EVOLUTIONS OF A BLACK-GRASS POPULATION SUBMITTED TO DIVERSE CROP SYSTEMS.

Black-grass is a common grass weed, widely spread in Northern Europe and also in Belgium. The first case of resistance in Belgium was reported by Eelen et al. (1996). Since then, monitoring showed that resistant black-grass was not confined to restricted areas anymore and that all usually effective modes of action could be subject to resistance issue (Henriet and Maréchal, 2009). There is no report that agrochemical companies will soon bring a new mode of action effective against grasses on the market, in a close future. It is therefore important to preserve the still effective actives by integrating them into global weeding strategies. A long-lasting trial was set up in order to study the evolution of a black-grass population when submitted to diverse crop systems. Several factors were studied such as rotation (quadri-annual-bisannual-monoculture winter wheat), sowing date (standard date or delayed), cultivation (inversion tillage or not) and herbicide treatments. During three years, each time winter wheat occurred in the rotation, each plot gets the same factorial combination (rotation excepted). In untreated plots, black-grass head counting's showed no differences between tillage or not and bisannual or quadri-annual rotation. On the other hand, number of black-grass heads was higher in standard sowing date and monoculture than in delayed sowing date and other rotations, respectively. The general efficacy of the herbicide treatments was decreasing over the years.

Integrated management of wild chamomile (Matricaria chamomilla L.) populations by tillage.

Nowadays, environmental, health and economic concerns encourage reviewing our weed management in agriculture. Integrated pest management is one key element in the development of weed management strategies less dependent from herbicides. To reach this goal, impact of different methods of tillage (Combinations of stubble cultivator and moldboard plow) on biology and dynamic of wild chamomile populations was studied in experimental plots of experimental farm of Gembloux Agro-Bio Tech. In summer 2012, wild chamomile densities were significantly lower in plots tilled with a moldboard plow. The use of a stubble cultivator did not significantly affect M. chamomilla density. In addition, we found higher wheat yields in plowed plots, indicating that the decrease in M. chamomilla densities reduces competition for wheat. These results show well long run impact of plowing and his effect on densities of wild chamomile and the seedbank.

Silky bent grass resistance to herbicides: one year of monitoring in Belgium.

Silky bent grass (Apera spica-venti (L.) P. Beauv.) is a common weed of cereal crops widely spread in Northern and Easthern Europe (Germany, Czech Republic,...), Northern Asia, Siberia and Canada. Up to now, no resistant case has been detected in Belgium but some chemical weeding failures have been observed in Wallonia fields. During summer 2011, 37 seed samples of Apera spica-venti were collected in Wallonia and submitted to resistance tests in controlled conditions. Three modes of action were tested: acetyl coenzyme-A carboxylase inhibitors (pinoxaden and cycloxydim), acetolactate synthase inhibitors (mesosulfuron+iodosulfu-ron, pyroxsulam and sulfometuron) and photosynthesis inhibitors (isoproturon). One susceptible standard population was included in the test in order to validate it and to permit wild populations classification according to "R" rating system developed by Moss et al (2007). Most of populations were susceptible but some populations showed resistance to at least one of the three tested modes of action.

Study of factors influencing the germination of Apera spica-venti(L.) P. Beauv. in controlled conditions.

Apera spica-venti (L.) P. Beauv. is a common weed of cereal crops widely spread in Northern and Eastern Europe, Northern Asia, Siberia and Canada. Given the ability of Apera spica-venti to grow and develop in Wallonia and lack of scientific knowledge of its biology, germination tests were performed on four populations. These laboratory tests were carried on according to the standards of the International Seed Testing Association (ISTA) in order to study three key factors (composition of water solution, presence/absence of pre-chilling and temperature) influencing the germination of the seeds. Germination tests showed that a solution containing 2 g/L of KNO3 and alternating night/day temperatures between 10 degrees C and 30 degrees C favorably influence the germination of Apera spica-venti. These results in controlled conditions are a premise for greenhouse and field tests.

Effect of mechanical weeding on wild chamomile (Matricaria chamomilla L.) populations in winter wheat crop (Triticum aestivum L.).

Currently, economic, agronomic and environmental concerns lead to reduce the use of herbicides. Mechanical weeding can help to reach this objective. Dynamics and biology of wild chamomile (Matricaria chamomilla L.) populations were assessed as well as dynamic of winter wheat (Triticum aestivum L.) for four level of application of a weeder-harrow (0, 1, 2, 3 treatment(s)). After each treatment, an effect of mechanical weeding on wild chamomile density was observed. Density of wild chamomile decreased significantly with intensification of mechanical weeding. A third treatment allowed eliminating late emerged plants.

Dynamic of black-grass populations depending on the sowing time of winter wheat.

Currently, economic, agronomic and environmental concerns, lead to reduce use of herbicides. This reduction can be help by cultural measures like delay of the sowing date. Four sowing dates of winter wheat from 15th of October to 26th of November were tested. Dynamic of black-grass (Alopecurus myosuroides Huds.) populations and their reproduction rate were assessed as well as dynamic of winter wheat for each date. Delay of sowing could significantly reduce reproduction rate of black-grass. It was shown that the emergence rate (pl/m2), but also number of ears per plant and number of seeds per ear of black-grass decreased significantly with the sowing date. This reduction of seeds production already is from sixty per cent for a delay of two weeks sowing.

Crossing abilities of ALS target-site resistant black-grass (Alopecurus myosuroides Huds.).

Black-grass crossings were conducted outdoor for two successive years in a way to determine the percentage of resistant black-grass that can be engendered in one generation, after the introduction of one target-site resistant individual. After germination, plants were genotyped and couples of resistant and susceptible individuals were formed, then planted outdoor and enclosed with a pollen-proof cloth to avoid any external pollen contamination. Seeds were collected and tested in glasshouses with herbicide spraying. DNA analyses were also performed on surviving plants to distinguish hetero- from homozygotes. The resistance level of the progeny seems to be fairly high and Mendelean distribution is generally confirmed. The offspring of self-crossings was also observed to assess the facultative autogamy ability, according to the genotype and the presence of mutant ALS alleles (position W-574).

Black-grass resistance to herbicides: three years of monitoring in Belgium.

Black-grass (Alopecurus myosuroides HUDS.) is a common weed of cereal crops widely spread in Northern Europe. Even if the first Belgian case of resistance was reported in 1996, until now, Belgium was quite spared of this problem and only a few restricted areas were concerned: the Polders, the marshland of the Escaut River and the Fosses-la-Ville region. About 90 seed samples were collected trough the South part of Belgium and in the Polders during July 2006, 2007 and 2008. These populations were tested in greenhouse conditions by spraying plantlets with herbicides of three modes of action. The herbicides used were photosynthesis inhibitor, ACCase inhibitors and ALS inhibitors. Susceptible and resistant standard populations (Rothamsted and Peldon) were included in the test in order to validate it and to permit wild populations classification according to "R" rating system. Populations showed differences of susceptibility to photosynthesis inhibitor, ACCase inhibitors and ALS inhibitors. For each herbicide mode of action, it was possible to find at least one population in each resistance class of the "R" rating system. Furthermore, it appeared that resistance was not confined to restricted areas listed above anymore.

Detection of ACCase target-site resistant Alopecurus myosuroides Huds. (black-grass) in Belgian populations.

Black-grass is a common grass weed, widely spread in Northern Europe and also in Belgium. For ages, it has been an increasing problem in industrial crops, especially winter cereals. The first case of resistance in Belgium was reported in 1996 by Robert Bulcke (Eelen et al., 1996). Yet the resistance mechanism was not specified. Since then, no more information was published about the evolution Belgium, while research continued in the United Kingdom and in France. Moreover, during the last decade, progress in molecular biology allowed to highlight the mechanism of target-site resistance. A simple PCR method allows to detect the mutation conferring resistance to herbicide. After two years of resistance monitoring in Belgium, mostly in the Walloon part, some populations have been clearly identified as highly resistant to ACCase inhibitor. These populations have been tested by molecular biology so as to detect the single nucleotide polymorphism (SNP) involved in this case. The method employed was the Polymerase Chain Reaction Allele Specific Assays (PASA: Délye, 2002a) for the mutation Ile-1781-Leu that confers a target-site resistance to ACCase inhibitors. Those analyses were performed on plant material issued from bioassays, either in glasshouses or in Petri dishes. Leaves have been collected from plants which survived a fenoxaprop-P treatment applied in a glasshouse single dose assay. Seedlings from resistant populations grown in Petri dishes containing either fenoxaprop-P or cycloxydim provided the second type of sample. Ile1781 mutants were discovered within three populations. Each mutant plant was heterozygote. Five of those samples have been sequenced to confirm PASA results and everyone was matching. Moreover, they were all issued from Petri dishes containing cycloxydim, known to be unaffected by enhanced metabolism, confirming that theses populations are indeed target-site resistant.

Treatment influence on herbicide resistance level of Belgian Alopecurus myosuroides populations (black-grass).

Black-grass is a common grass weed, widely spread in Northern Europe and also in Belgium. For ages, it has been an increasing problem in industrial crops, especially winter cereals. Therefore, farmers started to spray herbicide intensively and soon cases of failure occurred for different molecules and different modes of action. Black-grass populations have been tested in greenhouses to assess the influence of an herbicide treatment as to the resistance level regarding three different herbicides: chlortoluron, fenoxaprop-P and mesosulfuron+iodosulfuron. Black-grass seeds were collected in field trials in six locations in Belgium, on individuals which have survived the herbicide treatment. Each population comes from trial plots, measuring 2 meters wide by 5 meters long and characterized by a single or a combination of products. Herbicides sprayed were isoproturon, flufenacet+diflufenican, ACCase inhibitors and ALS inhibitors. Seeds were also collected in the untreated plots. The population present in these last ones corresponds to the former population, before the herbicide selection pressure was applied. In the glasshouse assay, this population was used as the standard population to compare with other populations issued from the same field. The 'R' rating system was set up with this population to assess the evolution of resistance level, year in, year out. Rothamsted and Peldon populations were also included as cross-reference. Each field population presented different behaviours towards herbicide applied in greenhouses and some cases of resistance can be highlighted. Generally, a reduction of treatment efficiency between field and greenhouse results was clearly visible for the whole of studied active ingredients. Indeed, a distribution shift of the populations towards higher resistance classes could be observed. This is particularly remarkable for active ingredients sharing the same mode of action. For example, it has been found that populations already sprayed with fenoxaprop-P on the field showed a higher resistance level to fenoxaprop-P than to mesosulfuron in the greenhouse test.

Influence of the spraying system on fungicides distribution on wheat plants.

Three trials were carried out during springs 2003 and 2004 to compare the distribution of fungicides on the different leaf layers of wheat plants. Mixtures of 1 L/ha of Amistar (SC, 250 g/L of azoxystrobin) and 1 L/ha of Opus (SC, 125 g/L of epoxiconazol) were applied using two experimental sprayers carried by hand and three farmer's sprayers (including a Hardi TwinFlow one). Working pressure, speed, boom length, nozzles, volume of mixture per hectare were specific to each material. One to six days after the treatments, leaf samples were collected at each canopy level and the amount of both active ingredients was determined using gas chromatography with electron capture detection (GC-ECD). The distribution pattern of the fungicides on the different leaf layers was not affected by the spraying system. In the same way, neither the used equipments, nor the mixture volume per hectare, nor the air flow of the Hardi TwinFlow sprayer did not significantly influence the distribution of fungicide.

Fate of epoxiconazole and kresoxim-methyl in wheat according to time of application.

During the 2000-2001 season a field trial was conducted with the aim of quantifying the distribution and persistence of epoxiconazole and kresoxlm-methyl in the different leaf layers of winter wheat plants. In the case of applications before flag leaf emergence, the redistribution of the two active ingredients in the newly formed leaves following the applications was also measured. Allegro (125 g/L epoxiconazole and 125 g/L kresoxim-methyl, SC) was applied at the manufacturer's recommended rate (1 L/ha) either in a single treatment at stages GS32, GS39 and GS59 or in 2, 3 or 4 split applications. Following spraying, leaf samples were collected over time, from each leaf layer, and the two active ingredients were quantified by gas chromatography with electron capture detection (GC-ECD). Fungicide distribution varies according to time of application. A descending gradient through the leaves was observed in the case of application at GS59. When sprayed at stage GS39, on the other hand, the second leaf intercepted more fungicide than the flag leaf. Kresoxim-methyl was found to degrade faster than epoxiconazole. With split treatments, the last spraying appears to be very significant in terms of final fungicide quantities. Redistribution appears possible, especially in the case of epoxiconazole, though in very small quantities.