Guidelines for improving statistical analyses of validation datasets for plant pest diagnostic tests

peer reviewedAppropriate statistical analysis of the validation data for diagnostic tests facilitates the evaluation of the performance criteria and increases the confidence in the conclusions drawn from these data. A comprehensive approach to analysing and reporting data from validation studies and inter-laboratory comparisons such as test performance studies is described. The proposed methods, including statistical analyses, presentation and interpretation of the data, are illustrated using a real dataset generated during a test performance study conducted in the framework of the European project, VALITEST. This analytical approach uses, wherever possible and whenever applicable, statistical analyses recommended by international standards illustrating their application to plant health diagnostic tests. The present work is addressed to plant health diagnosticians and researchers interested and/or involved in the validation of plant diagnostic tests, and also aims to convey the necessary information to those without a statistical background. Detailed statistical explanations are provided in the Appendices

What did we achieve with VALITEST an EU project on validation in plant pest diagnostics?

peer reviewedEnsuring the reliability of diagnostic activities is an essential cornerstone of Plant Health strategies to reduce the risk of entry and spread of plant pests in a region and ultimately their impacts. Diagnostic tests should be validated to ensure that they are fit for purpose. Validation is usually done by diagnostic laboratories although companies commercializing diagnostic kits also produce validation data for their products. Due to the high number of pest , matrix and method combinations and given the significant resources required to validate tests, it is essential that validation data are shared with the entire diagnostic community and produced in a harmonized way to facilitate their use by different stakeholders. Indeed, the selection of tests to be used in specific contexts is not the sole responsibility of diagnostic laboratories and also involve National Plant Protection Organizations. The VALITEST EU project (2018-2021) was established to tackle all these issues. New validation data for tests targeting important pests for the EPPO region were produced. Guidelines to improve and harmonize the validation framework were developed. Sharing of validation data and experience was ensured through the development of new or existing databases, the organization of training courses and the dissemination of the project outputs in scientific publications and Standards. Finally, the involvement of researchers, diagnosticians, policy makers, inspectors, industries etc. and the establishment of the European Plant Diagnostic Industry Association were important actions to strengthen the interactions between Plant Health stakeholders

Early Detection of the Root-Knot Nematode Meloidogyne hapla Through Developing a Robust Quantitative PCR Approach Compliant With the Minimum Information for Publication of Quantitative Real-Time PCR Experiments Guidelines

International audienceRoot-knot nematodes (RKNs) are major threats to crops through attacking the roots, which induces an abnormal development of the plant. Meloidogyne hapla Chitwood is of particular concern as it is currently expanding its distribution area and displays a wide host range. Effective plant protection against this RKN requires early detection as even a single individual can cause severe economic loses on susceptible crops. Molecular tools are of particular value for this purpose and among them, qPCR presents all the advantages, i.e. sensitivity, specificity, rapidity of diagnosis at a reduced cost. Although few studies already proposed detecting M. hapla through this technique, they lack experimental details and performance testing, and suffered from low taxonomic resolution and/or required expensive hydrolysis probes. Here we propose a qPCR detection method that uses SybrGreen with developed primers amplifying a fragment of COI mitochondrial region. The method is developed and evaluated following the MIQE guidelines to ensure its quality, i.e. sensitivity, specificity, repeatability, reproducibility, robustness. The results demonstrate that the newly developed method fulfills its goals as it proved specific to M. hapla and allowed for a reproductible detection level as low as 1.25 equivalent of a juvenile individual. All criteria associated with the MIQE guidelines were also met, what makes it of general use for the reliable early detection of M. hapla

Two decades of epidemiological surveillance of the pine wood nematode in France reveal its absence despite suitable conditions for its establishment

Abstract Key message This study takes stock of the first 20 years (2000–2019) of monitoring the pine wood nematode (PWN) in metropolitan France. While PWN was never found in the wild during this period, it was reported in some wood-based commodities entering or circulating on French territory. This stresses the importance of remaining extremely vigilant, as the conditions found in France, especially weather conditions, could be particularly suitable for the pest’s establishment. Context The pine wood nematode (PWN) Bursaphelenchus xylophilus, responsible for pine wilt disease (PWD), is one of the most important forest tree pests worldwide. It is thus the focus of many monitoring programmes. In the European Union, for example, it is categorised as a priority quarantine pest, so each member state is obliged to monitor the PWN on its territory. Aims The first objective of this paper was to describe PWN monitoring in metropolitan France, namely how it is organised and whether it has led to the nematode’s detection. Secondly, we wished to investigate what the levels of PWD expression for host pines infected by B. xylophilus would be in France. Thirdly, we wanted to find out whether other Bursaphelenchus species had been found on French territory during these two decades of PWN monitoring. Methods We analysed data from samples collected in the framework of the monitoring programme between 2000 and 2019 to track the PWN in its host pines, its insect vector (Monochamus spp.) and in wood-based commodities imported into or circulating in metropolitan France. We also generated risk maps of PWD expression based on an evapo-transpiration model using climate data for the period 2000–2019. Results This monitoring, which was regularly reinforced from 2000 to 2019, consisted of sampling and analysing around 18,000 wood samples and 66,000 insects over this period. Although the PWN was not detected in pine stands or within its insect vector, some wood-based commodities were found to be contaminated. Risk maps of PWD expression show that in the most recent years (2015–2019), the weather conditions in a large fraction of metropolitan France were suited to PWD expression, mostly with a delay (i.e., latency) between infection and observable wilt symptoms. PWN monitoring has also revealed the presence of other Bursaphelenchus species, most of which were discovered for the first time in metropolitan France and are described herein. Conclusion While metropolitan France is still free of the PWN, this study emphasises the need to remain cautious as the French territory appears particularly suitable for this pest’s establishment. Furthermore, our research has led us to propose some ideas on how to improve PWN monitoring

PDTM-36. WHOLE EXOME SEQUENCING (WES) OF DIPG PATIENTS FROM THE BIOMEDE TRIAL REVEALS NEW PROGNOSTIC SUBGROUPS WITH SPECIFIC ONCOGENIC PROGRAMMES

International audienceAbstract BACKGROUND & METHODS. The BIOlogical MEdicines for DIPG Eradication (BIOMEDE) trial is a randomized multi-arm multi-stage program (drop-the-loser adaptive design). The first trial was an open-label phase-II trial comparing three drugs (everolimus, dasatinib, erlotinib) combined with irradiation, allocated according to the presence of their specific targets. All patients underwent a biopsy at diagnosis to confirm the DIPG molecularly and establish the expression of pre-specified biomarkers. Patient samples were explored by WES with a sequencing depth average >100X for the tumor and >60X for the blood. RESULTS. Unsupervised clustering of copy-number-variations (CNV) identified 4 groups corresponding to a new stratification of DIPG: cluster 1 (n=42) with high CNV, cluster 2 (n=23) with chromosome 1q + chromosome 2 gains, cluster 3 (n=9) with low CNV and cluster 4 (n=21) with isolated 1q gain. Clusters 1 had higher hazard-ratio for death than the others (mean HR 1.959, range 1.054–3.643, p< 0.0001, adjusted Cox model). Extensive structural rearrangements/chromotrypsis were significantly more frequent in TP53-mutated samples. Indeed, when stratiftying patients in 4 groups based on type of histone H3 mutated at K27 and the presence of a TP53 pathway alteration the subgroup with TP53 altered pathway had a significantly higher hazard-ratio for death than the others (mean HR 3.450, p=0.0017, adjusted Cox model). Additional drug targets were identified, especially in the DNA repair machinery that could be exploited for new targeted therapies. CONCLUSION. WES at diagnosis was feasible in most patients and brings new prognostic and theranostic informations. This allows a better patients stratification and the development of personalized medicine in DIPG

Mythologies urbaines

Comment une ville s’empare des éléments de son quotidien et de son passé pour inventer sa propre légende, ses mythes protecteurs, ses rêves et ses dieux… De Dunkerque à Londres, de New York à Coblence, de Saint-Malo à Paris, à partir d’exemples géographiquement très divers et choisis sur une longue durée, les auteurs de cet ouvrage ont tenté de comprendre les conditions dans lesquelles une communauté donnée s’approprie un personnage, historique ou non, un événement né de la ville, afin de construire les symboles qui éclaireront son devenir et dépasseront les tragédies humaines. C’est dans le cadre des événements culturels qui ont ponctué le Tricentenaire de la mort de Jean Bart en 2002 que la Ville de Dunkerque et la Communauté Urbaine, en partenariat avec l’Université du Littoral Côte d’Opale, ont organisé les Rencontres Mythologies Urbaines. Faisant suite à la parution des actes du colloque Jean Bart et son Temps - autre rendez-vous prestigieux de cette année du Tricentenaire - voici donc la somme des interventions de ces Rencontres réunies dans le présent ouvrage, exemplaire d’une collaboration universitaire à l’échelle de l’Hexagone puisque ce sont les Presses Universitaires de Rennes qui en ont assuré l’édition

Prognostic relevance of clinical and molecular risk factors in children with high-risk medulloblastoma treated in the phase II trial PNET HR+5

International audienceAbstract Background High-risk medulloblastoma is defined by the presence of metastatic disease and/or incomplete resection and/or unfavorable histopathology and/or tumors with MYC amplification. We aimed to assess the 3-year progression-free survival (PFS) and define the molecular characteristics associated with PFS in patients aged 5–19 years with newly diagnosed high-risk medulloblastoma treated according to the phase II trial PNET HR+5. Methods All children received postoperative induction chemotherapy (etoposide and carboplatin), followed by 2 high-dose thiotepa courses (600 mg/m2) with hematological stem cell support. At the latest 45 days after the last stem cell rescue, patients received risk-adapted craniospinal radiation therapy. Maintenance treatment with temozolomide was planned to start between 1–3 months after the end of radiotherapy. The primary endpoint was PFS. Outcome and safety analyses were per protocol (all patients who received at least one dose of induction chemotherapy). Results Fifty-one patients (median age, 8 y; range, 5–19) were enrolled. The median follow-up was 7.1 years (range: 3.4–9.0). The 3 and 5-year PFS with their 95% confidence intervals (95% CI) were 78% (65–88) and 76% (63–86), and the 3 and 5-year OS were 84% (72–92) and 76% (63–86), respectively. Medulloblastoma subtype was a statistically significant prognostic factor (P-value = 0.039) with large-cell/anaplastic being of worse prognosis, as well as a molecular subgroup (P-value = 0.012) with sonic hedgehog (SHH) and group 3 being of worse prognosis than wingless (WNT) and group 4. Therapy was well tolerated. Conclusions This treatment based on high-dose chemotherapy and conventional radiotherapy resulted in a high survival rate in children with newly diagnosed high-risk medulloblastoma

NEMATOOLS : Développement d’outils pour la maîtrise durable du risque nématodes en plant de pomme de terre et cultures en rotation

Nematools project aimed at reinforcing prophylactic methods to prevent introduction and disseminationof regulated phytoparasites nematodes Meloidogyne chitwoodi, M. fallax, Globodera pallida and G.rostochiensis which are serious threats for potato crops and to improve the control strategies in order to preserve the privileged health status of the French territory. This project made it possible to developnew molecular tools for the detection of root-knot nematodes in soil samples, and later their validationon a collection of nematode populations established during the project. The risk analysis on thedissemination of nematodes related to agricultural practices have led firstly to the development of aconceptual model for the identification of management strategies for the use of agricultural equipment atrisk and on the other hand to highlight and to quantify the importance of cyst nematodes dispersal bythe soil practices and by soil-adhering to agricultural machinery at the field and at the farm scale. Theactions carried out in partnership with industrials and with experimental stations made it possible toproduce a user-friendly risk analysis tool characterizing the critical phases of different waste and effluenttreatment processes and providing answers on the sanitizing performances of different decontaminationprocesses such as composting, lagooning, anaerobic digestion, chlorination, and heat treatment. Thisproject also allowed to screen, under controlled conditions, plant species which are not conducive to thedevelopment and propagation of root-knot nematodes such as M. chitwoodi and M. fallax and may beproposed as rupture plants on contaminated sitesLes actions conduites dans le cadre du projet de recherche Nematools visaient à renforcer lesméthodes prophylactiques pour prévenir l’introduction et la dissémination des nématodesphytoparasites réglementés Meloidogyne chitwoodi, M. fallax, Globodera pallida et G. rostochiensis.Ces nématodes constituent des menaces sérieuses pour diverses cultures, dont la pomme de terre, etconduisent à repenser les stratégies de lutte afin de préserver l’état sanitaire privilégié du territoirenational. Le projet a permis le développement de nouveaux outils moléculaires de détection desnématodes à galle dans des échantillons de sol, outils validés sur une collection de populations denématodes constituée pendant la durée du projet. Les travaux portant sur l’analyse du risque dedissémination des nématodes, associés aux pratiques agricoles, ont conduit (1) à l’élaboration d’unmodèle conceptuel permettant d’identifier les stratégies de gestion du matériel agricole à risque et (2) àmettre en évidence et à quantifier l’importance de la dispersion des nématodes à kyste par les travauxdu sol et par le matériel agricole à l’échelle parcellaire et inter-parcellaire. Les travaux menés enpartenariat avec des industriels ainsi qu’en stations expérimentales ont permis la production d’un outilinteractif d’analyse de risque caractérisant les phases critiques de différents process de traitements desdéchets et d’effluents. Il apporte des éléments de réponse sur les performances d’assainissement dedifférents procédés de décontamination comme le compostage, le lagunage, la méthanisation, lachloration et le traitement thermique. Ce projet a aussi permis de cribler, en conditions contrôlées, desespèces végétales non favorables à la multiplication des nématodes à galle M. chitwoodi et M. fallax etpouvant être proposées comme plantes de rupture sur des sites contaminés