Marco Beyer

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. graminearum, 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. graminearum, 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.

Quantifying the effects of previous crop, tillage, cultivar and triazole fungicides on the deoxynivalenol content of wheat grain — a review

Deoxynivalenol (DON) is a mycotoxin produced by fungi of the genus Fusarium. Maximum tolerable DON concentrations for grain and selected wheat products were laid down by national and European legislative institutions to protect consumers from health risks associated with the intake of this mycotoxin. Among the factors that can be controlled by growers, previous crop, tillage, wheat cultivar and fungicide regimes were identified as the most important variables for DON contamination of wheat. Beyond the identification of the risk factors, their quantitative impact needs to be known for integrated control strategies. The purpose of this review was to extract information on the magnitude of DON reductions caused by agronomic methods or fungicide applications from recent publications. Efficacies of fungicides containing triazoles as active ingredient differed significantly (P= 0.004) in the studies published recently. This effect was solely based on a poor performance of fenbuconazole. No significant differences between triazole fungicide efficacies were observed after excluding fenbuconazole from the analysis. On average, triazole fungicides applied around the time of wheat anthesis reduced the DON content to 53 ± 4% of the grain obtained from untreated control plots. Planting wheat after crops other than maize reduced the DON content to 33 ± 11% compared to maize as the pre-crop. Using a mouldboard plough lowered the DON content to 33 ± 7% compared to minimal or no tillage plots. Cultivating commercially available moderately susceptible cultivars decreased the DON content down to 24 ± 7% compared to susceptible cultivars. Hence, careful choice of cultivar is currently the most effective agronomic method to decrease DON contamination levels in wheat followed by ploughing, avoiding maize as pre-crop and applying triazole fungicides at wheat anthesis. If DON contents of reference scenarios (for instance long term average DON contamination levels for particular cultural practices and regions) are known, the factors extracted in this review may be useful to estimate how many risk factors need to be avoided to decrease DON contents below the limits given in the actual legal regulations.ZusammenfassungDeoxynivalenol (DON) ist ein von Pilzen der Gattung Fusarium gebildetes Mykotoxin. Gesetzgebende nationale und europäische Institutionen haben maximal zulässige Höchstmengen für DON in Getreide und ausgewählten Weizenprodukten festgelegt, um Verbraucher vor den gesundheitlichen Risiken, die mit der Aufnahme von Mykotoxinen verbunden sind, zu schützen. Unter den Faktoren, die von Getreideproduzenten beeinflusst werden können, wurden die Vorfrucht, die Art der Bodenbearbeitung, die Weizensorte und Fungizidanwendungen als die wichtigsten Einflussfaktoren für die Belastung von Weizen mit DON identifiziert. über die rein qualitative Identifikation der Risikofaktoren hinaus sind Informationen über die quantitativen Auswirkungen der Vermeidung der Risikofaktoren für integrierte Mykotoxinreduktionsstrategien notwendig. Es war das Ziel dieses Übersichtsartikels, den quantitativen Nutzen der Vermeidung von Risikofaktoren und des Fungizideinsatzes auf der Basis der veröffentlichten Literatur zu schätzen. Die Effizienz eingesetzter Triazolfungizide zur Verringerung des DON-Gehaltes im Weizen war in den veröffentlichten Studien signifikant unterschiedlich (P= 0.004). Dieser Effekt basierte auf einer geringen Wirkung von Fenbuconazol. Ein Test ohne Fenbuco-nazol ergab, dass es keine signifikanten Wirksamkeitsunterschiede zwischen den anderen Triazolfungiziden gab. Im Mittel der Studien wurde der DON-Gehalt im Erntegut durch den Einsatz von Triazolfungiziden im Zeitraum um die Weizenblüte auf 53 ± 4% im Vergleich zu unbehandelten Kontrollen gesenkt. Der DON-Gehalt von Weizen, der auf Flächen mit anderen Vorfrüchten als Mais angebaut wurde, lag bei 33 ± 11% der Flächen, bei denen Mais die Vorfrucht war. Durch den Einsatz des Wendepfluges wurde der DON-Gehalt im Mittel der Studien auf 33 ± 7% der Gehalte bei fehlender oder minimaler Bodenbearbeitung gesenkt. Durch den Anbau wenig anfälliger Sorten sank der DON-Gehalt auf bis zu 24 ± 7% im Vergleich zu anfälligen Sorten. Damit ist zurzeit der Anbau wenig anfälliger Sorten die effizienteste pflanzenbauliche Strategie, um DON-Gehalte im Weizen zu senken, gefolgt vom Einsatz des Wendepfluges, der Vermeidung der Vorfrucht Mais und Triazolfungizidapplikationen zur Weizenblüte. Wenn DON-Gehalte aus Referenzszenarien (z.B. langjährige Mittelwerte für bestimmte Anbausysteme und Regionen) bekannt sind, können die hier extrahierten Faktoren nützlich sein, um abzuschätzen, wie viele und welche Risikofaktoren vermieden werden müssen, um die jeweils geltenden Grenzwerte einhalten zu können.

Differences between the succinate dehydrogenase sequences of isopyrazam sensitive Zymoseptoria tritici and insensitive Fusarium graminearum strains

Forty-one Zymoseptoria tritici strains isolated in Luxembourg between 2009 and 2010 were highly sensitive towards the new succinate dehydrogenase inhibitor (SDHI) isopyrazam, with concentrations inhibiting fungal growth by 50% (EC50) ranging from 0.0281 to 4.53μM, whereas 41 Fusarium graminearum strains isolated in Europe and Northern America between 1969 and 2009 were insensitive with the average rate of inhibition converging towards 28% with increasing isopyrazam concentration. Seven isolates of both species covering the range of isopyrazam sensitivities observed in the present study were selected for the sequencing of the subunits B, C and D of the succinate dehydrogenase (sdh) gene. Predicted sdh amino acid sequences of subunits B, C and D were identical among F. graminearum strains. By comparing with fungal strains where resistance towards SDHIs was previously reported, three variations were unique to F. graminearum; B-D130N located in the iron-sulfur cluster [2Fe-2S], B-A275T located in the [3Fe-4S] cluster and an additional S at amino acid position 83-84 of sdhC, probably modifying structurally the ubiquinone binding site and therefore the biological activity of the fungicide. No variation was found among the Z. tritici strains in subunits B and D. Two variations were observed within the subunit C sequences of Z. tritici strains: C-N33T and C-N34T. The difference in EC50 values between Z. tritici strains with the NN and TT configuration was non-significant at P=0.289. Two outliers in the Z. tritici group with significantly higher EC50 values that were not related to mutations in the sdhB, sdhC, or sdhD were detected. The role of isopyrazam for the control of F. graminearum and Z. tritici in Luxembourg is discussed.

Characterizing Meteorological Scenarios Favorable for Septoria tritici Infections in Wheat and Estimation of Latent Periods

Septoria tritici epidemics were monitored on winter wheat cv. Ritmo between 1995 and 2003 at 8 to 11 locations per year in Northern Germany (area between 53.70 and 54.38°N latitude and 8.83 and 10.88°E longitude) by counting the number of pycnidia on the leaves of plants obtained from plots under natural infection. Furthermore, meteorological data (leaf wetness, temperature, and precipitation) were recorded within the same period at the same locations. Groups of similar meteorological events were identified by hierarchical cluster analysis. The temporal distance of those clusters from the point of time when an increase of more than 70 S. tritici pycnidia was observed per leaf within 1 week was calculated for all epidemiological case studies and meteorological clusters. A cluster with average temperature = 13.62 ± 2.30°C, leaf wetness = 92.39 ± 4.15%, and precipitation = 0.04 ± 0.10 mm per day was consistently observed at 20.35 ± 4.15 days before epidemic outbreaks. This estimate of a latent period was significantly affected by geographic latitude, average temperature during infection, average temperature during the latent period, year, and precipitation, but not by leaf layer and longitude. On average, an increase in temperature during the infection period by 1°C decreased latent periods by 0.95 day. Latent periods were decreased by 0.2 day upon an increase of the average temperature by 1°C during the latent period. Average latent periods decreased by 1.7 days per degree of north latitude. Latent period estimates had lower coefficients of variation than temperature sums accumulated over latent periods. The usefulness of the approach described above for general epidemiology and for increasing fungicide efficacy by improving the timing of applications is discussed.

FcStuA from Fusarium culmorum controls wheat foot and root rot in a toxin dispensable manner.

Fusarium culmorum is one of the most harmful pathogens of durum wheat and is the causal agent of foot and root rot (FRR) disease. F. culmorum produces the mycotoxin deoxynivalenol (DON) that is involved in the pathogenic process. The role of the gene FcStuA, a StuA ortholog protein with an APSES domain sharing 98.5% homology to the FgStuA protein (FGSG10129), was determined by functional characterisation of deletion mutants obtained from two F. culmorum wild-type strains, FcUk99 (a highly pathogenic DON producer) and Fc233B (unable to produce toxin and with a mild pathogenic behavior). The ΔFcStuA mutants originating from both strains showed common phenotypic characters including stunted vegetative growth, loss of hydrophobicity of the mycelium, altered pigmentation, decreased activity of polygalacturonic enzymes and catalases, altered and reduced conidiation, delayed conidial germination patterns and complete loss of pathogenicity towards wheat stem base/root tissue. Glycolytic process efficiency [measured as growth on glucose as sole carbon (C) source] was strongly impaired and growth was partially restored on glutamic acid. Growth on pectin-like sources ranked in between glucose and glutamic acid with the following order (the lowest to the highest growth): beechwood xylan, sugarbeet arabinan, polygalacturonic acid, citrus pectin, apple pectin, potato azogalactan. DON production in the mutants originating from FcUK99 strain was significantly decreased (−95%) in vitro. Moreover, both sets of mutants were unable to colonise non-cereal plant tissues, i.e. apple and tomato fruits and potato tubers. No differences between mutants, ectopic and wild-type strains were observed concerning the level of resistance towards four fungicides belonging to three classes, the demethylase inhibitors epoxiconazole and tebuconzole, the succinate dehydrogenase inhibitor isopyrazam and the cytochrome bc1 inhibitor trifloxystrobin. StuA, given its multiple functions in cell regulation and pathogenicity control, is proposed as a potential target for novel disease management strategies.

A survey on some factors potentially affecting losses of managed honey bee colonies in Luxembourg over the winters 2010/2011 and 2011/2012

Summary The beekeepers of Luxembourg were asked for: 1. the honey bee (Apis mellifera) colony losses they experienced over winter; and 2. aspects of their beekeeping practice in May 2011 and 2012. Results on the magnitude of the winter losses in the managed bee colonies of Luxembourg are reported for the first time, and some factors potentially related to the losses such as varroa control methods, agricultural land use at the level of municipalities and the hive types employed were investigated. The number of managed honey bee colonies lost in Luxembourg was 938 out of 5,580 in winter 2010/11 and 1,172 out of 5,382 in winter 2011/12, corresponding to national loss rates of 16.8% and 21.8%, respectively. Elevated loss rates were observed in the Northern municipalities in both years. The losses reported from apiaries employing the hive types Dadant 10 or Dadant 12 were significantly lower compared to apiaries employing the hive type Deutsch Normal and losses with Dadant 12 were lower compared to hive type Alberti. Varroa control methods were grouped according to similarity concerning timing and treatment combinations. Differences of colony losses (%) between the four largest groups of similar treatment combinations were nonsignificant at P = 0.72, indicating that none of the most common varroa control method combinations reduced colony losses significantly below the average percentage of colonies lost per apiary. Evidence for an effect of the interaction between varroa control methods and hive type as well as for agricultural land use on colony losses is presented and discussed.

Estimating deoxynivalenol contents of wheat samples containing different levels of Fusarium-damaged kernels by diffuse reflectance spectrometry and partial least square regression.

Fusarium head blight is a fungal disease causing yield losses and mycotoxin contamination in wheat and other cereals. Wheat kernels (cultivar Ritmo) were sampled in 2001, 2002, 2003, and 2006 and Fusarium-damaged kernels were separated from sound grain based on visual assessment. Subsequently, grain lots containing 0, 20, 40, 60, 80, and 100% of damaged kernels were compiled. Each lot was split and the spectrometric reflectance (wavelengths 350-2500nm) was measured using subgroup one, while the concentration of the mycotoxin deoxynivalenol (DON) was determined by high-performance liquid chromatography in subgroup two. DON concentrations in batches classified as sound were not significantly different from 0. Estimating DON contents from the percentage of Fusarium-damaged kernels was impeded by vast variability, resulting in a coefficient of determination of 0.49. Using spectrometric data subjected to partial least square regression allowed estimating DON contents with higher accuracy, in particular at elevated percentages of damaged kernels. The coefficient of determination was 0.84 for the relationship between DON contents estimated based on spectrometric data and the DON contents measured. The intercept of a regression line fitted through a plot of estimated versus measured DON contents was 0.89+/-3.61mg/kg. Since intercept+standard error was larger than the actual legal limit (1.25mg DON per kg dry grain in the European Union), the spectrometric procedure was still not precise enough to allow a reliable separation of grain samples with DON contents below 1.25mg/kg from samples with DON contents above the limit. However, spectrometric data also allowed estimating the DON content of the average damaged kernel within a given lot composed of sound and damaged kernels, which is probably the reason for the reduction of the fraction of unexplained variance by 35% compared to the visual approach and illustrates that spectrometric approaches can make a contribution to reducing DON contents of wheat grain.

Correlations between land covers and honey bee colony losses in a country with industrialized and rural regions

High levels of honey bee colony losses were recently reported from Canada, China, Europe, Israel, Turkey and the United States, raising concerns of a global pollinator decline and questioning current land use practices, in particular intense agricultural cropping systems. Sixty-seven crops (data from the years 2010-2012) and 66 mid-term stable land cover classes (data from 2007) were analysed for statistical relationships with the honey bee colony losses experienced over the winters 2010/11-2012/13 in Luxembourg (Western Europe). The area covered by each land cover class, the shortest distance between each land cover class and the respective apiary, the number of plots covered by each land use class and the size of the biggest plot of each land cover class within radii of 2 km and 5 km around 166 apiaries (2010), 184 apiaries (2011) and 188 apiaries (2012) were tested for correlations with honey bee colony losses (% per apiary) experienced in the winter following the season when the crops were grown. Artificial water bodies, open urban areas, large industrial facilities including heavy industry, railways and associated installations, buildings and installations with socio-cultural purpose, camping-, sports-, playgrounds, golf courts, oilseed crops other than oilseed rape like sunflower or linseed, some spring cereals and former forest clearcuts or windthrows were the land cover classes most frequently associated with high honey bee colony losses. Grain maize, mixed forest and mixed coniferous forest were the land cover classes most frequently associated with low honey bee colony losses. The present data suggest that land covers related to transport, industry and leisure may have made a more substantial contribution to winter honey bee colony losses in developed countries than anticipated so far. Recommendations for the positioning of apiaries are discussed.

Evidence for a reversible drought induced shift in the species composition of mycotoxin producing Fusarium head blight pathogens isolated from symptomatic wheat heads.

Fusarium species are fungal plant pathogens producing toxic secondary metabolites such as deoxynivalenol (DON), 15-acetyl-deoxynivalenol (15AcDON) and nivalenol (NIV). In Luxembourg, the Fusarium species composition isolated from symptomatic winter wheat heads was dominated by Fusarium graminearum sensu stricto strains (genetic 15AcDON chemotype) between the years 2009 and 2012, except for 2011, when Fusarium culmorum strains (genetic NIV chemotype) dominated the pathogen complex. Previous reports indicated that F. graminearum sensu stricto (genetic 15AcDON chemotype) was also most frequently isolated from randomly sampled winter wheat kernels including symptomatic as well as asymptomatic kernels in 2007 and 2008. The annual precipitation (average of 10 weather stations scattered across the country) decreased continuously from 924.31mm in 2007 over 917.15mm in 2008, to 843.38mm in 2009, 736.24mm in 2010, and 575.09mm in 2011. In 2012, the annual precipitation increased again to 854.70mm. Hardly any precipitation was recorded around the time of wheat anthesis in the years 2010 and 2011, whereas precipitation levels >50mm within the week preceding anthesis plus the week post anthesis were observed in the other years. The shift to genetic NIV chemotype F. culmorum strains in 2011 was accompanied by a very minor elevation of average NIV contents (2.9ngg(-1)) in the grain. Our data suggest that high NIV levels in Luxembourgish winter wheat are at present rather unlikely, because the indigenous F. culmorum strains with the genetic NIV chemotype seem to be outcompeted under humid in vivo conditions by F. graminearum DON producing strains on the one hand and seem to be inhibited - even though to a lower extent than DON producing strains - under dry in vivo conditions on the other hand.

Postponing First Shoot Topping Reduces Grape Cluster Compactness and Delays Bunch Rot Epidemic

Field trials investigating the impact of the timing of first shoot topping in a vertical shoot positioning (VSP) trellis system on (i) the cluster morphology, (ii) the canopy structure in the cluster-zone, and (iii) the progress of the bunch rot disease severity, were conducted in two white Vitis vinifera (Pinot gris, Riesling) cultivars in the years 2012 and 2013 in Luxembourg. The first shoot topping was carried out at seven different times from one week prior to beginning of flowering (BBCH 57) to four weeks after the end of flowering (BBCH 75–79) at approximately weekly intervals. Late first shoot topping reduced cluster compactness, delayed bunch rot epidemics, and thereby prolonged the potential ripening period compared to the standard treatment, in which the first shoot topping was performed one week postbloom. First shoot topping four weeks postbloom delayed the moment when 5% disease severity was reached up to eleven days and increased total soluble solids by 0.77 to 2.24 Brix compared to the standard treatment, with no significant impact on yield. Postponing the first shoot topping to the latest technically possible moment is recommended to improve grape architecture and health as well as to optimize grape maturity and potential wine quality under cool climate conditions. Late first shoot topping can be implemented in bunch rot control strategies without imposing additional costs and might support the reduction of pesticide use.