Nutrient Composition and Distance from Point Placement to the Plant Affect Rice Growth

Accepted Manuscript

Title: Nutrient Composition and Distance from Point Placement to the Plant Affect Rice Growth

Author: HU Fengqin, WANG Huoyan, MOU Pu, ZHOU Jianmin

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S1002-0160(17)60393-X 10.1016/S1002-0160(17)60393-X NA

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Please cite this article as: HU Fengqin, WANG Huoyan, MOU Pu, ZHOU Jianmin, Nutrient Composition and Distance from Point Placement to the Plant Affect Rice Growth, Pedosphere (2017), 10.1016/S1002-0160(17)60393-X.

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ACCEPTED MANUSCRIPT NUTRIENT COMPOSITION AND DISTANCE AFFECTS RICE GROWTH 2

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PEDOSPHERE Pedosphere ISSN 1002-0160/CN 32-1315/P

doi:10.1016/S1002-0160(17)60393-X

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State Key Laboratory of Soil and Sustainable Agriculture, Institute of Soil Science, Chinese Academy of Sciences, Nanjing 210008 (China) 2 Ministry of Education Key Laboratory for Biodiversity Sciences and Ecological Engineering, College of Life Sciences, Beijing Normal University, Beijing 100875 (China) ∗Corresponding author. E-mail: [email protected].

ABSTRACT

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Point placement of urea is an efficient technology to improve urea use efficiency in transplanted rice (Oryza sativa L.), but it is largely unknown how nutrient composition in the point placement and the distance between point site to the plant influence rice root distribution and growth, nutrient uptake and rice grain yield. A controlled greenhouse experiment was conducted using both N and P deficient soil with point placement of N alone or N+P together at distance close to or far from the plant, with N spilt application or no N control treatment. Both nutrient composition and distance significantly affected rice root growth. Compared to N alone treatment, N+P point placement had smaller root length density and mass density, higher specific root length (SRL) around placement site, smaller whole root system, higher straw mass and grain yield and higher N and P uptake. The difference between N+P and N alone point placement was greater close to the plant compared to far from the plant. It is suggested that higher SRL around the placement site is essential for improving nutrient uptake and rice grain yield and simultaneously point placement of N and P had a synergetic positive effect on rice growth.

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Nutrient Composition and Distance from Point Placement to the Plant Affect Rice Growth HU Fengqin1, WANG Huoyan1,*, MOU Pu2, ZHOU Jianmin1

Key Words: Nutrient uptake, phosphate, rice yield, root growth, urea

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INTRODUCTION

Rice (Oryza sativa L.) has been cultivated in China for thousands of years (Fuller et al., 2009), and now is the most important food crop of the country (Guo et al., 2005). It has contributed 40% of total calorie intake of Chinese people (Cheng et al., 2007). To sustain the productivity of rice, the use of fertilizers has been continuously increased in China during the past decades (Zhang et al., 2011;Fan et al., 2012). However, the efficiency of fertilizer use in the irrigated rice system has been very low (Peng et al., 2006). The apparent recovery efficiency of applied N (REN) has been estimated only about 25% (Jin, 2012). Another report of farmers’ practice REN from Zhejiang province was as low as 15-21% mainly because of high N input and large N losses (Wang et al., 2001). Measures have been proposed to improve the nutrient use efficiency of fertilizer through using

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right product at right time and place with right placement method (Roberts, 2007; Ryan et al., 2012). A simple yet effective method that increases N use efficiency is deep-point placement of urea (Bowen et al., 2004). Point placement of urea in wetland rice cultivation would largely prevent N loss from ammonia volatilization and nitrification–denitrification as compared with other conventional approaches such as N split application (Savant and Stangel, 1990; Kapoor et al., 2008). Studies have shown improved fertilizer use efficiency in transplanted rice system using point placement (Savant and Stangel, 1990; Mohanty et al., 1999; Bowen et al., 2004). However, the influence of point placement of fertilizer on rice roots has yet not been examined. Root growth and root system establishment play a crucial role in the capture and acquisition of resources, thus strongly influence nutrient use efficiency (López-Bucio et al., 2003; Jing et al., 2010).Plants respond to soil nutrient heterogeneous environment by exhibiting root morphological and physiological plasticity in nutrient foraging, whereby plants proliferate roots in patches of high nutrient concentration, and increase uptake rates as well (Drew and Saker, 1975; Gersani and Sachs, 1992; Mou et al., 1997; Wang et al., 2006; Grossman and Rice, 2012; Mou et al., 2013). Roots systems are modular (de Kroon, 2005) that allows them to be highly plastic to cope with the heterogeneous nature of soil nutrients (Hodge, 2004; 2006). The plasticity of root nutrient foraging underpins a potential approach to enhance plant nutrient capture and nutrient use efficiency through manipulation of root morphology (Jing et al., 2012). Nutrient composition in patches may have a vital influence on root plasticity (Einsmann et al., 1999; Rajaniemi, 2007). When nutrients were applied in patches to barley (Hordeumvulgare L. cv. Proctor), high concentration of phosphate, nitrate and ammonium but not potassium promoted laterals proliferation and elongation (Drew and Saker, 1975). Grain yield and leaf tissue P concentration in winter wheat increased significantly in dual knife N-P applications than in N knifed only (Leikam et al., 1983). As N and P are both present in the fertilized zone, corn (Zea mays L.) roots significantly proliferated and P uptake increased, while they did not when N applied alone (Duncan and Ohlrogge, 1958; Miller and Ohlrogge, 1958). Localized supply of P and ammonium together markedly stimulated root proliferation (especially of fine roots) of maize (Z. mays) by ammonium-induced rhizosphere acidification, thus significantly increased N and P uptake (Jing et al., 2010). The positive interaction of N and P banded together could be explained by the following two reasons: 1) P is one of the least available nutrients to plants due to high levels of phosphate fixation (Raghothama, 1999; Vance et al., 2003). Localized application of P reduced soil-fertilizer contact thus reduced the extent of P fixation (He et al., 2003). 2) Roots stimulated in the bands provided greater root surface area which could increase nutrient uptake (McConnell et al., 1986; Ma et al., 2013). The distance between plant and fertilized location influences the probability of detecting nutrient by the crop roots, and eventually affects nutrient uptake of crops and fertilizer use efficiency (Sander and Eghball, 1999). As 32P-labeled ammonium polyphosphate solution was injected into circles that 16, 32, 48 or 64 cm away from the corn seeds, the distance to P application spot affected plant dry weight at early stages, the closer the P was applied, the greater the dry matter production would be. While little effect of P application distance on dry weight was observed at maturity (Eghball and Sander, 1989). As the solution of 15N-labeled ammonium nitrate were injected at distance of 1, 5, 10 or 15 cm from spring wheat (Triticumae stivum L. cv. Dragon) rows, the distance effect was more pronounced in the elongation phase, but diminished at maturity (Petersen, 2001; Petersen and Mortensen, 2002). When a 15-15-15 NPK fertilizer was

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applied at 10 and 15 cm from maize individuals, plant growth and yield were both significantly higher than at 20 cm distance (Adekayode, 2010).The effects of distance between plant and fertilizer location on plant growth can easily be observed at early stages when most roots were concentrated in the nutrient-rich patch (Eghball and Sander, 1989; Li et al., 2012). When plant roots fully established at late stages, the proportion of roots in the nutrient patches decreased, so the differences may gradually weakened (Petersen, 2001; Petersen and Mortensen, 2002). The objective of this study was to investigate the influences of nutrient composition (N or N+P) in point placement and the distance from placement site to plant on 1) the growth and distribution of rice roots, 2) the grain yield, and 3) fertilizer accumulation in plants to evaluate N use efficiency. The difference of fertilizer application methods (point placement vs. N split supply) on rice roots, plant growth and N use efficiency were also examined. Compared to homogenous environment, most plants can respond to soil nutrient heterogeneity by proliferating roots in the patches (Robinson, 1994; Hodge, 2004). Point placement of fertilizer in the soil is one way to create heterogeneous supply of nutrients available to plants. Banded N and P together has been proved to increase nutrient uptake and plant growth by massive proliferation of roots. So we predict that 1) comparing with N split supply, point placement can promote rice growth and nutrient uptake through inducing root proliferation, especially N and P together point placement, 2) closer distance will increase rice growth and nutrient uptake.

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MATERIALS AND METHODS

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Experimental design

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The experiment was conducted in a greenhouse at Jiangsu Academy of Agricultural Sciences in 2014. The silt loam soil used in the experiment was collected from a long-term experiment with no fertilization for 5 years (Luhe, Nanjing, 32° 40' N, 118° 84' E). It contained(per kg) 9.5 g organic carbon, 10.6mg available N, 2.8 mg Olsen-P and 86 mg available K, with an initial pH of 8.3 (soil to water ratio of 1:5). The soil was air-dried and passed through a 3 mm sieve prior to use. Seeds of rice (O. sativa L. cv. Wuyunjing 24) were sowed in germination pans containing cleaned moist perlite10-cm deep with 1 cm of perlite on top of the seeds. Two seedlings were transplanted per PVC box(LWH =30×15×35cm3without drainage holes) as a hill when they had 4-5 leaves (about 30 d after sowing) at the center of one third along the box (Fig. 1). The experiment consisted of six treatments: (1) CN: point placement of N close (2.5 cm) to the plant, (2) CN+P: simultaneous point placement of N and P close to the plant, (3) FN: point placement of N far (10cm) from the plant, (4) FN+P: simultaneous point placement of N and P far from the plant. In addition of the 2 x 2 factorial combination of nutrient compositions and distances, two more treatments were included: (5) NS: conventional farmers’ practice of N split application with basal N broadcasting and two topdressings, and(6) NC: N control for efficiency calculation, with no N applied. Each treatment was replicated six times. The pots were completely randomized in the greenhouse. Plants per box received the following fertilizers as a basal dose (except N):0.9 g N as urea powder, 0.6 g P2O5 as monocalcium phosphate, 0.6 g K2O as potassium chloride and trace elements Mn as MnCl2•4H2O, Zn as ZnSO4•7H2O and Cu as CuSO4•5H2O in solution (equivalent

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to N-P2O5-K2O: 60-40-40 mg.kg-1 and Mn-Zn-Cu: 0.4-8×10-3-8×10-3 mg.kg-1 soil). The dose of N was split into three and applied as follows: 40% as a basal dose by mixing it with the soil; 20% at the tillering stage, 10 days after transplanting, by spreading the urea powder into the water; and 40% at the jointing stage, 25 days after transplanting. The basal dose was given as follows: all the fertilizers were mixed thoroughly with 15 kg soil and then the mix was slowly poured into each experimental box along with 4 kg tap water. After one day recompaction the soil depth in the pots was about 27 cm. Point placements of fertilizers was applied only once with full dose at one day before transplanting by removing a 5 cm × 2.5 cm diameter soil core, then nutrient was put into the hole, and refilling it with displaced soil. Thus the point of placement was 5cm below the soil surface (vertical distance) and either 2.5 cm or 10 cm away from the seedlings (lateral distance) (Fig. 1).

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Rice management

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Harvest and measurements

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The plants were harvested at maturity(130 d after transplantation). Rice was cut at 5 cm above the soil surface, and the aboveground parts were then separated into grain and straw. The values mentioned later were the total of two plants the made up each hill. To evaluate fertilizer placement on root proliferation, soil cubes of 10 × 10 × 10 cm3 were taken around the point of placement in such a way that the placement site formed the midpoint of the cubes. In NS and NC treatment, two soil cubes were removed from each box in such a way that one cube corresponded to the cube that represented the close placement and the other cube represented the far placement. Soil cubes were washed with tap water on a 20-mesh sieve and roots were hand-picked. The roots in the remaining soil of each box were collected the same way. The roots were detached at the connection point from the stubble if there was any and the remaining stubble was added to the straw mass. All the cube root samples were scanned on an Epson Expression 11000XLscanner (Epson Corporation, Nagano, Japan)with threshold setting of 174 and resolution of 300 dpi. Root length from images was analyzed using Win RHIZO 2013a software (Regent Instruments, Quebec City, Quebec, Canada). All the grain, straw (including stubbles) and root (in the cubes and remaining soil) of each box were oven-dried to constant weight at 75°C and weighed. They were then ground to pass through 80-mesh sieve and about 0.2 g samples were digested by 5 ml 98% sulphuric acid and about 6 ml of 30% v/v hydrogen peroxide (Johnson and Ulrich, 1959). N and P concentration in the digested solution was determined by SmartChem 200 auto-analyzer (Westco scientific instruments inc., Brookfield, Connecticut, USA).

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Transplanted rice was grown under waterlogging conditions (4-5cm deep water). Fungicide (phosethyl-Al) and insecticide (avermectin-hexaflumuron) were sprayed throughout the experiment monthly. The boxes were re-randomized before each spraying. All boxes were maintained weed-free by hand weeding. Irrigation was stopped 10 days before harvest. The greenhouse was set an overall temperature fluctuation between 22 and 40 °C, relative humidity of 40-80% with daytime of 11-14 hours. Light intensity at top of plants was approximately 800 µmol·m−2·s−1photosynthetic photon flux density (PPFD) during the day.

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For roots in the cubes, four variables were computed, they were root length density, root mass density (root length/ root mass in the cube divided by the volume of the cube), specific root length (SRL, the ratio of the root length to their dry mass) and root mass percentage (root mass in the cube as a percentage of the total root mass).Apparent recovery efficiency of N was evaluated by using the formula: (rice N uptake of fertilized – rice N uptake of unfertilized) / N fertilizer applied (Ladha et al., 2005). Data analysis Root density decreased strongly as distance from plant increased, so root length density, root mass density, SRL and root mass percentage in the close and far cube were analyzed separately using one-way ANOVA. In order to detect the effects of nutrient composition and nutrient point distance from the plant on rice growth (grain, straw and root), root/shoot ratio, N,P accumulation and REN, two-way ANOVA was carried out for the first four treatments. Then one-way ANOVA was carried out for all the six treatments. To meet the requirement of homoscedasticity, root mass percentage was square-root-transformed, and the other parameters were ln-transformed when necessary. Duncan’s multiple comparison was used for post-hoc comparison. All statistical analyses were performed in SPSS v. 20 (IBM, Chicago, Illinois, USA).

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RESULTS

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Roots in the close and far cube

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In the closer cube, nutrient treatment significantly influenced root length density, root mass density, specific root length (SRL) and root mass percentage (P< 0.001, Table 3). Compared toNS, CN and CN+P increased root proliferation, but the trends in root length density and root mass density under N and N+P treatment differed (Fig. 2A, B). The highest root length density was recorded in point placement of N followed by point placement of N+P; they were all significantly higher compared to NS and NC(Fig. 2A). Root mass density in CN was significantly higher compared to other treatments. No significant difference was detected in root mass density under CN+P and NS. Root mass density in CN,CN+P and NS were all significantly higher compared to NC(Fig. 2B). SRL in CN+P was significantly higher relative to CN; they were all significantly higher compared to NS and NC(Fig. 2C). CN and CN+P all had higher root mass percentage compared to NS, especially CN+P in which root mass percentage was significantly higher than that in NS. They were all significantly higher relative to NC(Fig. 2D). In the far cube, FN and FN+P significantly increased root growth than NS and NC did. The trends between root length density and root mass density were similar; with both significantly higher in FN and FN+P compared to NS and NC(Fig. 2E, F). SRL in the far cube did not differ among treatments (Fig. 2G). FN and FN+P significantly increased root mass percentage than NC and NS did (Fig. 2H).

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Grain, straw, root growth and root/shoot ratio Two-way ANOVA showed that nutrient composition significantly affected grain yield, root

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mass and root/shoot ratio. Straw and root mass were significantly influenced by distance of the nutrient point to plant. The interaction of nutrient composition and distance had significant effects on root mass and root/shoot ratio (Table 1). N+P point placement had higher grain yield, less root mass, and smaller root/shoot ratio than N alone point placement had. A significant difference in rice grain yield was detected among treatments. The highest grain yield was recorded in CN+P followed by FN+P, with the former was significantly higher than CN, FN, NS and NC(Fig. 3). There was no significant difference between FN+P, CN, FN and NS. Rice grain yield was significantly higher in all five treatments compared to that in NC(Fig. 3). Straw mass were significantly higher in CN+P compared to FN and FN+P (Fig. 3). There was no significant difference in straw mass between four treatments of nutrient point placement in comparison with NS. Straw mass was significantly higher in all five treatments relative to that in NC (Fig. 3). The largest root mass was recorded in CN and followed by FN, and both were significantly higher compared to two treatments with N+P point application (Fig. 3). In Comparison with NS, root mass increased by 11.4% in FN and decreased 8.3% to 8.6% in FN+P and CN+P, respectively, but their difference was insignificant. Root mass in NC was significantly lower relative to other five treatments (Fig. 3). Root/shoot ratio showed the same pattern as root mass. Two N point placement significantly increased rice root/shoot ratio compared to the two N+P point placement and NS(Fig. 4). Root/shoot ratio decreased by 21.3% and 4.3% in CN+P and FN+P respectively, compared to NS but the differences were insignificant.

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N and P uptake and N efficiency

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Two-way ANOVA showed that N uptake was significantly influenced by distance of nutrient point from the plant; while nutrient composition significantly affect P uptake (Table 1). CN+P significantly increased uptake of N compared to NS and NC (Table 2).CN, FN+P and FN all had higher N uptake compared to NS but their difference was insignificant. All five treatments significantly increased N uptake compared to the NC. P uptake increased significantly in CN+P relative to NS and NC(Table 2). FN+P, CN, FN and NS differed insignificantly in P uptake. N recovery efficiency in CN+P was significantly increased relative to FN, FN+P and NS(Table 2). Compared to NS, CN increased N recovery efficiency by 22.4%, FN+P by 11.1%, FN by 5.9%, but their difference was insignificant.

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DISCUSSION

Plant roots show a high degree of plasticity in their development in response to soil nutrient heterogeneity by strong lateral root proliferation (Robinson,1994, 2001; Hodge, 2004). Many studies have reported that nutrient rich patches significantly stimulated root proliferation, including increased lateral root length, biomass, and lateral root numbers (Drew and Saker, 1975;Wang et al., 2006; Mou et al., 2013; Hu et al., 2014). In this study, CN, CN+P, FN and FN+P all proliferated large amounts of fine roots in the 1,000 cm3 cubes around the fertilizer points, leading to increased root length density and mass density (Fig. 2A, B, E, F). Others reported similar findings with N+P (roots was stimulated by N and P present together) (Duncan 7

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and Ohlrogge, 1958; Jing et al., 2010; Jing et al., 2012; Ma et al., 2013; Li et al., 2014), but failed to detect root proliferation in only N rich patches (Duncan and Ohlrogge, 1958; Miller and Ohlrogge, 1958; Li et al., 2014), probably because of relatively high background N supply. Whether plant roots respond to patches and the magnitude of their response depends on the overall plant nutrient status and the contrast of nutrient concentration in patches relative to the background (Lamb et al., 2004; Blouin and Puga-Freitas, 2011; Ruffelet al., 2011, Hu et al., 2014). In this study, the background soil provided limited N (rice total biomass in N control treatment was only 45.7-55.2% of that in other treatments, Fig. 3), and all N fertilizer was point placed at one time, which caused high contrast of available N concentration around the nutrient point and the background soil. This high contrast induced a massive root proliferation. The results also showed that the magnitude of root proliferation in the nutrient-rich cubes was dependent on the nutrients presented. Root mass and length densities were greater in the cubes around N placement point than in the cubes around N+P point, especially in point placement close to the plant (Fig. 2A, B). SRL may play a vital role in this difference (Fig. 2C). SRL characterizes economic aspects of the root system (Fitter, 1991) with long thin roots (high SRL) being less expensive to construct (Eissenstat, 1991). High SRL increases the volume of soil exploited per unit biomass invested, thus has high nutrient-uptake efficiency in the patch (Ostonenet al., 2007). Higher SRL in cubes of CN+P thus had better nutrient-uptake efficiency relative to cubes of N alone, resulting in less root mass had even higher nutrient uptake. The mechanisms underlying the variable SRL responses to point placement N vs. N+P need to be elucidated. N point placement of this study increased not only the root growth around the N point but also the entire root system, while N+P point placement only increased root growth around the site of placement (Fig. 3). The background soil had also limited available P (rice total biomass in P control was 42.0-50.7% of that in other treatments, data not shown), and P mobility is very slow (diffusion coefficient ranges 10-13 to 10-15 m2s-1, Tinker and Nye, 2000). Roots in N point treatments were possibly extended out of the point cube to uptake P, whereas roots in the N+P point cube could uptake both N and P. Therefore the balance nutrient supply of the patch is important for plant to save carbon input in constructing new roots to search for the nutrient not in the patch (Eissenstat, 1992). And our results showed that the localized modifications of root growth and acquisition of N and P in nutrient-rich patches largely compensated for the deficient supply of N and P to the remainder of the root system (Robinson 1994, 1996; Ma et al., 2009), which indicated that rice can meet the demand for nutrient by only part of the roots reaching the high-nutrient patch, as partial root-zone irrigation can fully meet the water requirement of the crops (Li et al., 2007). This means nutrient uptake efficiency around the fertilizer placement site had increased, our another 15N experiment showed that the percentage of total N in plant tissue derived from the 15N-labeled urea in point placement is significantly higher compared to urea split treatment (data not shown). This had practical implications that point placement of fertilizer can increase uptake efficiency from the fertilizers (He et al., 2003).The possible reason was that point placement reduced N losses. Many studies reported that broadcasting and split application of urea by farmers in transplanted rice system is very inefficient due to several N loss processes (Savant and Stangel, 1990; Mohanty et al., 1999). Point placement of urea has been proven to reduce N losses through ammonia volatilization, denitrification and runoff (Mohanty et al., 1999; Bowen et al., 2004).Deep point placement of urea in transplanted rice system has been proved to be an agronomically efficient technology (Savant and Stangel, 1990; Bowen et al., 2004; Kapoor et al.,

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2008). This study provided convincing evidence from the aspects of rice roots, but how to point place nutrients with precision in the field is challenging. Fertilizers can be placed manually or with applicators (Mohanty et al., 1999). Manual application is drudgery and requires lots of labor (Savant and Stangel, 1990). Several machines have been developed to facilitate the placement, but they have not been adopted widely due to mechanical problems when working in muddy conditions (Bsutista et al., 2001). Precision drilling machine with direct-seeding developed by Luoet al. (2007, 2008) and Baimba et al. (2014) seems to be a promising solution. It should be noted that farmers’ practice usually needs two to three splits other than basal broadcasting, while in point application full dose of fertilizer was applied at transplanting or sowing, thus it’s labor-saving in this aspect (Bsutista et al., 2001; Baimba et al., 2014). Urea in point placement was usually shaped as supergranules (USG) or briquettes (UB) which weighed more than 1 g per granule (Savant and Stangel, 1990; Bowen et al., 2004). In this study, urea powder was used because small amount urea was to broadcast evenly to the soil surface in the box. Point placement of urea powder decreased proportionately the N contact with soil and therefore may slow the urea hydrolysis and resulted in a steep gradient of ammonium concentration at the placement site (Savant and Stangel, 1990). The spatial heterogeneity of essential resource availability can affect placement and growth of roots, the growth of whole plants and the yield (Fransen et al., 2001; Day et al., 2003; Hutching et al., 2004). This study showed that point placement of N increased root growth and maintained identical grain yield compared to NS. When N and P were simultaneously point placed, rice grain yield increased, especially placement site was close to the plant which had significantly higher grain yield than NS. It is noteworthy that CN+P resulted in the highest grain yield, straw growth, as a result of the largest N and P uptake with the smallest root investment(except N control treatment). Plant nutrients rarely work independently that interactions among nutrients are usually significant because a deficiency of one restricts the uptake and use of another (Roberts, 2008). The synergetic effect of N (especially ammonium) and P in localized supply has been reported to increased corn and wheat yield also (Leikam et al., 1983; Ma et al., 2013). Another reason N and P together point placed had higher yield than other treatments was that the alkaline soil used in this study promoted P fixation hence reduced P availability (Busman et al., 2009). Compared to P broadcast in other treatments, point placement of P reduced soil-P contact thus may provide more available P to rice (He et al., 2003). The yields obtained in this study (from 13 g/hill in NC to 27 g/hill in CN+P) were relatively lower than the yields of our field study (30 to 40 g/hill, data not shown). There were two main reasons: 1) the soil used in this study was deficient in both N and P; 2) the fertilization level we chosen was relative low compared to the filed study (N-P2O5-K2O: 60-40-40 mg.kg-1 soil in the present study vs. N-P2O5-K2O 225-150-180 kg.ha-1 in the field). One shortcoming which may affect the understanding of rice growth in the field was that competition among rice plants had not been considered in this study. In competition, plant roots may alter their plastic responses, including increased or decreased root growth (Cahill et al., 2010; Wang et al., 2012). Thus experiments integrating information about fertilizer point placement and competition among rice plants are needed to derivate more accurate management recommendations. Root/shoot partition in homogenous environment is often predicted by optimal partitioning and ontogenetic theory (Gedroc et al.,1996), but the effect of heterogeneity on R/S ratio is complex. Patch quality, scale and the contrast between patches and background all affect R/S ratio

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(Hutchings and John, 2004). We showed that point placement of N led to an excessive root growth around the placement site and the entire root system that R/S ratio significantly increased compared to NS. R/S ratio was relatively unresponsive to N+P point placement because more root growth around N+P placement site may be accompanied by compensatory decreases in root growth elsewhere with no nutrients applied. Such root growth regulations offer advantage in plant adaptation to the heterogeneous soil environment (Li et al., 2014). N uptake was significantly influenced by distance between plants and fertilizer point (Table 1). N uptake in treatments close was higher than in treatments far (Table 2) indicated that the timing of roots encountering a patch may be important to rice N accumulation (Li et al., 2012). P uptake was significantly influenced by nutrient composition (Table 1). N+P point placement had higher P uptake compared to N point placement. Application of N and P together has been reported to increase P uptake in crops (Duncan and Ohlrogge, 1958; Miller and Ohlrogge, 1958; Leikam et al., 1983). Fertilizer too close to the plant may be toxic, too far may not be reached by plant roots in early establishment stages, so that appropriate distance of fertilizer to plant is as important as fertilization rates to crops (Havlin et al., 1999). The most efficient placement of fertilizer is that provides an adequate supply in soil occupied by plant roots at critical periods of growth (Shen et al., 2013). We showed that root growth in CN was significantly higher compared to FN, and the straw mass in CN+P was significantly higher relative to FN+P. Grain yield in point treatments close were higher compared to that of point treatments far, but with insignificant differences (Fig. 3). The effects of distance between plant and fertilizer site on plant growth can easily be observed at early stages where plant roots proliferated in the patches while whole root system has not established fully (Eghball and Sander, 1989). When plant roots penetrated most of the soil at late stages, the differences may gradually diluted (Petersen, 2001; Petersen and Mortensen, 2002). There is a time lag before point fertilizer becoming available to plant roots (Savant and Stangel, 1990). Generally, the greater the distance from the point of fertilizer to the plant, the slower the fertilizer is available to them (Savant et al., 1982). Therefore there is a possibility that more fertilizer placement sites at different distances to plants may meet demands at different developmental stages. In this study, the differences between N+P and N point placement in grain yield, straw and root growth and N, P uptake were greater close to the plant than far from the plant(Fig. 3, Table 2). Two reasons may explain: 1) root mass percentage in the close cube was greater than in the far cube; and 2) SRL between N+P and N alone in the close cube significantly differed. Thus roots proliferated in the close cube played greater roles than in the far cube. The results of this study illustrated that effective fertilizer placement had improved both rice yield and nutrient use efficiency. The importance of right fertilizer placement deserves more attention. Wang and Zhou (2013) proposed the concept of root-zone fertilization which emphasized the overlap of the fertisphere (the special nutrient environment around the localized placement site of fertilizers, Su et al., 2011) and root zone by applying the proper fertilizers to the active root zone. Root zone fertilization technique was apromising approach in increasing nutrient use efficiency. More research is needed on this area.

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ACKNOWLEDGMENTS

This work is supported by Major State Basic Research Development Program of China (2013CB127401), the National Science Foundation of China (41271309), and Postdoctoral Science Foundation of Jiangsu (140064C). We thank JD Wang, ST Jiang, XW Liu, ZM Chen, YZ Liu, XL Zhao, YS Jia, and YL Wang for their help with the experiments, and three anonymous reviewers for helpful comments on the manuscript.

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Nutrient composition significantly influenced rice root distribution and root growth. N+P point placement reduced whole root mass, had smaller root mass and length densities around the placement site, had higher SRL than N alone point placement, thus higher straw mass, grain yield, and N, P uptake. The differences between N+P and N point placement in grain yield, straw and root growth, and N, P uptake were greater when close to the plant than far. Point placement of fertilizers significantly promoted root growth around placement site, increased uptake efficiency from the fertilizers. And point placement significantly increased rice yield or maintained identical yield with full dose of fertilizer applied only once compared to three to four splits of farmers’ practice. The results also demonstrated that rice can meet the demand for nutrient by only part of the root system reaching the high-nutrient zone. These findings may have substantial practical implications for agricultural production that N+P point application at appropriate distance from the plant could be vital for improving rice yield, N and P uptake from the fertilizer. More research about root zone fertilization needs to be done.

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plants. Chinese J Plant Ecol(in chinese).36: 1184–1196. Zhang F, Cui Z, Fan M, Zhang W, Chen X, Jiang R. 2011.Integrated soil–crop system management: reducing environmental risk while increasing crop productivity and improving nutrient use efficiency in China. J Environ Qual.40:1051–1057.

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Table 1 Results of two-way ANOVA for grain, straw, root mass, root/shoot ratio, N and P uptake

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Distance (D) N×D F-valu F-value P-value P-value F-value P-value e Grain 4.68 0.04* 0.70 0.41 1.64 0.21 Straw 0.09 0.76 6.53 0.02* 0.16 0.69 Root 37.34 <0.01*** 8.61 <0.01** 8.88 <0.01** Root/shoot ratio 47.03 <0.01*** 0.56 0.46 10.10 <0.01** N uptake 1.28 0.27 4.98 0.04* 0.39 0.54 P uptake 12.31 <0.01** 0.71 0.41 0.65 0.43 REN 1.28 0.27 4.98 0.04* 0.39 0.54 *, **, *** indicate significant at the 0.05, 0.01 and 0.001 probability levels, respectively. Nutrient represents nutrient composition in the point placement (N or N+P); distance represents the distance from the point placement to the plant (close or far).

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Rice nitrogen (N), phosphorus (P) uptake and recovery efficiency of N (REN) in different nutrient supply treatments. Treatments N Uptake (mg/pot) P Uptake (mg/pot) REN (%) CN 688 ±31ab 69 ±3bc 52 ±3ab CN+P 756 ±50 a 102 ±12 a 60 ±6 a FN 625 ±45ab 70 ±6bc 45 ±5 b FN+P 645 ±26 ab 87 ±5ab 47 ±3 b NS 603 ±19 b 79 ±7 b 43 ±2 b NC 225 ±8 c 56 ±4 c --Each value is the mean of six replicates (±SE). Different letters in each column denote significant difference among treatments at 5% level according to Duncan’s multiple-range test. CN: point placement of N close to the plant, CN+P: point placement of N+P close to the plant, FN: point placement of N far from the plant, FN+P: point placement of N+P far from the plant, NS: N split application, NC: no N.

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Results of one-way ANOVA for root length density, root mass density, specific root length (SRL) and root mass percentage (% of root mass in the cube to total mass of the entire root system) in 10 × 10 × 10 cm3 soil cube around the close and far nutrient placement point.

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Fig. 1 Illustration of locations of rice plants and point placement in the experimentalbox. X marks the location of the rice plants. Dot A and B represent nutrient point placement site close to and far from the plants respectively, both were 5cm below soil surface.

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Fig. 3 Effects of different nutrient supply treatments on grain, straw and root growth in rice. Values are means of six replicates (+SE). Different letters denote significant difference among treatments at 5% level according to Duncan’s multiple-range test. Different letters with ‘ and ‘’represent significant difference in straw and root mass respectively. CN: point placement of N close to the plant, CN+P: point placement of N+P close to the plant, FN: point placement of N far from the plant, FN+P: point placement of N+P far from the plant, NS: N split application, NC: no N.

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Fig. 4 Effects of different nutrient supply treatments on rice root/shoot ratio. Values are means of six replicates (+SE). Different letters denote significant difference among treatments at 5% level according to Duncan’s multiple-range test. CN: point placement of N close to the plant, CN+P: point placement of N+P close to the plant, FN: point placement of N far from the plant, FN+P: point placement of N+P far from the plant, NS: N split application, NC: no N.

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