morphological and physiological responses of rice roots and shoots to varying water regimes and soil...

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This article was downloaded by: [North Dakota State University] On: 21 October 2014, At: 19:49 Publisher: Taylor & Francis Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK Archives of Agronomy and Soil Science Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/gags20 Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities Abha Mishra a & Norman Uphoff b a School of Environment, Resources and Development , Asian Institute of Technology , Klong Luang , Pathumthani , Thailand b Cornell International Institute for Food, Agriculture and Development , Ithaca , USA Accepted author version posted online: 22 Mar 2012.Published online: 28 May 2012. To cite this article: Abha Mishra & Norman Uphoff (2013) Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities, Archives of Agronomy and Soil Science, 59:5, 705-731, DOI: 10.1080/03650340.2012.669474 To link to this article: http://dx.doi.org/10.1080/03650340.2012.669474 PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms &

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Page 1: Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities

This article was downloaded by: [North Dakota State University]On: 21 October 2014, At: 19:49Publisher: Taylor & FrancisInforma Ltd Registered in England and Wales Registered Number: 1072954 Registeredoffice: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK

Archives of Agronomy and Soil SciencePublication details, including instructions for authors andsubscription information:http://www.tandfonline.com/loi/gags20

Morphological and physiologicalresponses of rice roots and shootsto varying water regimes and soilmicrobial densitiesAbha Mishra a & Norman Uphoff ba School of Environment, Resources and Development , AsianInstitute of Technology , Klong Luang , Pathumthani , Thailandb Cornell International Institute for Food, Agriculture andDevelopment , Ithaca , USAAccepted author version posted online: 22 Mar 2012.Publishedonline: 28 May 2012.

To cite this article: Abha Mishra & Norman Uphoff (2013) Morphological and physiological responsesof rice roots and shoots to varying water regimes and soil microbial densities, Archives of Agronomyand Soil Science, 59:5, 705-731, DOI: 10.1080/03650340.2012.669474

To link to this article: http://dx.doi.org/10.1080/03650340.2012.669474

PLEASE SCROLL DOWN FOR ARTICLE

Taylor & Francis makes every effort to ensure the accuracy of all the information (the“Content”) contained in the publications on our platform. However, Taylor & Francis,our agents, and our licensors make no representations or warranties whatsoever as tothe accuracy, completeness, or suitability for any purpose of the Content. Any opinionsand views expressed in this publication are the opinions and views of the authors,and are not the views of or endorsed by Taylor & Francis. The accuracy of the Contentshould not be relied upon and should be independently verified with primary sourcesof information. Taylor and Francis shall not be liable for any losses, actions, claims,proceedings, demands, costs, expenses, damages, and other liabilities whatsoever orhowsoever caused arising directly or indirectly in connection with, in relation to or arisingout of the use of the Content.

This article may be used for research, teaching, and private study purposes. Anysubstantial or systematic reproduction, redistribution, reselling, loan, sub-licensing,systematic supply, or distribution in any form to anyone is expressly forbidden. Terms &

Page 2: Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities

Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions

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Page 3: Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities

Morphological and physiological responses of rice roots and shoots to

varying water regimes and soil microbial densities

Abha Mishraa* and Norman Uphoffb

aSchool of Environment, Resources and Development, Asian Institute of Technology,Klong Luang, Pathumthani, Thailand; bCornell International Institute for Food, Agriculture and

Development, Ithaca, USA

(Received 26 September 2011; final version received 17 February 2012)

To understand the responses of rice roots and shoots to non-continuously-floodedsoil–water regimes together with varied levels of soil microbial density, studieswere conducted evaluating three different water regimes: intermittent flooding(IF) through the vegetative stage (IF-V), IF extended into the reproductive stage(IF-R), and soil maintained with no standing water (NSW); and three levels ofsoil microbial density: normal, reduced (autoclaved), and enhanced (EMS). Atflowering, EMS-treated plants were found to have increased root-length densityand greater root oxidizing activity rates under both IF-V and IF-R and moreavailable soil nitrogen (ASN) under IF-V. Earlier senescence of plants grown withEMS was also observed under all water treatments. Water regime was seen tohave a major effect on grain yield with a number of causal mechanisms involved.Significant relationships were observed between root oxidizing activity rate(ROA) and ASN, and between ROA and the chlorophyll content of lower leaves.The EMS effects were apparently not caused by microorganisms directly, butrather by differences in ASN facilitated by interaction between water regime andsoil microbial activity. The latter would probably vary with different levels of soilorganic carbon, but this additional parameter was not investigated. Further studyon the role of soil organic matter under similar conditions is warranted for betterunderstanding of these relationships, which have significant effects on rice grainyield.

Keywords: microbial activities; moisture; plant plasticity; root-length density; rootoxidizing activity

Introduction

Sustainable intensification of rice production under conditions of increasing waterscarcity, deteriorating soil fertility and increasing costs of energy presents a majorchallenge to rice growers, particularly in Asia. To address these constraints,attention is being given to altering the currently predominant practice of continuousflooding of rice paddies, either through intermittent irrigation or by keeping paddysoil moist, but not always inundated (Belder et al. 2002; Bouman et al. 2007). At thesame time, there is a growing interest in utilizing plant-growth-promoting

*Corresponding author. Email: [email protected]

Archives of Agronomy and Soil Science

Vol. 59, No. 5, May 2013, 705–731

ISSN 0365-0340 print/ISSN 1476-3567 online

� 2013 Taylor & Francis

http://dx.doi.org/10.1080/03650340.2012.669474

http://www.tandfonline.com

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microorganisms (PGPM) as biofertilizers to enhance soil and plant productivity(Vessey 2003; Banerjee et al. 2005). There is an expectation that these can enhancenutrient availability for crops grown under such non-flooded soil conditions (Khanet al. 2007). Although the complexity of interactions among PGPMs and theirsynergistic effects are poorly understood, there is evidence that microbial diversityand activity may influence plant growth (van der Heijden et al. 2008; Anas et al.2011).

Regarding the cessation of continuous flooding, some previous reports havesuggested that under unsaturated soil moisture conditions, there is a significantdecrease in dry matter production and grain yield for rice (McCauley 1990; Mishraet al. 1997). This could be due to a rapid rate of loss of nitrogen resulting fromnitrification and denitrification (Sah and Mikkelsen 1983). However, others havereported higher yields associated with intermittent irrigation during the vegetativestage (Ceesay et al. 2006), and even under post-anthesis water-deficit conditionswhen organic matter has been applied to the soil (Yang et al. 2003; Yang et al. 2004),attributable to enhanced soil microbial activity.

These and other reports on the effects of intermittent irrigation/non-floodedwater regimes and of enhanced soil microbial activity and density are not consistent,leaving some important questions unanswered. In particular, they did not assesshow rice plants’ roots and shoots will grow and perform when subjected todifferent soil moisture conditions in conjunction with different levels of soil microbialactivity.

It is believed that the promotion of vigorous root growth for the purpose ofstrengthening plants’ drought tolerance or enhancing their ability to enduremoisture stress can improve rice productivity on paddy soils that are notcontinuously flooded (Tajima 1995). Several research groups have shown thatwhen soil dries due to a cessation of flooding, water deficits first appear near thesurface and then progress to the deeper layers, making increased depth of plantrooting common under these conditions (Proffitt et al. 1985; Newell and Wilhelm1987; Mishra and Salokhe 2008). Gallardo et al. (1994) found in a pot study thatdrought effects increased root length and weight in the lower soil layer where onlythe upper third of the soil had been allowed to dry. Such changes in rootmorphology induced by soil moisture variability have been often observed, but theirrelationships with shoot growth and to other physiological responses such as yieldhave not been explored in detail.

It is known that water and nitrogen supply generally interact with eachother and are important factors affecting higher rice yield and root development(Skinner et al. 2004). One of the important root parameters that reflectsroot morphology and which is known to be affected by soil nitrogen level isroot-length density (RLD). This influences not only the amount of root–microbialinteraction, but also the physiological activity of the roots themselves, i.e., rootoxidizing activity (ROA), which plays an important role in increasing plants’photosynthetic capacity (Mitsui and Tensho 1953; Ookawa et al. 2003; Mishraet al. 2006).

It is also known that cytokinin production is regulated by root activity and isgreatly influenced by soil moisture condition, available nitrogen and the NO3

7/NH4þ

ratio. The level of NO37 in the soil increases the auxin/cytokinin balance required for

tiller and biomass production, while availability of NH4þ shifts this ratio in favor of

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cytokinin production (Mercier et al. 1997), which probably delays root senescence atlater growth stages (Mishra et al. 2006).

Researchers have demonstrated that rice plants with a higher ROA rate duringtheir later growth stages have higher grain yield (Osaki et al. 1997; Zhang et al.2008). In this context, a recent study has shown that paddy yield was increasedwhen rice plants were grown with intermittent irrigation during their vegetativestage when having reduced intra-hill competition (Mishra and Salokhe 2010). Thisincrement in yield was attributable to delayed senescence of roots and shoots,along with a higher ROA rate of the plants. However, the mechanisms of delayedsenescence and the relevance of delayed senescence for increases in grainproduction are not fully understood, especially in the context of varying nitrogenavailability. The latter is affected in both amount and form by both soil waterconditions and microbial activities.

In addition, the roles of soil microbes in influencing important root and shoottraits, and the associated effects of these traits on yield, are seldom examined,although there are now many different microbial biofertilizers available foragricultural use. One biofertilizer that has received a lot of attention is the microbialinoculum called Effective Microorganisms (EM), developed by Teruo Higa at theUniversity of the Ryukyus in Okinawa, Japan (Higa 1991). This has been describedas a combination of * 80 co-existing beneficial microorganisms (Higa and Parr1995). The main species included in EM preparation are: Lactobacillus plantarum,Lactobacillus casei, Streptococcus lactis; the photosynthetic bacteria Rhodopseudo-monas palustris and Rhodobacter sphaeroides; the yeasts Saccharomyces cerevisiaeand Candida utilis; the actinomycetes Streptomyces albus and Streptomyces griseus;and finally the fungi Aspergillus oryzae and Mucor hiemalis (Xu 2000).

It is not clear whether EM contains microorganisms that are acting as PGPM,but Higa (2001, 2003) has suggested a broad range of applications for EMpreparations, with reports of beneficial effects in quite different environments such assoils, plant and water. Very few reports are available, however, on the effects of EMon crop yield, plant development or soil processes. There is some evidence that EMapplications increase microbial density and activity in the soil (Dilly and Blume1996; Schenck zu Schweinsberg-Mickan and Muller 2009). Therefore, we decided toexamine the effects of soil microbial density and activity on root–shoot traits underalternative soil water regimes using soil applications of EM microorganisms,assessing also soil nitrogen availability and any associated effects on plant growthand performance. This could illuminate the types and range of adaptivephysiological and morphological responses of plants to different levels of microbialactivity under such different soil–water conditions.

Here, we report on research findings that explored several causal relationshipsaffecting the morphological and physiological responses of rice roots and shootsunder alternative water regimes, with concurrent investigation of the effects withdifferent soil microbial levels, using EM application as one of the treatments. Theseparameters are investigated to get a better understanding of the effects thatalternative water regimes and different soil microbial densities can have on the soil’savailable nitrogen status and on the morphological and physiological developmentof rice plants’ roots and shoots. We were looking for any causal relationships and forany evidence that these could significantly affect yield-contributing parameters andgrain yield.

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Methods

Experiments were conducted from 2005 to 2006 in an open-sided greenhouse withplastic roof (light transmissivity 93%) at the Asian Institute of Technology, Bangkok(Thailand), at 148040N and 1008370E with 2.27 m altitude. Average temperaturerange was 30–358C and relative humidity (RH) was 75–85%.

Black clay soil (soil series: ongkharak; taxonomy: very fine, mixed acid, isohypersulfic tropaquepts) was collected from the rice research farm of the Asian Institute ofTechnology where the previous crop grown was rice. Soil was sampled from a depthof 0–10 cm, and the samples were analyzed for soil pH, cation exchange capacity(CEC), total N (Kjeldahl method) and plant available K (atomic absorptionspectrophotometer), plant available P (Bray-2) and organic C (automatic nitrogenand carbon analyzer-mass spectrometer). Average composition of the soil was 10.2%sand, 23.2% silt and 66.6% clay, with pH (1:1) of 5.0. Organic C was 1.38%; total N0.14%; available P 11 mg kg71; and available K (potassium) 212 mg kg71. CECwas 22.6 cmol kg71.

After air drying, the soil was crushed and crop residues were removed by hand;65 kg of soil (fresh weight) were placed in plastic pots, 60 cm high with 50 cm topand 40 cm bottom diameters, with a drainage hole of 1 cm diameter at 7 cm heightfrom the bottom of the pot. Pots’ drainage holes were fitted with a stopper, whichmaintained water except during the drainage period. Three soil and three watertreatments were evaluated respectively and jointly.

Soil treatments

(1) Normal soil (NS): pots were filled with the soil described above, having noamendments.

(2) Autoclaved soil (AUS): pots were filled with soil that was prepared usingstandard steam-sterilization methods to achieve a minimal microbialpopulation and the least microbial activity in the soil. Soils used in thesetrials were placed in plastic bags and autoclaved at 1218C with 0.103 MPapressure for 20 min.

(3) Soil supplied with EM solution (EMS): soil in these pots had enhancedabundance of microbial populations and increased microbial activity fromapplying to normal soil an EM solution (details on this are given below).

Before starting the experiments, all three soil treatments were tested for their soilrespiration rate (Anderson 1982). This assessed the microbial activity rates in therespective soils under evaluation. The respiration rates measured were: for NS,1.01+ 0.001 mg CO2 day

71 20 g soil71; for EMS, 1.53+ 0.002 mg CO2 day71

20 g71 soil; and for AUS treatment, 0.03+ 0.009 mg CO2 day71 20 g71 soil. The

EMS soil thus had heightened microbial activity (and populations) as expected,whereas AUS soil had negligible populations, also as intended.

After this measurement, all pots were flooded by adding distilled water to a depthof 3–4 cm for a week before transplanting. They were dressed with 138 mg N kg71

and 12.3 mg P kg71, applied in NPK fertilizer (16:16:0) as a basal application; thenurea (46:0:0) was added at 15 and 45 days after transplanting (DAT). Nitrogen wasapplied in an excess amount to ensure the availability of nitrogen during soil analysiseven after repeated alternate wetting and drying cycles. This ensured that relative

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comparisons could be made among the treatments. No potassium (K) amendmentswere made as the soils exhibited amply supply of this macronutrient.

Single 15-day-old seedlings grown in a dry seedbed were transplanted when theyhad two fully expanded leaves, within 2 h of uprooting from the nursery seedbed andat a depth of 1.5 cm. The rice cultivar planted was Pathumthani, a photoperiod non-sensitive, high-yielding modern variety with an average maturity period of 120–130days.

Water treatments

Seven days after transplanting, when transplanting shock had disappeared, threedifferent water treatments were introduced to each soil treatment in order to introducedifferent degrees and kinds of water stress to the plants at various growth stages:

(1) Intermittent flooding through the vegetative growth phase (IF-V): pots weremaintained with the following water regime: 5 cm depth of water wasmaintained every day for 12 days; then the pots were drained for 3 days andthen reflooded with the same depth of standing water. Three drying periodsof 3 days each were provided at 19, 34 and 50 DAT, followed by continuousflooding with 5 cm water depth until crop maturity. In this treatment, thedraining cycle continued during the vegetative stage only.

(2) Intermittent flooding into the reproductive phase (IF-R): a similar procedurewas followed as with IF-V, but with draining done five times for 3-dayperiods, at 19, 34, 50, 66 and 82 DAT; this was followed by flooding withstanding water of 5 cm depth until maturity. Here the flooding and drainingcycle was continued into the reproductive stage.

(3) No standing water (NSW) on the soil surface – the soil in these pots wasmaintained with soil moisture at field capacity (FCp) at 235 mm soil depth,keeping the top soil in a non-flooded, unsaturated condition. To maintainthe soil at lower depths at field capacity (with 0.378 m3 m73 volumetric soilwater content), a moisture meter (wet sensor, type WET-2, Delta-T DeviceLtd, Cambridge, UK) was used, calibrated for clay (experimental) soil, witha sensor and rooting depth of 235 mm. Water was added at intervals of24 h to maintain the status quo by checking the soil moisture percentageevery day.

Application of EMS

A commercial EM preparation known as ‘Bio EM’ was obtained from EMROThailand. The Bio EM was prepared by using a concentrated stock solution of EM,EM-1. The formulation of EM-1 is kept secret, although according to one of theEMRO centers (Bionova Hygiene GmbH, Stans, Switzerland), EM-1 contains1.36 107 colony forming units (cfu) of lactic acid bacteria mL71, 3.36 104 cfu ofphotosynthetic bacteria mL7l, and 1.36 104 cfu of yeast mL7l. Bio EM wasprocessed from EM-1 by fermentation under anaerobic conditions with water andsugarcane molasses for 7 days.

In the EMS–soil pots, the Bio EM solution was first applied at 7 DAT, with6.75 mL of concentrated EM solution mixed in 4.5 L of water. Before the start ofany irrigation of these trials with EM-treated soil (EMS), 0.5 L of this mixed

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solution was applied. After that, water levels were maintained in all EMS potsaccording to the treatment schedules. The EM application was repeated at weeklyintervals until one month before harvesting, unless a draining period coincided withthe EM application. EM application was avoided during draining periods and wasapplied with the next scheduled irrigation, immediately following a drainage period.

Soil moisture and pH status during experimentation

During the flooding phase in the two intermittent flooding treatments (IF-V and IV-R), the average volumetric soil water content (m3 m73) was 0.575+ 0.0009, whereasafter a 3-day draining period, this value reached 0.0682+ 0.0031 m3 m73. In thenon-flooded water treatments (NSW), the average soil water content varied from0.378+ 0.0008 m3 m73 (just after water application) to 0.217 + 0.0021 m3 m73

(before water application). These values were recorded with a sensor at rooting depthof 235 mm.

Soil pH was affected primarily by the degrees and timing of flooding. In thenon-flooded water treatments, pH was always 55, whereas under intermittentirrigation, soil pH increased favourably after flooding to above pH 6 and thendecreased to below pH 5 after draining. Under the IF-V water regime, after thecompletion of three draining periods, soil pH stayed steady and even rose afterflooding – remaining above pH 6 for the rest of the crop growth period. With theIF-R management, pH continued to cycle according to the drainage cycle.No significant effect on soil pH from the application of EM solution wasobserved.

Plant parameter measurements

Rice-yield-affecting parameters were assessed by counting the number of tillers perplant, spikelets per panicle and filled grains per panicle, and single grain weight.Grain-filling period was calculated by observing the number of days to flowering(from date of sowing), and days to physiological maturity (from date of sowing).Grain-filling rate (G) was calculated by using the equation:

W ¼ A=ð1 þ Be�ktÞ1=N and

G ¼ AKBe�kt=ð1 þ Be�ktÞ1=N

where W¼ grain weight (mg), A¼ final grain weight, t¼ time after anthesis (d) andB, k, and N are coefficients determined by regression. K is the maximum observablevalue of W. The active grain-filling period was defined as the days when W wentfrom 5% (t1) to 95% (t2) of A. An average grain-filling rate during this period wastherefore calculated from t1 to t2.

The root parameters evaluated were RLD and the ROA. Soil–root core sampleswere taken from 15–20 cm and 35–40 cm soil depth at flowering stage (F) and at20 days after flowering (20 DAF) using cylindrical tubes (5 cm diameter by 90 cmlength) 7 cm away from the hill. Since the number of days to flowering variedsomewhat among the treatments, average dates were used, i.e., 87 DAS forflowering, and 107 DAS for 20 DAF.

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Roots after being washed with water were cut into small pieces. The root lengthwas calculated using the line intersection method described by Tennant (1975). RLDwas then calculated by using the formula:

RLD¼RL=V;

where RL¼ root length and V¼ volume of the soil core soil.ROA rate was measured according to the methods described by Zhang et al.

(1994) by assaying the oxidation of a-naphthylamine. Five grams of fresh roots weretransferred into a 150 mL flask containing 100 mL of 20 mg L71 a-naphthylamine.The flask was incubated for 4 h at room temperature (25+ 18C) in an end-over-endshaker. After incubation, the aliquots was filtered, and 2-mL aliquots were reactedwith 10 mL of 0.1% sulfanilic acid, and then with 2 mL of 50 mg L71 NaNO3. Theresultant color was measured by spectrophotometer at 530 nm, and the value isexpressed as mg (g fw)71 h71

Levels of soil available/exchangeable nitrogen (NH4þ-N/NO3

7-N) were measuredboth at flowering and at 20 DAF, assessing soil (mixed samples) from the same potsthat were used for root study. Soil cores were drawn at the corresponding dates whenthe pots were maintained with 5 cm water depth from the soil surface in the IF-Vand IF-R treatments, and at field capacity in the third set of treatments. Fresh soildrawn from the pots was extracted using 2N KCl solution, followed by distillationusing MgO for NH4

þ and Devarda’s alloy for NO37, and titration with standard

sulfuric acid. Values are expressed on a dry soil basis.Chlorophyll content of the flag leaf and of the third leaf was recorded at intervals

of 7 days from flowering to physiological maturity stages, using a chlorophyll meter(SPAD-502; Minolta Corp; Tokyo) calibrated by using spectrophotometric assays inorder to determine the exponential equation to directly convert its output to leafchlorophyll concentration (Markwell et al. 1995). These data were collected fromundisturbed pots for each treatment combination, which had not been used for rootstudy and nitrogen estimation.

Statistical analysis

Experiments were set up in a completely randomized block design with factorialcombinations of 363 with six replications and were repeated twice, with 12replicates for each treatment combination. Data were collected from fourundisturbed pots at flowering, 20 DAF and harvesting, two from each experimentfor each treatment combination. Data from both experiments were subjected toLevene’s test for account of homogeneity of variance. There was no significantdifference found between both experiments. Hence the data were pooled before beinganalyzed. Analysis was performed using SigmaPlot.11 to determine single-factor andinteraction effects. Computed ‘MS’ and ‘F’ values for some of the important plantparameters are presented in Tables 1 and 2.

Whenever significant interaction was observed between the factors, the level ofone factor was compared with each level of the other factor by using all PairwiseMultiple Comparison Procedures (Tukey test). Whenever significant interaction wasnot observed, the significant major effect of either of the factors or of both factors ispresented by pooling and analyzing data using one-way ANOVA. All data arepresented as means+ SE (standard error of the mean). A significance level of 0.05was used for evaluating all analyses.

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Table

1.

ComputedMSandFvalues

from

ANOVA

ofsomeoftheyield-contributing,above-groundrice

plantparameters.

ComputedMS(F)

Source

Number

of

tillers

Plantheight

Activegrain

fillingperiod

Spikelets

per

panicle

Filledgrains

per

panicle

Daysto

physiological

maturity

Grain

fillingrate

Grain

weight

per

pot

Soilmicrobial

treatm

ent(S)

76.27

(7.84*)

44.06

(4.89*)

25.33

(62.18**)

3502.26

(130.34**)

3194.96

(219.48**)

248.44

(745.33**)

0.00974

(49.37**)

952.58

(14.07**)

Watertreatm

ent(W

)517.02

(53.19**)

331.06

(36.74**)

117.00

(287.18**)

5931.09

(220.73**)

8250.87

(566.81**)

149.77

(449.33**)

0.0211

(106.90**)

17585.16

(259.75**)

S6

W63.89

(6.57**)

15.23

(1.69ns )

181.33

(199.63**)

40.84

(1.52

ns )

228.66

(15.70**)

25.77

(77.33**)

0.0296

(150.10**)

164.28

(2.42

ns )

p¼0.18

p¼0.22

p¼0.07

Note:df¼2,35,forsoilmicrobialandwatertreatm

ents,and4,35,forsoilmicrobialtreatm

ents6

watertreatm

ents.ns,

notsignificant;

significantat*p50.05and

**p50.001,respectively.

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Table

2.

ComputedMSandFvalues

from

ANOVA

ofsomerootparameters(R

LD

andROA)andavailable

soilN.

ComputedMS(F)

Atflowering

At20DAF

Source

RLD

(15–20cm

)RLD

(35–40cm

)ROA

Available

soilN

RLD

(15–20cm

)RLD

(35–40cm

)ROA

Available

soilN

Soilmicrobialstatus(S)

418.10

(55.99**)

34.89

(17.97**)

94.19

(28.69**)

176.07

(13.83**)

123.77

(26.97**)

31.42

(24.00**)

318.72

(111.44**)

1280.20

(204.31**)

Waterregim

e(W

)3925.11

(525.71**)

115.58

(59.53**)

415.77

(126.66**)

3136.31

(246.52*)

498.12

(108.54**)

58.09

(44.36**)

665.33

(232.64**)

3128.74

(499.34**)

S6

W91.70

(12.28**)

7.79

(4.01*)

5.95

(1.81

ns )

151.87

(11.93**)

19.59

(4.27*)

6.36

(4.86*)

83.13

(29.07**)

192.64

(30.74**)

p¼0.15

Note:RLD,rootlength

density;ROA,rootoxidizngactivity.df¼2,35,forsoilmicrobialandwatertreatm

ents,and4,35,forsoilmicrobialtreatm

ents6

watertreatm

ents.

ns,notsignificant;significanytat*p5

0.05and**p5

0.001,respectively.

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Results

ANOVA results

Significant interaction effects of soil microbial treatments (S)6water treatments (W)were found for tiller number, days to physiological maturity, active grain-fillingperiod, filled grains per panicle and grain-filling rate (p 5 0.001). Spikelets perpanicle and final grain weight per pot were significantly influenced by both the soiland water treatments, but without any interaction effect (Table 1). Similarly, RLDand soil available/exchangeable nitrogen (NH4

þ-N/NO37-N) were significantly

influenced by the interaction of soil and water treatments. At the flowering stage,the interaction effect of S6W was not significant for ROA rate; however, at thelater growth stage, this effect was highly significant (p 5 0.001) (Table 2).

Effects on above-ground morphological and physiological yield-contributing parameters

Tiller number

Rice plants grown with intermittent flooding through the vegetative stage (IF-V)and with intermittent flooding extending into the reproductive stage (IF-R)had higher tiller numbers compared to the non-flooded (NSW) water regime(IF-V � IF-R4NSW; Table 3). Within the IF-V water treatments, plants grownwith autoclaved soil (IF-V/AUS) had 27% higher tiller numbers than plants grownin normal soil (IF-V/NS), whereas plants grown with EM-soil treatment (IF-V/EMS) showed 9% higher tiller number compared with normal soil (IV-V/AUS4 IF-V/EMS4 IF-V/NS). This difference, a soil microbial treatment effect,was significant only under IF-V water conditions, not under IF-R or NSW waterconditions.

Plant height

We note that the effect of water treatments was more significant for this parameterthan for soil microbial treatments (Table 1). Plants were taller under both of theintermittent water conditions compared to the no-flooding (NSW) treatment (IF-V� IF-R4NSW).

Spikelets per panicle

These were affected by water regime (p 5 0.001) as a greater number of spikeletswas observed under IF-V compared with NSW and IF-R, with no differenceobserved between the IF-R and NSW water treatments (IF-V4NSW � IF-R).Regarding soil microbial status, a significant effect of EM application was observed(p 5 0.001). The application of EM solution increased the number of spikeletsper panicle compared with autoclaved (AUS) and normal (NS) soil treatments(Table 3).

Filled grains per panicle

By contrast, the parameter of filled grains per panicle was significantly increased withthe application of EM solution under all three water conditions (Table 3). This

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increment was highest under IF-V water conditions, followed by NSW4 IF-R (IF-V/EMS4NSW/EMS4 IF-R/EMS).

Grain-filling rate

Grain-filling rate was accelerated in the plants grown with EM-treated soil underintermittent flooding. However, soil maintained in wet condition but withoutstanding water (NSW) had the lowest grain-filling rate in the plants that were grownin EM-treated soil (IF-V/EMS � IF-R/EMS4NSW/EMS). Under normal soil

Table 3. Effects of water treatments (IF-V, IF-R and NSW) and different soil microbialtreatments (NS, EMS and AUS) on yield contributing parameters of rice plant.

Water regimes

Soil types

Normal soil EM-treated soil Autoclaved soil

No. of tillersIF-V 51.25+ 1.43 cA 56.5+ 2.17 bA 65.18+ 1.48 aAIF-R 51.5+ 2.5 aA 53.23+ 1.1 aA 53.25+ 1.43 aBNSW 44.56+ 1.31 aB 45.34+ 0.77 aB 44.01+ 1.01 aC

Active grain-filling period (days)IF-V 37.01+ 0.57 bA 31.5+ 0.28 cB 43.5+ 0.28 aAIF-R 32.01+ 0.01 aC 30.5+ 0.28 bB 31.5+ 0.27 abCNSW 33.5+ 0.28 bB 39.5+ 0.27 aA 34.5+ 0.28 bB

Filled grains per panicleIF-V 211.75+ 2.6 bA 234.25+ 1.2 aA 209.75+ 3.03 bAIF-R 149.56+ 0.93 bC 180.36+ 1.43 aC 151.75+ 0.62 bCNSW 187.62+ 2.24 bB 195.75+ 2.17 aB 169.12+ 1.43 cB

Grain-filling rate (mg � d71 � grain71)IF-V 0.56+ 0.01 bC 0.70+ 0.002 aA 0.50+ 0.004 cBIF-R 0.61+ 0.006 bB 0.71+ 0.006 aA 0.67+ 0.006 cANSW 0.70+ 0.006 aA 0.58+ 0.005 bB 0.66+ 0.006 cA

Days to physiological maturityIF-V 107.5+ 0.28 bB 103.5+ 0.28 cB 116.5+ 0.28 aAIF-R 103.5+ 0.28 bC 103.5+ 0.28 bB 107.5+ 0.28 aBNSW 112.5+ 0.28 bA 106.5+ 0.28 cA 116.5+ 0.28 aA

Plant height (cm)IF-V 106.65+ 2.87 A 104.84+ 0.77 A 109.33+ 1.27 AIF-R 102.55+ 0.95 A 100.81+ 1.19 A 107.05+ 0.95 ANSW 97.63+ 1.87 B 96.68+ 1.38 B 96.36+ 1.04 B

Spikelet per panicleIF-V 227.50+ 3.22 bA 244.75+ 2.05 aA 203.75+ 3.1 cAIF-R 186.75+ 0.85 bB 197.28+ 3.08 aB 168.12+ 1.78 cBNSW 191.37+ 2.65 bB 205.81+ 3.56 aB 173.97+ 1.63 cB

Grain weight (g pot71)IF-V 165.01+ 4.56 bA 185.75+ 4.67 aA 190.75+ 4.69 aAIF-R 101.87+ 5.34 bC 115.2+ 4.64 aC 103.5+ 3.22 aCNSW 116.5+ 1.75 bB 134.5+ 3.57 aB 125.62+ 3.33 aB

Note: IF-V, intermittent flooding through the vegetative growth phase; IF-R, intermittent flooding intothe reproductive phase; NSW, non-flooded soil condition; NS, normal soil; EMS, Effective Microorgan-isms solution; AUS, autoclaved soil. Values are shown as mean+ SE. Means with similar lower caseletters within a row and upper case letters within a column are not significantly different (p 5 0.05, Tukeytest SigmaPlot.11).

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conditions (NS), the grain-filling rate was higher under non-flooded water conditions(NSW) compared to either intermittent flooding condition (NWS/NS4 IF-R/NS4 IF-V/NS).

Grain-filling period

Similarly, it was observed that the active grain-filling period was shorter in riceplants grown with EM soil under intermittent flooding; whereas under non-floodedsoil conditions (NSW), plants had a prolonged duration of grain filling with EM-treated soil. Among all of the treatment combinations, it was interesting to note thatplants with the IF-V/AUS treatment combination had the longest grain-fillingperiod, while also the lowest grain-filling rate. This compensatory relationship raisesquestions worth investigation.

Grain weight

It was observed that the water treatments had more pronounced effects on grainweight compared with soil microbial treatments (Table 1) without any interactioneffects. The highest grain weight was recorded under the IF-V water conditionsfollowed by NSW4 IF-R. This increment under IF-V was 31 and 42% higher thanNSW and IF-R, respectively. The yield difference between NSW and IF-R was of18% (using one-way ANOVA; data not presented). Likewise, there was nosignificant difference between autoclaved and EMS-applied soil, but relatively lowergrain weight was observed with the NS soil treatment (Table 3). Soil microbialtreatment showed no significant effect when one-way ANOVA was calculated,however data not shown. While there was no significant major effect of soil microbialtreatments on final grain weight, it was nevertheless observed that plants grown withEM-applications matured faster compared to plants grown in autoclaved soil(Table 3).

Changes in chlorophyll content

The chlorophyll content of the flag leaves and the third leaves decreased as ripeningproceeded, as expected (Figure 1). However, the rate of decrease varied under thedifferent water treatments and with different soil microbial treatments. The rate ofdecrease in chlorophyll content was seen to be lower under the IF-V water treatment,followed by NSW4 IF-R. It was observed that at the later growth stage, in plantsgrown in EM-treated soil, the chlorophyll content of both the flag leaf and the lowerleaf (third leaf) decreased, but at an accelerating rate. While this trend was observedunder all water regimes, the rate of decrease in chlorophyll levels was greater underIF-R water conditions. Plants grown with the combination of IF-V/AUS had theirsenescence most delayed compared with all other combinations (Figure 1 andTable 3).

Effects on root growth pattern and root physiological activity

At flowering stage, it was observed that root growth was higher in both of theintermittent water treatments (IF-V and IF-R) compared with non-flooded watertreatment (NSW) (Figure 2). In the intermittent water treatments, 475% of the

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Figure

1.

Changes

inchlorophyllcontentofupper

leaf(flagleaf)andlower

leaf(3rd

leaf)ofrice

plants

duringreproductivephase

grownin

pots

under

semi-fieldconditionswithdifferentwatertreatm

ents:interm

ittentfloodingthroughthevegetativegrowth

phase

(IF-V

),interm

ittentfloodinginto

the

reproductivephase

(IF-R

),andnon-flooded

soilconditions(N

SW)withdifferentsoilmicrobialtreatm

ents:norm

alsoil(N

S),soiltreatedwithEffective

Microorganismssolution(EMS),andautoclaved

soil(A

US).Verticalbars

show

standard

error.

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Page 16: Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities

roots were distributed in the upper soil depth, whereas in the non-flooded (NSW)pots, the roots were more evenly divided between upper and lower soil depths. In theNSW pots, a larger proportion of the roots was found at the lower soil depthcompared with IF-V and IF-R pots (Figure 2).

Root length density

It was observed that the application of EM solution increased root growth (RLD) atthe flowering stage under both of the intermittent water conditions (IF-V and IF-R),

Figures 2(a)–(b). Effects on root length density (RLD) (cm cm73) at 15-20 cm depth (Fig.2a) and at 35-40 cm depth (Fig. 2b) in rice plants grown in pots under semi-field conditionswith different water treatments: intermittent flooding through the vegetative growth phase (IF-V), intermittent flooding into the reproductive phase (IF-R), and non-flooded soil condition(NSW), with different soil microbial treatments: normal soil (NS), soil treated with EffectiveMicroorganisms solution (EMS), and autoclaved soil (AUS). Bars show mean+ s.e. Bars withsimilar letters are not significantly different (p5 0.05, Tukey test SigmaPlot11).

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but not at a later plant growth stage. At a later stage, the RLD of rice plants wassignificantly reduced when plants grew in EM-treated soil under intermittent waterconditions.

By contrast, RLD was relatively higher in plants grown with autoclaved soil(AUS) under all three water regimes. Importantly, root degeneration was fasterunder IF-R water conditions compared with IF-V and NSW, and was fastest withthe IF-R/EMS combination.

A similar effect was seen at lower soil depth at the later growth stage (Figure 2b).The least root distribution was observed under IF-R compared with IF-V and NSW.At this depth also, plants grown in autoclaved soil had a greater root distributioncompared with plants grown in EM-treated soil and normal soil (NS) when receivingintermittent water treatments (IF-V and IF-R). However, with the non-floodedwater regime (NSW), the differences attributable to soil microbial activity were notsignificant, either at flowering or at 20 DAF stage.

Root oxidizing activity

When ROA rates were compared at the flowering stage, it was seen that this rate washigher under both of the intermittent water conditions compared with non-floodedwater conditions (IF-V � IF-R4NSW; Figure 3a). Further, plants grown in EM-treated soil had a higher root activity rate compared with the plants grown in normalsoil (NS) (EMS4NS � AUS), with no significant difference observed betweennormal soil and autoclaved soil.

At a later growth stage, 20 DAF, a higher rate of root activity was recorded inthe plants grown under IF-V water conditions, followed by NSW (Figure 3b); lowestroot activity rate was under IF-R water conditions (IF-V4NSW4 IF-R). Plantsgrown in autoclaved soil (AUS) under all water conditions showed a higher rootactivity rate compared with EMS and NS soil treatments, and the highest rootactivity rate was observed in those plants grown under the IF-V/AUS treatmentcombination (IF-V/AUS4NSW/AUS4 IF-R/AUS).

Effects on available soil nitrogen (NH4þ-N/NO3

7-N)

At flowering, a significant increase in the concentration of NH4þ/NO3

7 nitrogen (13–14%), and of NH4

þ-N in particular, was found in the EM-treated soil that receivedintermittent water treatments, whereas under non-flooded water conditions (NSW),autoclaved soil had a slightly higher concentration of available inorganic soil N (6%more) compared with EM-treated soil (Table 4), with no difference betweenautoclaved and normal soil.

At a later growth stage, i.e., at 20 DAF, the availability of NH4þ/NO3

7 nitrogenwas higher in autoclaved soil under both IF-V and NSW water conditions (Table 4).Under the IF-R water treatment, there was no difference in available nitrogencontent in autoclaved and normal soil; however, in this case also, the autoclaved soilhad a higher nitrogen content than the EM-treated soil. This indicated that enhancedsoil microbial activity had reduced N availability at this later growth stage.

Assessment of the NH4þ/NO3

7 nitrogen in AUS and NS soils treatments underintermittent water conditions at flowering stage indicated further that almost half ofthe nitrogen was present in NO3

7 form, whereas with EMS, the percentage of NH4þ

was greater (71%). This suggests that soil organic nitrogen was being mineralized at

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an increasing rate and with higher resulting availability under anaerobic conditions(Table 4).

The kinetics of available nitrogen and NH4þ-N were somewhat different under

non-flooded soil conditions (NSW). Total NH4þ/NO3

7 nitrogen was similar to that inthe IF-V treatment, but N was present mainly in NO3

7 form, indicating thedominance of autotrophs under aerobic soil condition and thereby the conversion ofammonium to nitrate. Also, less availability of nitrogen in the EM-treated soilcompared with AUS and NS soil treatments under non-flooded water condition,both at flowering stage and at 20 DAF, indicated that probably the nitrogenimmobilization rate was higher in the EM-treated soils.

Causal relationships between root and shoot growth and available soil nitrogen

Our analysis of the morphological responses of roots and shoots, seeking to assesspossible causal relationships if any, showed a positive correlation between thechlorophyll content of the lower leaves and the ROA rate, and also an associationbetween the chlorophyll content of the flag leaf and the duration of grain filling.These responses were observed under all water conditions and for all soil microbialdensities (Figure 4). It is known that the lower leaves provide most of the roots’supply of photosynthate to support metabolism (Tanaka 1958), so this observedrelationship was expected.

Further, the relationships between total NH4þ/NO3

7 nitrogen and ROA showed asignificant positive association between available soil nitrogen and the ROA rate atthe later growth stage, i.e., at 20 DAF (Figure 5). However, this relationship wassignificant at the flowing stage with NH4

þ nitrogen only, not with total NH4þ/NO3

7

nitrogen (not shown).

Figures 3(a)–(b). Effects on root-oxidizing activity rate (mg (g Fw)71 h71) at flowering (Fig.3a) and at 20 days after flowering (Fig. 3b) in rice plants grown in pots under semi-fieldconditions with different water treatments: intermittent flooding through the vegetativegrowth phase (IF-V), intermittent flooding into the reproductive phase (IF-R), and non-flooded soil conditions (NSW) with different soil microbial treatments: normal soil (NS), soiltreated with Effective Microorganisms solution (EMS), and autoclaved soil (AUS). Figure 3ashows the effects of water regimes (N ¼ 12, one-way ANOVA) and soil microbial conditions(N ¼ 12, one-way ANOVA). Bars show mean+ s.e. Bars with similar letters are notsignificantly different (p5 0.05, Tukey test SigmaPlot11).

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Discussion

Soil nitrogen, yield-contributing parameters and grain yield

A major yield-contributing parameter of rice plants is tiller number. Thismorphological trait was seen to be directly related to the growth rate of plantsduring their initial stage, which was faster under IF-V and IF-R compared withNSW water conditions. Less plant height and an initially slower rate of plant growthunder non-flooded soil water conditions suggested a lower rate of nutrientabsorption during the early vegetative stage of plant growth. This could be due toshallower rooting depth at the initial stage. This differentiation could be relevantespecially for NO3

7 since under aerobic soil conditions, NO37 would be the dominant

form of nitrogen nutrient available.Higher leaching rate of the more highly mobile nitrate to lower soil depths under

the non-flooded (NSW) soil conditions could have resulted in a lower uptake ofnutrients by the plants in their initial growth stage, when their roots had not grownso much into deeper soil layers, which in turn would contribute to lesser tillergrowth. However, even with a lesser number of tillers, grain yield was 18% higherfrom non-flooded soil conditions compared with the IF-R water condition.

This was mainly due to relatively deeper root systems and higher ROA rate at latergrowth stage with NSW soil–water conditions that enabled the plants to withdrawmore nutrients and water at the reproductive stage. This resulted into a higher numberof productive panicles, more number of filled grains per panicle, prolonged duration ofgrain filling and thus higher grain yield. For grain yield, having more productivepanicles is more important than having just a higher tiller number.

We found that the plants grown under IF-R water conditions, i.e., withintermittent water applications continuing into the reproductive stage, even thoughthey had a higher tiller number, had many tillers that were unproductive. Moreover

Table 4. Effects of different water (IF-V, IF-R and NSW) and different soil microbialconditions (NS, EMS and AUS) on available nitrogen (NH4

þ þ NO37) content of pot soil

grown with rice plants under semi-field conditions.

Water treatment

(IF-V) (IF-R)Non-floodedsoil (NSW)

Available soil N (NH4þ-N þ NO3

7-N) at flowering (mg kg71)NS 112.69+ 2.13 aB 84.67+ 2.06 bB 112.38+ 2.47 aBAEMS 128.28+ 2.37 aA 94.00+ 1.2 cA 109.34+ 0.57 bBAUS 113.29+ 1.24 aB 84.75+ 1.88 bB 116.56+ 1.03 aAAvailable soil N (NH4

þ-N þ NO37-N) at 20 days after flowering (mg kg71)

NS 43.62+ 0.49 aB 10.66+ 1.31 bBA 36.04+ 1.49 bBEMS 23.38+ 1.73 aC 8.10+ 0.68 bB 24.24+ 0.52 aCAUS 55.35+ 1.86 aA 13.72+ 1.5 cA 48.48+ 0.63 bAPercentage of soil NH4

þ-N on total available soil N at floweringNS 50.02+ 1.99 aC 51.32+ 1.43 aB 33.31+ 2.24 bAEMS 71.25+ 1.10 aA 72.01+ 1.88 aA 39.25+ 2.17 bAAUS 57.21+ 2.13 aB 56.28+ 1.65 aB 34.15+ 1.78 bA

Note: IF-V, intermittent flooding through the vegetative growth phase; IF-R, intermittent flooding intothe reproductive phase; NSW, non-flooded soil conditions; NS, normal soil; EMS Effective Microorgan-isms solution; AUS, autoclaved soil. Values are shown as mean+ SE. Means with similar lower caseletters within a row and upper case letters within a column are not significantly different (p 5 0.05, Tukeytest SigmaPlot.11).

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in IF-R, reduced spikelet numbers, fewer filled grains per panicle, a shorter activegrain-filling period, a lower root activity rate, and consequently lower grain weightwas observed mainly due to limited nitrogen availability during the reproductivestage associated with several periods of drainage and losses of nitrogen thatcoincided with the plants’ flowering and grain-filling stages. Some reduction in theseyield-influencing parameters might also, or instead, be due to water deficit-inducedpollen abortion (Nguyen and Sutton 2009).

Another possible reason for lower productivity could be a decreasedphotosynthetic rate in rice plants grown under IF-R water conditions due to theirstomatal closure in the afternoon. A decreasing rate of root activity, evident with IF-R (seen in Figure 3), reduces the stomatal aperture and lead to closure of thestomata. The depression of the stomatal aperture in aged leaves is thought to be aresponse to reduced water uptake capacity of the related roots (Ishihara et al. 1971).Further studies are needed to understand the close relationships between stomatalaperture and root activity rate in both young and aged plants.

Morphological responses of roots under varying water regimes and soil microbialdensities

It was evident from the experimental results that morphological and physiologicalresponses of rice roots and shoots were greatly affected by the availability of N andthe ratio of soil NH4

þ/NO37 nitrogen as affected by water regimes and soil microbial

activity. Under intermittent water regimes, the increased activity of soil microbesplayed an evident role in influencing RLD, most likely by influencing the forms ofsoil inorganic nitrogen available. The NH4

þ/NO37 nitrogen concentration, and NH4

þ-N in particular, was greater at the flowering stage in the EM-treated soils that hadreceived intermittent irrigation (Table 4) possibly due to the impact of drying andrewetting the soil on microbial populations in the soil (reported by Birch 1958),which was relatively higher in EMS soil treatments compared with others.

Such intermittent irrigation events might provide plants with an opportunity tobenefit from roots having less competition for nitrogen when the microbial populationwas decreased, and having also access to additional nitrogen becoming available viamicrobial-cell lysis. Therefore, greater RLD in the upper soil (Figure 2a) could be theresult of an enhanced concentration of nitrogen as earlier reported by severalresearchers (Shukla and Sen 1974; Maurya and Ghildyal 1975; Fitter 1987).

However at this growth stage, we did not find any significant relationshipsbetween total available soil nitrogen and root growth (regression slope not shown).Probably, the higher percentage of NH4

þ-N would have favored cytokininproduction, and in turn more cytokinin would have favored root elongation in theupper soil layer. This assumption is based on the earlier findings of Mercier et al.

Figures 4(a)–(b). Regression slopes between the chlorophyll content of flag leaf and theduration of grain filling (Fig. 4(a), on left), and root oxidizing activity rate and chlorophyllcontent of lower leaf (Fig. 4(b), on right) at later growth stage of rice plants grown in pots withdifferent water treatments: intermittent flooding through the vegetative growth phase (IF-V),intermittent flooding into the reproductive phase (IF-R), and non-flooded soil condition(NSW) and with different soil microbial conditions: normal soil (NS), soil treated withEffective Microorganisms solution (EMS), and autoclaved soil (AUS). SigmaPlot 11.(Adopted from Mishra and Salokhe, 2011)

"

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Figure 5. Regression slope between available soil N and root oxidizing activity rate at thelater growth stage of rice plants grown in pots with different water treatments: intermittentflooding through the vegetative growth phase (IF-V), intermittent flooding into thereproductive phase (IF-R), and non-flooded soil condition (NSW) and with different soilmicrobial conditions: normal soil (NS), soil treated with Effective Microorganisms solution(EMS), and autoclaved soil (AUS). SigmaPlot 11. (Adopted from Mishra and Salokhe, 2011)

(1997) and Debi et al. (2005), who have shown, respectively, that cytokininproduction is favored by NH4

þ ions, and cytokinin favors root elongation.By contrast, it was seen that under non-flooded water conditions (NSW), EM

application reduced the available nitrogen, and there was also less total root growthaccordingly. The lower availability of inorganic nitrogen in NSW/EMS potsindicated that the nitrogen requirement of microorganisms that decompose organicmatter in non-flooded soils is higher than for decomposers in flooded soils. Thisresults in higher net immobilization of N in aerobic soils than in flooded soils. Asimilar finding has been reported previously by Broadbent (1979).

At a later growth stage, the lower RLD in EMS treatments compared with NSand AUS treatments under intermittent water conditions (Figure 2) might be due toa lower content of available soil inorganic nitrogen (Table 4) probably because ofcompetition for nitrogen between plant roots and soil microbes, which wouldincrease at later growth stages due to higher microbial populations and thus to ahigher rate of immobilization of NH4

þ. Earlier research findings have shown thatnitrogen mineralization decreased with an increase in total root growth and with anincrease in the number of microorganisms around the plant roots (Goring and Clark1949; Kuzyakov 2002). Another possible reason could be an increased rate ofdenitrification after reflooding as a result of reduced soil conditions, with a relativelyhigher rate of oxygen demand by soil microorganisms.

Nevertheless, lower inorganic nitrogen availability and reduced RLDs in the IF-R/EMS treatments, compared with the IF-V/EMS combination, confirmed the

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contribution that available soil nitrogen makes toward the maintenance of rootgrowth during the late maturity stage. This also suggests that there is no direct effectof EM application as such, but rather that the observed response was due toincreased versus decreased soil nitrogen availability, mediated either by an increasedmineralization rate or by an increased rate of immobilization, respectively.

It is important to note that under non-flooded water conditions (NSW), totalroot growth as characterized by RLD in this study was less compared with the effectswith intermittent flooding. Generally, it is believed that NO3

7 nutrition stimulatesroot branching and has significant effects on lateral root length and numbers(Wiersum 1958; Friend et al. 1990). It appears that the effect of NO3

7 nutrition onroot growth is to some extent contingent on environmental conditions such as thenitrate percentage in the soil nitrogen supply. In this regard, Duan et al. (2007)found higher root growth in plants when the nitrogen supplied to them was only25% NO3

7 compared with 50% and 75% NO37. This might be one reason for

reduced root growth under non-flooded water conditions (NSW) in our study, sincethe percentage of nitrate nutrition in the NSW treatments was 60–70%.

Another reason could be the limited availability of nitrogen to plants at their initialvegetative growth stage because of the downward movement of nitrate under non-flooded soil conditions. In this case, the nitrogen absorbed by the roots appeared toreact with carbohydrates present in the roots because much of it was used in the rootsfor their growth in the deeper soil layer required for water and nutrient acquisition.That would explain why root growth was greater in deeper soil layers.

Consequently, this would have affected initial shoot growth as evident from aslower growth rate and lesser tiller number in non-flooded soil conditions. When theplants’ supply of nitrogen increased due to deeper root growth, more of the nitrogenreached the shoot and support the use of carbohydrates there for shoot growth.Accordingly, less carbohydrate remained for translocation to the roots, and thegrowth of roots was then limited.

This was probably another reason for the lower less root growth under the non-flooded soil condition. It also appeared that these responses would be visible onlywhen the soil had limited nitrate-holding capacity. Obviously, the plants’ root andshoot response under the non-flooded condition would differ when growing in soilwith higher organic matter/humus content.

Further, greater root growth at lower soil depth under non-flooded soil conditions(NSW) (almost 50% of total roots) reflects roots’ orientation of their growth towardaccessing available water (and nutrients) as evident above. Moreover, it also seemsthat the ratio of NH4

þ to NO37 in the nitrogen supply, which is influenced by different

water regimes and soil microbial density, also plays an important role in tailoring rootstructure, having a greater effect than total available soil nitrogen.

RLD is accordingly be affected primarily by the varying ratio of NH4þ/NO3

7

nitrogen, by their availability at different growth stages and at different soil depths,and by total available nitrogen. These findings are in agreement with earlier studies(Goring and Clark 1949; Hodge et al. 1999; Duan et al. 2007).

Physiological response of roots under varying water regimes and with different soilmicrobial densities

It was evident from the results that ROA, which was significantly influenced by soilwater conditions and microbial activity, was influenced by NH4

þ nitrogen. HigherROA appeared to be due to a higher ratio of NH4

þ nitrogen compared with

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NO37 nitrogen under intermittent water conditions, and in EM-applied soil in

particular. Higher NH4þ nitrogen favors cytokinin production (Mercier et al. 1997),

and ROA is regulated by cytokinin, favoring higher ROA with higher cytokininconcentration.

Moreover, at a later growth stage, ROA was higher under IF-V and NSW waterconditions compared with IF-R, probably also due to the higher availability ofNH4

þ/NO37 nitrogen indicating that this response was primarily governed by the

available soil inorganic N status and by the chlorophyll content of the lower leaves(Figures 4b and 5). These leaves are the primary source of photosynthate for theplants’ roots. It also appeared that the combination of higher ROA, higheravailability of NH4

þ/NO37 nitrogen, and higher chlorophyll content of the lower

leaves at the later growth stage was one of the reasons for having higher yield underthese two water conditions compared with IF-R. This finding is in agreement withearlier findings of Mishra and Salokhe (2010).

We also noted that plants grown in autoclaved soil, either with IF-V or NSW,had higher ROA rates compared with the other soil treatments. This increment didnot result in higher grain yield than with the other soil conditions, however. Evenwith EM application, the ROA rate at a later growth stage was reduced significantly,even so, grain yield was similar to the NS and AUS soil treatments (using one-wayANOVA, data not shown).

It seems that this physiological response of roots, i.e., their ROA rate, dependson the relative costs and benefits to the plant, as discussed in Mishra and Salokhe(2011). If the supply of photosynthate to the roots, which comes mostly from thelower leaves of the plant, is restricted, or if the soil is limited in its nutrientavailability and roots are unable to supply sufficient nutrients to the above-groundparts, the plasticity of response of plants’ roots – either morphological proliferationor higher physiological activity – will be a burden for the plant.

Ultimately, the cost to the plant will depend on what is actually limiting itsgrowth, whether nutrients or photosynthate supply. This is evident from therelationships presented in Figures 4 and 5.

If the soil has limited N availability, decomposition of chloroplasts will take placein the lower leaves in order to supply sufficient N to the flag leaf. Consequently, thephotosynthetic ability of the lower leaves will decrease, they will become less able tosupply sugar and oxygen to the roots, and the ROA rate will accordingly decrease.This is apparently why EM-applied soil showed lower ROA at a later growth stage.

This means that higher ROA rate at a later growth stage is not the sole factorthat influences final grain yield, although it is important during the late maturitystage which contributes to greater yield by delaying senescence. This factor, inparticular, can result in increased photosynthetic efficiency of the plant, which inturn can lead to higher grain yield. This was particularly evident in plants grown inthe IF-V/AUS treatment. However, longer growth duration is not necessarilydesirable if more accelerated grain filling is compensatory.

Another yield-determining factor is the rate of dry matter partitioning andtranslocation to the grains, which was most evident in plants grown with EM-appliedsoil. Enhanced sink capacity (higher tiller number, greater root growth and morespikelets in plants grown with the IF-V/EM treatment) coupled with an enhancedgrain-filling rate appeared to be major contributing factors for a similar yield gain inIF-V/EMS plants as in IF-V/AUS plants, despite the earlier senescence seen in theIF-V/EMS plants.

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In general, partitioning of assimilates between panicles, leaves, culms and storageis under hormonal control. It is not clear in this study whether the higher grain-fillingrate might be due to some hormonal changes facilitated by EM application.However, based on the lower soil available nitrogen status and the earlier senescenceof roots in the EM treatments, it can be inferred that final grain yield in thesetreatments was mainly governed by the better partitioning and translocation of drymatter to the grain, whereas in autoclaved soil this was mainly governed by theprolonged duration of photosynthesis. The mechanisms involved need not be thesame, or of the same relative importance, for the processes of grain formation andgrain filling under different environmental conditions.

Effectiveness of Effective Microorganisms

Under the intermittent water regimes, EMS influenced RLD, as discussed above, andalso affected the plants’ sink capacity and the allocation of dry matter to the sink,i.e., to the grains. Moreover, under intermittent/EMS treatments higher ammoniumconcentration with a higher root oxidizing activity rate at the flowering stage, alongwith higher root growth overall, resulted in a higher grain-filling rate. However, sucheffects were not seen the same way under non-flooded (NSW) water conditions.

It appeared that the response of plants to EM application was affected by the soilnitrogen input facilitated by mineralization and immobilization rates, the rate ofnutrient demand (slower vs. faster growth rate of plant) and the ability of plants toabsorb nutrients (larger vs. smaller root surface area). Therefore, the observed effectswere due more to increased soil nitrogen input availability and to the plants’ demandfor N than to the presence and amount of EM microorganisms as such.

This was more evident at the later growth stage where plants grown in EM-treatedsoil showed earlier senescence of roots and shoots, along with lower availability of soilnitrogen. It was not clear whether this response was triggered only by limited soilnitrogen availability facilitated by a greater rate of immobilization at the later growthstage, and/or due to a higher uptake and higher rate of partitioning and translocationof dry matter to the seeds that was evident from the higher grain-filling rate in thesetreatments. Further study of plant N uptake at different growth stages under similarwater and soil conditions is needed for a better understanding of these mechanisms.

It is known that most soil microorganisms are usually limited by the supply ofeasily decomposed carbon (C) in the soil, and most soil N transformations arecarried out by microbial heterotrophs that depend upon the supply of availableorganic C. In this study, we did not apply any additional organic matter to the soil,and that might be another reason for a lower rate of mineralization and lessavailability of soil N at a later growth stage in EM-applied soil, along with higherplant demand at the earlier growth stage due to higher shoot and root activity.Determination of hot-water-extractable C might be useful for getting some ideaabout C supply and N mineralization.

There is also evidence that nitrogen mineralization decreases with increasinglignin content (Peevy and Norman 1948) and with decreasing water-soluble nitrogencontent (Iritani and Arnold 1960). In general, the level of lignin in root cells increaseswith the age of the plants. Therefore, it might be possible that at a later growth stage,a higher concentration of lignin from degenerating roots, along with decreasedcontent of water-soluble nitrogen, decreased the mineralization rate. However, it hasalso been suggested in the past that with the addition of soluble carbohydrates to the

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soil, this immobilization could be redirected to mineralization of soil nitrogen morerapidly (Black 1968), thereby making soil nitrogen more available for plant growth.

Therefore, it seems important to have sufficient, easily decomposed carbon sourcesin the soil to capitalize on the benefits associated with soil microbial enhancementwhen using Effective Microorganisms and/or other biofertilizers for increasing grainyield. A similar finding has been reported by Javaid et al. (2008). This research foundthat there was no increase in yield when EM solution was applied alone; however,Javaid and associates found that the yield of wheat was significantly increased in a potstudy when EM was applied in combination with farmyard manure.

Because our research did not assess different soil levels of organic matter thatcould in turn affect soil microbial levels and activity as reported by others. Therefore,further research with different level of soil organic matter is needed to constructmore comprehensive understandings of these relationships for exploiting soilmicrobial enhancement through Effective Microorganisms and/or biofertilizersapplication to increase grain yield.

Relevance of physiological and morphological responses of roots and shoots for cropmanagement

On the basis of above discussed results, it can be put forward that an understandingof the mechanisms of root–shoot response is crucial for predicting changes in cropproductivity under altered soil environments. In particular, with water andbiofertilizer management strategies for irrigated rice production now changing inresponse to the spreading physical and economical scarcity of water and todeteriorating soil fertility, understanding these responses is crucial for makingoptimizing adjustments in crop management to avoid potential yield losses underalternative water regimes.

The three sets of water management practices evaluated in this study are withinthe range of recommendations for the System of Rice Intensification (SRI) as analternative rice crop management system (Thakur et al. 2010). SRI recommenda-tions also call for the promotion of soil microbial populations (Anas et al. 2011). Butlittle is known about interactions among different kinds of reduced waterapplications, replacing conventional continuous flooding of rice paddies, andvarious soil microbial densities.

Our data indicate that the phenotypic and physiological responses of plantsprompted by altered soil water and soil microbial conditions are not simple,unimodal or monotonic plastic responses. These responses could be different withdifferent soil water regimes and with different soils’ abiotic and biotic factors.

Agronomic practices that seek to optimize water and fertilizer input must takeaccount of the multiple factors involved, of stimuli induced by insufficient wateravailability as well as by other interacting factors such as nutrient status, soilcharacteristics (organic and inorganic), microbial activity, aerobic conditions andeven hypoxia.

Conclusions

The above results and ensuing discussion give evidence that rice root morphologyand physiology are significantly affected by variations in soil–water and microbialconditions and interactions. These variations are greatly affected by the availability

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and the NH4þ/NO3

7 ratio of soil available nitrogen. There are strong relationshipsbetween ROA rate and soil available nitrogen, ROA rate and the chlorophyllcontent of lower leaf, the chlorophyll content of the flag leaf and duration of grainfilling, and soil available nitrogen and RLD at the late reproductive stage.

These relationships govern the soil’s ability to supply nutrients to the roots, theplant’s ability to supply photosynthate to the roots, and the roots’ ability to supportplant growth and productivity. These results are highly interactive and can varyconsiderably with different soil and water dynamics. Higher root oxidizing activityrate at a later growth stage is not the sole factor that influences final grain yield, butthis is important at the late maturity stage because it contributes to plant yield bydelaying senescence and prolonging the duration of grain filling.

The effects observed with EM application were apparently not caused by EMmicroorganisms per se, but rather by the effects of nitrogen nutrient input facilitatedby the interaction between water regime and soil microbial activity. It is important tounderstand these complicated, multiply intermediated processes.

Further study on the role and impact of soil organic matter amendments and theirinteraction with soil nitrogen at different growth stages under alternative waterregimes, when promoting soil microbial enhancement through biofertilizer applica-tions, is warranted for better understanding these relationships and theirimplications for achieving higher crop productivity in the context of dealing withimpending water constraints and deteriorating soil fertility so as to sustain andfurther enhance rice crop productivity.

References

Anas I, Rupela OP, Thiyagarajan TM, Uphoff N. 2011. A review of studies on SRI effects onbeneficial organisms in rice soil rhizospheres. Paddy Water Environ. 9:53–64.

Anderson JPE. 1982. Soil respiration. In: Miller ALRH, Keeney DR, editors. Methods of soilanalysis. 2nd ed. Part II. Madison (WI): American Society of Agronomy and Soil ScienceSociety of America. p. 831–872.

Banerjee MR, Yesmin, L, Vessey JK. 2005. Plant-growth-promoting rhizobacteria asbiofertilizers and biopesticides. In: Rai MK, editor. Handbook of microbial biofertilizers.New York: Food Products Press. p. 137–182.

Belder P, Bouman BAM, Spiertz JHJ, Lu G, Quilang EJP. 2002. Water use of alternatelysubmerged and nonsubmerged irrigated lowland rice. In: Bouman BAM et al., editors.Water-wise rice production. Los Banos (Philippines): International Rice ResearchInstitute. p. 51–61.

Birch HF. 1958. The effect of soil drying on humus decomposition and nitrogen. Plant Soil.10:9–31.

Black CA. 1968. Soil–plant relationships. 2nd ed. New York: Wiley.Bouman BAM, Lampayan RM, Tuong TP. 2007. Water management in irrigated rice: Coping

with water scarcity. Los Banos (Philippines): International Rice Research Institute. 54 pp.Broadbent FE. 1979. Mineralization of organic nitrogen in paddy soils. In: Brady NC, editor.

Nitrogen and rice. Los Banos (Philippines): International Rice Research Institute. p. 105–118.

Ceesay M, Reid SW, Fernandes ECM, Uphoff N. 2006. The effect of repeated soil wetting anddrying on low land rice yield with System of Rice Intensification (SRI) methods. Intl J AgrSust. 4:5–14.

Debi BR, Taketa S, Ichii M. 2005. Cytokinin inhibits lateral root formation but stimulateslateral root elongation in rice (Oryza sativa). J Plant Physiol. 162:507–515.

Dilly O, Blume HP. 1996. Indicators to assess sustainable land use with reference to soilmicrobiology. In: Blume HP, Eger H, Fleischhauer E, Hebel A, Reij C, Steiner KG,editors. 9th Conference of the International Soil Conservation Organisation. Bonn(Germany): Catena Verlag. p. 29–36.

Archives of Agronomy and Soil Science 729

Dow

nloa

ded

by [

Nor

th D

akot

a St

ate

Uni

vers

ity]

at 1

9:49

21

Oct

ober

201

4

Page 28: Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities

Duan YH, Zhang YL, Wang SW, Shen QR. 2007. Effect of NH4þ to NO37 ratio (NH4þ/NO37) on biological characteristics of rice with different nitrogen use efficiency. J NanjingAgr Univ. 30(3):73–77.

Fitter AH. 1987. An architectural approach to the comparative ecology of plant root systems.New Phytol. 106:61–77.

Friend AL, Eide MR, Hinckley TM. 1990. Nitrogen stress alters root proliferation in Douglasfir seedlings. Can J For Res. 20:1524–1529.

Gallardo M, Turner NC, Ludwig C. 1994. Water relations, gas exchange and abscisic acidcontent of Lupinus cosentinii leaves in response to drying different proportion of the rootsystem. J Exp Bot. 45:909–918.

Goring CAI, Clark FE. 1949. Influence of crop growth on mineralization of nitrogen in thesoil. Soil Sci Soc Am Proc. 13:261–266.

Higa T. 1991. Effective microorganisms: A biotechnology for mankind. In: Parr JF, HornickSB, Whitman CE, editors. Proceedings of the 1st International Conference on KyuseiNature Farming. Washington (DC): US Department of Agriculture. p. 8–14.

Higa T. 2001. Effective microorganisms in the context of Kyusei nature farming: a technologyfor the future. In: Senanayake YDA, Sangakkara UR, editors. 6th InternationalConference on Kyusei Nature Farming; Pretoria, South Africa. p. 40–43.

Higa T. 2003. Kyusei nature farming and environmental management through effectivemicroorganisms – the past, present and future. 7th International Conference on KyuseiNature Farming; Christchurch, New Zealand. Available from: http://www.infrc.or.jp/english/KNF Data Base Web/7th Conf KP 2.html.

Higa T, Parr J. 1995. Beneficial and effective microorganisms in a sustainable agriculture andenvironment. Technol Trend. 9:1–5.

Hodge A, Robinson D, Griffiths BS, Fitter AH. 1999. Nitrogen capture by plant grown in N-rich organic patches of contrasting size and strength. J Exp Bot. 50:1243–1252.

Iritani WM, Arnold CY. 1960. Nitrogen release of vegetable crop residues during incubationas related to their chemical composition. Soil Sci. 89:74–82.

Ishihara K, Ishida Y, Ogura T. 1971. The relationship between environmental factors andbehaviour of stomata in rice plants. 2: On the diurnal movement of the stomata.Proceedings of the Crop Science Society of Japan 40:497–504. In: Matsuo T, KumazawaK, Ishi, R., Ishihara K., Hirata H, editors. Science of the rice plant: Physiology. Vol. 2.Tokyo: Ministry of Agriculture Forestry and Fisheries. p. 501–511.

Javaid A, Bajwa R, Anjum T. 2008. Effect of heat-sterilization and EM (effectivemicroorganisms) application on wheat (Triticum aestivum L.) grown in organic amendedsandy loam soil. Cereal Res Commun. 36:489–499.

Khan MS, Zaidi A, Wani PA. 2007. Role of phosphate-solubilizing microorganisms insustainable agriculture: a review. Agron Sustain Dev. 27:29–43.

Kuzyakov Y. 2002. Review: factors affecting rhizosphere priming effects. J. Plant Nutr. SoilSci. 165:382–396.

Markwell J, Osterman JC, Mitchell JL. 1995. Calibration of the Minolta SPAD-502 leafchlorophyll meter. Photosynth Res. 46:467–472.

Mauraya, PR, Ghildyal BP. 1975. Root distribution patterns of rice varieties evaluated underupland and flooded soil conditions. Il Riso 24(3):239–244.

McCauley GN. 1990. Sprinkler vs. flood irrigation in traditional rice production regions ofsoutheast Texas. Agron J. 82:677–683.

Mercier H, Kerbauy BG, Sotta B, Miginiac E. 1997. Effects of NO3, NH4 and urea nutritionon endogenous levels of IAA and four cytokinins in two epiphytic bromeliads. Plant CellEnviron. 20:387–392.

Mishra A, Salokhe VM. 2008. Seedling characteristics and the early growth of transplantedrice under different water regimes. Exp Agr. 44:365–383.

Mishra A, Salokhe VM. 2010. The effects of planting pattern and water regime on rootmorphology, physiology, and grain yield of rice. J Agron Crop Sci. 196:368–378.

Mishra A, Salokhe VM. 2011. Rice root growth and physiological responses to SRI watermanagement and implications for crop productivity. Paddy Water Environ. 9:41–52.

Mishra A, Whitten M, Ketelaar JW, Salokhe VM. 2006. The System of Rice Intensification(SRI): a challenge for science, and an opportunity for farmer empowerment towardssustainable agriculture. Int J Agr Sustain. 4:193–212.

730 A. Mishra and N. Uphoff

Dow

nloa

ded

by [

Nor

th D

akot

a St

ate

Uni

vers

ity]

at 1

9:49

21

Oct

ober

201

4

Page 29: Morphological and physiological responses of rice roots and shoots to varying water regimes and soil microbial densities

Mishra HS, Rathore, TR, Pant RC. 1997. Root growth, water potential and yield of irrigatedrice. Irrig Sci. 17:69–75.

Mitsui S, Tensho K. 1953. Dynamic studies on the nutrient uptake by crop plants. 3: thereducing power of roots of growing plants as revealed by nitrite formation in the nutrientsolution. J Sci Soil Manure (Japan). 22:301–307.

Newell RL, Wilhelm WW. 1987. Conservation tillage and irrigation effects on corn rootdevelopment. Agron J. 79:160–165.

Nguyen GN, Sutton BG. 2009. Water deficit reduced fertility of young microspores resultingin a decline of viable mature pollen and grain set in rice. J Agron Crop Sci. 195:11–18.

Ookawa T, Naruoka Y, Yamazaki T, Suga J, Hirasawa T. 2003. A comparison ofaccumulation and partitioning of nitrogen in plants between two rice cultivars(Akenohoshi and Nipponbare) at the ripening stage. Plant Prod Sci. 6:172–178.

Osaki M, Shinano T, Matsumoto M, Zheng T, Tadano T. 1997. A root-shoot interactionhypothesis for high productivity of field crops. Soil Sci Plant Nutr. 43:1079–1084.

Peevy WJ, Norman AC. 1948. Influence of composition of plant materials on properties of thedecomposed residues. Soil Sci. 65:209–226.

Proffitt, APB, Berliner PR, Oosterhuis DM. 1985. A comparative study of root distributionand water extraction efficiency by wheat grown under high and low frequency irrigation.Agron J. 77:655–662.

Sah RN, Mikkelsen DS. 1983. Availability and utilization of fertilizer nitrogen by rice underalternate flooding. Plant Soil. 75:221–226.

Schenck zu Schweinsberg-Mickan M, Muller T. 2009. Impact of effective microorganisms andother biofertilizers on soil microbial charakteristics, organic-matter decomposition andplant growth. J Plant Nutr Soil Sci. 172:704–712.

Shukla LM, Sen B. 1974. Studies on root distribution pattern of some high-yielding paddyvarieties. J Indian Soc Soil Sci. 23:266–267.

Skinner RH, Hanason JD, Benjamin JG. 2004. Root distribution following spatial separationof water and nitrogen supply in furrow irrigated corn. Plant Soil. 199:187–194.

Tajima K. 1995. Occurrence and mechanism of drought tolerance. 2: wind and drought. In:Matsuo T, Kumazawa K, Ishi R, Ishihara K, Hirata H, editors. Science of the rice plant:Physiology. Vol 2. Tokyo: Ministry of Agriculture Forestry and Fisheries. p. 838–849.

Tanaka A. 1958. Studies on the characteristics of physiological functions of the leaf at adefinite position on a stem of the rice plant. J Sci Soil Manure (Japan). 2:291–294.

Tennant D. 1975. A test of modified line intersected method of estimating root length. J Ecol.63:995–1001.

Thakur AK, Uphoff N, Antony E. 2010. An assessment of physiological effects of System ofRice Intensification (SRI) practices compared to recommended rice cultivated practices inIndia. Exp Agr. 46:77–98.

van der Heijden MGA, Bardgett RD, Straalen, NMV. 2008. The unseen majority: soilmicrobes as drivers of plant diversity and productivity in terrestrial ecosystems. Ecol Lett.11:296–310.

Vessey JK. 2003. Plant growth promoting rhizobacteria as biofertilizers. Plant Soil. 255:571–586.

Wiersum LK. 1958. Density of root branching as affected by substrate and separate ions. ActaBot Neerl. 7:174–190.

Xu H. 2000. Effects of a microbial inoculant and organic fertilizers on the growth,photosynthesis and yield of sweet corn. J Crop Prod. 3:183–214.

Yang C, Yang L, Yang Y, Ouyang Z. 2004. Rice root growth and nutrient uptake asinfluenced by organic manure in continuously and alternately flooded paddy soils. AgrWater Manag. 70:67–81.

Yang J, Zhang J, Wang Z, Liu L, Zhu O. 2003. Post-anthesis water deficit enhance grain fillingin two-line hybrid rice. Crop Sci. 43:2099–2108.

Zhang X, Tan G, Huang Y. 1994. Experimental technology of plant physiology. Shenyang(China): Liaoning Science and Technology Press. p. 51–75.

Zhang Z, Zhang S, Yang J, Zhang J. 2008. Yield, grain quality and water use efficiency of riceunder non-flooded mulching cultivation. Field Crops Res. 108:71–81.

Archives of Agronomy and Soil Science 731

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