- Email: [email protected]
Effect of nitrogen supply on leaf appearance, leaf growth, leaf nitrogen economy and photosynthetic capacity in maize (Zea mays L.)
Field Crops Research 93 (2005) 64–73 www.elsevier.com/locate/fcr
Effect of nitrogen supply on leaf appearance, leaf growth, leaf nitrogen economy and photosynthetic capacity in maize (Zea mays L.) J. Vosa,*, P.E.L. van der Puttena, C.J. Birchb a
Group Crop and Weed Ecology, Plant Sciences, Wageningen University and Research Centre, Haarweg 333, 6709 RZ Wageningen, The Netherlands b The University of Queensland, Gatton 4343, Qld., Australia Received 24 May 2004; received in revised form 8 September 2004; accepted 9 September 2004
Abstract Leaf area growth and nitrogen concentration per unit leaf area, Na (g m2 N) are two options plants can use to adapt to nitrogen limitation. Previous work indicated that potato (Solanum tuberosum L.) adapts the size of leaves to maintain Na and photosynthetic capacity per unit leaf area. This paper reports on the effect of N limitation on leaf area production and photosynthetic capacity in maize, a C4 cereal. Maize was grown in two experiments in pots in glasshouses with three (0.84– 6.0 g N pot1) and five rates (0.5–6.0 g pot1) of N. Leaf tip and ligule appearance were monitored and final individual leaf area was determined. Changes with leaf age in leaf area, leaf N content and light-saturated photosynthetic capacity, Pmax, were measured on two leaves per plant in each experiment. The final area of the largest leaf and total plant leaf area differed by 16 and 29% from the lowest to highest N supply, but leaf appearance rate and the duration of leaf expansion were unaffected. The N concentration of expanding leaves (Na or %N in dry matter) differed by at least a factor 2 from the lowest to highest N supply. A hyperbolic function described the relation between Pmax and Na. The results confirm the ‘maize strategy’: leaf N content, photosynthetic capacity, and ultimately radiation use efficiency is more sensitive to nitrogen limitation than are leaf area expansion and light interception. The generality of the findings is discussed and it is suggested that at canopy level species showing the ‘potato strategy’ can be recognized from little effect of nitrogen supply on radiation use efficiency, while the reverse is true for species showing the ‘maize strategy’ for adaptation to N limitation. # 2004 Elsevier B.V. All rights reserved. Keywords: Nitrogen supply; Nitrogen concentration; Leaf growth; Leaf appearance rate; Photosynthesis; Gramineae; Broad leaf species
1. Introduction * Corresponding author. Tel.: +31 317 245795; fax: +31 317 485572. E-mail address: [email protected] (J. Vos).
Yield response to larger nitrogen, N, supply is positive until factors other than nitrogen limit higher production. The positive response is brought about
0378-4290/$ – see front matter # 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.fcr.2004.09.013
J. Vos et al. / Field Crops Research 93 (2005) 64–73
either by (i) a larger amount of radiation intercepted over the crop growth period, or (ii) a higher average daily rate of photosynthesis, or a combination of (i) and (ii). Option (i) arises from effects on the dynamics of production and senescence of leaves and option (ii) from effects on the radiation use efficiency (RUE) and the nitrogen economy of the crop. Over the last decade, the responses of potato (Solanum tuberosum L.) were analyzed at the level of plant and organ (Vos and Biemond, 1992; Biemond and Vos, 1992) and for individual leaves (Vos and Van der Putten, 1998). The most important findings were that larger supply of nitrogen enhances apical branching, meaning that more leaves are formed per plant over a longer period of time. The timing of developmental events, e.g. the rate of leaf appearance, was not affected by N. Prolonged production of vegetative organs, rather than retarded rate of leaf senescence, appeared responsible for increase in the crop duration with more N (Vos and Biemond, 1992). Under N limitation potato reduces the size of the mature leaf to such an extent that maintenance of the N concentration and photosynthetic capacity per unit leaf area is achieved (Vos and Van der Putten, 1998). ‘Downregulation’ of leaf size is achieved by reduced rate of leaf expansion rather than reduced duration of expansion, identifying an effect of N on the rate of cell division or on cell size as possible mechanisms (Walter et al., 2003). The ‘potato strategy’ of adapting leaf size results in little effect of N supply on radiation use efficiency in potato (Millard and Marshall, 1986). In potato, for a given leaf insertion number, mature leaf size can differ up to a factor 4, depending on the supply of N. Previous work by Muchow (1988) showed that leaf size in maize (Zea mays L.) was responsive to N supply, but compared to potato, the differences were relatively small. Muchow and Davis (1988) reported that RUE of maize was more responsive to N supply than was radiation interception, the latter being related to leaf area index of the canopy. Such observations lead to the hypothesis that the strategy of response to N limitation differs fundamentally between potato and maize. The strategy of potato is to maintain productivity per unit leaf area rather than maximizing leaf area and light interception per plant. The hypothesis is that in maize leaf area and light interception are maintained to the detriment of the concentration of nitrogen and the
65
photosynthetic capacity per unit leaf area (referred to by ‘maize strategy’), also with relatively little effect of N on the kinetics of leaf appearance (Muchow, 1988). It is the main objective of this paper to test that hypothesis. To that end, maize plants were grown under experimental conditions similar to the ones used in the previous potato work (Vos and Van der Putten, 1998) and subjected to a range of N treatments. Leaf dynamics and growth, photosynthetic capacity and N content of individual leaves were analyzed over time.
2. Materials and methods 2.1. Arrangement of experiments Two pot experiments, Expt 1 and Expt 2 of basically similar design, were conducted in naturally lit glasshouses. Supplemental light from 400 W SONT Agro Philips lamps (0.5 lamps m2) switched on when global solar radiation outside the glasshouse dropped below 400 W m2 during daytime, and switched off again when it exceeded 500 W m2. Daylength was limited to 12 h (the glasshouse was externally covered with a light tight roof on rails from 6 h after solar noon to 6 h before solar noon). The fraction of the daily total of sunlight (as given in standard meteorological records) received at plant level in the glasshouse varied with daylength outside the glasshouse and cloudiness, but measurements showed that this fraction was about 0.5–0.6 for photosynthetically active radiation (PAR). Day/night temperatures set at 23/18 8C (12 h/12 h). Twelve litre pots were filled with sand. In Expt 1, the initial N content of the medium was not measured, but the substrate was richer than in Expt 2 (Expt 2: approx. 20 mg N kg1 of medium and estimated mineralization during the experimental period of 8 mg kg1). Pots with one plant each were initially spaced at 3 pots m2. As plants were harvested, density declined to 1 pot m2, i.e. plants were treated as single, spaced plants. Expt 1 was planted on 4 August 1997 and plants emerged on 9 August. Expt 2 was planted on 27 March 1998; emergence was on 1 April. In Expt 1, plants with three nitrogen treatments (Table 1) were grown over 101 days from emergence. In Expt 2, a wider range of N supply was used over five treatments (Table 1) and plants were grown for 78 days
66
J. Vos et al. / Field Crops Research 93 (2005) 64–73
Table 1 Amounts of nitrogen added in nutrient solution per treatment in both experiments Expt code
Code of nitrogen treatment N1
N2
N3
N4
N5
Expt 1 Expt 2
0.84 0.50
2.5 1.0
6.0 2.5
4.0
6.0
Units: g N per plant.
after emergence. The cultivar was Lincoln (three-way cross; see Birch et al., 2003a for comparative performance). Silking occurred at 47 days from emergence. The topmost ear was found at leaf 7 or 8. Nutrient solutions were given from 1 week after planting onwards at weekly intervals (six applications in Expt 1 and 11 times at intervals of 7 days in Expt 2). The first two applications were half the standard rates. The solutions contained Ca(NO3)2, NH4NO3, K2SO4, KH2PO4, MgSO4, FeEDTA, ZnSO4, and other micronutrients. Total additions of P, K, Mg, Ca were 1.2, 8.0, 1.3, 1.45 g pot1, respectively. In Expt 2, a foliar application of MgSO47H2O was applied twice (in total 250 mg Mg pot1). Single pots (plants) were the experimental units. The experimental design was a split plot with six blocks and harvests as main plots and nitrogen treatments as sub plots. The six blocks occupied the available glasshouse space; hence, within each plant sample (n = 6) the variation is represented that existed in climate conditions within the glasshouse. 2.2. Data collection 2.2.1. Leaf number and leaf area Rates of leaf appearance (tips and ligules) and the rate of leaf senescence were recorded twice weekly. Leaf expansion was monitored non-destructively on leaf insertion numbers 8 and 10 in Expt 1 and leaf numbers 7 and 9 in Expt 2. Appeared area of each leaf, from whorl or ligule to leaf tip, was estimated from (Sanderson et al., 1981): Leaf area ¼ leaf length maximum width k where k is a shape factor with value 0.75, as determined in Birch et al. (2003a). Photosynthetic rates were measured using portable equipment [LCA2 from Analytical Development Company (ADC), Hoddesdon, UK]. The measure-
ments were made in a temperature-controlled room. Cuvette temperature ranged between 20 and 24 8C and inlet CO2 concentration was typically 350 mmol mol1. Airflow rate was 6–8 ml s1 and adjusted to limit the CO2 depletion to maximally 75 mmol mol1. With an external, heat-filtered light source, constant irradiance was maintained. In Expt 1, 1550 mmol m2 s1 (PAR) was achieved at the level of the leaf in the cuvette. This was less than intended and probably less than needed for full light saturation of photosynthesis in relatively young leaves. Therefore, this paper does not report photosynthesis data from Expt 1. In Expt 2, the illumination was 2300 mmol m2 s1 (PAR) at leaf level. Subsidiary measurements of the photosynthesis–light response curve of relatively young leaves ascertained that light saturation was obtained in Expt 2. A photosynthesis reading was taken after 20–25 min after closing the cuvette when the rate of CO2 exchange had been steady for 5 min. Water vapour pressure deficit in the cuvette ranged between ca. 300 and 1000 Pa, depending on the rate of transpiration. Measurements were made halfway along the length of a leaf, midway between leaf edge and midrib. The enclosed leaf area was 11 cm2 in Expt 1 and 6.25 cm2 in Expt 2. 2.2.2. Plant dry weight sampling In Expt 1, laminae of leaf numbers 8 and 10, and in Expt 2 of leaf numbers 7 and 9, were harvested and processed separately to determine their leaf area, fresh and dry weight and the concentrations of total nitrogen and nitrate in the dry matter. In Expt 1, samplings were done on 15, 37 and 57 days after tip emergence for leaf 7 and on 9, 31 and 51 days after tip appearance for leaf 9. In Expt 2, leaf sampling was done at intervals of 2 weeks, starting on 13 days after tip appearance for leaf 8 and at 6 days after tip appearance for leaf 10. To examine the degree of N limitation, whole plants were harvested on 87 and 78 days after emergence (DAE) in Expt 1 and Expt 2, respectively, for the determination of dry weights and nitrogen concentrations in leaf blades (all blades on a plant bulked together) stems plus sheaths and cob. 2.3. Chemical analyses Total nitrogen concentration was determined in dried and ground (mesh sieve 1 mm) plant material in
J. Vos et al. / Field Crops Research 93 (2005) 64–73
two steps by digestion with sulphuric acid using salicylic acid, H2O2 and selenium as additives (Novozamsky et al., 1983), followed by colorimetric determination of nitrogen in an auto-analyzer using the Berthelot reaction (Technicon Auto-analyzer II, Industrial Method No. 334-74B/W+, released January 1976, revised March 1977; Technicon Industrial Systems, Tarrytown, New York). Nitrate was extracted by shaking 500 mg of dried and ground leaf material in 50 ml water for 30 min, followed by filtration. The nitrate concentration in the extract was determined with an auto-analyzer (Auto-Analyzer II Method NL 211-91WT; Technicon Industrial Systems, Tarrytown, New York). Organic nitrogen is defined as the difference between total nitrogen and nitrate-N. 2.4. Calculations and statistics Accumulation of thermal time was calculated from the average of the daytime and nighttime using a base temperature Tb of 8 8C for this cultivar (Birch et al., 2003a); hence, 1 day equals 12.5 8C d. The association between organic nitrogen concentration per unit leaf area, Noa, and Pmax was described with the logistic equation (Sinclair and Horie, 1989; Vos and Van der Putten, 1998): 2 1 (1) Pmax ¼ Am 1 þ exp ðsðN N ÞÞ oa
o
where Am is the asymptote of Pmax (mmol CO2 m2 s1) for infinite value of Noa; s the parameter determining the steepness of the slope; and No the value for Noa (g m2) for Pmax = 0. Specific or surface leaf weight (SLW) is defined as the ratio between leaf dry weight and area per leaf. Areal nitrogen concentration is the ratio between the amount of nitrogen in the leaf and its surface area.
3. Results 3.1. General treatment effects In both experiments the objective to create treatments encompassing the range from severe N limitation to non-limiting supply (Table 2) was achieved. Plant analyses at 87 DAE in Expt 1 and
67
Table 2 Treatment effect on total dry weight per plant (g) and total N content per plant (mg) determined on 87 and 78 days after emergence in Expts 1 and 2, respectively Expt code Plant attribute Code of nitrogen treatment N1
N2
N3
Expt 1 Expt 1
Dry weight N content
112 204 199 850 2370 3470
Expt 2 Expt 2
Dry weight N content
95 480
N4
N5
L.S.D. 14 142
130 219 239 248 19 840 2180 3580 4430 370
Least significant differences, L.S.D.: P = 0.05.
78 DAE in Expt 2 show higher dry weight and N content per plant for the higher N rates. However, whereas N content continued to increase over the whole range of N supply, dry matter production did not differ between N2 and N3 in Expt 1 and between N4 and N5 in Expt 2, indicating that there was no N limitation in the treatments with the highest rates of N. The N1 treatment in Expt 2 was more severely N limited than the N1 treatment in Expt 1 (Table 2) due to lower application and use of a less fertile root medium. For non-limiting N supply dry matter production and N uptake per plant were larger in Expt 2 than in Expt 1. This was associated with the decline in daily solar radiation as time progressed in Expt 1 and the opposite in Expt 2. 3.2. Leaf dynamics and leaf expansion There was no effect of nitrogen treatments on the rates of appearance of leaf tips and ligules (Fig. 1a). The average rate of leaf tip appearance was 0.022 leaf (8C d)1; the plastochron was 45.7 8C d per leaf. The thermal duration of leaf expansion, i.e. the horizontal distance between leaf tip appearance and ligule appearance (Fig. 1a), depended on leaf number, but was not systematically affected by N supply and did not differ between experiments. Leaf numbers 7 and 9, included in detailed analyses in Expt 2, showed durations of expansion of ca. 200 and 170 8C d, respectively. Leaf senescence started at ca. 430 8C d after emergence in all treatments (Expt 2). Leaf senescence preceded faster the lower the rate of N supply. For leaf numbers 8–12 in Expt 1, the thermal duration of life spans of N1 leaves was 16–10% shorter than for N3 leaves (Fig. 1b).
68
J. Vos et al. / Field Crops Research 93 (2005) 64–73
Fig. 1. Effect of nitrogen supply on the kinetics of leaf production and life spans: (a) leaf tip appearance vs. thermal time (base 8 8C), ligule appearance and the change in the number of dead leaves as a function of thermal time (8C d) (one day = 12.5 8C d); data from Expt 2. (b) Life span from leaf tip emergence till leaf death (8C d) for insertion numbers 8–12 in Expt 1; bars represent L.S.D. (P = 0.05).
Logistic equations fitted to data on the change with time after tip appearance of visible leaf area (Fig. 2a) showed no effect of N treatment on the point in time of inflection of leaf area growth rate (details not shown), indicating that the duration of phases of expansion were not affected by N supply. The maximum rate of expansion increased with N supply, resulting in larger mature area with more N supplied (ca. 16% difference between N1 and N5 leaves). Similar results were found in Expt 1 with 14% difference among N treatments in mature area of leaf number 8, the largest leaf (data not shown). Fig. 2b shows that leaves up to number 6 grew to similar mature sizes, irrespective of N application (Expt 2; the same result was found in Expt 1—data not
Fig. 2. Effect of nitrogen supply on leaf size (data from treatments N1–N5 in Expt 2). (a) The change in area of leaf insertion number 9 vs. thermal from tip appearance. (b) Full-grown areas of all leaf insertion numbers. Bars: L.S.D. (P = 0.05).
shown). For higher leaf numbers, mature area was greater with larger the amounts of N. The relative effect of N supply on leaf size increased with leaf rank number. The cumulative area of full-grown leaves increased from 3094 cm2 per plant in N1 to 3992 cm2 per plant in N5 plants of Expt 2 (i.e. 29% difference between extreme N treatments in Expt 2; the comparable figure in Expt 1 was 15%—data not shown). 3.3. Leaf attributes In all leaves examined, leaf blade weight increased even after expansion had stopped (Fig. 3a). Therefore, specific leaf weight (SLW) not only increased with time during leaf expansion but also continued to increase afterwards (Fig. 3b). In all cases, except leaf 10 in Expt 1, there was an N effect on SLW in that
J. Vos et al. / Field Crops Research 93 (2005) 64–73
69
Fig. 3. Effect of nitrogen supply on the change in leaf properties after leaf tip appearance. (a) Leaf dry weight, (b) specific leaf weight, (c) nitrogen concentration in leaf dry matter, (d) absolute amount of nitrogen per leaf, (e) nitrogen concentration per unit leaf area, (f) nitrate-N concentration per unit leaf area. Broken lines: data from Expt 1, full-drawn lines: data from Expt 2. Symbols: circles: lowest rate of N supply, i.e. N1 treatments of Expts 1 and 2; squares: highest rate of N supply, i.e. N3 in Expt 1 and N5 in Expt 2; open symbols: leaf 10 in Expt 1 and leaf 9 in Expt 2; closed symbols: additional data points from leaf rank 7 in Expt 2. Thin bars: L.S.D. (P = 0.05) in Expt 1; bold bars: L.S.D. for Expt 2.
SLW was highest in the treatments with the largest N supply (12–17% difference). The concentration of nitrogen in leaf dry matter, Ndm, (Fig. 3c) differed substantially between N treatments from the moment the tip appeared from the whorl. For the highest rate of N supply, generally
similar maximum values of Ndm and similar change with time were observed in Expts 1 and 2. In these treatments Ndm increased till shortly after full leaf expansion was reached. Ndm was lower in N1 leaves of Expt 2 than in corresponding leaves in Expt 1; this is in accordance with a lower N supply in the N1 treatment
70
J. Vos et al. / Field Crops Research 93 (2005) 64–73
in Expt 2 than in Expt 1. The absolute amount of nitrogen in the leaf blade differed approximately fourfold between the treatments with the highest and lowest N supply in Expt 2 (Fig. 3d), and less so in Expt 1, in accordance with a narrower range of N supply. The absolute amount of nitrogen per leaf reached its maximum near or shortly after the time of ligule appearance and gradually declined thereafter, implying a continuous export of N from the leaf. Peak values occurred later in a leaf’s life when more N was supplied. Differences in leaf size explain the relative differences in absolute amounts of N as depicted in Fig. 3d for leaf 10 from Expt 1 (456 cm2) and leaf 9 from Expt 2 (608 cm2). In expanding leaves (<200 8C d from tip appearance) areal total nitrogen concentration, Nat, ranged from 0.56 g m2 N at the lowest N rate to 1.37 g m2 N at the highest (Fig. 3e). Peak values were 1.9 g m2 N in high-N treatments, occurring 125 8C d after full leaf expansion. After the peak, Nat declined gradually during the leaf’s life, maintaining differences of a factor 3–4 between the extremes of the N treatments. Peak nitrate concentrations, expressed per unit leaf area, occurred during the phase of leaf expansion and amounted to 0.08 g m2 nitrate-N (Fig. 3f). Upon completion of leaf expansion values declined from ca. 0.05 g m2 nitrate-N to negligible levels. In expanding leaves nitrate-N represented at maximum 6% of the total nitrogen; thus, any error incurred for not accounting for nitrate would be negligible for most of the leaf’s life.
Fig. 4. Nitrogen and the light-saturated rate of photosynthesis, Pmax, (carbon dioxide assimilation); data from Expt 2. (a) Change in Pmax in time after tip appearance. Symbols: circles: N1 treatments; squares: N5 treatments; open symbols: leaf 9; closed symbols: leaf 7. (b) Pmax as a function of the nitrogen concentration per unit leaf area; symbols: circles: N1; triangles: N2; squares: N5. Bars: L.S.D. (P = 0.05).
3.4. Photosynthesis There was a large effect of N treatment on Pmax (Fig. 4a). Already during the phase of leaf expansion Pmax differed by 38% between the extremes of the N treatments. With non-limiting N supply, Pmax increased from a first measured value of 41 mmol m2 s1 to a peak value of 45 mmol m2 s1 1 at 30 days (375 8C d) after leaf tip emergence. In the low-N treatment Pmax gradually declined after cessation of leaf expansion. Data points from all Pmax recordings made in Expt 2 were plotted against Noa (Fig. 4b). Except for two deviating points, obtained on 51 days after leaf tip appearance in the high-N treatment, the data from all treatments fell on one common curve, and were described well by Eq. (1).
4. Discussion In both potato and maize the association between Pmax and areal N concentration, Na could be described well by Eq. (1). Maize was characterized by lower No (estimated at 0.25 g m2 versus 0.5 g m2 in potato), steeper slope s (2.90 versus 1.74 for potato) and higher asymptote of maximum photosynthesis rate (45e mmol m2 s1 CO2 fixation versus 19 mmol m2 s1 for potato). The current maximum rate for maize fits well with literature reports showing values between 48 and 57 mmol m2 s1 (2.1 and 2.5 mg m2 s1) (Sinclair and Horie, 1989; Wong et al., 1985; Muchow and Sinclair, 1994). The current data support the view
J. Vos et al. / Field Crops Research 93 (2005) 64–73
that there is a strong association between Na and photosynthetic capacity, regardless whether variation in Na arises from variation in rate of N supply and differences in leaf age (e.g. Connor et al., 1993; McCullough et al., 1994b; Vos and Van der Putten, 1998), or differences in illumination among leaves. Also it is established that the coefficients of Eq. (1) differ much between plant species, the largest segregation being between C3 and C4 species (Sinclair and Horie, 1989; Grindlay, 1997). The study of Gastal and Nelson (1994) in the C3 grass tall fescue (Festuca arundinacea Schreb.) showed large differences in nitrogen deposition rates and ribulose 1,5-biphosphate carboxylase content in the early stages of leaf growth, i.e. before the leaf tip appears. This supports the current finding of large differences in N concentration in young expanding maize leaves (Fig. 3), although C3 and C4 species differ fundamentally in their N economy (e.g. Brown, 1978). With respect to leaf dynamics, both current experiments produced similar results, the most important of which are: (i) in maize, the rate of leaf appearance, the duration of leaf expansion and the number of leaves are not affected by nitrogen supply; (ii) the cumulative area of mature leaves on a plant differed up to ca. 30% when N supply increased from severely limiting to non-limiting rates. That implies that maize showed a conservative response of leaf size in comparison to potato where leaf size varied three- to four-fold in response to N supply, when measured under largely similar experimental conditions (Vos and Van der Putten, 1998); (iii) among the extremes in N treatments, areal N concentration, Na, differed by a factor of ca. 2 in expanding leaves (<17 days or 210 8C d from tip appearance) (Fig. 3e). At and shortly after ligule appearance three- to four-fold differences were found among extreme N treatments as regards N concentration in leaf dry matter (Fig. 3c), absolute amount of N per leaf (Fig. 3d) and areal N concentration (Fig. 3e). This is in clear contrast to potato with small differences only in Na and its decline with leaf age (Fig. 3f in Vos and Van der Putten, 1998). Points (i)–(iii) are in support of the hypothesis posed: potato and maize show contrasting strategies to deal with N limitation. Potato adapts leaf size and avoids compromizing leaf N economy and the associated photosynthetic capacity, whereas maize strives for maintenance of leaf area per leaf at the
71
expense of decreased N concentration per unit leaf area and decreased photosynthetic capacity. With some exceptions (Grindlay, 1997; McCullough et al., 1994a) it is the general finding that, over a broad range of supply rates, there is little effect of N on the rate of leaf appearance and the duration of leaf expansion. This holds true for monocots, e.g. in maize and sorghum (Sorghum bicolor L. Moench) (Muchow, 1988), perennial ryegrass (Lolium perenne L.) (Van Loo, 1993), leek (Allium porrum L.) (Biemond, 1995b) as well as for dicots, e.g. potato (Vos and Biemond, 1992) and sunflower (Helianthus annuus L.) (Tra´ pani and Hall, 1996). As there are few studies addressing area expansion and Na at leaf level, it is not straightforward to verify whether the ‘maize strategy’ is common in Gramineae, and the ‘potato strategy’ in broadleaf species. Radin (1983) hypothesized that leaf elongation rate of broadleaf species was much more sensitive to N limitation than in Gramineae and attributed this to differences in water potential in the elongation zone, because leaves of broadleaf species expand in open air, whereas the elongation zone of cereals is protected by the sheath of preceding leaves. In Muchow (1988) the area of the largest leaf of maize differed by about 33% among N treatments, i.e. comparable to the N effect observed in the current study, while Van Loo (1993) reported similar effects in perennial ryegrass. McCullough et al. (1994b) observed similar effects of nitrogen supply on Na and leaf area of maize as in the current study. Dicot species showing a bigger effect of nitrogen limitation on sizes of individual leaves than on leaf nitrogen concentration include sunflower (Tra´ pani and Hall, 1996) and Brussels sprouts (Brassica oleracea L. var. gemmifera DC) (Biemond, 1995a; Biemond et al., 1995). In Radin’s (1983) observations C3 and C4 Gramineae behaved similar. The cited studies support the hypothesis that the ‘maize strategy’ is more common in Gramineae (both C3 and C4) and the ‘potato strategy’ more common in dicot species. Adaptation of leaf size and Na are the primary options for response to N limitation in maize, Brussels sprouts and sunflower as these species commonly do not tiller or branch. Small grain C3 Gramineae, but also several C4 Gramineae, e.g. pearl millet (Coaldrake, 1985), show a variable degree of tillering, commonly very responsive to nitrogen supply (Longnecker et al., 1993). Vos and Biemond (1992) showed
72
J. Vos et al. / Field Crops Research 93 (2005) 64–73
the degree of basal and apical branching to be responsive to nitrogen supply in potato. Some of these mechanisms to adapt plant architecture to conditions come into play concurrently, e.g. tillering, and some sequentially, e.g. apical branching. The plant needs to ‘decide’ on the number of primary tillers while the main stem leaves expand. With apical branching in potato, the plant ‘decides’ on the next order of branching at the time when the expansion of the lower orders of branching is completed (Vos and Biemond, 1992). The hierarchy and interplay of these mechanisms of adaptation determine the overall response at the level of the plant and canopy. There is growing interest in and need for architectural modelling of plants (e.g. Birch et al., 2003b). Such models can only accommodate effects of N limitation if these hierarchies and the underlying mechanism of regulation are better understood. Na at leaf level is a determinant of radiation use efficiency (RUE) at canopy level (Gimenez et al., 1994; Hammer and Wright, 1994; Bange et al., 1997). Since N limitation has relatively little effect on Na in species showing the ‘potato strategy’, it follows that RUE of intercepted radiation in these species should be relatively non-responsive to N limitation. For potato itself this was confirmed by Millard and Marshall (1986), while Booij et al. (1996) found no effect of nitrogen nutrition on RUE in expanding crops of Brussels sprouts. In species showing the ‘maize strategy’ the sensitivity of Na to N limitation should also show RUE being dependent on the degree of N limitation, as observed in Muchow and Davis (1988) and Muchow and Sinclair (1994). Be´ langer et al. (1992) concluded that in tall fescue swards there was little effect of N limitation on cumulative light interception, unless the nitrogen concentration dropped below 50% of the optimum concentration, but RUE increased with N rate over the entire range of supply. The degree of sensitivity of RUE to nitrogen limitation could therefore serve as an indicator to decide which of the two is dominating the adaptation: leaf area or areal nitrogen concentration.
5. Conclusion Faced with nitrogen limitation, plants can reduce leaf area and show relatively little change in leaf
nitrogen concentration, or alternatively, show conservative reaction in leaf area, but adapt leaf nitrogen concentration to N availability. The first strategy results in reduction in light interception per plant under N limitation, but little effect of N on radiation use efficiency. In the second strategy light interception is maximized to the detriment of radiation use efficiency. Previous work showed that the broadleaf species potato shows the first strategy. This paper presents and discusses evidence that maize exhibits the second strategy: for a large range of N supply, covering severe limitation to supra-optimal, leaf area per leaf differed by ca. 15% for the largest leaves, whereas leaf nitrogen concentration differed by a factor of about 2 in expanding leaves and more in fully grown leaves. Previous work showed the reverse for potato. In potato and maize, the light-saturated rate of leaf photosynthesis can be described as a logistic function of the nitrogen concentration per unit leaf area, irrespective whether variation in the latter arises from changes related to leaf age, or from different N supply to the plant.
Acknowledgements Thanks are due to Mr. J. Hinderink for conducting Expt 1 as an M.Sc. student, to Unifarm personnel for nursing the plants and to Mr. H. Halm for chemical analyses.
References Bange, M.P., Hammer, G.L., Rickert, K.G., 1997. Effect of specific leaf nitrogen on radiation use efficiency and growth of sunflower. Crop Sci. 37, 1201–1207. Be´ langer, G., Gastal, F., Lemaire, G., 1992. Growth analysis of a tall fescue sward fertilized with different rates of nitrogen. Crop Sci. 32, 1371–1376. Biemond, H., 1995a. Effects of nitrogen on development and growth of the leaves of vegetables. 1. Appearance, expansion growth and life span of leaves of Brussels sprouts plants. Neth. J. Agric. Sci. 43, 217–232. Biemond, H., 1995b. Effects of nitrogen on development and growth of the leaves of vegetables. 2. Appearance, expansion growth and life span of leaves of leek plants. Neth. J. Agric. Sci. 43, 233–246.
J. Vos et al. / Field Crops Research 93 (2005) 64–73 Biemond, H., Vos, J., 1992. Effects of nitrogen on the development and growth of the potato plant. 2. The partitioning of dry matter, nitrogen and nitrate. Ann. Bot. 70, 37–45. Biemond, H., Vos, J., Struik, P.C., 1995. Effects of nitrogen on accumulation and partitioning of dry matter and nitrogen of vegetables. 1. Brussels sprouts. Neth. J. Agric. Sci. 43, 419–433. Birch, C.J., Vos, J., Van der Putten, P.E.L., 2003a. Plant development and leaf area production in contrasting cultivars of maize grown in a cool temperate environment in the field. Eur. J. Agron. 19, 173–188. Birch, C.J., Andrieu, B., Fournier, C., Vos, J., Room, P., 2003b. Modelling kinetics of plant canopy architecture—concepts and applications. Eur. J. Agron. 19, 519–523. Booij, R., Kreuzer, A.D.H., Smit, A.L., Van der Werf, A., 1996. Effect of nitrogen availability on dry matter production, nitrogen uptake and light interception of Brussels sprouts and leeks. Neth. J. Agric. Sci. 44, 3–19. Brown, R.H., 1978. A difference in N use efficiency in C3 and C4 plants and its implications in adaptation and evolution. Crop Sci. 18, 93–98. Coaldrake, P.D., 1985. Leaf area accumulation of pearl millet as affected by nitrogen supply. Field Crops Res. 11, 185–192. Connor, D.J., Hall, A.J., Sadras, V.O., 1993. Effect of nitrogen content on photosynthetic characteristics of sunflower leaves. Aust. J. Plant Physiol. 20, 251–263. Gastal, F., Nelson, C.J., 1994. Nitrogen use within the growing leaf blade of tall fescue. Plant Physiol. 105, 191–197. Gimenez, C., Connor, D.J., Rueda, F., 1994. Canopy development, photosynthesis and radiation-use efficiency in sunflower in response to nitrogen. Field Crops Res. 38, 15–27. Grindlay, D.J.C., 1997. Towards an explanation of crop nitrogen demand based on leaf nitrogen per unit leaf area. J. Agric. Sci. Camb. 128, 377–396. Hammer, G.L., Wright, G.C., 1994. A theoretical analysis of nitrogen and radiation effects on radiation use efficiency in peanut. Austr. J. Agric. Res. 45, 575–589. Longnecker, N., Kirby, E.J.M, Robson, A., 1993. Leaf emergence, tiller growth, and apical development of nitrogen-deficient spring wheat. Crop Sci. 33, 154–160. McCullough, D.E., Girardin, P., Mihajlovic, M., Aguilera, A., Tollenaar, M., 1994a. Influence of N supply on development and dry matter accumulation of an old and a new maize hybrid. Can. J. Plant Sci. 74, 471–477. McCullough, D.E., Aguilera, A., Tollenaar, M., 1994b. N uptake, N partitioning, and photosynthetic N-use efficiency of an old and a new maize hybrid. Can. J. Plant Sci. 74, 479–484. Millard, P., Marshall, B., 1986. Growth, nitrogen uptake and partitioning within the potato (Solanum tuberosum L.) crop, in
73
relation to nitrogen application. J. Agric. Sci., Camb. 107, 421–429. Muchow, R.C., 1988. Effect of nitrogen supply on the comparative productivity of maize and sorghum in a semi-arid tropical environment. I. Leaf growth and leaf nitrogen. Field Crops Res. 18, 1–16. Muchow, R.C., Davis, R., 1988. Effect of nitrogen supply on the comparative productivity of maize and sorghum in a semi-arid tropical environment. II. Radiation interception and biomass accumulation. Field Crops Res. 18, 17–30. Muchow, R.C., Sinclair, T.R., 1994. Nitrogen response of leaf photosynthesis and canopy radiation use efficiency in fieldgrown maize and sorgum. Crop. Sci. 34, 721–727. Novozamsky, I., Houba, V.J.G., Van Eck, R., Van Vark, W., 1983. A novel digestion technique for multi-element plant analysis. Comm. Soil Sci. Plant Anal. 14, 239–249. Radin, J.W., 1983. Control of plant growth by nitrogen: differences between cereals and broadleaf species. Plant Cell Environ. 6, 65–68. Sanderson, J.B., Daynard, T.B., Tollenaar, M., 1981. A mathematical model of the shape of corn leaves. Can. J. Plant Sci. 61, 1009–1011. Sinclair, T.R., Horie, T., 1989. Leaf nitrogen, photosynthesis, and crop radiation use efficiency: a review. Crop Sci. 29, 90– 98. Tra´ pani, N., Hall, A.J., 1996. Effects of leaf position and nitrogen supply on the expansion of leaves of field grown sunflower (Helianthus annuus L.). Plant Soil 184, 331–340. Van Loo, E.N., 1993. On the relation between tillering, leaf area dynamics and growth of perennial ryegrass (Lolium perenne L.). Thesis Wageningen Agricultural University, Wageningen. p. 169. Vos, J., Biemond, H., 1992. Effects of nitrogen on the development and growth of the potato plant. 1. Leaf appearance, expansion growth, life span of leaves and stem branching. Ann. Bot. 70, 27–35. Vos, J., Van der Putten, P.E.L., 1998. Effect of nitrogen supply on leaf growth, leaf nitrogen economy and photosynthetic capacity in potato. Field Crops Res. 59, 63–72. Walter, A., Roggatz, U., Schurr, U., 2003. Expansion kinematics are an intrinsic property of leaf development and are scaled from cell to leaf level at different nutrient availabilities. Plant Biol. 5, 642– 650. Wong, S., Cowan, I.R., Farquhar, G.D., 1985. Leaf conductance in relation to rate of CO2 assimilation. I. Influence of nitrogen nutrition, phosphorus nutrition, photon flux density, and ambient partial pressure of CO2 during ontogeny. Plant Physiol. 78, 821– 825.
Effect of nitrogen supply on leaf appearance, leaf growth, leaf nitrogen economy and photosynthetic capacity in maize (Zea mays L.) J. Vosa,*, P.E.L. van der Puttena, C.J. Birchb a
Group Crop and Weed Ecology, Plant Sciences, Wageningen University and Research Centre, Haarweg 333, 6709 RZ Wageningen, The Netherlands b The University of Queensland, Gatton 4343, Qld., Australia Received 24 May 2004; received in revised form 8 September 2004; accepted 9 September 2004
Abstract Leaf area growth and nitrogen concentration per unit leaf area, Na (g m2 N) are two options plants can use to adapt to nitrogen limitation. Previous work indicated that potato (Solanum tuberosum L.) adapts the size of leaves to maintain Na and photosynthetic capacity per unit leaf area. This paper reports on the effect of N limitation on leaf area production and photosynthetic capacity in maize, a C4 cereal. Maize was grown in two experiments in pots in glasshouses with three (0.84– 6.0 g N pot1) and five rates (0.5–6.0 g pot1) of N. Leaf tip and ligule appearance were monitored and final individual leaf area was determined. Changes with leaf age in leaf area, leaf N content and light-saturated photosynthetic capacity, Pmax, were measured on two leaves per plant in each experiment. The final area of the largest leaf and total plant leaf area differed by 16 and 29% from the lowest to highest N supply, but leaf appearance rate and the duration of leaf expansion were unaffected. The N concentration of expanding leaves (Na or %N in dry matter) differed by at least a factor 2 from the lowest to highest N supply. A hyperbolic function described the relation between Pmax and Na. The results confirm the ‘maize strategy’: leaf N content, photosynthetic capacity, and ultimately radiation use efficiency is more sensitive to nitrogen limitation than are leaf area expansion and light interception. The generality of the findings is discussed and it is suggested that at canopy level species showing the ‘potato strategy’ can be recognized from little effect of nitrogen supply on radiation use efficiency, while the reverse is true for species showing the ‘maize strategy’ for adaptation to N limitation. # 2004 Elsevier B.V. All rights reserved. Keywords: Nitrogen supply; Nitrogen concentration; Leaf growth; Leaf appearance rate; Photosynthesis; Gramineae; Broad leaf species
1. Introduction * Corresponding author. Tel.: +31 317 245795; fax: +31 317 485572. E-mail address: [email protected] (J. Vos).
Yield response to larger nitrogen, N, supply is positive until factors other than nitrogen limit higher production. The positive response is brought about
0378-4290/$ – see front matter # 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.fcr.2004.09.013
J. Vos et al. / Field Crops Research 93 (2005) 64–73
either by (i) a larger amount of radiation intercepted over the crop growth period, or (ii) a higher average daily rate of photosynthesis, or a combination of (i) and (ii). Option (i) arises from effects on the dynamics of production and senescence of leaves and option (ii) from effects on the radiation use efficiency (RUE) and the nitrogen economy of the crop. Over the last decade, the responses of potato (Solanum tuberosum L.) were analyzed at the level of plant and organ (Vos and Biemond, 1992; Biemond and Vos, 1992) and for individual leaves (Vos and Van der Putten, 1998). The most important findings were that larger supply of nitrogen enhances apical branching, meaning that more leaves are formed per plant over a longer period of time. The timing of developmental events, e.g. the rate of leaf appearance, was not affected by N. Prolonged production of vegetative organs, rather than retarded rate of leaf senescence, appeared responsible for increase in the crop duration with more N (Vos and Biemond, 1992). Under N limitation potato reduces the size of the mature leaf to such an extent that maintenance of the N concentration and photosynthetic capacity per unit leaf area is achieved (Vos and Van der Putten, 1998). ‘Downregulation’ of leaf size is achieved by reduced rate of leaf expansion rather than reduced duration of expansion, identifying an effect of N on the rate of cell division or on cell size as possible mechanisms (Walter et al., 2003). The ‘potato strategy’ of adapting leaf size results in little effect of N supply on radiation use efficiency in potato (Millard and Marshall, 1986). In potato, for a given leaf insertion number, mature leaf size can differ up to a factor 4, depending on the supply of N. Previous work by Muchow (1988) showed that leaf size in maize (Zea mays L.) was responsive to N supply, but compared to potato, the differences were relatively small. Muchow and Davis (1988) reported that RUE of maize was more responsive to N supply than was radiation interception, the latter being related to leaf area index of the canopy. Such observations lead to the hypothesis that the strategy of response to N limitation differs fundamentally between potato and maize. The strategy of potato is to maintain productivity per unit leaf area rather than maximizing leaf area and light interception per plant. The hypothesis is that in maize leaf area and light interception are maintained to the detriment of the concentration of nitrogen and the
65
photosynthetic capacity per unit leaf area (referred to by ‘maize strategy’), also with relatively little effect of N on the kinetics of leaf appearance (Muchow, 1988). It is the main objective of this paper to test that hypothesis. To that end, maize plants were grown under experimental conditions similar to the ones used in the previous potato work (Vos and Van der Putten, 1998) and subjected to a range of N treatments. Leaf dynamics and growth, photosynthetic capacity and N content of individual leaves were analyzed over time.
2. Materials and methods 2.1. Arrangement of experiments Two pot experiments, Expt 1 and Expt 2 of basically similar design, were conducted in naturally lit glasshouses. Supplemental light from 400 W SONT Agro Philips lamps (0.5 lamps m2) switched on when global solar radiation outside the glasshouse dropped below 400 W m2 during daytime, and switched off again when it exceeded 500 W m2. Daylength was limited to 12 h (the glasshouse was externally covered with a light tight roof on rails from 6 h after solar noon to 6 h before solar noon). The fraction of the daily total of sunlight (as given in standard meteorological records) received at plant level in the glasshouse varied with daylength outside the glasshouse and cloudiness, but measurements showed that this fraction was about 0.5–0.6 for photosynthetically active radiation (PAR). Day/night temperatures set at 23/18 8C (12 h/12 h). Twelve litre pots were filled with sand. In Expt 1, the initial N content of the medium was not measured, but the substrate was richer than in Expt 2 (Expt 2: approx. 20 mg N kg1 of medium and estimated mineralization during the experimental period of 8 mg kg1). Pots with one plant each were initially spaced at 3 pots m2. As plants were harvested, density declined to 1 pot m2, i.e. plants were treated as single, spaced plants. Expt 1 was planted on 4 August 1997 and plants emerged on 9 August. Expt 2 was planted on 27 March 1998; emergence was on 1 April. In Expt 1, plants with three nitrogen treatments (Table 1) were grown over 101 days from emergence. In Expt 2, a wider range of N supply was used over five treatments (Table 1) and plants were grown for 78 days
66
J. Vos et al. / Field Crops Research 93 (2005) 64–73
Table 1 Amounts of nitrogen added in nutrient solution per treatment in both experiments Expt code
Code of nitrogen treatment N1
N2
N3
N4
N5
Expt 1 Expt 2
0.84 0.50
2.5 1.0
6.0 2.5
4.0
6.0
Units: g N per plant.
after emergence. The cultivar was Lincoln (three-way cross; see Birch et al., 2003a for comparative performance). Silking occurred at 47 days from emergence. The topmost ear was found at leaf 7 or 8. Nutrient solutions were given from 1 week after planting onwards at weekly intervals (six applications in Expt 1 and 11 times at intervals of 7 days in Expt 2). The first two applications were half the standard rates. The solutions contained Ca(NO3)2, NH4NO3, K2SO4, KH2PO4, MgSO4, FeEDTA, ZnSO4, and other micronutrients. Total additions of P, K, Mg, Ca were 1.2, 8.0, 1.3, 1.45 g pot1, respectively. In Expt 2, a foliar application of MgSO47H2O was applied twice (in total 250 mg Mg pot1). Single pots (plants) were the experimental units. The experimental design was a split plot with six blocks and harvests as main plots and nitrogen treatments as sub plots. The six blocks occupied the available glasshouse space; hence, within each plant sample (n = 6) the variation is represented that existed in climate conditions within the glasshouse. 2.2. Data collection 2.2.1. Leaf number and leaf area Rates of leaf appearance (tips and ligules) and the rate of leaf senescence were recorded twice weekly. Leaf expansion was monitored non-destructively on leaf insertion numbers 8 and 10 in Expt 1 and leaf numbers 7 and 9 in Expt 2. Appeared area of each leaf, from whorl or ligule to leaf tip, was estimated from (Sanderson et al., 1981): Leaf area ¼ leaf length maximum width k where k is a shape factor with value 0.75, as determined in Birch et al. (2003a). Photosynthetic rates were measured using portable equipment [LCA2 from Analytical Development Company (ADC), Hoddesdon, UK]. The measure-
ments were made in a temperature-controlled room. Cuvette temperature ranged between 20 and 24 8C and inlet CO2 concentration was typically 350 mmol mol1. Airflow rate was 6–8 ml s1 and adjusted to limit the CO2 depletion to maximally 75 mmol mol1. With an external, heat-filtered light source, constant irradiance was maintained. In Expt 1, 1550 mmol m2 s1 (PAR) was achieved at the level of the leaf in the cuvette. This was less than intended and probably less than needed for full light saturation of photosynthesis in relatively young leaves. Therefore, this paper does not report photosynthesis data from Expt 1. In Expt 2, the illumination was 2300 mmol m2 s1 (PAR) at leaf level. Subsidiary measurements of the photosynthesis–light response curve of relatively young leaves ascertained that light saturation was obtained in Expt 2. A photosynthesis reading was taken after 20–25 min after closing the cuvette when the rate of CO2 exchange had been steady for 5 min. Water vapour pressure deficit in the cuvette ranged between ca. 300 and 1000 Pa, depending on the rate of transpiration. Measurements were made halfway along the length of a leaf, midway between leaf edge and midrib. The enclosed leaf area was 11 cm2 in Expt 1 and 6.25 cm2 in Expt 2. 2.2.2. Plant dry weight sampling In Expt 1, laminae of leaf numbers 8 and 10, and in Expt 2 of leaf numbers 7 and 9, were harvested and processed separately to determine their leaf area, fresh and dry weight and the concentrations of total nitrogen and nitrate in the dry matter. In Expt 1, samplings were done on 15, 37 and 57 days after tip emergence for leaf 7 and on 9, 31 and 51 days after tip appearance for leaf 9. In Expt 2, leaf sampling was done at intervals of 2 weeks, starting on 13 days after tip appearance for leaf 8 and at 6 days after tip appearance for leaf 10. To examine the degree of N limitation, whole plants were harvested on 87 and 78 days after emergence (DAE) in Expt 1 and Expt 2, respectively, for the determination of dry weights and nitrogen concentrations in leaf blades (all blades on a plant bulked together) stems plus sheaths and cob. 2.3. Chemical analyses Total nitrogen concentration was determined in dried and ground (mesh sieve 1 mm) plant material in
J. Vos et al. / Field Crops Research 93 (2005) 64–73
two steps by digestion with sulphuric acid using salicylic acid, H2O2 and selenium as additives (Novozamsky et al., 1983), followed by colorimetric determination of nitrogen in an auto-analyzer using the Berthelot reaction (Technicon Auto-analyzer II, Industrial Method No. 334-74B/W+, released January 1976, revised March 1977; Technicon Industrial Systems, Tarrytown, New York). Nitrate was extracted by shaking 500 mg of dried and ground leaf material in 50 ml water for 30 min, followed by filtration. The nitrate concentration in the extract was determined with an auto-analyzer (Auto-Analyzer II Method NL 211-91WT; Technicon Industrial Systems, Tarrytown, New York). Organic nitrogen is defined as the difference between total nitrogen and nitrate-N. 2.4. Calculations and statistics Accumulation of thermal time was calculated from the average of the daytime and nighttime using a base temperature Tb of 8 8C for this cultivar (Birch et al., 2003a); hence, 1 day equals 12.5 8C d. The association between organic nitrogen concentration per unit leaf area, Noa, and Pmax was described with the logistic equation (Sinclair and Horie, 1989; Vos and Van der Putten, 1998): 2 1 (1) Pmax ¼ Am 1 þ exp ðsðN N ÞÞ oa
o
where Am is the asymptote of Pmax (mmol CO2 m2 s1) for infinite value of Noa; s the parameter determining the steepness of the slope; and No the value for Noa (g m2) for Pmax = 0. Specific or surface leaf weight (SLW) is defined as the ratio between leaf dry weight and area per leaf. Areal nitrogen concentration is the ratio between the amount of nitrogen in the leaf and its surface area.
3. Results 3.1. General treatment effects In both experiments the objective to create treatments encompassing the range from severe N limitation to non-limiting supply (Table 2) was achieved. Plant analyses at 87 DAE in Expt 1 and
67
Table 2 Treatment effect on total dry weight per plant (g) and total N content per plant (mg) determined on 87 and 78 days after emergence in Expts 1 and 2, respectively Expt code Plant attribute Code of nitrogen treatment N1
N2
N3
Expt 1 Expt 1
Dry weight N content
112 204 199 850 2370 3470
Expt 2 Expt 2
Dry weight N content
95 480
N4
N5
L.S.D. 14 142
130 219 239 248 19 840 2180 3580 4430 370
Least significant differences, L.S.D.: P = 0.05.
78 DAE in Expt 2 show higher dry weight and N content per plant for the higher N rates. However, whereas N content continued to increase over the whole range of N supply, dry matter production did not differ between N2 and N3 in Expt 1 and between N4 and N5 in Expt 2, indicating that there was no N limitation in the treatments with the highest rates of N. The N1 treatment in Expt 2 was more severely N limited than the N1 treatment in Expt 1 (Table 2) due to lower application and use of a less fertile root medium. For non-limiting N supply dry matter production and N uptake per plant were larger in Expt 2 than in Expt 1. This was associated with the decline in daily solar radiation as time progressed in Expt 1 and the opposite in Expt 2. 3.2. Leaf dynamics and leaf expansion There was no effect of nitrogen treatments on the rates of appearance of leaf tips and ligules (Fig. 1a). The average rate of leaf tip appearance was 0.022 leaf (8C d)1; the plastochron was 45.7 8C d per leaf. The thermal duration of leaf expansion, i.e. the horizontal distance between leaf tip appearance and ligule appearance (Fig. 1a), depended on leaf number, but was not systematically affected by N supply and did not differ between experiments. Leaf numbers 7 and 9, included in detailed analyses in Expt 2, showed durations of expansion of ca. 200 and 170 8C d, respectively. Leaf senescence started at ca. 430 8C d after emergence in all treatments (Expt 2). Leaf senescence preceded faster the lower the rate of N supply. For leaf numbers 8–12 in Expt 1, the thermal duration of life spans of N1 leaves was 16–10% shorter than for N3 leaves (Fig. 1b).
68
J. Vos et al. / Field Crops Research 93 (2005) 64–73
Fig. 1. Effect of nitrogen supply on the kinetics of leaf production and life spans: (a) leaf tip appearance vs. thermal time (base 8 8C), ligule appearance and the change in the number of dead leaves as a function of thermal time (8C d) (one day = 12.5 8C d); data from Expt 2. (b) Life span from leaf tip emergence till leaf death (8C d) for insertion numbers 8–12 in Expt 1; bars represent L.S.D. (P = 0.05).
Logistic equations fitted to data on the change with time after tip appearance of visible leaf area (Fig. 2a) showed no effect of N treatment on the point in time of inflection of leaf area growth rate (details not shown), indicating that the duration of phases of expansion were not affected by N supply. The maximum rate of expansion increased with N supply, resulting in larger mature area with more N supplied (ca. 16% difference between N1 and N5 leaves). Similar results were found in Expt 1 with 14% difference among N treatments in mature area of leaf number 8, the largest leaf (data not shown). Fig. 2b shows that leaves up to number 6 grew to similar mature sizes, irrespective of N application (Expt 2; the same result was found in Expt 1—data not
Fig. 2. Effect of nitrogen supply on leaf size (data from treatments N1–N5 in Expt 2). (a) The change in area of leaf insertion number 9 vs. thermal from tip appearance. (b) Full-grown areas of all leaf insertion numbers. Bars: L.S.D. (P = 0.05).
shown). For higher leaf numbers, mature area was greater with larger the amounts of N. The relative effect of N supply on leaf size increased with leaf rank number. The cumulative area of full-grown leaves increased from 3094 cm2 per plant in N1 to 3992 cm2 per plant in N5 plants of Expt 2 (i.e. 29% difference between extreme N treatments in Expt 2; the comparable figure in Expt 1 was 15%—data not shown). 3.3. Leaf attributes In all leaves examined, leaf blade weight increased even after expansion had stopped (Fig. 3a). Therefore, specific leaf weight (SLW) not only increased with time during leaf expansion but also continued to increase afterwards (Fig. 3b). In all cases, except leaf 10 in Expt 1, there was an N effect on SLW in that
J. Vos et al. / Field Crops Research 93 (2005) 64–73
69
Fig. 3. Effect of nitrogen supply on the change in leaf properties after leaf tip appearance. (a) Leaf dry weight, (b) specific leaf weight, (c) nitrogen concentration in leaf dry matter, (d) absolute amount of nitrogen per leaf, (e) nitrogen concentration per unit leaf area, (f) nitrate-N concentration per unit leaf area. Broken lines: data from Expt 1, full-drawn lines: data from Expt 2. Symbols: circles: lowest rate of N supply, i.e. N1 treatments of Expts 1 and 2; squares: highest rate of N supply, i.e. N3 in Expt 1 and N5 in Expt 2; open symbols: leaf 10 in Expt 1 and leaf 9 in Expt 2; closed symbols: additional data points from leaf rank 7 in Expt 2. Thin bars: L.S.D. (P = 0.05) in Expt 1; bold bars: L.S.D. for Expt 2.
SLW was highest in the treatments with the largest N supply (12–17% difference). The concentration of nitrogen in leaf dry matter, Ndm, (Fig. 3c) differed substantially between N treatments from the moment the tip appeared from the whorl. For the highest rate of N supply, generally
similar maximum values of Ndm and similar change with time were observed in Expts 1 and 2. In these treatments Ndm increased till shortly after full leaf expansion was reached. Ndm was lower in N1 leaves of Expt 2 than in corresponding leaves in Expt 1; this is in accordance with a lower N supply in the N1 treatment
70
J. Vos et al. / Field Crops Research 93 (2005) 64–73
in Expt 2 than in Expt 1. The absolute amount of nitrogen in the leaf blade differed approximately fourfold between the treatments with the highest and lowest N supply in Expt 2 (Fig. 3d), and less so in Expt 1, in accordance with a narrower range of N supply. The absolute amount of nitrogen per leaf reached its maximum near or shortly after the time of ligule appearance and gradually declined thereafter, implying a continuous export of N from the leaf. Peak values occurred later in a leaf’s life when more N was supplied. Differences in leaf size explain the relative differences in absolute amounts of N as depicted in Fig. 3d for leaf 10 from Expt 1 (456 cm2) and leaf 9 from Expt 2 (608 cm2). In expanding leaves (<200 8C d from tip appearance) areal total nitrogen concentration, Nat, ranged from 0.56 g m2 N at the lowest N rate to 1.37 g m2 N at the highest (Fig. 3e). Peak values were 1.9 g m2 N in high-N treatments, occurring 125 8C d after full leaf expansion. After the peak, Nat declined gradually during the leaf’s life, maintaining differences of a factor 3–4 between the extremes of the N treatments. Peak nitrate concentrations, expressed per unit leaf area, occurred during the phase of leaf expansion and amounted to 0.08 g m2 nitrate-N (Fig. 3f). Upon completion of leaf expansion values declined from ca. 0.05 g m2 nitrate-N to negligible levels. In expanding leaves nitrate-N represented at maximum 6% of the total nitrogen; thus, any error incurred for not accounting for nitrate would be negligible for most of the leaf’s life.
Fig. 4. Nitrogen and the light-saturated rate of photosynthesis, Pmax, (carbon dioxide assimilation); data from Expt 2. (a) Change in Pmax in time after tip appearance. Symbols: circles: N1 treatments; squares: N5 treatments; open symbols: leaf 9; closed symbols: leaf 7. (b) Pmax as a function of the nitrogen concentration per unit leaf area; symbols: circles: N1; triangles: N2; squares: N5. Bars: L.S.D. (P = 0.05).
3.4. Photosynthesis There was a large effect of N treatment on Pmax (Fig. 4a). Already during the phase of leaf expansion Pmax differed by 38% between the extremes of the N treatments. With non-limiting N supply, Pmax increased from a first measured value of 41 mmol m2 s1 to a peak value of 45 mmol m2 s1 1 at 30 days (375 8C d) after leaf tip emergence. In the low-N treatment Pmax gradually declined after cessation of leaf expansion. Data points from all Pmax recordings made in Expt 2 were plotted against Noa (Fig. 4b). Except for two deviating points, obtained on 51 days after leaf tip appearance in the high-N treatment, the data from all treatments fell on one common curve, and were described well by Eq. (1).
4. Discussion In both potato and maize the association between Pmax and areal N concentration, Na could be described well by Eq. (1). Maize was characterized by lower No (estimated at 0.25 g m2 versus 0.5 g m2 in potato), steeper slope s (2.90 versus 1.74 for potato) and higher asymptote of maximum photosynthesis rate (45e mmol m2 s1 CO2 fixation versus 19 mmol m2 s1 for potato). The current maximum rate for maize fits well with literature reports showing values between 48 and 57 mmol m2 s1 (2.1 and 2.5 mg m2 s1) (Sinclair and Horie, 1989; Wong et al., 1985; Muchow and Sinclair, 1994). The current data support the view
J. Vos et al. / Field Crops Research 93 (2005) 64–73
that there is a strong association between Na and photosynthetic capacity, regardless whether variation in Na arises from variation in rate of N supply and differences in leaf age (e.g. Connor et al., 1993; McCullough et al., 1994b; Vos and Van der Putten, 1998), or differences in illumination among leaves. Also it is established that the coefficients of Eq. (1) differ much between plant species, the largest segregation being between C3 and C4 species (Sinclair and Horie, 1989; Grindlay, 1997). The study of Gastal and Nelson (1994) in the C3 grass tall fescue (Festuca arundinacea Schreb.) showed large differences in nitrogen deposition rates and ribulose 1,5-biphosphate carboxylase content in the early stages of leaf growth, i.e. before the leaf tip appears. This supports the current finding of large differences in N concentration in young expanding maize leaves (Fig. 3), although C3 and C4 species differ fundamentally in their N economy (e.g. Brown, 1978). With respect to leaf dynamics, both current experiments produced similar results, the most important of which are: (i) in maize, the rate of leaf appearance, the duration of leaf expansion and the number of leaves are not affected by nitrogen supply; (ii) the cumulative area of mature leaves on a plant differed up to ca. 30% when N supply increased from severely limiting to non-limiting rates. That implies that maize showed a conservative response of leaf size in comparison to potato where leaf size varied three- to four-fold in response to N supply, when measured under largely similar experimental conditions (Vos and Van der Putten, 1998); (iii) among the extremes in N treatments, areal N concentration, Na, differed by a factor of ca. 2 in expanding leaves (<17 days or 210 8C d from tip appearance) (Fig. 3e). At and shortly after ligule appearance three- to four-fold differences were found among extreme N treatments as regards N concentration in leaf dry matter (Fig. 3c), absolute amount of N per leaf (Fig. 3d) and areal N concentration (Fig. 3e). This is in clear contrast to potato with small differences only in Na and its decline with leaf age (Fig. 3f in Vos and Van der Putten, 1998). Points (i)–(iii) are in support of the hypothesis posed: potato and maize show contrasting strategies to deal with N limitation. Potato adapts leaf size and avoids compromizing leaf N economy and the associated photosynthetic capacity, whereas maize strives for maintenance of leaf area per leaf at the
71
expense of decreased N concentration per unit leaf area and decreased photosynthetic capacity. With some exceptions (Grindlay, 1997; McCullough et al., 1994a) it is the general finding that, over a broad range of supply rates, there is little effect of N on the rate of leaf appearance and the duration of leaf expansion. This holds true for monocots, e.g. in maize and sorghum (Sorghum bicolor L. Moench) (Muchow, 1988), perennial ryegrass (Lolium perenne L.) (Van Loo, 1993), leek (Allium porrum L.) (Biemond, 1995b) as well as for dicots, e.g. potato (Vos and Biemond, 1992) and sunflower (Helianthus annuus L.) (Tra´ pani and Hall, 1996). As there are few studies addressing area expansion and Na at leaf level, it is not straightforward to verify whether the ‘maize strategy’ is common in Gramineae, and the ‘potato strategy’ in broadleaf species. Radin (1983) hypothesized that leaf elongation rate of broadleaf species was much more sensitive to N limitation than in Gramineae and attributed this to differences in water potential in the elongation zone, because leaves of broadleaf species expand in open air, whereas the elongation zone of cereals is protected by the sheath of preceding leaves. In Muchow (1988) the area of the largest leaf of maize differed by about 33% among N treatments, i.e. comparable to the N effect observed in the current study, while Van Loo (1993) reported similar effects in perennial ryegrass. McCullough et al. (1994b) observed similar effects of nitrogen supply on Na and leaf area of maize as in the current study. Dicot species showing a bigger effect of nitrogen limitation on sizes of individual leaves than on leaf nitrogen concentration include sunflower (Tra´ pani and Hall, 1996) and Brussels sprouts (Brassica oleracea L. var. gemmifera DC) (Biemond, 1995a; Biemond et al., 1995). In Radin’s (1983) observations C3 and C4 Gramineae behaved similar. The cited studies support the hypothesis that the ‘maize strategy’ is more common in Gramineae (both C3 and C4) and the ‘potato strategy’ more common in dicot species. Adaptation of leaf size and Na are the primary options for response to N limitation in maize, Brussels sprouts and sunflower as these species commonly do not tiller or branch. Small grain C3 Gramineae, but also several C4 Gramineae, e.g. pearl millet (Coaldrake, 1985), show a variable degree of tillering, commonly very responsive to nitrogen supply (Longnecker et al., 1993). Vos and Biemond (1992) showed
72
J. Vos et al. / Field Crops Research 93 (2005) 64–73
the degree of basal and apical branching to be responsive to nitrogen supply in potato. Some of these mechanisms to adapt plant architecture to conditions come into play concurrently, e.g. tillering, and some sequentially, e.g. apical branching. The plant needs to ‘decide’ on the number of primary tillers while the main stem leaves expand. With apical branching in potato, the plant ‘decides’ on the next order of branching at the time when the expansion of the lower orders of branching is completed (Vos and Biemond, 1992). The hierarchy and interplay of these mechanisms of adaptation determine the overall response at the level of the plant and canopy. There is growing interest in and need for architectural modelling of plants (e.g. Birch et al., 2003b). Such models can only accommodate effects of N limitation if these hierarchies and the underlying mechanism of regulation are better understood. Na at leaf level is a determinant of radiation use efficiency (RUE) at canopy level (Gimenez et al., 1994; Hammer and Wright, 1994; Bange et al., 1997). Since N limitation has relatively little effect on Na in species showing the ‘potato strategy’, it follows that RUE of intercepted radiation in these species should be relatively non-responsive to N limitation. For potato itself this was confirmed by Millard and Marshall (1986), while Booij et al. (1996) found no effect of nitrogen nutrition on RUE in expanding crops of Brussels sprouts. In species showing the ‘maize strategy’ the sensitivity of Na to N limitation should also show RUE being dependent on the degree of N limitation, as observed in Muchow and Davis (1988) and Muchow and Sinclair (1994). Be´ langer et al. (1992) concluded that in tall fescue swards there was little effect of N limitation on cumulative light interception, unless the nitrogen concentration dropped below 50% of the optimum concentration, but RUE increased with N rate over the entire range of supply. The degree of sensitivity of RUE to nitrogen limitation could therefore serve as an indicator to decide which of the two is dominating the adaptation: leaf area or areal nitrogen concentration.
5. Conclusion Faced with nitrogen limitation, plants can reduce leaf area and show relatively little change in leaf
nitrogen concentration, or alternatively, show conservative reaction in leaf area, but adapt leaf nitrogen concentration to N availability. The first strategy results in reduction in light interception per plant under N limitation, but little effect of N on radiation use efficiency. In the second strategy light interception is maximized to the detriment of radiation use efficiency. Previous work showed that the broadleaf species potato shows the first strategy. This paper presents and discusses evidence that maize exhibits the second strategy: for a large range of N supply, covering severe limitation to supra-optimal, leaf area per leaf differed by ca. 15% for the largest leaves, whereas leaf nitrogen concentration differed by a factor of about 2 in expanding leaves and more in fully grown leaves. Previous work showed the reverse for potato. In potato and maize, the light-saturated rate of leaf photosynthesis can be described as a logistic function of the nitrogen concentration per unit leaf area, irrespective whether variation in the latter arises from changes related to leaf age, or from different N supply to the plant.
Acknowledgements Thanks are due to Mr. J. Hinderink for conducting Expt 1 as an M.Sc. student, to Unifarm personnel for nursing the plants and to Mr. H. Halm for chemical analyses.
References Bange, M.P., Hammer, G.L., Rickert, K.G., 1997. Effect of specific leaf nitrogen on radiation use efficiency and growth of sunflower. Crop Sci. 37, 1201–1207. Be´ langer, G., Gastal, F., Lemaire, G., 1992. Growth analysis of a tall fescue sward fertilized with different rates of nitrogen. Crop Sci. 32, 1371–1376. Biemond, H., 1995a. Effects of nitrogen on development and growth of the leaves of vegetables. 1. Appearance, expansion growth and life span of leaves of Brussels sprouts plants. Neth. J. Agric. Sci. 43, 217–232. Biemond, H., 1995b. Effects of nitrogen on development and growth of the leaves of vegetables. 2. Appearance, expansion growth and life span of leaves of leek plants. Neth. J. Agric. Sci. 43, 233–246.
J. Vos et al. / Field Crops Research 93 (2005) 64–73 Biemond, H., Vos, J., 1992. Effects of nitrogen on the development and growth of the potato plant. 2. The partitioning of dry matter, nitrogen and nitrate. Ann. Bot. 70, 37–45. Biemond, H., Vos, J., Struik, P.C., 1995. Effects of nitrogen on accumulation and partitioning of dry matter and nitrogen of vegetables. 1. Brussels sprouts. Neth. J. Agric. Sci. 43, 419–433. Birch, C.J., Vos, J., Van der Putten, P.E.L., 2003a. Plant development and leaf area production in contrasting cultivars of maize grown in a cool temperate environment in the field. Eur. J. Agron. 19, 173–188. Birch, C.J., Andrieu, B., Fournier, C., Vos, J., Room, P., 2003b. Modelling kinetics of plant canopy architecture—concepts and applications. Eur. J. Agron. 19, 519–523. Booij, R., Kreuzer, A.D.H., Smit, A.L., Van der Werf, A., 1996. Effect of nitrogen availability on dry matter production, nitrogen uptake and light interception of Brussels sprouts and leeks. Neth. J. Agric. Sci. 44, 3–19. Brown, R.H., 1978. A difference in N use efficiency in C3 and C4 plants and its implications in adaptation and evolution. Crop Sci. 18, 93–98. Coaldrake, P.D., 1985. Leaf area accumulation of pearl millet as affected by nitrogen supply. Field Crops Res. 11, 185–192. Connor, D.J., Hall, A.J., Sadras, V.O., 1993. Effect of nitrogen content on photosynthetic characteristics of sunflower leaves. Aust. J. Plant Physiol. 20, 251–263. Gastal, F., Nelson, C.J., 1994. Nitrogen use within the growing leaf blade of tall fescue. Plant Physiol. 105, 191–197. Gimenez, C., Connor, D.J., Rueda, F., 1994. Canopy development, photosynthesis and radiation-use efficiency in sunflower in response to nitrogen. Field Crops Res. 38, 15–27. Grindlay, D.J.C., 1997. Towards an explanation of crop nitrogen demand based on leaf nitrogen per unit leaf area. J. Agric. Sci. Camb. 128, 377–396. Hammer, G.L., Wright, G.C., 1994. A theoretical analysis of nitrogen and radiation effects on radiation use efficiency in peanut. Austr. J. Agric. Res. 45, 575–589. Longnecker, N., Kirby, E.J.M, Robson, A., 1993. Leaf emergence, tiller growth, and apical development of nitrogen-deficient spring wheat. Crop Sci. 33, 154–160. McCullough, D.E., Girardin, P., Mihajlovic, M., Aguilera, A., Tollenaar, M., 1994a. Influence of N supply on development and dry matter accumulation of an old and a new maize hybrid. Can. J. Plant Sci. 74, 471–477. McCullough, D.E., Aguilera, A., Tollenaar, M., 1994b. N uptake, N partitioning, and photosynthetic N-use efficiency of an old and a new maize hybrid. Can. J. Plant Sci. 74, 479–484. Millard, P., Marshall, B., 1986. Growth, nitrogen uptake and partitioning within the potato (Solanum tuberosum L.) crop, in
73
relation to nitrogen application. J. Agric. Sci., Camb. 107, 421–429. Muchow, R.C., 1988. Effect of nitrogen supply on the comparative productivity of maize and sorghum in a semi-arid tropical environment. I. Leaf growth and leaf nitrogen. Field Crops Res. 18, 1–16. Muchow, R.C., Davis, R., 1988. Effect of nitrogen supply on the comparative productivity of maize and sorghum in a semi-arid tropical environment. II. Radiation interception and biomass accumulation. Field Crops Res. 18, 17–30. Muchow, R.C., Sinclair, T.R., 1994. Nitrogen response of leaf photosynthesis and canopy radiation use efficiency in fieldgrown maize and sorgum. Crop. Sci. 34, 721–727. Novozamsky, I., Houba, V.J.G., Van Eck, R., Van Vark, W., 1983. A novel digestion technique for multi-element plant analysis. Comm. Soil Sci. Plant Anal. 14, 239–249. Radin, J.W., 1983. Control of plant growth by nitrogen: differences between cereals and broadleaf species. Plant Cell Environ. 6, 65–68. Sanderson, J.B., Daynard, T.B., Tollenaar, M., 1981. A mathematical model of the shape of corn leaves. Can. J. Plant Sci. 61, 1009–1011. Sinclair, T.R., Horie, T., 1989. Leaf nitrogen, photosynthesis, and crop radiation use efficiency: a review. Crop Sci. 29, 90– 98. Tra´ pani, N., Hall, A.J., 1996. Effects of leaf position and nitrogen supply on the expansion of leaves of field grown sunflower (Helianthus annuus L.). Plant Soil 184, 331–340. Van Loo, E.N., 1993. On the relation between tillering, leaf area dynamics and growth of perennial ryegrass (Lolium perenne L.). Thesis Wageningen Agricultural University, Wageningen. p. 169. Vos, J., Biemond, H., 1992. Effects of nitrogen on the development and growth of the potato plant. 1. Leaf appearance, expansion growth, life span of leaves and stem branching. Ann. Bot. 70, 27–35. Vos, J., Van der Putten, P.E.L., 1998. Effect of nitrogen supply on leaf growth, leaf nitrogen economy and photosynthetic capacity in potato. Field Crops Res. 59, 63–72. Walter, A., Roggatz, U., Schurr, U., 2003. Expansion kinematics are an intrinsic property of leaf development and are scaled from cell to leaf level at different nutrient availabilities. Plant Biol. 5, 642– 650. Wong, S., Cowan, I.R., Farquhar, G.D., 1985. Leaf conductance in relation to rate of CO2 assimilation. I. Influence of nitrogen nutrition, phosphorus nutrition, photon flux density, and ambient partial pressure of CO2 during ontogeny. Plant Physiol. 78, 821– 825.