have proven to be of outstanding interest as therapeutics against human diseases and pathogens including viruses, but their use against phytopathogens remains limited. Many plant viruses including Grapevine fanleaf virus (GFLV), a nematode-transmitted icosahedral virus and causal agent of fanleaf degenerative disease, have worldwide distribution and huge burden on crop yields representing billions of US dollars of losses annually, yet solutions to combat these viruses are often limited or inefficient. Here, we identified a Nb specific to GFLV that confers strong resistance to GFLV upon stable expression in the model plant Nicotiana benthamiana and also in grapevine rootstock, the natural host of the virus. We showed that resistance was effective against a broad range of GFLV isolates independently of the inoculation method including upon nematode transmission but not againstits close relative, Arabis mosaic virus. We also demonstrated that virus neutralization occurs at an early step of the virus life cycle, prior to cell-to-cell movement. Our findings will not only be instrumental to confer resistance to GFLV in grapevine, but more generally they pave the way for the generation of novel antiviral strategies in plants based on Nbs

Avirulence factors are critical for the arm's race between a virus and its host in determining incompatible reactions. The response of plants to viruses from the genus Nepovirus in the family Secoviridae, including Grapevine fanleaf virus (GFLV), is well characterized, although the nature and characteristics of the viral avirulence factor remain elusive. By using infectious clones of GFLV strains F13 and GHu in a reverse genetics approach with wild-type, assortant and chimeric viruses, the determinant of necrotic lesions caused by GFLV-F13 on inoculated leaves of Nicotiana occidentalis was mapped to the RNA2-encoded protein 2AHP , particularly to its 50 C-terminal amino acids. The necrotic response showed hallmark characteristics of a genuine hypersensitive reaction, such as the accumulation of phytoalexins, reactive oxygen species, pathogenesis-related protein 1c and hypersensitivity-related (hsr) 203J transcripts. Transient expression of the GFLV-F13 protein 2AHP fused to an enhanced green fluorescent protein (EGFP) tag in N. occidentalis by agroinfiltration was sufficient to elicit a hypersensitive reaction. In addition, the GFLV-F13 avirulence factor, when introduced in GFLV-GHu, which causes a compatible reaction on N. occidentalis, elicited necrosis and partially restricted the virus. This is the first identification of a nepovirus avirulence factor that is responsible for a hypersensitive reaction in both the context of virus infection and transient expression.

La Presse Medicale - In Press.Proof corrected by the author Available online since mercredi 31 janvier 2018

Grapevine fanleaf virus (GFLV) and arabis mosaic virus (ArMV) are nepoviruses responsible for grapevine degeneration. They are specifically transmitted from grapevine to grapevine by two distinct ectoparasitic dagger nematodes of the genus Xiphinema. GFLV and ArMV move from cell to cell as virions through tubules formed into plasmodesmata by the self-assembly of the viral movement protein. Five surface-exposed regions in the coat protein called R1 to R5, which differ between the two viruses, were previously defined and exchanged to test their involvement in virus transmission, leading to the identification of region R2 as a transmission determinant. Region R4 (amino acids 258 to 264) could not be tested in transmission due to its requirement for plant systemic infection. Here, we present a fine-tuning mutagenesis of the GFLV coat protein in and around region R4 that restored the virus movement and allowed its evaluation in transmission. We show that residues T258, M260, D261, and R301 play a crucial role in virus transmission, thus representing a new viral determinant of nematode transmission.

Grapevine fanleaf virus (GFLV) is a picorna-like plant virus transmitted by nematodes that affects vineyards worldwide. Nanobody (Nb)-mediated resistance against GFLV has been created recently and shown to be highly effective in plants including grapevine, but the underlying mechanism is unknown. Here we present the high-resolution cryo-EM structure of the GFLV-Nb23 complex which provides the basis for the molecular recognition by the nanobody. The structure reveals a composite binding site bridging over 3 domains of the capsid protein (CP) monomer. The structure provides a precise mapping of the Nb23 epitope on the GFLV capsid in which the antigen loop is accommodated through an induced fit mechanism. Moreover, we uncover and characterize several resistance-breaking GFLV isolates with amino acids mapping within this epitope, including C-terminal extensions of the CP, which would sterically interfere with Nb binding. Escape variants with such extended CP fail to be transmitted by nematodes linking Nb-mediated resistance to vector transmission. Together, these data provide insights into the molecular mechanism of Nb23-mediated recognition of GFLV and of virus resistance loss.SignificanceGrapevine fanleaf virus (GFLV) is a picorna-like plant virus that severely impacts vineyards worldwide. While Nanobodies (Nb) confer resistance to GFLV in plants the underlying molecular mechanism of action is unknown. Here we present the high-resolution cryo-EM structure of the GFLV-Nb complex. It uncovers the conformational epitope on the capsid surface which is a composite binding site into which the antigen loop is accommodated through an induced fit mechanism. Furthermore, we describe several resistance-breaking isolates of GFLV with reduced Nb binding capacity. Those that carry a C-terminal extension also fail to be transmitted by nematodes. Together, these data provide structure-function insights into the Nb-GFLV recognition and the molecular mechanism leading to loss of resistance.

The mechanisms underlying host plant symptom development upon infection by viruses of the genus Nepovirus in the family Secoviridae, including grapevine fanleaf virus (GFLV), are poorly understood. In the systemic host Nicotiana benthamiana, GFLV strain GHu produces characteristic symptoms of vein clearing in apical leaves, unlike other GFLV strains such as F13, which cause an asymptomatic infection. In this study, we expanded on earlier findings and used reverse genetics to identify residue 802 (lysine, K) of the GFLV-GHu RNA1-encoded RNA-dependent RNA polymerase (1EPol) as a modulator of vein-clearing symptom development in N. benthamiana. Mutations to this site abolished (K to G, A, or Q) or attenuated (K to N or P) symptom expression. Noteworthy, residue 802 is necessary but not sufficient for vein clearing, as GFLV-F13 RNA1 carrying K802 remained asymptomatic in N. benthamiana. No correlation was found between symptom expression and RNA1 accumulation, as shown by reverse transcription-quantitative polymerase chain reaction. Additionally, the involvement of RNA silencing of vein clearing was ruled out by virus-induced gene silencing experiments and structure predictions for protein 1EPol suggested that residue 802 is flanked by strongly predicted stable secondary structures, including a conserved motif of unknown function (805LLKT/AHLK/RT/ALR814). Together, these results reveal the protein nature of the GFLV-GHu symptom determinant in N. benthamiana and provide a solid basis for probing and determining the virus-host proteome network for symptoms of vein clearing.

Introduction Peganum harmala, aussi appele rue sauvage, est une plante herbacee vivace, a l’origine rencontree dans les milieux semi-arides du bassin mediterraneen et en Asie centrale mais repandue aujourd’hui en Afrique, Amerique du Sud et Australie. Graines, feuilles et fleurs sont utilisees pendant des rites, pour leurs effets medicaux, comme drogues ou comme teinture des tapis. Les graines de Peganum harmala contiennent les alcaloides Harmine et Harmaline. Il s’agit d’inhibiteurs de la monoamine oxidase A qui potentialisent ou demasquent les effets psychoactifs de certaines drogues. Melangees avec de la dimethyltryptamine (DMT, extrait de Psychotria Viridis) dans une infusion, elles sont utilisees en medecine traditionnelle [1] , [2] . Nous decrivons un cas d’automedication survenu a Mayotte avec apparition d’effets indesirables ayant necessite une prise en charge aux urgences. Observation Pour traiter l’enuresie nocturne d’un garcon de 12 ans, ses parents ont prepare une infusion en melangeant une poignee de graines d’harmal dans un litre d’eau. Le garcon a bu plusieurs gorgees le matin avant de partir a l’ecole. Arrive sur place, il a presente des nausees, des vomissements, une vision floue et un malaise avec perte de connaissance. Transporte aux urgences via le SAMU, l’examen clinique retrouvait une somnolence. La gazometrie montrait une acidose respiratoire avec hypercapnie (pH 7,27, pCO2 = 60 mmHg). L’ECG et le reste du bilan etaient normaux. Le medecin des urgences a contacte le centre antipoison de Marseille pour avis concernant la prise en charge. Une hydratation intraveineuse associee a une surveillance avec controle de la gazometrie et de l’ECG a ete proposee. L’etat clinique du garcon s’est ameliore apres le traitement, le controle de la gazometrie dans l’apres-midi se revelait normal ainsi que le deuxieme ECG. L’enfant a pu regagner son domicile le jour meme. Conclusion Les parents ont utilise les graines de Peganum harmala pour traiter l’enuresie nocturne de leur enfant. Or, la revue de la litterature n’a pas retrouve l’enuresie nocturne comme indication de ce traitement. Au contraire, les inhibiteurs de la monoamine oxidase A harmine et harmaline provoquent l’effet oppose, soit une polyurie. Par ailleurs, les parents, non conscients des effets indesirables potentiels de leur traitement, n’ont pas surveille leur enfant et l’ont envoye a l’ecole ou il a perdu connaissance. Le cas clinique illustre la dangerosite de l’automedication en medecine traditionnelle par des usagers n’ayant pas forcement les connaissances requises.

The mechanisms underlying host plant symptom development upon infection by viruses of the genus

Dear Editor,Ascidians of Microcosmus taxa are eaten all along the French Mediterranean coast. It is a delicacy eaten during winter, especially during Christmas fests. French people call them sea fi...

Resume Il existe tres peu de donnees sur les intoxications par champignons macroscopiques recoltes sur l’Ile de la Reunion dans la litterature scientifique. Les auteurs decrivent le cas d’un patient âge de 43 ans qui a developpe un syndrome resinoidien une heure trente apres avoir mange 3 specimens de Chlorophyllum molybdites cuits, qu’il avait recoltes en ete dans le Sud Ouest de l’ile en pensant cueillir des volvaires. Il a developpe des douleurs abdominales severes ainsi que des diarrhees glairo-sanglantes d’evolution favorable en 48 heures apres traitement symptomatique en service d’urgences medicales. Ce cas montre d’une part, que C. molybdites , champignon que l’on retrouve habituellement dans les zones tropicales et subtropicales du globe, est present sur l’Ile de la Reunion et d’autre part, confirme que sa consommation peut engendrer un syndrome gastro-intestinal severe necessitant l’hospitalisation du patient.

International audience; One of the greatest hindrances to the study of grapevine fanleaf virus (GFLV) is the dearth of robust protocols for reliable, scalable, and cost-effective inoculation of host plants, especially methods which allow for rapid and targeted manipulation of the virus genome. Agroinoculation fulfills these requirements: it is a relatively rapid, inexpensive, and reliable method for establishing infections, and enables genetic manipulation of viral sequences by modifying plasmids. We designed a system of binary plasmids based on the two genomic RNAs [RNA1 (1) and RNA2 (2)] of GFLV strains F13 (F) and GHu (G) and optimized parameters to maximize systemic infection frequency in Nicotiana benthamiana via agroinoculation. The genomic make-up of the inoculum (G1–G2 and reassortant F1–G2), the identity of the co-infiltrated silencing suppressor (grapevine leafroll associated virus 2 p24), and temperature at which plants were maintained (25 °C) significantly increased systemic infection, while high optical densities of infiltration cultures (OD600nm of 1.0 or 2.0) increased the consistency of systemic infection frequency in N. benthamiana. In contrast, acetosyringone in the bacterial culture media, regardless of concentration, had no effect. Plasmids in this system are amenable to rapid and reliable manipulation by one-step site-directed mutagenesis, as shown by the creation of infectious RNA1 chimeras of the GFLV-F13 and GHu strains. The GFLV agroinoculation plasmids described here, together with the optimized protocol for bacterial culturing and plant maintenance, provide a robust system for the establishment of systemic GFLV infection in N. benthamiana and the rapid generation of GFLV mutants, granting a much-needed tool for investigations into GFLV-host interactions.