Effects of free air CO2 enrichment on root growth of barley, sugar beet and wheat grown in a rotation under different nitrogen supply

Europ. J. Agronomy 63 (2015) 36–46

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Effects of free air CO2 enrichment on root growth of barley, sugar beet and wheat grown in a rotation under different nitrogen supply Andreas Pacholski ∗ , Remigius Manderscheid, Hans-Joachim Weigel Institute of Biodiversity, The Thünen Institute, German Federal Research Institute for Rural Areas, Fisheries and Forestry, Bundesallee 50, D-38116 Braunschweig, Germany

a r t i c l e

i n f o

Article history: Received 1 February 2014 Received in revised form 27 October 2014 Accepted 28 October 2014 Keywords: Free air carbon dioxide enrichment Winter wheat Winter barley Root dry weight Root length density Shoot root ratio CN ratio N supply

a b s t r a c t Elevated atmospheric CO2 concentrations [CO2 ] are known to change plant growth by stimulation of C3 photosynthesis and by reduction of transpiration of both C3 and C4 crops. In comparison to the information on above ground plant responses only limited knowledge exists on the response of root growth of arable crops to elevated [CO2 ] which is particularly true for temperate crop species under real field conditions. A free air CO2 enrichment (FACE) study (550 ppm at daylight hours) was carried out in a crop rotation of winter barley, sugar beet and winter wheat repeated twice in the course of six years on a sandy loam soil at Braunschweig, Northern Germany. Winter barley and sugar beet were included for the first time in a FACE study. A possible interaction with restricted nitrogen (N) supply was studied by fertilizing the CO2 treatment plots with adequate and 50% of adequate N supply. Fine root samples were taken in the plough layer and below at 3–4 sampling dates during the vegetation period and root dry matter (excluding sugar beet storage root), shoot root ratio, root length density, specific root length and root tissue composition (CN ratio) were determined. Main effects of elevated [CO2 ] on the investigated variables were slightly significant. Significant CO2 effects were observed in interaction with the sampling date. In most cases elevated [CO2 ] increased root dry matter early in the vegetation period with a maximum growth stimulation of up to 54% as compared to ambient [CO2 ]. Concomitantly, root length densities were increased in both winter wheat and sugar beet. For winter barley also a significant decrease in root dry weight and significant increase of shoot root ratio was detected at final harvest while such an effect was not significant for sugar beet. Specific root length as an indicator of root morphology was mainly influenced by crop species. As a result, there was no consistent overall effect of elevated [CO2 ] on biomass partitioning in this study as changes in shoot root ratio only occurred at specific sampling dates indicating a similar stimulation of roots and above-ground biomass due to elevated [CO2 ]. Nitrogen supply did not alter the effect of elevated [CO2 ] on any of the root variables apart from CN ratios. A significant increase of root CN ratios in wheat and sugar beet was observed under elevated [CO2 ], but this effect was much smaller than the effect of N supply. © 2014 Elsevier B.V. All rights reserved.

1. Introduction The rapidly increasing atmospheric CO2 concentrations [CO2 ] are considered as the key driver of global climate change and an increase of [CO2 ] to about 550 ppm is expected for the middle of the

∗ Corresponding author. Present address: Innovation-Incubator – Graduate School, Associated to Institute of Sustainable and Environmental Chemistry, Leuphana-University Lüneburg, Scharnhorststr. 1, Lüneburg 21335, Germany. Tel: +49 4131 6772906; fax: +49 4131 6772411. E-mail addresses: [email protected] (A. Pacholski), [email protected] (R. Manderscheid), [email protected] (H.-J. Weigel). http://dx.doi.org/10.1016/j.eja.2014.10.005 1161-0301/© 2014 Elsevier B.V. All rights reserved.

21st century (Meehl et al., 2007). Apart from its physical effect on the global radiation balance and thus global temperature, elevated [CO2 ] exerts direct effects on plant and crop growth. Photosynthesis and consequently growth of C3 crop species are known to be positively affected by elevated [CO2 ] which mostly goes along with modified water and nutrient turnover in the plants (Ainsworth and Long, 2005; Kimball et al., 2002). For C4 crops beneficial growth effects of elevated [CO2 ] seem to result solely from an improved water economy of the plants (Leakey et al., 2009; Manderscheid et al., 2014). Driven by concerns about potential effects of climate change on global food security most studies on effects of elevated [CO2 ] have focussed on above-ground biomass growth and yields of agricultural crops with the majority of studies conducted under artificial growth conditions of e.g. growth chambers, field

A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

tunnels and greenhouses mostly using potted-plants (Ainsworth and McGrath, 2010). However, the limitations of such experiments for realistic assessments of the size of elevated [CO2 ] effects have recently been discussed (Leakey et al., 2006; Long et al., 2006). Free air CO2 enrichment (FACE) techniques have thus been developed (Hendrey and Miglietta, 2006; McLeod and Long, 1999) and applied in experiments with important staple crops (wheat, rice, soybean, barley, sugar beet) in China, Japan, USA and Germany (Kim et al., 2003; Kimball et al., 2002; Long et al., 2005; Ma et al., 2007; Weigel and Manderscheid, 2012). FACE systems leave the natural environment of a crop plot nearly fully undisturbed during the CO2 exposure and allow the investigation of in-situ water and nutrient turnover at the agro-ecosystem level. Collectively, these FACE studies point to lower crop growth and yield stimulations by elevated CO2 levels as compared to chamber studies (Long et al., 2006). In contrast to the amount of information available on effects of elevated [CO2 ] on above-ground crop growth, effects on the soil root systems have been studied less intensively (Kimball et al., 2002; Madhu and Hatfield, 2013). According to Kimball et al. (2002) elevated [CO2 ] can result in a stimulation of root growth and root length density in excess of the stimulation of above-ground biomass. From earlier studies (summarized e.g. by Rogers et al., 1994), there is evidence, although highly variable, that elevated [CO2 ] may stimulate total root biomass and lead to changes in root structural characteristics. However, much of this previous information results from studies where root growth was observed under non-natural soil conditions and where measurements were performed only at a single point in time. Such single point measurements do not consider transient changes in carbon allocation pattern between above- and below-ground structures in the course of the plant’s ontogeny (Gregory et al., 1997). For example, root growth of most annual crops shows a strong seasonal dynamics. It has been observed in earlier studies that elevated CO2 may stimulate root growth to a larger extent at the vigorous early vegetative growth stages than in the phase between the beginning of the vegetation period and stem elongation (Madhu and Hatfield, 2013; Pritchard et al., 2006). The few more recent FACE studies with C3 crops where root growth has been assessed were limited to one study with paddy rice (Ma et al., 2007) and two wheat studies (spring wheat, Wall et al., 2006; Wechsung et al., 1999; winter wheat, Ma et al., 2007). For both crops significantly increased root biomass and root growth rates were observed under elevated [CO2 ], with maximum increases of 37% for spring wheat (550 ppm CO2 ), 75% for winter wheat (590 ppm) and 68% for paddy rice (590 ppm), respectively. There is no such field information available for important temperate crops like winter barley or sugar beet. Effects of elevated [CO2 ] on root characteristics such as total root dry matter, root length density or stem-root ratio may have strong repercussions on total crop growth and adaptation mechanisms and on carbon (C) and nutrient turnover in the agro-ecosystem. Moreover, along with an improved soil water status by reduced transpiration under elevated [CO2 ] (Burkart et al., 2011) a deeper and denser root system may further improve water availability for crop growth (Wechsung et al., 1999). Changes in above-ground plant tissue composition, e.g. increased CN ratios in crop residues, have frequently been observed under elevated CO2 conditions (Loladze, 2002, 2014; Taub et al., 2008). If this holds true for crop roots, it may have effects on nitrogen turnover and availability in agro-ecosystems. Therefore, the quantification of root growth and quality under elevated CO2 is of general importance particularly for the modelling of overall climate change effects on agricultural crop production and agro-ecosystem properties. For example, in most models partitioning of carbon between different sink pools is one of the most important features of total crop development (Brisson et al., 2006) which also applies to the distribution of nitrogen (N) within the plant. Effects of elevated [CO2 ]

37

are mostly modified by the nutrient availability to a crop, which is particularly true for nitrogen. A number of studies have shown CO2 × N interactions (Stitt and Krapp, 1999; Kimball et al., 2002) and a common presumption assumes that N deficiency acts as a growth inhibitory factor which may decrease the relative response to elevated [CO2 ] (Ainsworth and Long, 2005). Nitrogen deficits are known to decrease the shoot root ratio (Gastal and Lemaire, 2002) and exploration of the soil can extent faster to deeper soil layers than under sufficient N supply (Svoboda and Haberle, 2006). Hence, it is important to test whether elevated [CO2 ] particularly stimulates root growth under limited N supply as observed in earlier studies (Stulen and den Hertog, 1993). The aim of the present study was to investigate the seasonal fine root growth dynamics of winter barley, sugar beet (excluding storage root) and winter wheat grown in a crop rotation in Central Europe repeated twice over a total period of six years under FACE conditions with adequate and growth limiting N fertilization. We addressed the following questions: (i) to what extent is root growth stimulated by elevated [CO2 ] under the conditions of the crop rotation? (ii) do the crop species behave differently with respect to elevated [CO2 ]?, (iii) how is the seasonal time course of CO2 effects on root growth? and (iv) are relative CO2 effects on root growth particularly evident under limited N supply and are there interactive effects of elevated [CO2 ] and different levels of N supply on functional root variables such as specific root length? To our knowledge this is the first experiment reporting on effects of elevated [CO2 ] on root growth of sugar beet and winter barley under real agronomic growth conditions. With respect to root growth of winter wheat it is the first FACE study under growth conditions of central Europe. 2. Materials and methods 2.1. Experimental site The 22-ha experimental field was located at the Thuenen Institute (TI) in Braunschweig, south-east Lower Saxony, Germany (52◦ 18 N, 10◦ 26 E, 79 m a.s.l.). The soil is a luvisol of a loamy sand texture (69% sand, 24% silt, 7% clay) in the plough horizon. The profile has a depth of about 0.6 m (0–0.3 m plough horizon, 0.3–0.45 m Eb clay eluviation layer, 0.45–0.6 m Bt clay illuviation layer, >0.6–0.7 m CII, secondary parent material). The lower layers, in particular >0.7 m, are characterized by a coarser soil texture (almost pure sand) and are structured by the succession of thin silt/clay layers. The plough layer has a pH of 6.5 and a mean organic matter content of 1.4%. The soil has a volumetric plant available water content of ca. 18% in the plough layer, which decreases slightly with increasing soil depth. Overall, the soil is of low to intermediate fertility and provides a comparatively shallow effective rooting zone (0–0.6 m). 2.2. Crop management The FACE experiment was carried out in a typical North German crop rotation consisting of winter barley, ryegrass as a cover crop, sugar beet and winter wheat (Table 1). The rotation cycle was repeated twice, resulting in two growing seasons for winter barley (Hordeum vulgare cv. ‘Theresa’ 1999/2000 and 2002/2003), sugar beet (Beta vulgaris cv. “Wiebke” 2001 and cv. ‘Impuls’ 2004) and winter wheat (Triticum aestivum cv. “Batis” 2001/2002 and 2004/2005). The winter cereals were spaced in rows of 0.12 m distance while distance between sugar beet rows was 0.45 m. Agricultural management measures were carried out according to local farm practices. Amounts of added mineral nutrients were based on analysis of soil nutrient contents (K, Mg, N, P, S) determined in early

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A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

Table 1 Crop cultivars, crop management and performance of the FACE system in the crop rotation studied in 1999–2005 at Braunschweig, Germany. Year

1999–2000

Crop

Winter barley ‘Teresa’ Rye grass

Seeding/harvest

N-fertilization N100/N50

Precipitation (seeding–harvest)

Irrigation CO2 fumigation

System off Reasons for system off

Date/seeding density

kg N ha−1

Doses

mm

mm

h

h

25.09.1999 (280 kernel/m2 ) 26.06.00 27.07.2000 09.11.00

264/105

(4: slurry autumn, 3 spring)

445

69

2943

985

Temperature, wind

109

44

915

37

System, wind Wind, CO2 conc.

197/99

2001

Sugar beet ‘Wiebke’

10.04.2001 (11 plants/m2 ) 23.10.2001

126/63

2, spring

464

107

2086

38

2001–2002

Winter wheat ‘Batis’

07.11.2001 (360 kernel/m2 ) 31.07.2002

251/114

5, spring

501

60

2445

429

Temperature, wind

2002–2003

Winter barley ‘Theresa’

24.09.2002 (270 kernel/m2 )

179/105

3, spring

478

77

2955

1 112

Rye grass

26.06.2003 20.08.2003 08.11.2003

72/36

206

–

600

34

System (winter), temperature System, wind

2004

Sugar beet ‘Impuls’

14.04.2004 (11 plants/m2 ) 15.10.2004

156/78

2, spring

404

84

2078

20

Temperature

2004–2005

Winter wheat ‘Batis’

26.10.2004 (360 kernel/m2 ) 27.07.2005

168/84

3, spring

387

121

2557

681

Temperature, wind

springtime. Just before the first sowing of winter barley in 1999 the field was fertilized with slurry. Subsequently, only mineral N was used (urea ammonium nitrate or calcium ammonium nitrate) and added on two dates to sugar beet (April and June). Winter cereals were fertilized with N on three to five dates from March until May (barley) or until June (wheat). Additional details of the crop management are presented by Weigel and Manderscheid (2012). In order to avoid interacting effects of elevated [CO2 ] and drought stress, the field was irrigated using a linear irrigation system to keep the soil water content above 50% of maximum plant available soil water content. This was achieved by modelling soil water budget with the AMBAV model (Kersebaum et al., 2005). The model runs were continually controlled and re-parameterized by regularly measuring of the soil water content by soil coring and with time domain reflectometry probes at 0.0–0.4 m depth (probes at 0–0.2 m and 0.2–0.4 m, amounts of irrigation see Table 1).

2.3. FACE system and experimental design A FACE system consisting of 4 rings, each 20 m of diameter, engineered by Brookhaven National Laboratory (Lewin et al., 1992) was operated as described previously (Weigel et al., 2005; Weigel and Manderscheid, 2012). Fumigation treatments included 2 rings equipped with blowers and enriched with CO2 (FACE, pure CO2 mixed with ambient air) and 2 control rings operated with blowers and ambient air (amb). Rings were arranged in 2 rows of 100 m distance between rows and 100 m distance in a row. A FACE ring and an ambient ring were placed opposite to each other between rows. In order to investigate possible interactions between CO2 enrichment and nitrogen supply to the plants nitrogen fertilization was restricted to 50% (N50) of adequate N (N100) in one half of each ring. For ease of N application by conventional fertilization machinery the same N level was established at the same side of the FACE and the ambient ring within a row and kept the same during the two rations. So, arrangement of N level and [CO2 ] level combinations was partly randomized. Each N level and [CO2 ] level

combination was repeated twice (n = 2). Each ring half covered an area of 100 m2 . Plant sampling for crop analyses did not take place within a 2 m distance from the vertical vent pipes for the CO2 release. The target CO2 concentration in the enriched rings was set to 550 ppm, as expected for the middle of the 21st century (Meehl et al., 2007), during daylight hours. No-enrichment criteria for CO2 were wind speeds >6.5 m/s and air temperatures <5 ◦ C. This procedure was used because crop growth is low and the reaction of photosynthesis to increased atmospheric CO2 concentrations is negligible below 5 ◦ C (Long et al., 2005). The target CO2 concentration was controlled by measuring CO2 -concentrations in air samples taken every second at the centre of each experimental ring (just above plant canopy) and continually modifying the amount of CO2 added to the air blown into the rings based on CO2 concentrations and actual wind speed values. During more than 97% of total CO2 fumigation time the CO2 concentration was within <±10% of the target concentration (1 min average values). Cut off times compared to maximum fumigation times (daylight hours) varied between 1 and 38%, mainly in winter due to low temperatures (<5 ◦ C) but also system errors (Table 1). While average daytime CO2 concentrations were increased to 550 ppm in FACE rings mean daytime CO2 concentration in the ambient rings averaged over the six years period amounted to 376 ppm and ranged from 371 to 378 ppm. Total mineral N added to the respective experimental areas, CO2 fumigation times as well as major crop management events are summarized in Table 1.

2.4. Root extraction and root dry weight Sampling was done at 3–4 dates during the vegetation period after winter. Sampling dates and corresponding EC stages are given in Table 2. Root sampling for winter barley could only be carried out in the growing period 1999/2000. Sugar beet was partly affected by rhizomania (beet necrotic yellow vein virus) at later growth stages

A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

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Table 2 Root sampling dates, crop phenological (EC) stages and weather characteristics during the two rotations (1st, 2nd) of the FACE experiment 1999–2005, values in brackets indicate deviations of weather data from long term average values (1961–1990). Crop

Winter barley

Sugar beet

Winter wheat

Temperature (◦ C)

Radiation (MJ/m2 ) Rotation

Root harvest (date/EC stage)

Precipitation (mm)

1st

2nd

1st

2nd

1st

2nd

1st

2nd

20.03.2000 (28) 17.04.2000 (35) 15.05.2000 (69) 22.06.2000 (>92)

–

195 (−29)

–

1849 (+139)

–

12.2 (+2.1)

–

28.06.2001 (38) 21.08.2001 (45) 29.09.2001 (49)

07.07.2004 (38) 17.08.2004 (45) 423 (+73) 28.09.2004 (49)

413 (+62)

2811 (+107)

2905 (+155)

14.6 (+0.5)

14.6 (+0.6)

10.04.2002 (25) 13.05.2002 (34) 19.06.2002 (68)

26.04.2005 (31) 18.05.2005 (39) 468 (+186) 06.06.2005 (64) 29.06.2005 (72)

214 (−68)

2158 (−79)

2439 (+203)

12.7 (+1.2)

12.5 (+1)

in summer 2001. Therefore, only the first sampling date could be used for sugar beet analyzes in 2001 (Table 3). Above ground biomass was harvested in each quarter per ring from a sampling area of ca. 1 m2 (cereals) up to 5 m2 (sugar beet) and immediately afterwards roots were extracted from the same area. While for the cereals above ground biomass was harvested by

chopping the plants directly at the soil surface, in the case of sugar beet this also included the storage root. Root samples were extracted in between the rows but directly besides the point of shoot emergence by means of an Eijkelkamp bi-partite root auger (0.15 m depth, 0.08 m diameter). In sugar beet fine root samples, excluding storage root, were taken from beet

Table 3 Root characteristics in the plough layer (0–0.3 m) of winter barley, sugar beet and winter wheat grown in 1999–2005 without and with elevated [CO2 ] (factor CO2 : amb = 375 ppm and FACE = 550 ppm) and 2N levels (factor N: N100 = sufficient, N50 = 50% of N100), Braunschweig, Germany; mixed effects model,** = significant (p < 0.01), * = significant (p < 0.05), (*) = marginally significant (0.05 < p < 0.1),. = slightly significant (p < 0.2), ns: non-significant. RDW, root dry weight; RLD, root length density; SRR, shoot root ratio; SRL, specific root length; % = difference between amb and FACE treatment. Variable

Date

N50 amb

Winter barley RDW (g m−2 ) SRR Sugar beet RDW (g m−2 ) 2001 2004 SRR 2001 2004

RLD (cm cm−3 )

SRL (m g−1 )

CN ratio

Winter wheat RDW (g m−2 ) SRR RLD (cm cm−3 ) SRL (m g−1 )

CN ratio

N100 FACE

%

amb

Mixed effects model FACE

%

All dates All dates All dates 28.06.2001 All dates All dates 28.06.2001 All dates 07.07.2004 17.08.2004 28.09.2004 All dates 28.06.2001 28.09.2004 All dates 28.06.2001 28.09.2004 All dates 07.07.2004 17.08.2004 28.09.2004 All dates All dates All dates (2002) All dates (2002) 10.04.2002 13.05.2002 19.06.2002 06.06.2005 All dates (2005) 26.04.2005 18.05.2005 06.06.2005 29.06.2005

38.9

53.4

+37

28.4

39.0

+37

11.1

9.0

−19

18.8

13.5

−28

6.7 20.0 27.2

6.9 17.3 38.9

+3 −14 +43

10.8 27.0 32.6

8.3 20.8 39.4

−23 −23 +21

1.8 2.6

2.9 2.3

+61 −12

2.1 2.7

2.3 2.9

+10 +7

135.6 110.0

155.6 116.7

+15 +6

222.2 112.7

186.4 136.1

−16 +21

17.4 15.5 16.2

18.0 18.3 16.8

+3 +18 +4

15.3 17.1 14.5

15.5 15.4 14.8

+1 −10 +2

110.5 119.4 52.9 –

114.0 140.5 35.6 –

+3 +18 −33 –

114.3 113.1 61.0 80.0

119.7 114.2 51.5 70

+5 +1 −16 −13

48.9 49.1 56.0 53.9

49.1 50.1 58.6 55.9

0 +2 +5 +4

42.4 41.2 49.4 44.4

46.3 47.0 56.1 48.9

+9 +14 +14 +10

CO2

N

CO2 × N

date

CO2 × date

. *

(*) *

ns ns

* *

* *

. ns (*) ns . ns ns . . ns ns ns ns ns ns . ns ns ns

. ns * . . ns ** ns ns ns ns (*) (*) ** ns * * . **

ns ns * ns ns ns ns ns ns ns ns ns ns ns ns . ns ns ns

*

(*)

(*) *

ns *

*

*

(*)

ns

*

*

ns

ns

. . . ns ns ns ns – ns ns . ns ns

ns (*) ns ns ns ns ns ns * * * . (*)

ns ns ns ns ns ns ns ns (*) ns ns ns ns

* * * *

. ns * ns

*

ns

40

A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

plants directly neighbouring the sampling area for above ground biomass. Generally root samples were collected in the plough layer (0–0.3 m). At two dates in winter wheat (2004–2005) samples were taken to a depth of 0.6 m. Three samples were extracted at each sampling position and mixed for each sampling depth (0–0.3 m, 0.3–0.6 m). Roots were analyzed separately for each position and mean values for each ring half were used for further statistical analysis per sampling date. Soil samples for root extraction were mixed thoroughly and subsamples of 1 kg were taken for root washing. For determination of root dry weight (RDW, g m−2 ) root samples were removed from debris and dried until constant weight at 75 ◦ C. In order to obtain root dry matter, a subsample was incinerated. The weight of the ash corresponding soil particles adherent to the roots, were subtracted from the weight of dried root samples. Shoot (stems + leaves + ears) root ratio (SRR) was calculated as the fraction of shoot dry weight and RDW extracted at the same sampling dates: SRR =

m−2 )

shoot (g RDW (g m−2 )

2.5. Root length density and specific root length Root length density (RLD, m cm−3 ) and specific root length (SRL, m g−1 ) were determined at one sampling each in 2001 and 2004 in sugar beet and at three samplings in 2002 and one sampling in 2005 in winter wheat. For determination of RLD, extracted roots were stored in ethanol and analyzed by means of root image analysis. Roots were submerged in water in flat transparent plates and pictures were taken by a scanner (Epson Expression 1680, Epson corp. Tokyo, Japan). These pictures were analyzed for root length by means of the WinRHIZO® software (WinRHIZO 2.0, Regent instruments Canada Inc., QC, Canada, 2000). SRL was calculated as the fraction of RLD per unit soil volume root dry weight (RDWv, g cm−3 ): SRL (m g−1 ) =

RLD (m cm−3 ) RDWv (g cm−3 )

RDWv was calculated by dividing RDW (g m−2 ) by soil layer volume (cm3 m−2 ).

Fig. 1. Root dry weight in the plough layer (0–0.3 m) of winter barley cv. Theresa grown in 1999/2000 without and with elevated [CO2 ] (amb 375 ppm, FACE 550 ppm) and 2 N levels (N100 sufficient, N50 50% of N100); Braunschweig, Germany; mixed effects model, n = 2; * = significant (p < 0.05), (*) = marginally significant (0.05 < p < 0.1),. = slightly significant (p < 0.2), ns: non-significant.

linear mixed effects model: y = CO2 × N, random = ∼1|Ring. Heteroscedasticity in the data was considered by weighted least square estimation method implemented in the R nlme package (weights = varIdent (form = ∼1|CO2 × N)) and residuals of each model fits were checked for even distribution. In some cases of temporarily unrepeated measurements two-way ANOVA was applied (n = 2). Because of the high inherent variability of root samples and the low number of independent replicates available in FACE studies statistical tests often yielded no significance at the p < 0.05 level. For highlighting results close to this level of significance, additional thresholds for addressing particular p-levels are used: p ≥ 0.20 non-significant (“ns”), 0.20 > p ≥ 0.10 slightly significant (“.”), 0.10 > p ≥ 0.05 for marginally significant (“(*)”) (see Klironomos et al., 1999, applied also in e.g. Treseder and Allen, 2002; Koerner et al., 2005), 0.05 > p ≥ 0.01 for significant (“*”), significant at p < 0.01 (“**”). 3. Results 3.1. Root dry weight, root length density and specific root length

2.6. Root tissue composition Root total C and N concentrations were determined with a LecoTruspec CN (St. Joseph, MI, USA). Dried root samples were milled to powder (<1 ␮m) by a mixer mill (Retsch MM 400, Retsch GmbH, Haan, Germany) and two subsamples (10 mg) of each milled root sample were analyzed. Data are reported as ratio of C and N (CN ratio). 2.7. Statistics Data and results are either depicted in figures or presented in tables. Mean values were calculated for each ring half as independent replicates. Due to replicated sampling in rings and the sub-plots with N treatments, a statistical analysis for repeated sampling by means of a linear mixed effects model approach was used. The analysis was done using the R software package (R Development Core Team, 2008) applying the nlme package. Data were analyzed covering both experimental years of RDW and SRR of for winter wheat by means of the linear mixed effects model: y = CO2 × N × date × year (fixed effects), random = ∼1|Ring/N (random effects), otherwise without the factor year. Data testing of each sampling date was done by the

For none of the three crops the full model showed significant main effects of [CO2 ] and N on root biomass (RDW), root length density (RLD) and specific root length (SRL) in the plough layer. However, for most of these variables a slightly significant effect (i.e. 0.20 > p ≥ 0.10; see Section 2.5) of elevated [CO2 ] was observed (Table 3). For winter wheat RDW was significantly different between years (not shown). All detected significant effects of the two factors occurred in interaction with sampling date for which a main effect was detected on all of the three variables. A significant interaction effect between sampling date and elevated [CO2 ] on RDW was observed for winter barley (Fig. 1) while the effects were slightly significant for winter wheat and sugar beet (Table 3, Figs. 2 and 3). For none of the crops a significant interaction between [CO2 ] and N supply for RDW was found in the full model. For all crops, at sampling dates when significant differences of RDW between treatments were observed, treatments with elevated [CO2 ] showed in most cases higher RDW as compared to ambient (Figs. 1–3). For winter barley higher RDW under elevated [CO2 ] compared to ambient occurred at early growth stages (Fig. 1) but it was significantly lower at the last harvest. A maximum stimulation was observed at tillering stage under low N supply with (+54%).

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Fig. 2. Root dry weight in the plough layer (0–0.3 m) of winter wheat cv. Batis grown in 2001/2002 and 2004/2005 under ambient and elevated [CO2 ] (amb 375 ppm, FACE 550 ppm) and 2N levels (N100 sufficient, N50 50% of N100); Braunschweig, Germany; mixed effects model, n = 2; . =slightly significant (p < 0.2), ns: non-significant.

At sampling dates with significant differences between treatments in winter wheat, RDW was higher under elevated [CO2 ] (Fig. 2) compared to ambient conditions with a maximum stimulation of 57% at early dough stage in 2002. For the 0.3–0.6 m layer no significant effect of elevated [CO2 ] on RDW was observed (Table 4). In 2001 RDW values of sugar beet were higher under elevated [CO2 ] than under ambient conditions, though not significant. In 2004 a marginally significant effect of elevated [CO2 ] on RDW was detected (Fig. 3, Table 3), there also existed a significant effect of N and a significant interaction of [CO2 ] and N supply (Table 3). The N × CO2 interaction in sugar beet becomes evident by crossing of the respective graphs between the second and the last sampling (Fig. 3). The maximum effect of elevated [CO2 ] was 37% in 2004. Similar to winter barley RDW was lower under elevated [CO2 ] compared to ambient conditions at the last sampling before harvest in 2004. The general statistical model on RLD for winter wheat showed a slightly significant main effect of [CO2 ] and a significant interaction with sampling date in the plough layer (0–0.3 m, Table 3). Overall, higher values were observed for elevated [CO2 ] irrespective of N supply compared to ambient conditions. The most

pronounced effect was observed during stem elongation (EC 34) in 2002 (+39%). In 2005 higher RLD was observed in the deeper layer (0.3–0.6 m) for N100 compared to N50 in interaction with elevated [CO2 ], which reduced RLD (Table 4) compared to ambient conditions. For sugar beet no main effect of elevated [CO2 ] on RLD was found (Table 3). At the sampling in 2001 (EC 45) RLD was higher (+61%) under elevated [CO2 ] compared to ambient conditions, though not significant. In the sampling at beet harvest in 2004 values under elevated [CO2 ] were non-significantly higher (N100) and lower (N50) than values under ambient conditions. For winter wheat no effect of elevated [CO2 ] on SRL was determined at either sampling depth (Tables 3 and 4). In the deeper layer a significant effect of N supply was detected with higher values for N100 compared to N50 (Table 4). For sugar beet also no significant effect of [CO2 ] on SRL was determined although a significant interaction between [CO2 ] and harvest date was observed (Table 3). A marginally significant effect of N supply on SRL of sugar beet was observed for the whole model and was highly significant at one individual sampling, SRL being smaller under low N supply than under N100 (Fig. 4).

Table 4 Root characteristics in 0.3–0.6 m and 0–0.6 m soil depth of winter wheat cv. Batis grown in 2004/05 without and with elevated [CO2 ] (factor CO2 : amb = 375 ppm and FACE = 550 ppm) and 2N levels (factor N: N100 = sufficient, N50 = 50% of N100), Braunschweig, Germany; mixed effects model, * = significant (p < 0.05), (*) = marginally significant (0.05 < p < 0.1),. = slightly significant, ns: non-significant. RDW, root dry weight; RLD, root length density; SRR, shoot root ratio; SRL, specific root length; %, difference between amb and FACE treatment. Variable

Date

N50

RDW 0.3–0.6 m (g m−2 )

All dates 18.05.2005 06.06.2005

12.2 22.6

14.2 25.2

+16 +12

17.9 19.2

RDW 0–0.6 m (g m−2 )

All dates 18.05.2005 06.06.2005

145.7 140.0

141.3 162.4

−3 +16

SRR (0–0.6 m)

All dates 18.05.2005 06.06.2005

3.1 6.0

3.1 5.4

06.06.2005

0.29

0.32

amb

RLD (0.3–0.6 m) (cm cm−3 ) −1

SRL (0.3–0.6 m) (m g CN ratio

)

06.06.2005 All dates 18.05.2005 06.06.2005

40.3 41.1 52.1

N100 FACE

39.7 43.0 51.5

%

Mixed effects model %

CO2

N

CO2 × N

date

CO2 × date

ns ns ns

ns

+64 +5

ns . ns

ns

29.3 20.2

ns ns ns

+16 +30

ns * ns

(*) (*) .

ns

169.6 172.7

ns ns ns

ns

146.5 132.6

3.6 6.2

0 −15

. . ns

ns ns ns

.

3.6 7.3

ns ns ns

*

0 −10 +10

0.40

0.34

−15

ns

*

(*)

−16

ns

*

ns

+22 +24

ns – –

ns – –

ns – –

−1 +5 −1

amb

61.5 34.8 40.0

FACE

51.8 42.3 49.6

42

A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

Fig. 3. Root dry weight in the plough layer (0–0.3 m) of sugar beet cv. Impuls grown in 2004 under ambient and elevated [CO2 ] (amb 375 ppm, FACE 550 ppm) and 2N levels (N100 sufficient, N50 50% of N100); Braunschweig, Germany; mixed effects model, n = 2; * = significant (p < 0.05), · =slightly significant (p < 0.2), ns: nonsignificant.

3.2. Shoot root ratio Significant effects of elevated [CO2 ] on SRR were observed in winter barley (Table 3, Fig. 5). While SRR under elevated [CO2 ] was lower as compared to ambient at the first sampling, SRR was significantly higher at the last sampling date. N supply also had significant effects on SRR, with generally higher values under high N supply compared to N50. At the second sampling date elevated [CO2 ] resulted in higher SRR compared to ambient conditions only under N100. For winter wheat a slightly significant effect of elevated [CO2 ] on SRR was determined (Table 3, Fig. 6) with lower values compared to ambient, in particular in 2002. As for winter barley, a significant effect was observed for the N treatments, with significantly lower SRR values under low N supply compared to N100. Taking also into account the data obtained for the 0.3–0.6 m, the overall model resulted in a slightly significant interaction between harvest date and [CO2 ] (Table 4). At the second sampling date SRR was lower for elevated [CO2 ] compared to ambient conditions while for

Fig. 5. Shoot root ratio between above-ground biomass and root dry matter in the plough layer (0–0.3 m) of winter barley cv. Theresa grown in 1999/2000 under ambient and elevated [CO2 ] (amb 375 ppm, FACE 550 ppm) and 2N levels (N100 sufficient, N50 50% of N100); Braunschweig, Germany; mixed effects model, n = 2; ** = significant (p < 0.01), * = significant (p < 0.05), (*) = marginally significant (0.05 < p < 0.1), ns: non-significant.

both samplings N50 showed smaller SRR values compared to the N100 level. In sugar beet a significant interaction between harvest date and [CO2 ] was determined for SRR in 2004 (Table 3). For most of the samplings SRR was lower for elevated [CO2 ] compared to ambient conditions excluded the last sampling date in 2004 with higher values. A significant effect of N supply with smaller SRR values under low N supply compared to N100 was observed for several sampling dates. For all crops no significant interaction for SRR between [CO2 ] and N was observed, however, there was a trend of a stronger effect of elevated [CO2 ] on SRR under high N supply. 3.3. Root CN ratio While there was no significant effect of harvest date on CN ratios in sugar beet, this effect was very pronounced in winter wheat with widening CN ratios at later samplings (Table 3). For both crops no significant main effect of elevated [CO2 ] was observed with respect to CN ratio, while N-supply was significant. Treatments with lower N supply were characterized by wider CN ratios compared to N100. For winter wheat there existed a marginally significant interaction between elevated [CO2 ] and N supply with higher CN ratios for elevated [CO2 ] treatments compared to ambient conditions under high N supply. Similar trends were observed for winter wheat roots in the 0.3–0.6 m layer (Table 4). Due to small amounts of root material, roots had to be pooled and no statistical testing was possible. An increased CN ratio was also only observed under ample N supply whereas low N supply had a stronger effect on CN ratios as compared to elevated [CO2 ]. 3.4. Comparison of crops

Fig. 4. Root length density in the plough layer (0–0.3 m) of winter wheat cv. Batis grown in 2001/2002 and 2004/2005 under ambient and elevated [CO2 ] (amb 375 ppm, FACE 550 ppm) and 2N levels (N100 sufficient, N50 50% of N100) Braunschweig, Germany; error bars indicate range of values (n = 2). Two-way ANOVA, Treatment with different letters are significantly different at the p < 0.05 level.

The three crops were compared with respect to the four main root variables RDW, RLD, SRR and SRL at crop specific phases in the vegetation period with maximum root expansion (Table 5). The factor crop resulted in a significant effect on all of the variables. Elevated [CO2 ] showed a significant effect on RDW and a slightly significant effect on RLD, stimulating root growth. Shoot root ratio was mainly affected by N supply and SRL by crop species. With respect to CN ratio in the root tissue sugar beet was significantly different to winter wheat and elevated [CO2 ] and N supply

A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

43

Fig. 6. Shoot root ratio between above-ground biomass and root dry matter in the plough layer (0–0.3 m) of winter wheat cv. Batis grown in 2001/2002 and 2004/2005 under ambient and elevated [CO2 ] (CO2 − 375 ppm and CO2 + 550 ppm) and 2N levels (N100 sufficient, N50 50% of N100) Braunschweig, Germany; mixed effects model, n = 2; ** = significant (p < 0.01), * = significant (p < 0.05), ns: non-significant.

showed significant main effects with elevated [CO2 ] and low N supply increasing CN ratio.

4. Discussion A FACE study was carried out to investigate the effects of elevated [CO2 ] and N supply on root growth and dynamics of typical arable crops grown in rotations in Central Europe, winter barley, sugar beet and winter wheat. While knowledge on the stimulating effects of elevated [CO2 ] on above-ground biomass is abundant much less information is available on below ground effects. The study was guided by the overarching questions whether stimulation of root biomass by elevated [CO2 ] deviates from the effect on above-ground biomass and whether effects of elevated [CO2 ] on root growth differ between arable crops depending on N supply. Potential stimulation of root biomass by elevated [CO2 ] can have strong effects on soil C storage, N turnover and water relationships in agro ecosystems.

The general magnitudes of RDW and RLD values in our study were similar as reported in other studies for winter wheat (Ma et al., 2007; Wall et al., 2006; Wechsung et al., 1999) and sugar beet (Brown and Biscoe, 1985; Vamerali et al., 2003). Winter wheat RDW was lower in the study by Ma et al. (2007). In this study, a Chinese winter wheat variety with lower biomass potential and a different root extraction method was used. Similar values compared to literature data were also obtained for winter barley (Lampurlanés et al., 2002; Sidiras et al., 2001). Root growth of the investigated crops responded in a similar manner to elevated [CO2 ]. However, according to the levels of significance applied in this study, the statistical significance of such effects varied. Significant and marginally significant stimulation of RDW of the crops under investigation due to elevated [CO2 ] were observed in interaction with sampling date. Maximum values of relative stimulation of RDW of winter wheat under elevated [CO2 ] (57%) were smaller than the 75% observed in a different winter wheat variety in China (Ma et al., 2007). As reported for other crops (Madhu and Hatfield, 2013; Kimball et al., 2002; Pritchard and

Table 5 Root characteristics of winter barley, sugar beet and winter wheat at maximum root expansion grown in 1999–2005 without and with elevated [CO2 ] (amb = 375 ppm and FACE = 550 ppm) and 2N levels (N100 sufficient, N50 50% of N100), Braunschweig, Germany; data of winter barley and winter wheat include data of two years; mixed effects model, * = significant (p < 0.05), (*) = marginally significant (0.05 < p < 0.1),. = slightly significant (p < 0.2), ns: non-significant. RDW, root dry weight; RLD, root length density; SRR, shoot root ratio; SRL, specific root length; %, difference between amb and FACE treatment. Crop Winter barley (EC 34)

CO2 −

N

N+ Sugar beet (EC 45)

N− N+

Winter wheat (EC 65)

N− N+

Statistics (mixed effects model)

Crop CO2 N CO2 :crop CO2 :N N:crop

N

RDW 0–0.3 m (g m−2 )

SRR

CO2 − CO2 + CO2 − CO2 +

77 88 (+14%) 78 84 (+ 8%)

4.9 4.4 (−9%) 6.2 6.5 (+5%)

RLD (cm cm−3 )

SRL (m g−1 )

CN ratio

CO2 − CO2 + CO2 − CO2 +

62 75 (+21%) 48 66 (+38)

14.2 14.1 (−1%) 23.0 17.5 (−24%)

1.8 2.9 (+61%) 2.1 2.4 (+14%)

135.6 155.6 (+15%) 222.3 186.4 (−16%)

15.6 18.3 (+17%) 17.1 15.4 (−10%)

CO2 − CO2 + CO2 − CO2 +

127 149 (+17%) 110 159 (+45%)

8.6 8.1 (−6%) 10.8 8.5 (−27%)

2.3 1.9 (−18%) 2.6 3.2 (+23%)

52.9 35.6 (−32%) 70.5 60.8 (−14%)

56.0 58.6 (+5%) 49.4 56.1 (+14%)

* * ns ns ns ns

* ns (*) ns ns ns

. . ns ns ns ns

* ns ns ns ns .

* * . * ns ns

44

A. Pacholski et al. / Europ. J. Agronomy 63 (2015) 36–46

Rogers, 2000), in our study maximum stimulation of RDW under elevated [CO2 ] (up to 60%) was in excess of the stimulation of above ground biomass observed in the same study (up to 18%, Weigel and Manderscheid, 2012). However, these stimulations were observed for specific growth stages only. Dynamics of biomass stimulation by elevated [CO2 ] were different for roots as compared to shoot biomass, with highest stimulation rates for roots at early growth stages, while above ground biomass, in particular of the winter cereals, showed the highest increase at final harvest (Weigel and Manderscheid, 2012). Consequently, for all three crops SRR was decreased at early growth stages, but increased at late stages by elevated [CO2 ]. Similarly, SRR was decreased by low N supply for all species, as theoretically expected. For winter barley a strong negative effect of elevated [CO2 ] on SRR at early growth stages turns into a positive effect at harvest. One explanation for this phenomenon might be that leaf senescence of barley was about one week earlier under elevated than under ambient [CO2 ]. Root senescence begins even earlier as compared to leaf senescence (Gregory et al., 1978). In sugar beet first a positive and then later, a negative CO2 effect on leaf area index as compared to ambient conditions in early and late summer, respectively, was observed in the same experiment (Manderscheid et al., 2010). This might also have contributed to the temporal changes in the [CO2 ] effect on root growth, since root growth and shoot root ratio is regulated by the nutrient demand of the canopy (Gastal and Lemaire, 2002; Svoboda and Haberle, 2006). Variable, crop specific responses have been reported for SRR in earlier studies under CO2 enrichment (Kimball et al., 2002; Madhu and Hatfield, 2013). The absolute values of stimulation of different root variables under elevated [CO2 ] in our study are characterized by a high degree of uncertainty. However, reduction in SRR for winter barley and winter wheat support the hypothesis that root biomass is stimulated more strongly by elevated [CO2 ] than above-ground biomass at early growth stages. The reasons for a specific stimulation of root growth are unclear. As the effect of elevated [CO2 ] on SRR is similar to the effect of limited N supply, a possible increased demand of nutrients under elevated [CO2 ] at early growth stages could trigger this effect. This is supported by the analysis of composition of root tissue. For high N supply a widening of the CN ratio is observed under elevated [CO2 ] compared to ambient conditions which agrees with a stronger stimulation of root biomass under elevated [CO2 ] at the same N-level. Although stimulation of root biomass was in excess of stimulation of shoot biomass for individual sampling dates, the overall effect of elevated [CO2 ] on root biomass of the winter cereals was probably not stronger than on shoot biomass. This is supported by the non-existence of a significant positive main effect of elevated [CO2 ] for all sampling dates and a variable effect on SRR. Vincent and Gregory (1989a,b) found a linear relationship between thermal time and intercepted photosynthetically active radiation and root dry matter development of winter wheat under ambient [CO2 ]. On this background, a similar total effect of elevated [CO2 ] on root dry matter of the crops investigated in our study to above-ground biomass is plausible. Nevertheless, maximum stimulation of belowground and above-ground biomass occurred at different growth stages indicating an increased resource demand and availability under elevated [CO2 ] which might be of importance for modelling of crop growth. Effects of elevated [CO2 ] on RLD were similar to the findings for RDW while SRL was not influenced by the [CO2 ] treatments. Thus, both crops produced higher RLD under elevated [CO2 ] compared to ambient conditions by an increased production of RDW and not due to changes in root morphology, i.e. SRL. Root morphology was mainly affected by crop type and slightly modified by N-supply. Changes of root phenology in the subsequent phases of root growth caused by a treatment are very hard to detect. Therefore

a comparison of root development of a crop between treatments at the same date within a vegetation period but at potentially different growth stages would lead to a biased interpretation of the data (Pritchard and Rogers, 2000). However, although there exists only limited information on root growth dynamics as part of the whole crop response to elevated [CO2 ], effects are not expected to differ markedly from above-ground changes (Pritchard and Rogers, 2000). With respect to above-ground processes no shifts in crop phenology (Weigel and Manderscheid, 2012) apart from a slightly earlier ripening of cereals under elevated CO2 were observed in the present FACE experiment. Previous studies showed that the main differences between root samples under ambient and elevated [CO2 ] conditions were observed in the plough layer (Rogers et al., 1994; Pritchard and Rogers, 2000). However, it was found that [CO2 ] elevation particularly stimulates root growth in deeper soil layers when plant growth is limited by nutrient or soil water availability (Burkart et al., 2004; Wechsung et al., 1999). In our study, effects on root growth of winter wheat in deeper soil layers were similar to those in the plough layer. The different findings in our study as compared with previous studies might be due to differences in the degree of the resource deficiencies of other plant growth factors. For example, halving the N supply in the present crop rotation compared to an optimum N supply decreased plant growth only slightly by 14% when averaged across all crop species (Weigel and Manderscheid, 2012). In many studies effects of elevated [CO2 ] on CN ratios in aboveground biomass was observed (Kimball et al., 2002; Taub et al., 2008). Although these results varied strongly, N concentrations in the various plant tissues investigated decreased by 10–15% due to the high [CO2 ] treatment. In our crop rotation study the CN ratio in roots was slightly increased (2–18%) under FACE indicating a decrease in N concentration. The [CO2 ] effect for winter wheat rather appeared under high N and for sugar beet under low N supply. However, the [CO2 ] effect was much smaller than the effect of N supply, and the crop species still had much higher influence on CN ratio. It can be hypothesized that in the arable system studied the effect of N supply overcompensated the effect of elevated [CO2 ] on this root variable. Nevertheless, changes in bacterial and fungal microbial biomass in soil and effects on soil collembola diversity and abundances were observed in the same field trial (Anderson et al., 2011; Sticht et al., 2008). These changes were mainly attributed to higher levels of plant residues and root biomass under elevated [CO2 ] compared to ambient conditions which supports the above stated hypothesis.

5. Conclusions The results of the present crop rotation study show that an increase of the atmospheric CO2 concentration from about 380 ppm to 550 ppm as expected for the middle of this century might have positive effects on the root growth of winter barley, winter wheat and sugar beet and that these effects may not depend on the plant N supply. However, stimulation of root growth due to elevated [CO2 ] may be transient during the vegetation period with maximum stimulation of RDW and a decrease of SRR early in the vegetation period while at the end of the growing season a CO2 effect on RDW may disappear and effects on SRR may even be reversed. The CN ratio of roots, which is an important determinant for overall soil C turnover and soil microbiology, respectively, was only slightly increased by elevated [CO2 ] in comparison to the influence of N supply and crop species. This result points to small direct effects of elevated [CO2 ] on soil processes governed by root composition. Our data nevertheless show that elevation of atmospheric [CO2 ] may enhance crop root and residue pools in soil with potential positive effects on soil

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C sequestration. While the present study only encompassed [CO2 ] and N effects on the crops, future climate change will in particular also increase air temperature and soil temperature and change rainfall patterns. Higher soil temperatures will probably enhance soil C turnover and thus mitigate the potential C sequestration effect of elevated [CO2 ] and in addition make more soil born N available to plants or increase N leaching risks. The temporal difference between the CO2 stimulation of root and shoot biomass may have effects on the temporal availability of water and nutrient resources under elevated [CO2 ], with higher availability of this resources at early growth stages under elevated [CO2 ] when crops are particularly vulnerable to environmental stresses. The dynamics of such effects should particularly be considered when modelling plant and crop biomass partitioning under future climate conditions and elevated [CO2 ]. Acknowledgements We thank Dr. Bernd Kleikamp, Prof. Dr. Mohamed Helal and Carina Trenkler for their support in sampling and extracting crop roots. We are much obliged to Dr. Keith Lewin, Dr. George Hendrey and Dr. John Nagy for setting up the FACE system in 1999 and further support in the following years as well as to Dr. Cathleen Frühauf for managing the FACE system in the years 1999–2002. The study was financially supported by the German Federal Ministry of Food, Agriculture and Consumer Protection. References Ainsworth, E.A., Long, S.P., 2005. What have we learned from 15 years of free-air CO2 enrichment (FACE)? A meta-analytic review of the responses of photosynthesis, canopy properties and plant production to rising CO2 . New Phytol. 165 (2), 351–371. Ainsworth, E.A., McGrath, J.M., 2010. Direct effects of rising atmospheric carbon dioxide and ozone on crop yields. In: Lobell, D., Burke, M. (Eds.), Climate Change and Food Security. Adapting Agriculture to a Warmer World. Adv. Global Change Res., vol. 37. Springer, New York, pp. 109–130. Anderson, T.-H., Heinemeyer, O., Weigel, H.-J., 2011. Changes in the fungal-tobacterial respiratory ratio and microbial biomass in agriculturally managed soils under free-air CO2 enrichment (FACE) – a six-year survey of a field study. Soil Biol. Biochem. 43, 895–904. Brisson, N., Wery, J., Boote, K., 2006. Fundamental concepts of crop models illustrated by a comparative approach. In: Wallach, D., Makowski, D., Jones, J.W. (Eds.), Working with Dynamic Crop Models. Elsevier, The Netherlands, pp. 257–279. Brown, K.F., Biscoe, P.V., 1985. Fibrous root-growth and water-use of sugar-beet. J. Agric. Sci. 105, 679–691. Burkart, S., Manderscheid, R., Weigel, H.J., 2004. Interactive effects of elevated atmospheric CO2 concentrations and plant available soil water content on canopy evapotranspiration and conductance of spring wheat. Eur. J. Agron. 21 (4), 401–417. Burkart, S., Manderscheid, R., Wittich, K.P., Löpmeier, F.J., Weigel, H.J., 2011. Elevated CO2 effects on canopy and soil water flux parameters measured using a large chamber in crops grown in free air CO2 -enrichment. Plant Biol. 13, 258–269. Gastal, F., Lemaire, G., 2002. N uptake and distribution in crops: an agronomical and ecophysiological perspective. J. Exp. Bot. 53 (370), 789–799. Gregory, P.J., McGowan, M., Biscoe, P.V., Hunter, B., 1978. Water relations of winterwheat.1. Growth of root-system. J. Agric. Sci. 91 (1), 91–102. Gregory, P.J., Palta, J.A., Batts, G.R., 1997. Root systems and root:mass ratio carbon allocation under current and projected atmospheric conditions in arable crops. Plant Soil 187, 221–228. Hendrey, G.R., Miglietta, F., 2006. FACE technology – past, present and future. In: Nösberger, J., Long, S.P., Norby, R.J., Stitt, M., Hendrey, G.R., Blum, H. (Eds.), Managed Ecosystems and CO2 , vol. 187. Springer, New York, pp. 15–46. Kersebaum, K.C., Friesland, H., Löpmeier, F.J., 2005. Irrigation and Pest and Disease Models: Comparison of Three Irrigation Models under German Conditions. Report COST Action 718., pp. 16–25. Kim, H.Y., Lieffering, M., Kobayashi, K., Okada, M., Mitchell, M.W., Gumpertz, M., 2003. Effects of free-air CO2 enrichment and nitrogen supply on the yield of temperate paddy rice crops. Field Crop Res. 83, 261–270. Kimball, B.A., Kobayashi, K., Bindi, M., 2002. Responses of agricultural crops to freeair CO2 enrichment. Adv. Agron. 77, 293–368. Klironomos, J.N., Rillig, M.C., Allen, M.F., 1999. Designing belowground field experiments with the help of semi-variance and power analyses. Appl. Soil Ecol. 12, 227–238. Koerner, C., Asshoff, R., Bignucolo, O., Haettenschwiler, S., Keel, S.G., Peláez-Riedl, S., Pepin, S., Siegwolf, R.T., Zotz, G., 2005. Carbon flux and growth in mature deciduous forest trees exposed to elevated CO2 . Science 309 (5739), 1360–1362.

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