Frédéric Giraud

Fusarium head blight and associated mycotoxin occurrence on winter wheat in Luxembourg in 2007/2008.

Fusarium head blight (FHB) is among the major causes of reduced quality in winter wheat and its products. In addition, the causal fungi produce a variety of toxins. A relatively high FHB infection rate in winter wheat was observed in 2007 and 2008 in Luxembourg. A fusariotoxin survey was carried out in 17 different geographical locations. Three groups of Fusarium mycotoxins (trichothecenes A and B and zearalenone) were analysed by a multi-detection HPLC–MS/MS method. Fusarium strains were also investigated by morphological and molecular methods. In addition, questionnaires relating to cultural practices were sent to the farmers managing the 17 fields investigated. FHB prevalence ranged from 0.3 to 65.8% (mean: 8.5%) in 2007 and from 0 to 24.5% (mean: 8.3%) in 2008. Results of morphological and molecular identification showed that the most common species isolated from diseased wheat spikes was F. graminearum (33.1%), followed by F. avenaceum (20.3%) and F. poae (17.8%). The chemical analysis revealed that 75% of the investigated fields were contaminated by deoxynivalenol (DON, range 0–8111 µg/kg). The preceding crop was highly and significantly correlated to the number of grains infected and had a significant impact on disease prevalence (p = 0.025 and 0.017, respectively, Fishers F-test). A trend was found for maize as the preceding crop (p = 0.084, Tukeys test) to predict the amount of DON in the fields. This is the first report on the occurrence of DON and ZON in naturally infected wheat grains sampled from Luxembourg.

Toxin Induction and Protein Extraction from Fusarium spp. Cultures for Proteomic Studies

Fusaria are filamentous fungi able to produce different toxins. Fusarium mycotoxins such as deoxynivalenol, nivalenol, T2, zearelenone, fusaric acid, moniliformin, etc... have adverse effects on both human and animal health and some are considered as pathogenicity factors. Proteomic studies showed to be effective for deciphering toxin production mechanisms (Taylor et al., 2008) as well as for identifying potential pathogenic factors (Paper et al., 2007, Houterman et al., 2007) in Fusaria. It becomes therefore fundamental to establish reliable methods for comparing between proteomic studies in order to rely on true differences found in protein expression among experiments, strains and laboratories. The procedure that will be described should contribute to an increased level of standardization of proteomic procedures by two ways. The filmed protocol is used to increase the level of details that can be described precisely. Moreover, the availability of standardized procedures to process biological replicates should guarantee a higher robustness of data, taking into account also the human factor within the technical reproducibility of the extraction procedure. The protocol described requires 16 days for its completion: fourteen days for cultures and two days for protein extraction (figure 1). Briefly, Fusarium strains are grown on solid media for 4 days; they are then manually fragmented and transferred into a modified toxin inducing media (Jiao et al., 2008) for 10 days. Mycelium is collected by filtration through a Miracloth layer. Grinding is performed in a cold chamber. Different operators performed extraction replicates (n=3) in order to take into account the bias due to technical variations (figure 2). Extraction was based on a SDS/DTT buffer as described in Taylor et al. (2008) with slight modifications. Total protein extraction required a precipitation process of the proteins using Aceton/TCA/DTT buffer overnight and Acetone /DTT washing (figure 3a,3b). Proteins were finally resolubilized in the protein-labelling buffer and quantified. Results of the extraction were visualized on a 1D gel (Figure 4, SDS-PAGE), before proceeding to 2D gels (IEF/SDS-PAGE). The same procedure can be applied for proteomic analyses on other growing media and other filamentous fungi (Miles et al., 2007).

Brown rust disease control in winter wheat: II. Exploring the optimization of fungicide sprays through a decision support system

A decision support system (DSS) involving an approach for predicting wheat leaf rust (WLR) infection and progress based on night weather variables (i.e., air temperature, relative humidity, and rainfall) and a mechanistic model for leaf emergence and development simulation (i.e., PROCULTURE) was tested in order to schedule fungicide time spray for controlling leaf rust progress in wheat fields. Experiments including a single fungicide treatment based upon the DSS along with double and triple treatment were carried out over the 2007–2009 cropping seasons in four representative Luxembourgish wheat field locations. The study showed that the WLR occurrences and severities differed according to the site, cultivar, and year. We also found out that the single fungicide treatment based on the DSS allowed a good protection of the three upper leaves of susceptible cultivars in fields with predominant WLR occurrences. The harvested grain yield was not significantly different from that of the double and triple fungicide-treated plots (P < 0.05). Such results could serve as basis or be coupled to cost-effective and environmentally friendly crop management systems in operational context.

First report of wheat leaf rust in the Grand Duchy of Luxembourg and the progress of its appearance over the 2003-2008 period.

Wheat leaf rust caused by Puccinia triticina Eriks. was identified for the first time in 2000 in the Grand Duchy of Luxembourg on the basis of orange-to-brown, round-to-ovoid, erumpent uredinia (1 to 1.5 mm in diameter) scattered on the upper and lower leaf surfaces and producing orange-brown urediniospores that are subgloboid, approximately 20 μm in diameter, and with up to eight germ pore scattered in thick, echinulate walls. In a second phase, wheat was monitored weekly (starting from Zadoks growth stage 30, pseudo stem erection) during the 2003-2008 cropping seasons for wheat leaf rust. Disease severity (percentage of leaf area with symptoms) was recorded in four, replicated field experiments located in three villages (Diekirch District: Reuler; and Grevenmacher District: Burmerange and Christnach), which are representative of the different agroclimatological zones of Luxembourg. A significant difference in severity was observed between the sites (P < 0.01) and the years (P < 0.05). Over the 6-year period, Burmerange and Reuler consistently showed the highest and lowest disease severity, respectively. In 2003 and 2007, Burmerange (a southern site with the highest average spring temperatures of 13.6 and 14.0°C, respectively) showed the highest disease severity with 66 and 57%, respectively, whereas the lowest severity (<1% for both years) was observed in the north at Reuler (site with the lowest average spring temperatures of 12.0 and 12.4°C, respectively). Christnach, located midway between Reuler and Burmerange, showed an intermediate disease severity with 7% (2003) and 22% (2007). The disease appeared at growth stages 77 (late milk) and 87 (hard dough) in the period 2003-2005, but at an earlier stage (45, boots swollen) for 2006-2008 (P < 0.001). In 2005, low severity was recorded due to a severe drought during May, June, and July. A reason for this earlier appearance of leaf rust occurrences in the two districts may be related to an increase in the average spring temperature (average March to May temperature for Luxembourg was 8.3°C for the 1971-2000 period, 9.5°C for the 2003-2005 period, 9.9°C for the 2006-2008 period, 2007 was exceptional with 11.9°C, P < 0.01). In the past, cereal disease management strategies were oriented toward the control of predominant and yield-reducing diseases such as that caused by Septoria tritici Desm. Because the succession of mild winters and warm springs during the last 5 years allowed the early occurrence and the fast development of wheat leaf rust in the Grand Duchy of Luxembourg, it is advisable to take this disease into account in fungicide application schemes.

First Report of Fusarium Head Blight on Winter Wheat in the Grand Duchy of Luxembourg

Following a comparatively mild winter (1.9°C above average [2000-2007]), Fusarium head blight (FHB) on winter wheat was observed during the 2007 season in 17 sites representing all three districts of Diekirch, Grevenmacher, and Luxembourg. The cultivars encountered were diverse and included Achat, Akteur, Aron, Bussard, Cubus, Enorm, Exclusiv, Flair, Rosario, Tommi, and Urban. The preceding crops were maize (six sites), rapeseed (three sites), and one site each of pea, triticale, winter barley, and winter wheat. Rainfalls recorded during the flowering period (June 1-23, mean June 12 for GS 65) ranged from 13 to 62 (mean 38) mm. An overall prevalence of FHB (percentage of infected spikes) of 8.9 ± 15.5% (mean ± SD) and a severity (percentage of infected grains per spike) of 21.0 ± 17.8% were recorded. A significant difference in FHB severity was observed between the cantons north and south of Luxembourg City, 13.4 ± 13.1% (range 0.01 to 46.4) and 35.1 ± 18.1% (range 6.2 to 61.9), respectively (Man-Whitney, P = 0.027), indicating the importance to take regional specificities such as topoclimatological aspects into account. Maize as a preceding crop resulted in significant higher prevalence of FHB as opposed to the other crops (5.9 ± 1.6% versus 3.3 ± 2.2%, Man-Whitney, P = 0.022).