Validation of a Quick PCR Method Suitable for Direct Sequencing: Identification of Fusarium Fungal Species and Chemotypes for Preventive Approaches in Food Safety

Za razvoj preventivnih mjera koje se provode radi poboljšanja sigurnosti hrane neophodno je odrediti toksičnost plijesni roda Fusarium, pa se sekvenciranjem i genetskom kemotipizacijom pomoću metode lančane reakcije polimerazom određuju vrste ovoga roda plijesni. U radu je predložen brzi protokol za standardni postupak utvrđivanja vrste plijesni sekvenciranjem faktora elongacije 1α i multipleks kemotipizacijom s pomoću gena Tri12. Provedena je i statistička obrada dobivenih podataka. Kao podloga za uzgoj plijesni upotrijebljen je filter papir Miracloth, a DNA je iz plijesni izdvojena pomoću mikrovalova. Ispitano je 75 sojeva plijesni Fusarium culmorum i Fusarium graminearum, te je zaključeno da se ovim postupkom mogu uspješno odrediti vrste roda Fusarium.Species determination by sequencing and PCR genetic chemotyping, used to determine the toxigenic potential of Fusarium strains, is fundamental for developing preventive strategies in food safety. Here we propose and statistically validate a quick protocol for standardizing the procedure of species determination by sequencing of the elongation factor 1-α and multiplex genetic chemotyping using the Tri12 gene, based on fungal growth on Miracloth tissue coupled with microwave extraction. The test was validated on 75 Fusarium culmorum and Fusarium graminearum strains

Ergeben sich Anhaltspunkte für einen Verlust von Biodiversität in der langjährigen Überwachung von Schaderregern?

Die Krefelder Studie zeigte 2017 einen Rückgang der Masse von fliegenden Insekten um etwa 75% innerhalb von 27 Jahren in einem geschützten Gebiet (HALLMANN et al. 2017). Um die Rolle der Landwirtschaft besser zu verstehen, wären Vergleichsdaten von ungeschützten landwirtschaftlichen Flächen hilfreich. Die Richtlinie 2009/128/EH des Europäischen Parlamentes und des Rates verlangt die Überwachung der Schaderreger in den wichtigsten Kulturpflanzen der Mitgliedsstaaten. Diese oft langjährigen Monitoringdaten von Agrarflächen können für Trendanalysen inklusive Tests auf einen potentiellen Verlust von Biodiversität im Bereich der Schädlinge und Krankheiten genutzt werden. Das Luxemburger Monitoring zeigte im Zeitraum 2007-2017 eine zunehmende Rolle von Gelbrost und eine abnehmende Rolle von Braunrost im Winterweizen. Bei Fusarium-Symptomen und Mehltau im Winterweizen sowie der Anzahl von Stängelrüsslern (gefangen mittels Gelbschalen im Winterraps) wurden sehr starke Schwankungen zwischen den Jahren beobachtet, ohne dass ein Trend in Richtung Aussterben einer Art gezeigt werden konnte. Septoria Blattdürre wurde in allen Jahren in hoher Dichte spätestens gegen Ende der Weizensaison gefunden. Die höchste pro Jahr in Luxemburg gefundene Anzahl von Rapsglanzkäfern pro Haupttrieb am Winterraps nahm zwischen 2007 und 2017 geringfügig aber statistisch absicherbar zu. Es wurde eine hohe Dynamik der Schaderreger zwischen den Jahren beobachtet ohne dass ein Verschwinden einer oder mehrerer der überwachten Arten auf den beobachteten Agrarflächen nachgewiesen werden konnte (DAM et al. 2020). Literatur DAM D, PALLEZ-BARTHEL M, EL JARROUDI M, EICKERMANN M, BEYER M (2020): The debate on a loss of biodiversity: can we derive evidence from the monitoring of major plant pests and diseases in major crops? Journal of Plant Diseases and Protection 127: 811-819. https://doi.org/10.1007/s41348-020-00351-9. HALLMANN CA, SORG M, JONGEJANS E, SIEPEL H, HOFLAND N, SCHWAN H, STENMANS W, MÜLLER A, SUMSER H, HÖRREN T, GOULSON D, DE KROON H (2017): More than 75 percent decline over 27 years in total flying insect biomass in protected areas. PLoS One 12:e0185809. https://doi.org/10.1371/journal.pone.0185809

A European Database of Fusarium graminearum and F. culmorum Trichothecene Genotypes

. Fusarium species, particularly Fusarium graminearum and F culmorum, are the main cause of trichothecene type B contamination in cereals. Data on the distribution of Fusarium trichothecene genotypes in cereals in Europe are scattered in time and space. Furthermore, a common core set of related variables (sampling method, host cultivar, previous crop, etc.) that would allow more effective analysis of factors influencing the spatial and temporal population distribution, is lacking. Consequently, based on the available data, it is difficult to identify factors influencing chemotype distribution and spread at the European level. Here we describe the results of a collaborative integrated work which aims (1) to characterize the trichothecene genotypes of strains from three Fusarium species, collected over the period 2000-2013 and (2) to enhance the standardization of epidemiological data collection. Information on host plant, country of origin, sampling location, year of sampling and previous crop of 1147 F graminearurn, 479 F culmorum, and 3 F cortaderiae strains obtained from 17 European countries was compiled and a map of trichothecene type B genotype distribution was plotted for each species. All information on the strains was collected in a freely accessible and updatable database (www.catalogueeu.luxmcc.lu), which will serve as a starting point for epidemiological analysis of potential spatial and temporal trichothecene genotype shifts in Europe. The analysis of the currently available European dataset showed that in F. grarninearum, the predominant genotype was 15-acetyldeoxynivalenol (15-ADON) (82.9%), followed by 3-acetyldeoxynivalenol (3-ADON) (13.6%), and nivalenol (NIV) (3.5%). In F culmorum, the prevalent genotype was 3-ADON (59.9%), while the NIV genotype accounted for the remaining 40.1%. Both, geographical and temporal patterns of trichothecene genotypes distribution were identified.</p

A European Database of Fusarium graminearum and F-culmorum Trichothecene Genotypes

Fusarium species, particularly Fusarium graminearum and F culmorum, are the main cause of trichothecene type B contamination in cereals. Data on the distribution of Fusarium trichothecene genotypes in cereals in Europe are scattered in time and space. Furthermore, a common core set of related variables (sampling method, host cultivar, previous crop, etc.) that would allow more effective analysis of factors influencing the spatial and temporal population distribution, is lacking. Consequently, based on the available data, it is difficult to identify factors influencing chemotype distribution and spread at the European level. Here we describe the results of a collaborative integrated work which aims (1) to characterize the trichothecene genotypes of strains from three Fusarium species, collected over the period 2000-2013 and (2) to enhance the standardization of epidemiological data collection. Information on host plant, country of origin, sampling location, year of sampling and previous crop of 1147 F graminearurn, 479 F culmorum, and 3 F cortaderiae strains obtained from 17 European countries was compiled and a map of trichothecene type B genotype distribution was plotted for each species. All information on the strains was collected in a freely accessible and updatable database (www.catalogueeu.luxmcc.lu), which will serve as a starting point for epidemiological analysis of potential spatial and temporal trichothecene genotype shifts in Europe. The analysis of the currently available European dataset showed that in F. grarninearum, the predominant genotype was 15-acetyldeoxynivalenol (15-ADON) (82.9%), followed by 3-acetyldeoxynivalenol (3-ADON) (13.6%), and nivalenol (NIV) (3.5%). In F culmorum, the prevalent genotype was 3-ADON (59.9%), while the NIV genotype accounted for the remaining 40.1%. Both, geographical and temporal patterns of trichothecene genotypes distribution were identified

Frequency of Deoxynivalenol Concentrations above the Maximum Limit in Raw Winter Wheat Grain during a 12-Year Multi-Site Survey

Mycotoxins such as deoxynivalenol (DON) in wheat grain pose a threat to food and feed safety. Models predicting DON levels mostly require field specific input data that in turn allow predictions for individual fields. To obtain predictions for entire regions, model results from fields commonly have to be aggregated, requiring many model runs and the integration of field specific information. Here, we present a novel approach for predicting the percentage of winter wheat samples with DON levels above the EU maximum legal limit (ML) based on freely available agricultural summary statistics and meteorological data for an entire region using case study data from Luxembourg and Switzerland. The coefficient of variation of the rainfall data recorded ±7 days around wheat anthesis and the percentage of fields with a previous crop of maize were used to predict the countrywide percentage of winter wheat grain samples with DON levels &gt; ML. The relationships found in the present study allow for a better assessment of the risk of obtaining winter wheat samples with DON contaminations &gt; ML for an entire region based on predictors that are freely available in agricultural summary statistics and meteorological data

Yellow rust does not like cold winters. But how to find out which temperature and time frames could be decisive in vivo?

Yellow rust epidemics caused by Puccinia striiformis f. sp. tritici were monitored in winter wheat grown without fungicides at four locations over the years 2010–2016 in the Grand Duchy of Luxembourg (GDL) and were observed at increased frequency since 2014. A total of 29 field case studies were subdivided into epidemic and non-epidemic cases based on the control threshold of the disease defined in the framework of integrated pest management (IPM). Significant air temperature differences were found between the time courses of epidemic and non-epidemic cases during seven periods and seven individual days. The longest periods with significantly higher temperatures for epidemic cases were found between 21 and 28 days after sowing (DAS) and between 132 and 134 DAS, corresponding approximately to the time of winter wheat emergence, when the disease may infect the newly sown crop, and to the coldest period of the year, respectively. Average daily temperatures were 7.33 ± 0.32 °C and 10.79 ± 0.26 °C between 21 and 28 DAS for non-epidemic and epidemic cases, respectively. Between 132 and 134 DAS, average daily temperatures were − 1.62 ± 0.74 °C and 1.58 ± 0.43 °C for non-epidemic and epidemic cases, respectively. Based on the significant temperature differences detected, up to 86.7% of correct classifications were obtained by leave-one out cross-validation, suggesting that some of the temperature differences identified here have considerable prognostic value for forecasting if an economically relevant yellow rust epidemic must be expected or not.Sentinell

The debate on a loss of biodiversity: can we derive evidence from the monitoring of major plant pests and diseases in major crops?

The European commission directive EC 128/2009 calls for monitoring pests and pathogens of major crops. The monitoring data may be analysed for trends over time, including tests for a potential loss of biodiversity in the domain of plant pests and pathogens. The monitoring programs carried out in Luxembourg since 2007 provided evidence for an increasing role of yellow rust and a decreasing role of brown rust on winter wheat. Vast inter-annual variability was observed at the level of Fusarium head blight and mildew symptoms on winter wheat as well as at the level of Ceutorhynchus counts in oilseed rape, but no trend towards extinction could be demonstrated. Septoria leaf blotch was present in winter wheat at high levels towards the end of all seasons. The maximum number of Brassicogethes aeneus individuals found per main stem and season on oilseed rape increased slightly but significantly between 2007 and 2017. Substantial evidence for highly dynamic changes in the pest populations was found, but no evidence for the vanishing of the monitored species could be demonstrated.Sentinell

Enhancing septoria leaf blotch forecasts in winter wheat I: the efect of temperature on the temporal distance between critical rainfall periods and the breaking of the control threshold

peer reviewedIn integrated pest management (IPM), pests are controlled when the costs of control correspond with the damage caused by a pest on a monetary scale, implying that low pest levels are left uncontrolled. Several forecast models have been developed in plant pathology to warn farmers before an epidemic occurs to allow timely control. Most of these models do not predict a control threshold (pest level at which action needs to be taken to prevent economic losses at the farm level) directly making an application in precision agriculture where pesticides and other inputs shall be used precisely where and when they are needed, difcult. Here, we quantifed the temporal distance between critical rainfall periods and the breaking of the control threshold of Z. tritici on winter wheat, as afected by temperature based on data from 52 feld experiments carried out in Luxembourg between 2005 and 2016. The highest frequency of hours with rain (≥0.1 mm/h) was observed approximately at 300 h before epidemic outbreaks at about 13 °C, at 350 h at 11.5 °C and at about 475 h at about 7.5 °C. A Q10 value of 2.8 was estimated. The knowledge generated here will be used to construct a model that directly forecasts the time at which the control threshold will be reached and thus, when fungicide use is needed according to the standards of IPM with direct applicability in precision agriculture.Sentinell