Nitrogen use efficiency in bread wheat: Genetic variation and prospects for improvement

Nitrogen (N) is one of the primary macronutrients required for crop growth and yield. This nutrient is especially limiting wheat yields in the dry and low fertile agro-ecologies having low N in the root zone soil strata. Moreover, majority of farmers in India and South Asia are small to marginal with meagre capacity to invest in costly nitrogen fertilizers. Therefore, there is an immense need to identify lines that use nitrogen efficiently. A set of 50 diverse wheat genotypes consisting of indigenous germplasm lines (05), cultivars released for commercial cultivation (23) and selected elite lines from CIMMYT nurseries (22) were evaluated in an alpha-lattice design with two replications, a six-rowed plot of 2.5m length for 24 agro morphological, physiological and NUE related traits during two consecutive crop seasons in an N-depleted precision field under two different N levels of 50%-N50 (T1) and 100%-N100 (T2) of recommended N, i.e., 100 kg/ha. Analysis of variance revealed significant genetic variation among genotypes for all the traits studied. About 11.36% yield reduction was observed at reduced N levels. Significant correlations among NUE traits and yield component traits were observed which indicated pivotal role of N remobilization to the grain in enhancing yield levels. Among N-insensitive genotypes identified based on their yielding ability at low N levels, UASBW13356, UASBW13358, UASBW13354, UASBW13357 and KRL1-4 showed their inherent genotypic plasticity toward N application. The genotypes with more yield and high to moderate NUtE can be used as parents for the breeding of N efficient genotypes for marginal agro-ecologies. Low N tolerant genotypes identified from the current investigation may be further utilized in the identification of genomic regions responsible for NUE and its deployment in wheat breeding programs. The comprehensive data of 24 traits under different nitrogen levels for diverse genotypes from India and global sources (mainly CIMMYT) should be useful for supporting breeding for NUE and thus will be of great help for small and marginal farmers in India and South Asia.

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Introduction
Nitrogen (N) is an essential nutrient for optimizing plant growth and reproduction and therefore, applying N fertilizer is an essential practice to secure productivity in diverse cropping systems.Plants take N from both atmospheric air and soil minerals, but crop plants are inefficient in the acquisition and utilization of applied nitrogen.The efficient use of N by plants is determined by their ability to absorb naturally available N or applied N fertilizers.Current global nitrogen (N) demand stands at about 110.19 million metric tons with a projected annual increase of ~1.3% in the future (FAO, 2019) which equates to global N fertilizer consumption of more than 110 Mt per annum of which half of the total is being used for the production of major cereal crops i.e., maize, rice, and wheat (Ladha et al., 2016).
Indian agriculture consumes over 17 million tons of N fertilizer per year.Studies have shown that more than 50% of applied N fertilizers are unused by crops (Raun and Johnson, 1999;Lassaletta et al., 2014) and this unabsorbed N fertilizer in the soil is lost to the environment by leaching, denitrification, runoff, and atmospheric release through volatilization and becomes a major environmental concern (Vitousek et al., 1997;Zafar and Muhammad, 2007).Surplus nitrogen also pollutes freshwater streams and air which is hazardous to most living species (Hickman et al., 2014;Russo et al., 2017).Besides, excess usage of N fertilizer not only decreases the efficiency of nutrient use but also affects the rate of economic returns per unit of chemical fertilizer applied.The effect of negative environmental and economic impacts could be reduced through better agronomic practices and also by utilizing N efficient lines with improved nitrogen use efficiency (NUE) (Raun et al., 1999).Hence, the development of crop plants with more N-use efficiency (NUE) with efficient nitrogen usage is, therefore, an important research goal in enhancing future food production capabilities to achieve greater agricultural sustainability (Hirel et al., 2007).
Wheat (Triticum aestivum L.) is one of the staple food crops that ensures food and nutritional security for the world's poorest people living in different agro-ecological zones.
In the Indian agriculture scenario, wheat is grown under diverse production conditions including low fertile and rainfall dependent central and peninsular areas often facing drought during different stages of crop growth.In such areas, farmers apply a high dose of N fertilizer for maximizing grain yield (GY), but excessive usage of N fertilizer leads to low NUE of the crop along with various environmental hazards.Further many farmers lack knowledge about the ideal dose of fertilizer to harvest the real yield potential.The majority (90%) of such farmers are small and marginal and do not have capacity to take risk of applying high inputs.Hence, yield improvement under low nitrogen input would be extremely beneficial for economic and environmentally sustainable cultivation of wheat crop.
Similar to grain yield, nitrogen use efficiency (NUE) is also a complex trait, which is associated with various morphological, physiological, molecular, and biochemical changes in plants throughout the life cycle.For a clear understanding of this complex nature, studies of various physiological traits and their close correlation with one or more economically important traits like GY are foremost critical.Nevertheless, it will help in selecting low N tolerant/high yielding lines at different N conditions (Monostori et al., 2016).In general, plant function is always associated with chlorophyll content, which directly indicates the N status of the leaf (Yang et al., 2014).Leaf chlorophyll content and photosynthetic capacity are appropriate benchmarks for identifying high NUE (HNUE) genotypes under low N conditions during field trials (Vijayalakshmi et al., 2015;Kiran et al., 2016).Leaf N status was usually measured by using a hand-held optical chlorophyll meter to monitor the leaf nitrogen status and chlorophyll content.
NUE has several definitions based on agronomic, genetic, or physiological studies (Fageria et al., 2008).NUE is the efficiency of nitrogen recovery from applied fertilizer, or the N available to the crop, and this gives rise to the 33% efficiency of crop recovery (Raun and Johnson, 1999;Zhang et al., 2015).Alternatively, it is often considered a productivity index and defined as the yield produced per unit of available N to the crop expressed in terms of kg yield per kg of available N (Moll et al., 1982;Barraclough et al., 2010).NUE included two components namely, N uptake efficiency (NUpE) and N utilization efficiency (NUtE) and is mathematically the product of NUpE x NUtE (Good et al., 2004;Hakeem et al., 2012;Vijayalakshmi et al., 2013).NUpE is the ability of the plant to take up N from the soil.
It is estimated as the ratio of N taken up by the plant per unit of N available from the soil and applied fertilizer and is expressed as kg N (in the crop) per kg N (available).NUtE is the ability to use N to produce gain yield which is estimated as the grain yield produced per unit of N taken up by the plant and is also expressed as kg (grain) per kg N (Ladha et al.,1998;Hirel et al., 2007).
A study identified 333 genomic regions associated to 28 traits related to NUE in winter wheat (Cormier et al., 2014).Cormier et al. (2016) has reviewed the molecular studies done for NUE.However, However, NUE and its related traits are polygenic and influenced by genetic background.
The high NUE/ N-insensitive genotypes (NIS-top grain yielders) give more or equal GY with minimal application of N fertilizer compared to recommended N fertilizer conditions (Hawkesford, 2017).The understanding of GY and its associated NUE traits performance is still lagging in wheat but few studies had explored genotypic variations for NUE at different N levels.A significant genetic variability was observed for NUtE than NUpE at low N inputs in wheat by Gaju et al., (2011).However, Maman et al., (2006) concluded that the NUtE trait was less responsive than the N uptake with increased N levels.In broader perspectives, NUE is an output of available N uptake, its efficient utilization, and ultimately remobilization to grain.Several studies in cereals suggested enhanced NUE through improved NUtE under low N conditions (Moll et al., 1982;Good et al., 2004).The nitrogen harvest index (NHI) is another useful measure of the efficiency of N use, which is the fraction of total N taken up by the crop which is partitioned to the grain.NHI is independent of yield and uptake efficiency, and a low yielding crop may have a high NHI (Hawkesford and Riche, 2020).
The green revolution period in India focused primarily on the responsiveness of wheat genotypes to high-input supply.In the present scenario of climate change, realizing consistent output (yield and quality) of the genotype under varying levels of nitrogen input is a major challenge to wheat improvement programmes.Identifying promising genotypes for enhanced NUE in wheat and estimating genetic variability can provide useful guidance to breeders (Gaju et al., 2011).Keeping high yield when N supply is reduced and/or increasing yield when N supply is continuous may be two strategies for NUE enhancement.Nitrogen production costs, environmental pollution owing to nitrate leaching (Pathak et al., 2008), and greenhouse gas volatilization necessitate improvement of wheat nitrogen use efficiency at a reduced N supply.Simultaneously, GY is a complex trait controlled by a network of multiple traits and their associations.Hence, uncovering the genetic basis of GY, and other related NUE traits under low and high N conditions is a prerequisite to understanding the mechanism and also identifying ideal NUE associated traits for selection.Nitrogen use efficient crop varieties can be a great choice for ensuring sustainability in farming systems and meeting future customer requirements, particularly in the face of changing environment caused by climate change.Thus, the current study aimed to evaluate a diverse set of wheat genotypes for their response to GY and NUE related traits under different N levels and also to determine the suitable NUE traits for selection.This study provides useful information for uncovering the physiological and genetic basis of GY and its related traits under low N which further facilitates the development of low N stress resilient wheat cultivars.

Experimental design, treatment, and crop management
The experiment was laid out in an Alpha-lattice design at two treatment levels of nitrogen (T1 and T2) with two replications.The T1 consisted of soil N + 50% of the recommended nitrogen dose (RND) of 100 kg N ha -1 which is equivalent to 50kg/ha -1 whereas T2 consisted of soil N +RND of 100 kg N ha -1 .The genotypes were randomly allotted to plots in each treatment in each replication and grown in 6 row plots of 2.5 meter length with 20 cm row spacing.The nitrogen fertilizer dose was calculated as per the treatments and was applied to each plot.The soil samples from the experimental sites were collected from 0 to 30 cm soil depth before the start of the field experiment and after harvest of the crop and N was analyzed using standard procedure.All other recommended agronomic package of practices was adopted to raise a good crop.

Observations
Data on 24 agro-morphological, physiological, and NUE related traits were recorded from phase 41 to phase 92 of the Zadoks scale (Zadoks et al., 1974) for wheat plants in the four central rows of each plot.The agro-morphological traits included grain yield (q ha-1), agromorphological traits namely, days to heading, days to flowering, plant height (cm), number of productive tiller per meter row length, spike length (cm), awn length (cm), spikelet number per spike, grains number per spike, 1000-grains weight (g), biomass yield (q ha-1) and harvest index (%); Physiological traits namely, chlorophyll content (SPAD) and normalized difference vegetation index (NDVI) were recorded at booting (SPAD 1, NDVI-1), anthesis (SPAD-2, NDVI-2) and grain filling (SPAD-3, NDVI-3) stages.The chlorophyll content was measured by a Chlorophyll meter or Spad-meter and NDVI was measured by the Trimble GreenSeeker handheld crop sensor using standard procedures.Nitrogen use efficiency (NUE) and its related parameters like nitrogen uptake efficiency (NUpE), nitrogen utilization efficiency (NUtE), nitrogen harvest index (NHI), and total nitrogen uptake (TNUp) were worked out according to Moll et al., (1982) and Ayadi et al., (2012).NHI was calculated as [Grain N/ (Grain N + straw N)] *100 and expressed in percentage.The grain N (%) and straw N (%) contents were analyzed by the Micro-Kjeldahl digestion method as per the FAO guide to laboratory establishment for plant nutrient analysis (FAO, 2008).TNUp (kg N ha-1) was determined as the sum of nitrogen in straw and grain at harvest.NUpE was calculated as total N uptake/crop N supply where crop N supply includes fertilizer N + soil mineral N at planting.NUtE was calculated as grain yield/total N uptake.NUE was measured as Grain yield/ total nitrogen supply from the soil and applied fertilizer.The values for NUE, NUtE and NUpE were expressed in kg kg -1 .

Statistical analysis
Before pooling the data across environments, the ANOVA assumption was tested for its homogeneity using the Bartlet test (Snedecor and Cochran, 1989).The ratio of the highest error means square and the smallest error mean square value was compared with the F table and it showed non-significant results for all the traits hence, two years of data across environments were pooled.From pooled data, the analysis of variance (ANOVA) was done as per Panse and Sukhatme (1967).Critical difference was estimated to see significant differences among the genotypes for various studied traits.The genotypic and phenotypic coefficients of variation [Burton 1952] and heritability (Lush, 1940;Hanson et al., 1956) were worked out and the traits were categorized as low, medium and high based on genetic parameters (Shivasubramanian and Menon, 1973;Robinson et al., 1949;).The correlation among all the traits studied under both the nitrogen levels was worked out using Karl Pearson's simple correlation coefficient method.
For understanding the effect of nitrogen doses on different genotypes concerning yield as well as NUE, various indexes namely, stress susceptibility index (Fischer and Maurer, 1978), percentage reduction (Chouken et al., 2006) and yield stability index (Bouslama and Schapaugh, 1984) were also estimated to identify promising genotypes.As grain yield and NUE measurement are reflective of each other in totality, grain yield was used to estimate these indices.

3.1Trait specific variability among genotypes
The computed mean sum of squares for various traits namely, chlorophyll content at booting stage (CC-1), anthesis stage (CC-2), grain filling stage (CC-3), NDVI at booting stage (NDVI-1), anthesis stage (NDVI-2), grain filling stage (NDVI-3), days to heading (DH), days to maturity (DM), plant height (PH), number of productive tiller per meter row length (TPM), spike length (SL), awn length (AL), number of spikelet's per spike (SPS), number of grains per spike (GPS), thousand grains weight (TGW), biomass yield (BMY), harvest index (HI), grain protein content (GPC), nitrogen harvest index (NHI), total nitrogen uptake (TNUp), nitrogen uptake efficiency (NUpE), nitrogen utilization efficiency (NUtE) and nitrogen use efficiency (NUE) showed that genotypes differed significantly for all the traits which indicated the existence of variation among genotypes for various traits studied.A wide range was observed for all the traits evaluated under T1 (soil N + 50 kg Nha -1 ) and T2 (Soil N + 100 kg Nha -1 ) as shown in Table .1.In general, the mean values for all the traits except HI, NHI, NUtE and NUE were higher at full/recommended fertilizer dose as compared to reduced nitrogen dose.A similar trend was also observed for trait specific maximum values where all the traits except NDVI-1, SL, HI and NUE showed higher values under T2 as compared to T1.

Yield and agro-morphological traits
The present investigation included 12 agro morphological traits including grain yield.A wide range was observed for these yield and agro-morphological traits under both the nitrogen levels.In T1 (N50), the average GY of the tested genotypes was 28.65 q/ha and ranged from 16.87 to 38.91q/ha whereas in T2 (N100), GY ranged from 20.40 q/ha to 40.36q/ha with a mean of 32.32 q/ha.Nitrogen in limited condition (N50) resulted in 11.36% reduction in GY compared to N100.A higher mean BMY of 105.9 q/ha with a range of 79.1 to 132.4q/ha was recorded in T2 as compared to 90.8 q/ha mean BMY with range of 73.0 to 108.7q/ha under the T1 condition that showed 14.25% reduction.Interestingly, 1.3% lower HI was observed in T2 (31.5%) as compared to T1 (31.9%) condition.TPM, GPS and TGW are major yield component traits that significantly contribute to higher yield realization.The results showed a higher mean of 97.3 TPM, 50.1 GPS and 37.0g TGW in the  reductions for TPM, GPS and TGW respectively under T1 condition.The lower mean values for other morphological traits DH, DM, PH, SL, AL and SPS were observed in T1 with 4.6, 5.3, 7.0, 4.2, 6.7 and 7.6 percent reduction, respectively as compared to mean values for these traits in the T2 condition.

Physiological parameters
Chlorophyll content and NDVI values are two physiological parameters that directly indicate the phenotypic nitrogen levels in crop plants based on the greenness of leaves.Chlorophyll content (CC-1, CC-2, CC-3) and NDVI values (NDVI-1, NDVI-2, NDVI-3) were measured at booting, anthesis, and grain filling stages, respectively which also showed a wide range in both the conditions.Higher mean values of 51.6, 57.7 and 51.0 chlorophyll content at all three stages were recorded in T2 as compared to 47.7, 49.2 and 47.9 in the T1 condition with a reduction of 7.6%, 14.6% and 6.1%, respectively.Higher chlorophyll content was observed at the anthesis stage in both T1 and T2 conditions.A similar trend was observed for NDVI at anthesis and grain filling stages with 5.0% and 1.8% reduction in the T1 but NDVI at booting stage was a little higher in T1 as compared to T2.

NUE associated traits
Nitrogen based six traits were estimated in the present investigation (Table .1).The NUE in T1 ranged from 9.4 to 21.5 with an average of 15.9 kg grain /kg N whereas it was 15.2 kg grain /kg N with a range of 9.6 to 19.0 in T2 which indicated 4.6% reduction as compared to T1. NUE is mainly dependent on TNUp, NupE and NutE.The results indicated an increase of TNUp with increased application of N fertilizer.It ranged from 71.9 to 113.1 with an average of 94.6 kg N/ha in T1 and decreased by 18.6% as compared with T2 (N100) where it ranged from 89.0 to 134.8 with a mean of 116.2 kg N/ha.Similarly, NupE was 0.52 and 0.54 with a range of 0.40 to 0.63 and 0.42 to 0.63 in T1 and T2, respectively indicating 3.7% reduction in T1.A large variation was observed for NUtE of genotypes which ranged from 22.1 to 35.2 with an average of 30.2 under T1 and 18.0 to 35.2 with a mean of 28.1 in T2.The overall mean showed 7.4% reduction in NUtE in T2 due to poor performance of some of the genotypes which was reflected in lower values of range in T2.The NHI varied from 70.0 to 78.6% with an average of 75.8% in the T1 condition whereas it ranged from 66.1 to 78.6% with a mean of 73.8% in T2 indicating 2.76% reduction as compared to T1. Grain protein content (GPC) is an important trait depending on N uptake and its utilization and in this study, it was approx.4.0% higher in T2 as compared to the mean of T1.The GPC ranged from 10.9-13.7% with a mean of 12.6% in T2 as compared to 12.1% in T1 with a range of 10.6 to 13.3%.

Genetic parameters
Genetic parameters namely phenotypic (PCV) and genotypic (GCV) coefficients of variation, heritability in the broad sense (h 2 bs) were worked out as shown in Table .1.

Coefficients of Variation
In this study, phenotypic (PCV) and genotypic (GCV) coefficients of variation were estimated in T1 and T2 conditions.In general, higher PCV values were observed as compared to GCV values at both the nitrogen levels.

Heritability
The heritability values in broad sense ranged from 34.05% for SL to 86.84% for DH in T1 and 37.81% for NHI to 93.84% for DM in T2 (

Phenotypic Correlation
Character association was studied among the genotypes using Karl Pearson's simple correlation coefficient method to identify the interrelation among the traits for both the N levels.Nitrogen use efficiency was measured as grain yield divided by available N soil (soil N + fertilizer N) due to which the correlation of nitrogen use efficiency and grain yield at both levels of nitrogen treatments showed similar results (Fig. 1).Therefore, the correlation values with NUE were estimated as shown in Table .2.The results indicated a positive and highly significant correlated response of nitrogen use efficiency with DM, HI, TGW, GPS, SPS, CC-1, CC-2, CC-3, NDVI-1, NDVI-2, NDVI -3 TNUp, NUpE, NUtE under both T1 and T2 conditions.In addition, similar correlation of NUE with TPM and SL in T1 and with DH and GPC in T2 was also observed.Among these correlations, NUE showed a very strong correlation with HI (0.82), NUpE (0.80) and NUtE (0.80) in the T1 condition.A significantly positive correlation of NUE was also recorded with PH in T1 and TPM, SL, and BMY in the T2 condition.
All the six physiological traits namely CC-1, CC-2, CC-3, NDVI-1, NDVI-2 and NDVI -3 showed highly significant and positive correlations with each other.Among NUE related traits, GPC had a significantly positive correlation with NHI, TNUp, and NUpE under both T1 and T2 conditions.Similarly, a highly significant and positive correlation of TNUp with NUpE and NUtE was observed in both conditions.NUpE has a highly significant and positive association with NUtE in T1 but negative in T2.For agro-morphological traits, NUE/grain yield showed highly positive correlations with DM, SPS, GPS, TGW and HI under both the nitrogen conditions.Traits SPS, GPS and TGW have a highly significant and positive correlation with each other under both conditions.DM and DH have highly significant and positive associations in T1 as well as T2 conditions.Besides these, highly significant and positive associations of SL with DM, TPM, SPS, GPS, TGW, HI; TPM with SPS, GPS, HI; HI with SPS, GPS, TGW and DH with PH were observed the in T1 condition whereas the similar correlation of DH and DM with SPS and HI were observed in the T2 condition.A highly significant but negative association of HI with BMY was also observed under both the nitrogen levels.
The NUE related traits showed highly significant and positive correlations with yield and its component traits among which GPC with TGW & HI; BMY with TNUp & NUpE and NUtE with SPS, GPS, TGW & HI are prominent.All the physiological traits namely chlorophyll content and NDVI at booting, anthesis and grain filling stages showed highly significant and positive correlations with NUE related traits GPC, TNUp, and NUpE under both conditions.
NUtE has similar associations with chlorophyll content at all the three stages under the T1 condition and with chlorophyll content and NDVI at the anthesis stage in the T2 condition.
Considering yield and physiological traits, SPS, GPS, TGW, and HI showed highly significant and positive associations with all the six physiological parameters under both T1 and T2.
Under both conditions, BMY showed significant negative associations with chlorophyll content at all three stages.Significantly positive associations of TPM with all the

Stress tolerance indices for NUE
The pooled data of grain yield in T1 and T2 for all the 50 genotypes were used to estimate different stress related indices for the identification of promising genotypes that have the least effects of stress conditions, i.e., reduced nitrogen levels (Table .3).Three different indices namely yield stability index (YSI), stress susceptibility Index (SSI), and percent reduction under stress (PR) were estimated among which higher values of YSI and lower values of SSI and PR are desirable for the identification of promising genotypes under stress conditions.Genotypes showed a mean YSI of 0.89 with a range of 0.66 to 1.05 whereas the mean SSI was 0.99 with a range of -0.44 to 2.97.PR among genotypes was 11.33 ranging from -4.97 to 33.86 among all the genotypes, exotic lines showed better performance for YSI (0.90), SSI (0.86), and PR (9.84) in comparison to indigenous Indian germplasm lines which showed mean YSI, SSI, and PR of 0.88, 1.10 and 12.5 respectively.The results indicated that two indigenous genotypes namely PBW175 and NP846 showed higher yield at reduced nitrogen levels which is also supported by negative SSI and PR values in PBW175 (-0.44 & -4.97) and NP846 (-0.03 & -0.37), respectively.The perusal of results of YSI, SSI, and PR indicated that PBW175, NP846, MP4010, UAS304, and UASBW10227 were promising genotypes that were least affected for grain yield under reduced nitrogen doses.In addition, the indigenous germplasm lines, lines, HI1500, DTW2011-56, KRL1-4, GW2013-540, and exotic germplasm lines UASBW13356, UASBW12878, UASBW13367, UASBW13363, UASBW13358, and UASBW13359 showed promising performance for these three stress indices.

Discussion
Nitrogen (N) is often a limiting factor for crop yield and is therefore considered an essential macronutrient required for plant growth.Globally, the adoption of N-responsive cultivars and extensive application of N fertilizer has augmented food production in the past 50 years of agriculture.This excessive application of N fertilizer is increasing the cost of cultivation and thereby reducing the net profitability at the farm level in addition to negative impacts on the environment (Raun et al., 2002;Rothstein, 2007;Pathak et al., 2011;Ali et al., 2018;Hawkesford and Griffiths, 2019).The present investigation was driven by a worldwide willingness, especially from a huge number of small and marginal farmers in developing countries, to understand genotype behaviour for the identification and development of genotypes (without compromising GY) with high NUE under low N conditions.As demand for wheat grain is still rising, there is a need to boost productivity and production for the amount of N applied and therefore, identification of genotypes with better NUE is crucial.
The immediate aim should, therefore, be to exploit the available variability for NUE in wheat cultivars through the appropriate breeding procedure.
The present study aimed to understand the variability and correlations for NUE traits with various yield and physiological parameters and identify the promising genotypes based on various parameters of nitrogen sensitivity, response to N fertilizers, and susceptibility indices to low N availability.Précised phenotyping under low N input is challenging and influenced by the genotype (G), environment (E), and G × E interactions (Chen et al., 2014;Rao et al., 2018) which makes it difficult to identify nitrogen insensitive genotypes under low N conditions at field level.Presently, very few wheat breeding programs are targeting the development of low N tolerance traits which are a must for sustainable agriculture with minimal negative impacts on the environment.
In this study, a total of 50 diverse wheat genotypes collected from indigenous germplasm and international nurseries from CIMMYT were characterized for 24 yield, yield component, physiological, and NUE related traits at two different N levels under field conditions.Analysis of variance indicated highly significant variation among genotypes at both treatments for all the traits studied and this existence of significant differences for the different attributes is very helpful to improve the trait of interest.Diverse responses have been observed among the genotypes for all the traits across N levels, despite similar growth conditions and an equal amount of N fertilizer in a given N level.The genotypic variations observed purely reveal trait specific genotype plasticity.These results concur with other reported field experiments on wheat (Sial et al., 2005;Belete et al., 2018).
Wheat is more responsive to a higher dose of nitrogen whereas, sensitive to less nitrogen availability like most cereal crops.Less soil N availability leads to reduced plant vegetative growth especially tillering which results in reduced grain yield.(Kaur et al., 2015).In this study, reduction in tillering and ultimately yield reduction to the tune of approx.11.3% was observed at reduced N levels as compared to the recommended dose.
Similarly, the low N availability is expressed phenotypically in the form of low chlorophyll content as indicated by lower SPAD and NDVI values which might be due to less N availability in the leaves at lower N levels.Similar results of increased chlorophyll content by increasing N application were reported by Kitajima and Hogan (2003), and Pramanik and Bera (2013).Almost all the traits studied except NDVI-1, HI, NUE, NUtE and NHI, showed reduced performance at lower N doses which reflects the genotypic sensitivity to low N availability.The traits viz., CC-1, CC-2, SPS, TGW, GY, BMY, and TNUp, were identified as the most sensitive traits to reduced N supply (Table 3).Paponov and Engels (2005) also elaborated the effect of low N supply on the grain filling process.When N supply was low, the remobilization of N from the tillers and leaves was efficient and converted into grain N indicating better NUtE.This suggested that few genotypes have the potential to use nitrogen more efficiently and thus exhibit high NUE.The NHI indicates the level of efficiency of plants to use acquired total N for grain formation (He et al., 2017) for which significant genotypic variation was observed at both the N levels.Varied response of genotypes to NUpE, NUtE, and NHI was observed by Ortiz-Monasterio et al., (1997); Le Gouis et al., (2000); Brancourt-Hulmel et al., (2003); Muurinen et al., (2007); Chen et al., (2011) and Monostori et al., (2017) which are in tune with the performance of NUE related traits at varying N levels in the present study.
To assess the nature and magnitude of diversity among the genotypes, the phenotypic and genotypic coefficients of variation were estimated.In general, higher PCV values were observed as compared to GCV values at both the nitrogen levels.High PCV and GCV were observed for harvest index whereas other major yield traits and NUE related traits showed moderate values.Similarly, previous studies have established significant genetic variation for NUE-related traits in wheat (Yousef et al., 2008;Kalimullah et al., 2012).
In general, moderate to high heritability and was recorded under both nitrogen levels.The heritability values in a broad sense ranged from 34.05% to 86.84% in T1 and 37.81% to 93.84% in T2.High heritability (>60%) were observed for CC-3, DH, DM, SPS, GY, BMY, HI, NUE, and NUtE under both the conditions.A similar trend was also observed for yield and NUE related traits by Majumder et al. (2008) and Nikolic et al. (2013).High heritability is useful in laying importance in choice for such traits during selection.
In the present study, nitrogen use efficiency was measured as grain yield divided by available N (soil N + fertilizer N) due to which the correlation of NUE and grain yield at both levels of nitrogen treatments showed similar results.The positive and highly significant correlated response of NUE with DM, HI, TGW, GPS, SPS, CC-1, CC-2, CC-3, NDVI-1, NDVI-2, NDVI -3 TNUp, NUpE, and NUtE under both the condition suggested improvement of these traits for better NUE in wheat genotypes (Rangare et al., 2010;Baranwal et al., 2012).
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years during rabi 2016-17 and 2017-18 at All India Coordinated Wheat Improvement Project, Main Agricultural Research Station, University of Agricultural Sciences, Dharwad located at 15.48 o N latitude and 74.98 o E longitude and falls in agro-climatic Zone 8 of Karnataka state in Peninsular India.