Perennial energy grasses have gained attention in recent years as a promising resource for the bioeconomy because of their benign environmental profile, high stress tolerance, high biomass yields and low input requirements. Currently, strong breeding efforts are being made to extend the range of commercially available miscanthus and switchgrass genotypes. In order to foster farmers’ acceptance of these crops, and especially of novel hybrids, more information is required about how they can be efficiently integrated into cropping rotations, how they can be removed at the end of their productive lifespan, and what effect they have on subsequently grown crops. Farmers in Europe are meanwhile increasingly constrained in the methods available to them to remove these crops, and there is a risk that the herbicide glyphosate, which has been used in many studies to remove them, will be banned in coming years. This study looks at the removal of  seven-year-old stands of miscanthus and switchgrass over one year at an experimental site in Southern-Germany. Three novel miscanthus genotypes were studied, alongside one variety of switchgrass, and the impact of each crop’s removal on the yield of maize grown as a follow-on crop was examined.  A combination of soil tillage and grass herbicides for maize cultivation was successful in controlling miscanthus regrowth, such that yields of maize grown after miscanthus did not differ significantly from yields of maize grown in monoculture rotation (18.1 t dry biomass ha^-1). Yields of maize grown after switchgrass (14.4 t dry biomass ha^-1) were significantly lower than maize in monoculture rotation caused by insufficient control of switchgrass regrowth by the applied maize herbicide. Although some regrowth of miscanthus and switchgrass was observed in the follow-on crop maize, complete eradication of both crops was achieved by subsequent winter wheat cultivation.

In order to test the effect of miscanthus and switchgrass removal on subsequent maize cropping, a field trial including three different miscanthus hybrids, switchgrass and maize was removed and the land resown with an annual crop, maize. The field trial was located at Ihinger Hof, a research station belonging to the University of Hohenheim in Southern-Germany. The field trial was established in 2013 as part of the EU project ‘OPTIMISC’ (Grant Agreement no. 289159) to assess the methane yield potential of three miscanthus hybrids and switchgrass by anaerobic digestion. Miscanthus and switchgrass were established at a planting density of 2 and 6 plants m^-2 respectively and grown alongside maize (annually sown at 10 seeds m^-2) in a randomized split-block design with 4 replicates (plot size 5 m x 15 m) of each genotype/crop. Miscanthus and switchgrass were harvested in three different harvest regimes (double cut in July and October, early single cut in August and late single cut in October). In the original field trial (2013-2018), the miscanthus and switchgrass plots were split according to the harvest regime (split plot size 5 m x 5 m). To avoid the harvest date treatment impacting the research performed in this study, the crop was allowed two seasons to regenerate in 2018 and 2019. All miscanthus and switchgrass plots were harvested in late October each year, resulting in uniform and vigorous crops by the end of 2019 growing season. Maize cultivation was continued in the plots from the initial experiment so that maize was continually grown on these plots since 2014. In 2019 all miscanthus, switchgrass, and maize plots were fertilized the day after sowing with 150 kg N (ha a)^-1 given as calcium ammonium nitrate. After the final green harvest in late October 2019, the field was cultivated by ploughing on November 22^nd 2019 to a depth of approx. 22 cm. In Table 1, the crop treatments in the initial field trial and the follow-on crop maize established after recultivation of the field are briefly characterized.

In April 2020 all plots were power-harrowed and maize ‘Kilomeris’ (KWS SAAT SE & Co. KGaA, Einbeck, Germany) was sown on April 27^th 2020 at a density of 9.3 seeds m^-2 with a row distance of 75 cm. Each plot of the recultivated field trial was sown with 6 rows of maize, resulting in 4.5 m x 15m maize plots. The crop protection strategy included two herbicide applications: The first treatment was carried out one week after sowing by application of a tank-mixture of soil active herbicides pendimethalin (1,365 g ha^-1, Stomp Aqua®, BASF SE, Ludwigshafen, Germany) and Dimethenamid-P (900 g ha^-1, Spektrum®, BASF SE, Ludwigshafen, Germany).  Both soil active herbicides are a standard treatment in maize cultivation and are known to also be suitable for miscanthus cultivation. For this reason, volunteer miscanthus and switchgrass regrowth appeared and needed to be suppressed with a second application of a leaf-active herbicide on May 26^th 2020 containing 63 g ha^-1 Foramsulfuron, 2 g ha^-1 Iodosulfuron, 20 g ha^-1 Thiencarbazone and 30 g ha^-1 Cyprosulfamide (MaisTer® power, Bayer Crop Science Deutschland GmbH, Monheim, Germany). For fertilization, 198 kg N ha^-1 was applied as stabilised Urea including Dicyandiamid und 1H-1,2,4 Triazol as a nitrification inhibitor (Alzon 46, SKW Stickstoffwerke Piesteritz GmbH, Wittenberg, Germany) in one application directly after sowing.

Physical measurements of crop growth

To assess the development of the maize crop, plant measurements were taken during the vegetation period on May 15^th 2020, June 5^th 2020 and August 6^th 2020. The measurements included number of maize plants m^-2, volunteer miscanthus and switchgrass shoots m^-2, and average canopy height of the maize crop and volunteer miscanthus and switchgrass regrowth. The number of maize plants and miscanthus and switchgrass shoots was measured in the centre of the maize plot over an area of approx. 1 m² (0.75 m x 1.33 m). The average canopy height of maize and volunteer miscanthus and switchgrass was estimated by using a measuring stick and was measured from the ground to the highest fully developed leaf. During these plant measurements the plots were also checked for volunteer growth of other weeds (i.e. neither miscanthus nor switchgrass), but due to the effective chemical weed management no significant quantity of weeds was found.

Weather data were collected from a weather station managed by the LTZ located at Ihinger Hof.

Yield and mineral offtake data collection
Two methods were used to assess the yield of maize and regrowing miscanthus and switchgrass.

Total harvestable fresh biomass yield was estimated on September 17^th 2020 using a BAURAL SF 2000 (Zürn Harvesting GmbH & Co. KG, Schöntal-Westernhausen, Germany) self-propelled plot forage harvester. The third and fourth maize rows, in the centre of the plot, were harvested at a cutting height of approx. 20 cm and along 13.6 m stretch resulting in a sampling area of 20.4 m². The harvested biomass was weighed during harvest by the harvester to assess the fresh biomass yield and a sample was taken for dry matter content analysis. Volunteer miscanthus and switchgrass biomass was not removed prior to harvesting with the forage harvester, but due to poor regrowth, their contribution to the overall yield was only minor. Yield calculated from this harvest was called harvestable yield, and reflected yields as they would be achieved in praxis.

A second yield determination was carried out by harvesting maize plants and regrowth by hand. A 1m² area (0.75 m x 1.33 m) was harvested manually in the fifth maize row at a cutting height of approx. 5 cm. During the hand harvest, maize and volunteer miscanthus and switchgrass biomass was collected separately and the fresh weight of the biomass samples was assessed by weighing on a scale. After weighing, the fresh biomass was chipped using a laboratory chipper and a representative sample was taken for further chemical analysis. This was called potential yield as it represented the amount of biomass that could potentially be taken from the field. Potential yield was calculated for maize and for switchgrass and miscanthus regrowth.

In both harvests, biomass samples were dried at 80 °C to a constant weight in a drying cupboard. The dry weight was then used to calculate the dry matter yield. Winter wheat was sown following the maize harvest, though data about the yield and growth of this crop was not collected.

Chemical analysis of harvested biomass

The dried subsample of pure maize, miscanthus or switchgrass was then milled in a cutting mill SM 200 (Retsch, Haan, Germany) using a 1-mm sieve. The milled subsample was then used for analysis of ash, nitrogen (N), phosphorus (P), potassium (K), magnesium (Mg) and calcium (Ca) content. The nitrogen content was analyzed using a Vario Macro Cube (Elementar Analysensysteme GmbH, Hanau, Germany) in which the samples are completely incinerated with oxygen and content of nitrogen compounds in the resulting off-gas was measured by a thermal conductivity detector. The P, K, Mg and Ca content was measured by microwave digestion of the biomass, followed by analysis of the extract using ICP-OES. For this analysis, 0.5 g of each sample was diluted with 8 ml HNO₃ and 6 ml H₂O and digested in an ETHOS.lab microwave (MLS GmbH, Leutkirch, Germany). The ICP-OES analysis of the extract was performed by Core Facility Hohenheim, which is the central laboratory of the University of Hohenheim.

Soil sampling

Soil samples were taken on four dates to examine the nutrient dynamics in the soil during the vegetation periods. On the first date, April 15^th, multiple samples were taken from the field and combined to use as a baseline for further calculations. Samples were then taken on May 18^th, June 24^th and after the harvest on September 22^nd 2020. Soil samples were taken with an auger to a depth of 90 cm and divided into three fractions (0-30 cm; 30-60 cm; 60-90cm) which were then analysed separately. Directly after extraction from the soil, the soil samples were cooled to avoid ammonium losses and stored frozen until analysis. Following extraction stones and other debris were removed from soil samples.

Plant available nitrogen in fresh soil (NO₃ and NH₄; referred to as N_min) was determined using a CaCl₂ extraction and FIA (flow-injection analysis) measurement (DIN ISO 14255:1998-11). The amount of plant available phosphorus and potassium was then analysed in dried soil via CAL-extraction followed by measurement with a flame photometer or FIA respectively (OENORM L 1087:2012-12-01).  Soil pH was determined with a glass electrode after CaCl2 extraction (DIN ISO 10390:2005).

Nitrogen balance in the soil
Soil samples taken from the beginning of the vegetation period until after maize was harvested were used to calculate a nitrogen balance. A mixed probe was used for the first value, hence for all treatments initial soil nitrogen is given as 18.15 kg ha^-1. Mineral offtake calculated from harvest data was compared with measured soil nitrogen. 

Statistical analysis

Statistical analysis was conducted in R (R Core Team 2020) and RStudio (Version 2022.02.3). The program ‘agricolae’ was used to perform an ANOVA and Tukey’s test using the following model.

Where:             y = yield of maize or regrowth of miscanthus or switchgrass

μ = general mean effect

prev = effect of the previous crop

B = effect of block

   = residual error

Homogeneity of variance was assessed visually. The effects were tested at a level of probability of α = 0.05.

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