Benaki Phytopathological Institute

Strengths and constraints of empirical and process-based models in predicting environmental parameters are a perennial topic of scientific dialogue, which is often conducted through comparisons. Two fundamentally different, yet comparable, empirical and physical models, used to estimate average erosion rates on a regional scale, are the (R)evised Universal Soil Loss Equation (R-USLE) and the Pan-European Soil Erosion Risk Assessment (PESERA), respectively. Comparisons of (R)USLE and PESERA in international literature are limited. Further studies are recommended to identify the key limitations of both models. This study aims to provide a deep understanding of the implementation and discover or confirm key differences between the models in one of Greece's largest wine-producing zones. PESERA estimated a very low or low (<1 t/ha/year) erosion risk in 91.69% of the study area. RUSLE overestimated erosion rates, classifying 50.02% of the area as having a high, very high or extreme (>10 t/ha/year) erosion risk and 40.91% as facing moderate (1–10 t/ha/year) erosion risk. 65.93% of the study area showed significant differences between the two models, mainly in agricultural land. PESERA showed a tendency for higher rates at steeper slopes, but RUSLE's rates were several orders of magnitude higher across all slopes. PESERA results were closer to conventional soil mapping recordings. This work paves the way for future studies to address other important issues, including the quantitative validation of models' results, the integration of additional laboratory soil data (e.g. soil erodibility and soil crusting) and the improvement of the accuracy of subjective and qualitative parameters derived from satellite products or coarse-scale statistics (e.g. vegetation cover and anti-erosion measures). • Estimated RUSLE values were several orders of magnitude higher than PESERA's. • Increases in slope degree were found to exacerbate RUSLE's overestimations. • PESERA results were closer to conventional soil mapping recordings. • The spatial distribution of erosion rate classes was similar in both models. • Spatial distribution of PESERA's results was similar in both runs (10 and 100m).

DNA methylation is a conserved regulatory mechanism of gene expression, genome stability, and development, and is highly associated with the effective induction of defense responses for plant priming. In the Green Deal era, the use of plant defense inducers (PDIs), compounds that activate defense and prime plants against imminent pathogen attacks, is a safe and environmentally sustainable approach to support plants against pathogens. Though efforts have succeeded at deciphering part of the mode of action of PDIs, more information is needed to understand the underlying pathways of their effectiveness. Here, salicylic acid (SA), loaded in chitosan nanoparticles, increased hypomethylation by more than 25% for 56 genomic regions that corresponded to defense-related genes, such as pectin lyases, defensins and leucine-rich repeat transmembrane protein kinases against the biotrophic fungal pathogen Podosphaera xanthii. A genomic region of the promoter of SKP1A, which is a core member of the SCF E3 ubiquitin ligase complex, was found to be a differentially methylated region (DMR), with 60% hypomethylation, both after PDI application and pathogen inoculation, possibly indicating a similar activation mechanism. Examination of this DMR revealed the presence of SA-, auxin-, and defense-related cis-elements. Investigation of the proteins associated with the above cis-elements showed significant upregulation in expression after PDI. Moreover, association of the identified DMR with transcriptomics showed enrichment of the SA pathway. Overall, these findings shed light on the epigenetic mechanisms that underlie SA-related defense priming in plants.

This review comprehensively examines the variability and uncertainty associated with test guideline (TG)-conform genotoxicity data and explores the respective implications for the integration of non-animal-methods (NAMs) into regulatory frameworks. Historical amendments to OECD TGs are mapped to reveal the method's evolution that improves the scientific quality of the data but also explains data heterogeneity within available databases. An analysis of the major genotoxicity databases ECVAM, ISSMIC, and OASIS demonstrates substantial variability in genotoxicity calls. Using the EFSA genotoxicity database, which currently harbours the best-curated (meta-) data, we estimate that 22-77% of compounds exhibit similarity of replicate results below 85%, depending on the assay. The potentially most important variables statistically explaining variability and sensitivity were analysed. The practical limitations to identify them with high reliability and to define their optimum needs to be accepted as a qualitative baseline uncertainty. These findings underscore the necessity of contextualizing NAM performance evaluations within the intrinsic variability and uncertainty of animal and in vitro reference data. We propose that this variability is explicitly considered in the development and validation of NAM-based Integrated Approaches for Testing and Assessment. This review provides a critical foundation for regulators and scientists aiming to enhance the acceptance and utility of NAMs in genotoxicity assessment.

Microplastics (MPs) enter terrestrial ecosystems through various pathways, including the use of plastic mulching films, treated sewage sludge, and chemical and organic fertilizers. Polypropylene (PP) and polyethylene (PE) are the dominant polymers found in both traditional and facility-based farmland soils. MPs negatively impact soil microbial communities and harm soil invertebrates such as earthworms, nematodes, and springtails. In plants, MPs can induce oxidative stress, damage cells and inhibit growth. Polystyrene (PS) is often identified as the most hazardous polymer, frequently linked to reduced plant growth, which is the most commonly reported effect of soil MP contamination. This review provides novel insights beyond those reported in the previous literature, revealing that greenhouse-based cultivation, vegetable crops, orchards, and vineyards are significant contributors to increased microplastic soil contamination. Furthermore, the findings underscore pronounced global heterogeneity in microplastic concentrations within paddy soils, with recorded levels varying widely from 16 to 10,300 items kg−1. Oxidative stress and additive leaching are the dominant mechanisms driving soil microplastic toxicity across exposed organisms. Quantitative studies of fungal-mediated microplastic biodegradation report mean degradation efficiencies of ~7.5% after 50 days, with mass losses of ~23.8% after 30 days and 35–38% after 90 days.

In December 2024, four samples of bean (Phaseolus vulgaris L.) seeds (500g /sample) were collected from four 0.6- to 1.2-ha field crops, in the Regional Unit of Florina (Western Macedonia, Greece) during the harvesting season by the local Phytosanitary Inspectors in the frame of the annual National Survey Program, and sent to the Laboratory of Bacteriology of the Benaki Phytopathological Institute for examination for the quarantine bacterium Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff). No symptoms of Cff, the cause of bacterial wilt in several species of the Fabaceae family (Osdaghi et al., 2020), had been observed on these crops or seeds. Each seed sample (500g or 1000 seeds) was soaked overnight (4 o C) in 2.5 x Thousand Seed Weight (in mL) sterile buffer (10 mM PB with 0.02 % v/v Tween 20), i.e. 1250 mL. An aliquot of 50 ml of each sample seed extract was used for DNA extraction, using the DNeasy mericon Food kit (Qiagen, Germany). Cff was detected in the DNA extract of one of the four tested seed samples by two qPCR assays, using: a) CffF3/CffB3 primers (Tegli et al. 2022), and b) Cff1-F/Cff1-R primers and Cff1-P probe (Venneman and Van Vaerenbergh 2024). Yellowish colonies were isolated on modified King's B medium (Volkers et al., 2025) from this positive sample, and one of these isolates was identified as Cff by qPCR (Tegli et al. 2022) and indirect immunofluorescence (IF) assay with a Cff-specific polyclonal antiserum (PRI, Netherlands). Cff was not isolated from the other three seed samples. The pathogenicity of the Cff isolate was confirmed via inoculation of five bean (cv. Borlotto) seedlings at the stage of two trifoliate leaves. A 10 μl drop of the isolate’s suspension (10 8 cfu/ml) was inserted by needle prick on the node of the first leaves to introduce the inoculum in the vascular strand of the stem. After 7 days of incubation at 25-28 o C/14 h daylight, the plants exhibited leaf flaccidity and wilt, typical symptoms of Cff infection. The pathogen was re-isolated from the main vein of a symptomatic leaf of an inoculated plant, and its identification was confirmed by IF and qPCR (Tegli et al. 2022), as before, fulfilling Koch’s postulates. The Cff strain NCPPB 559 and sterile distilled water were used as positive and negative controls respectively. Furthermore, partial sequencing (Eurofins, Austria) of the 16S rDNA, gyrB and atpD genes of the same isolate was performed using the primer pairs: pA/pH (EPPO 2021), 2F/6R (Richert et al. 2005) and aptD2F/aptD2R (Jacques et al. 2012), respectively. The obtained sequences for the 16S rDNA (1455 bp, PV770169), gyrB (897 bp, PV779358) and atpD (1016 bp, PV779359) genes were 100%, 98.77% and 99,70%, respectively, identical to those of the Cff type strain CFBP 3418 (NCPPB 1446; CP045852.1) and were deposited in the GenBank. This is the first confirmed report of the presence of Cff in Greece. This finding is added to three other recent Cff outbreaks reported in European countries: the Netherlands, Belgium and Switzerland (Volkers et al. 2025), and underlines the importance of seeds as a means of Cff introduction in an area. Currently official phytosanitary measures have been imposed in the affected area to eradicate Cff, whereas a wider monitoring of bean crops is ongoing to assess any possible Cff spread in Florina, which is one of the main bean-producing Regional Units in Greece, and the produced beans are registered as a ‘Protected Geographical Indication’ product.

Fungal pathogens pose a major threat to global food production, causing substantial yield losses and increasing the dependency on chemical controls. Advanced digital tools, incorporating Artificial Intelligence (AI) driven multi-spectral imaging, support phytopathologists in improving early disease diagnostics through rapid, scalable, and non-destructive methods. This study evaluates a dual AI-based diagnostic framework combining multi-spectral image segmentation with molecular validation to detect B. cinerea during early, asymptomatic stages. A Deep Learning (DL) segmentation model, derived from U-Net++ integrating a lightweight MobileViTV2-1.25 encoder and an augmentation technique for minority classes, was deployed on a newly captured dataset of 80 detached cucumber leaves, obtained from B. cinerea infected as well as control plants. The leaves were imaged at five discrete wavelengths and also subjected to quantitative PCR (qPCR) to measure fungal biomass as a biological ground truth. The model classified leaf regions into healthy, invisible and visible classes. The capability of the approach to detect grey mould was assessed by comparing the DL segmentation outputs with qPCR-based quantification of fungal colonization. Results confirmed the ability of the model to detect infection in tissues where symptoms were either minimal or invisible to the naked eye, achieving an overall segmentation accuracy of 91.9% and a mean IoU of 0.816. This integration of AI-based tools with molecular qPCR benchmarking and confirmation offers a robust methodology that complements phytopathological diagnostics, enabling more informed, early intervention and supporting sustainable crop protection strategies.

Sensitivity assessment of 300 Cercospora beticola isolates collected from North Greece revealed that 38% of the population was highly resistant to at least one of the demethylase inhibitors (DMIs) difenoconazole, epoxiconazole and flutriafol. Resistance factors greater than 50, 100 and 100 were calculated for the most resistant C. beticola isolates to flutriafol, epoxiconazole and difenoconazole, respectively. DMI-resistant isolates carried fitness penalties in terms of mycelial growth, sporulation and pathogenicity, probably accounting for their low frequencies in the field. Pearson correlation analysis revealed a positive cross-resistance relationship between all three DMIs tested suggesting the existence of a target-gene (CbCyp51) related resistance mechanism in the observed phenotypes. Sequence analysis of the CbCyp51 gene from sensitive and DMI-resistant isolates revealed a novel mutation (E149K) present only in C. beticola isolates resistant to all three fungicides tested. To the best of the author's knowledge, this is the first report of this target-site mutation associated with high resistance levels to all flutriafol, epoxiconazole and difenoconazole fungicides. No target-site mutations were found in the rest of DMI-resistance phenotypes. A compound-specific induced CbCyp51 overexpression was observed in C. beticola isolates with specific resistance to flutriafol, epoxiconazole, and difenoconazole, while no indication of constitutive overexpression was found. Overall, the results of the present study provide insights on the mechanisms responsible for the occurrence of high DMI-resistance in the field and a basis for establishing effective strategies for the control of C. beticola in sugar beet fields in Greece.

Abstract This document provides the conclusions of the pest survey card that was prepared in the context of the EFSA mandate on plant pest surveillance (M‐2020‐0114) at the request of the European Commission. The full pest survey card for the non‐EU isolates of Phytophthora ramorum is published and available online in the EFSA Pest Survey Card gallery at the following link and will be updated whenever new information becomes available: https://efsa.europa.eu/plants/planthealth/monitoring/surveillance/phytophthora-ramorum

Chios mastic gum (CMG) exhibits several pharmacological activities that have been confirmed by numerous studies. These include antibacterial, anti-inflammatory, hypolipidemic, hypoglycemic, antiatheromatic, and benefits for against gastrointestinal disorders. However, studies focusing on CMG’s safety are limited. The aim of the present study is to evaluate the genotoxicity of CMG using the limit test on the mammalian erythrocyte micronucleus assay, in male Wistar rats. CMG was administered by gavage to 5 rats at 2000 mg/kg bw for 3 days, while 5 rats received the vehicle and 5 rats received cyclophosphamide (positive control). Satellite groups of 3 rats were included for the negative control and CMG-treated groups to collect plasma and bone marrow for the chemical analyses. All rats were observed for mortality and clinical signs of toxicity during the dosing period. Rats were euthanatized 20 h after the last treatment, necropsied, and bone marrow was collected for smear preparation. No mortality, clinical signs of toxicity, gross organ pathology, or body weight changes were observed in CMG-treated rats compared to the negative control group. No statistically significant alteration of polychromatic erythrocytes (PCE) and the per-thousand incidences of micronucleated PCE in the bone marrow of CMG-treated rats were observed. Bone marrow exposure to CMG was unambiguously confirmed by the detection of the characteristic CMG triterpenic acids in both bone marrow extract and plasma of CMG-treated rats by UHPLC-HRMS/MS analysis. In conclusion, CMG exhibited no genotoxic effects on bone marrow erythrocytes at the tested limit dose of 2000mg/kg body weight.