Effects of nitrogen fertilization on nitrogen use efficiency of Coker (flue-cured) tobacco inoculated with Azotobacter chroococcum.
Due to the strong influence of nitrogen (N) on plant productivity, a vast amount of N fertilizers is used to maximize crop yield. Over-use of N fertilizers leads to severe pollution of the environment, especially the aquatic ecosystem, as well as reducing farmer's income [23, 19]. When N fertilizer is applied at greater rates than required for maximum yield, plant biomass and long-term soil organic carbon increase, but nitrogen use efficiency (NUE) decreases . NUE is a generic term that expresses some measures of plant yield or N as a ratio of N supply from soil or fertilizer or both . Improving NUE is an important target because it increases profitability, either through greater yields or reduced fertilizing costs . Results of Bertin and Gallais in 2000 showed that NUE was highly negatively related to N content . The three most common measures of NUE are agronomic N use efficiency (ANUE), N recovery efficiency (NRE) and physiological N use efficiency (PNUE) . ANUE is the product of the efficiency of N recovery from applied N and the efficiency with which the plant uses each additional unit of N acquired and can be increased by N, crop, and soil management practices . In fact, ANUE is a parameter representing the ability of the plant to increase yield in response to N applied . NRE is a measure of how much of N supply got into the plant. Typically, reports of NRE refer to applied fertilizer uptake, measured by actual N uptake and sometimes by yield . NRE in 160 rice experiments averaged 44% . Worldwide; single-season recovery of N in harvested crops is estimated at 33% of applied fertilizer N . Poor N recovery is a function of N flows to competing pathways such as gaseous N losses, leaching and biological immobilization [20, 17].
The observed patterns of N partitioning among plant parts at harvest depended on the amount of N additionally taken up by plants. Nitrogen harvest index (NHI) has been proposed as a criterion in selection of plants with high N use Efficiency . In fact, NHI is an index for N recovery in the harvested product. Farrokh et al. in 2012, studied effects of N and K fertilizers on performance and some of quantitative features of flue-cured tobacco K326 . They reported that N fertilizer had not any significance effect on NHI. Whereas, simultaneous application of N and K fertilizer had significance effect on NHI .
There are numerous studies of NUE in rice , wheat , maize , potato  and cotton . The results of Sifola and Postiglione  in Burley tobacco showed that application of N fertilization had significant effects on both NRE and ANUE, but not on PNUE. Ruiz et al. in 2006, with study on six commercial tobacco cultivars, it has been observed that grafting tobacco plants can be used as a quick and effective method to improve NUE, which has beneficial implications for human health and causes a reduction in environmental pollution . Yuan et al. in 2008, investigated NUE during different growth stages of flue-cured tobacco . Their results showed that tobacco plant mainly absorbed N from fertilizers in root elongation and vigorous growth stages and from soil in maturing stage. MacKown and Sutton in 1997, reported that fertilizer N use efficiency was 36.6% when N was broadcast and that total N in above-ground organs at harvest increased with increasing N supply, but N fertilizer recovery tended to decrease . Sisson et al. in 1991 were evaluated twelve popular cultivars spanning a period of development from the 1920s through the 1980s at fertilizer N rates of 47, 68, and 89 kg.[ha.sup.-1]at Research Station, Reidsville . They were found significant differences among cultivars and among N rates for all traits. Based on, values for NUE increased from the oldest to the newest cultivars and NUE of all cultivars examined decreased as the level of applied N increased and changes in NUE were the result of changes in both N uptake and the efficiency of utilization for dry matter accumulation . Santhi and Ponnuswamy  compared two levels of N fertilization (75 and 150 kg N.[ha.sup.-1]) on different cultivars of chewing tobacco and showed that NUE decreased with increasing amounts of N applied (it reached to maximum of 39% for the 75 kg N.[ha.sup.-1] level).
Despite numerous studies about N use efficiency in various plants, however, there are a few studies about tobacco, especially inoculated with Azotobacter chroococcum. The main purpose of this research was investigating the N use efficiencies of tobacco inoculated with Azotobacter chroococcum at various levels of N fertilization.
Material and Methods
Site description, weather conditions and soil properties:
A field experiment was conducted at the experimental farm of the Rasht Tobacco Research Station lying in 37[degrees] 16' northern latitude and 49[degrees] 31' eastern longitude, in Guilan province of Iran.
Mean annual precipitation at the nearest meteorological station (Rasht synoptic station) is 1359 mm without any dry season. Average of annual temperature is 16[degrees]C, average of annual relative humidity in percent is 81.5, and with high relative humidity especially in the summer (94%). Summery of meteorological information for Guilan Tobacco Research Center during the growth season of tobacco as month period in cropping season were recorded in years of 2009 and 2010 (Table 1).
The soil of experimental site has a sandy loam texture. That is poor in organic matter and CEC. The pH is 6.5 in saturated paste. Table 2 shows some physical and chemical characteristics of used soil.
Plant culture, seedling inoculation and laboratory analysis:
This study was conducted on flue-cured tobacco plant (Nicotiana tabacum L.) cultivar Coker 347. In order to prepare the soil for tobacco cultivation, the experimental site was ploughed at the depth of 30 cm. After application of Eradican herbicide in the level of 5 liters per hectare (2:1000) a rotary was applied. Seedlings of Coker 347 (flue-cured) tobacco cultivar were transplanted in experimental plots having 30 [m.sup.2] areas (5x6 m). The transplanting was accomplished when the tobacco seedlings had approximately 15 cm height. Seedlings were transplanted with population of 20000 plants per hectare (the space between rows was 100cm and between plants on rows was 50cm). Basal fertilizer was applied annually over the whole experimental area and consisted of 96 kg [P.sub.2][O.sub.5] [ha.sup.-1] and 200 kg [K.sub.2]O.[ha.sup.-1] and N fertilizer was applied in four levels (0, 15, 30 and 45 kg N [ha.sup.-1]). N and K fertilizer was divided by two stages (3 and 30 days after seedling). The commercial fertilizers used were ammonium nitrate (34.5% N), triple superphosphate (46% [P.sub.2][O.sub.5]) and potassium sulfate (50% [K.sub.2]O) and used in the middle of the ridge at 15 cm depth.
A mixture of fine calcium carbonate neutralized peat as a carrier was packed into polyethylene bags (200 g carrier per bag), then sealed and sterilized with gamma irradiation (5.0x [10.sup.6] rad/s). Azotobacter chroococcum strain AG22 was grown on the medium of Hegazy and Neimela (1976), incubated for 48 hr at 28[degrees]C to ensure population density of [10.sup.9] cfu.[ml.sup.-1] culture and then injected into the bags containing the sterilized carrier to have [10.sup.8] cell.[g.sup.-1] carrier. Ultimately, microbial inoculant was prepared in Soil and Water Research Institute of Iran as powdery form.
In addition, root segments of seedling from all treatments (except control treatment) were partially surface sterilized in absolute ethanol for 2 min and washed twice in sterilized distilled water. For applying Azotobacter, first a 20% solution of sucrose ([C.sub.12][H.sub.22][O.sub.11]) was made, afterwards, powder of Azotobacter in three levels (0, 1 and 2 kg.[ha.sup.-1]) was added to it and completely mixed, afterwards, seedling roots were placed in the solution for 30 minutes.
Leaves were harvested at five times in each year (2009 and 2010) from three leaf positions (priming, cutter and tip). Priming consist of the oldest, most mature leaves grown at the bottom of the stalk. Cutters are the middle leaves and are normally wider long than others leaves; and tips are narrow leaves from the top of the plant . First plants leaves area by leaf meter (model Ga-5 produced by Japan OSK company) were assessed. Total N was analyzed employing the Kjeldahl procedure . Nicotine was measured using CORESTA recommended method no. 35 (ISO/DIS 15152). In this method, an aqueous extract of the tobacco was prepared and the total alkaloids (as nicotine) content of the extract was determined by reaction with sulphanilic acid and cyanogen chloride. Cyanogen chloride was generated in situ by the reaction of potassium cyanide and chloramine T. The developed color was measured at 460 nm. . Reducing sugar was measured using CORESTA recommended method no. 38 (ISO/DIS 15154). In this method, samples (containing sucrose) were first hydrolyzed by invertase to form reducing monosaccharides. Reducing sugars reacted with phydroxybenzoic acid hydrazide (PAHBAH) in an alkaline media to form a yellow color measured at 410 nm. Calcium was used to enhance the color development .
Measurement of harvest index and nitrogen use efficiencies:
N harvest index (NHI) was calculated as the ratio of N in leaves (kg.[ha.sup.-1]) on the whole plant N amount (kg.[ha.sup.-1]). Nitrogen use efficiencies were calculated by equations of Dobermann . ANUE is ratio of yield to N supply and can be calculated by Eq. 1. NRE is the ratio of plant N to N supply and can be calculated by Eq. 2. PNUE is the ratio of yield to plant N and can be calculated by Eq. 3.
ANUE = [[Y.sub.N] - [Y.sub.0]/[F.sub.N]] (Eq. 1)
NRE = [[U.sub.N]1 [U.sub.0]/[F.sub.N]] (Eq. 2)
PNUE = [[Y.sub.N] - [Y.sub.0]/[U.sub.N] - [U.sub.0]] (Eq. 3)
Where, [Y.sub.N] is crop yield with applied N (kg.[ha.sup.-1]), [Y.sub.0] is crop yield (kg.[ha.sup.-1]) in a control treatment with no N, FN is amount of (fertilizer) N applied (kg.[ha.sup.-1]), UN is total plant N uptake in aboveground biomass at maturity (kg.[ha.sup.-1]) in a plot that received N and [U.sub.0] is the total N uptake in aboveground biomass at maturity (kg.[ha.sup.-1]) in a plot that received no N.
The experiment was performed as a 4x3x2 (Nitrogen levels x Bacterium levels x Years) factorial experiment in a randomized complete block design (RCBD), with three replications. Statistical analysis of data including normality test, analysis of variance, and comparisons of means was performed by using SAS program . Comparison of means was carried out using Tukey's tests at P < 0.05. Moreover, Pearson correlation and multivariate regression (stepwise) between parameters was performed by using SPSS program .
Results and Discussions
Quantitative and qualitative characteristics and nitrogen uptake:
The result of analysis of variance (ANOVA) (mean squares) on quantitative and qualitative characteristics and N concentration were presented in previous study by Sabeti et al. in 2012 . Summary of statistics information for quantitative characteristics (height, leaf length, leaf width, leaf number and yield) of tobacco were presented in Table 3. The plant height values ranged from 87 to 239 cm with mean of 133.1 (Table 3). The leaf length values ranged from 34 to 55 cm with mean of 44.7 and the leaf width ranged from 16 to 33 cm with mean of 20.8 (Table 3). Furthermore, the number of leaves ranged from 16 to 27 cm with mean of 21 (Table 3). The results of pervious study  revealed significant N fertilizer effect on plant height and leaf length that was similar to results of Haghighi et al. in 2011. The yield of cured leaf weight (mean) was higher in cutter in comparison with yield of cured leaf weight of priming and tip. Mean of yield in priming, cutter and tip was 319, 620 and 501, respectively (Table 3).
The nicotine and reducing sugar are two important compounds have been found in tobacco. They play an important role in the quality of tobacco. Mean of nicotine concentration in priming, cutter and tip was 1.96, 1.85 and 2.59, respectively. Nicotine concentration was higher in tip similar with N uptake (Table 4). Nicotine content interacts with N supply and increases with N content . Mean of reducing sugar concentration in priming, cutter and tip was 1.96, 1.85 and 2.59, respectively. Reducing sugar concentration showed a reverse trend to nicotine concentration and N concentration in plant . Mean of nitrogen concentration in priming, cutter and tip was 1.93, 2.37 and 2.38, respectively (Table 4). There is an increasing trend from bottom leaves (priming) to top leaves (tip). Mobilization of nitrogen from old leaves to meristems and young leaves leads to a diminished concentration of nitrogen in old, bottom leaves of plants . The amount of N in the plant increased in response to N fertilization. N uptake significantly increased with increasing of N fertilizer in priming, cutter and tip leaves .
Leaf area index (LAI):
Leaf area is vital and fundamental for light absorption by a plant and has an important effect on crop yield . The result of analysis of variance (ANOVA) on LAI showed that none of effects (year, N fertilizer and bacterium inoculation application) was statistically significant. Only, the interaction effect between N fertilizer and bacterium inoculation (N x B) had a significant effect (p<0.01) on LAI (Table 5). LAI increased with increasing of N fertilizer, although it was not statistically significant. The comparison of mean for the interaction effect between nitrogen and bacterium (N x B) on LAI showed that the N2B1 (with 15 kg.[ha.sup.-1] N and inoculation with without bacterium) and N1B1 (control treatment) treatments had maximum and minimum values of LAI, respectively (Figure 1a). LAI is an important factor affecting the photosynthesis rate of the plants. An early increase in leaf area could increase the potential crop yield . LAI had a significant positive correlation (p<0.01) with quantitative characteristics such as height, leaf length, leaf width, leaf number and yield. LAI also had a significant positive correlation (p<0.05) with N concentration, but it had not significant correlation with qualitative characteristics (nicotine and reducing sugar) (Table 7). The best model of LAI (model 1 in Table 8) was assigned. It showed that quantitative characteristics (plant height, leaf length, leaf width and leaf number) had important role in LAI.
Nitrogen harvest index (NHI):
The result of ANOVA on NHI showed that the years of experiment had significant effect on NHI in all of leaves of priming, cutter and tip (Table 6). NHI in priming and tip was lower in 2009 and increased in 2010, while, NHI in cutter was higher in 2009 year and decreased in 2010. Moreover, N fertilizer application had no significant affect on NHI (Table 6). This agrees with former studies. Farrokh et al. in 2012, reported that N fertilizing had not any significant effect on NHI of flue-cured tobacco variety K326 . Bacterium inoculation application only had significant effect on NHI in cutter (Table 6) and control treatment (without inoculation) had maximum NHI. Moreover, NHI had a significant positive correlation with leaf number in priming and yield in cutter and tip (Table 7). NHI had a significant negative correlation with leaf width in priming and tip, plant height in cutter and LAI, nicotine and reducing sugar contents in tip. The best models NHI in priming, cutter and tip (models No 2, 3 and 4 in Table 8) were assigned. With regard to the above equations, it is obvious that NHI is more related with physiological characteristics of tobacco in comparison with N use efficiencies.
Agronomic nitrogen use efficiency (ANUE):
The result of ANOVA on ANUE showed that the years of experiment had significant effect (p<0.01) on NUE in priming and cutter, but ANUE in tip did not change in years of experiment (Table 6). NUE in priming and cutter was higher in 2009 and decreased in 2010. Moreover, ANUE in priming and cutter were significantly affected by N fertilizer and bacterium inoculation application (Table 6), so that ANUE decreased by N fertilizer and bacterium inoculation application (Table 6). N fertilizer and bacterium inoculation application had no significant effect on ANUE in tip (Table 6). Changes in ANUE mainly reflected those in NRE and the efficiency of N uptake . Novoa and Loomis in 1981, reported that changes in AE due to increasing levels of N fertilization that had changed the NHI .
Jamaati-e-Somarin et al. in 2010, reported that with increasing the N levels, ANUE decreased in potato . Furthermore, Sifola and Postiglione in 2003, reported that ANUE decreased with increasing the N levels in Burley tobacco . The result of ANOVA (Table 6) showed that the interaction effect between year of experiment and bacterium inoculation (Y x B) had a significant effect on NUE in priming, based on [B.sub.2][Y.sub.1] treatment (with 1 kg.[ha.sup.-1] bacterium inoculant in 2009) and [B.sub.3][Y.sub.2] treatment (with 2 kg.[ha.sup.-1] bacterium inoculant in 2010) had maximum and minimum of ANUE in priming (Figure 1b). ANUE had a significant negative correlation with N and nicotine in all of leaves of priming, cutter and tip (except nicotine in tip). ANUE was higher in cutter in comparison with priming and tip. Delogu et al. in 1998, reported that ANUE was like in barley and wheat (8.7 and 9.2 kg.kg-1 of N applied, respectively), suggesting that both species respond equally to N fertilization . Moreover, ANUE had a significant positive correlation with yield and sugar content in all of leaves of priming, cutter and tip; and LAI in cutter (Table 7). Sisson et al. in 1991, reported that ANUE positively correlated with chemical quality of the cured leaf (especially the reducing sugar amount) . The best models of ANUE in priming, cutter and tip (models No 5, 6 and 7 in Table 8) were assigned. With regard to the above equations, it is obvious that with increase in yield, ANUE will increase, too; which shows yield is an important factor for increasing ANUE. In general, the main goal is reaching maximum ANUE, first by choosing a cultivar with high physiological efficiency, then avoiding N loss by making optimal fertilization .
Nitrogen recovery efficiency (NRE):
The efficiency of N absorption by the plant, expressed as NRE, was quite low and was like the results of Sifola and Postiglione  in Burley tobacco. Sifola and Postiglione in 2003, reported that the high amount of residual mineral N at the end of growing season and losses due to the topping of plants may explain the low NRE values .
The result of ANOVA on NRE was like ANUE, hence the years of experiment had a significant effect (p<0.01) on NRE in priming and cutter. This is in contrast with previous works made by Sifola and Postiglione  that have reported the year of study did not affect any of these efficiency indexes. NRE in tip did not change in years of experiment (Table 6). NRE in priming and cutter was higher in 2009 and decreased in 2010. Moreover, NRE in priming and cutter were significantly affected by N fertilizer and bacterium inoculation application (Table 6), so that NRE decreased by N fertilizer and bacterium inoculation application (Table 6). N fertilizer and bacterium inoculation application had no significant effect on NRE in tip (Table 6). The result of ANOVA (Table 6) showed that the interaction effect between year of experiment and bacterium inoculation (Y x B) for NRE was significant effect in priming, based on B2Y1 treatment (with 1 kg.[ha.sup.-1] bacterium inoculant in 2009) and [B.sub.2][Y.sub.2] treatment (with 1 kg.[ha.sup.-1] bacterium inoculant in 2010) had maximum and minimum of NRE in priming (Figure 1c). NRE had a significant negative correlation with N in cutter and nicotine content in priming. The results of Choudhury and Kennedy in 2005, showed NRE in rice decreased with increasing rate of N both surface broadcasting and injection fertilization . Moreover, NRE had a significant positive correlation with yield, LAI and leaf width in cutter and sugar content in all of leaves of priming, cutter and tip (except sugar in tip) (Table 7). The best models of NRE in priming, cutter and tip (models No 8, 9 and 10 in Table 8) were assigned. With regard to the above equations, it is obvious that with increase in yield content and decrease in nicotine, NRE will increase, too.
Physiological nitrogen use efficiency (PNUE):
The result of ANOVA on PNUE showed that the years of experiment had a significant effect on PNUE in all of leaves of priming, cutter and tip (Table 6). PNUE in priming, cutter and tip was higher in 2009 and decreased in 2010. Moreover, PNUE in all of leaves priming, cutter and tip were significantly affected by N fertilizer and bacterium inoculation application (Table 6), so that PNUE decreased by N fertilizer and bacterium inoculation application (Table 6). This agrees with former studies reporting that PNUE was affected by N fertilization and by the cultivar in flue-cured tobacco  and maize . Williams et al. in 2010, reported that PNUE is higher when N is low and it is low when N is abundant, affecting rate of N applied . Moreover, Jamaatie-Somarin et al. in 2010, reported that with increasing of N levels, PNUE was decreased in potato . This is in contrast with previous works made by Sifola and Postiglione  that reported the PNUE was not affected by neither irrigation nor N fertilizer treatment for Burley tobacco. The result of ANOVA (Table 6) showed that the interaction effect between year of experiment and bacterium inoculation (Y x B) for PNUE was significant in cutter, based on [B.sub.1][Y.sub.1] treatment (without bacterium inoculation in 2009) and B3Y1 treatment (with 2 kg.[ha.sup.-1] bacterium inoculant in 2009) had maximum and minimum of PNUE in cutter (Figure 1d). PNUE had a significant negative correlation with N in all of leaves and nicotine in priming. The negative correlation between PNUE and N fertilization could be explained by less N translocated to the stem and other organs. PNUE had a significant positive correlation with yield and sugar in all of leaves of priming, cutter and tip (Table 7). The best models PNUE in priming, cutter and tip (models No 11, 12 and 13 in Table 8) were assigned. With regard to the above equations, it is obvious that yield and N content are two important factors in all models.
As it is shown in this study, N use efficiencies in leaves of cutter were higher in comparison with leaves of priming and tip, but totally N use efficiencies are low in tobacco, especially at the time of increasing of N fertilizer and it is obvious that NUE must be increased with some methods such as biotechnology engineering or selection of desired cultivar. However, measurement of NUE requires careful experimentation and interpretation must consider potentially confounding factors. At low levels of N supply, rates of increase in yield and N uptake are high because N is the primary factor limiting crop growth and final yield. As the N supply increases, incremental yield becomes smaller. While the increase of chemical fertilizers application was very quick in Iran, unfortunately there is no attention to fertilizer efficiency. Solving this problem only needs promoting long-term management strategies. Strategies for improving N use efficiency should not be aimed at attaining the highest yield per unit of fertilizer applied, but rather at identifying the lowest fertilization rate required to achieve a satisfactory yield in a given environment.
We would like to gratefully thank all the members of Guilan Tobacco Research Center and Soil and Water Research Institute of Islamic Republic of Iran for providing the facilities to carry out this work and for their suggestions, comments and helps for preparing this paper.
ANUE--agronomic nitrogen use efficiency; LAI--leaf area index; N--nitrogen; NHI--nitrogen harvest index; NRE--nitrogen recovery efficiency; PNUE--physiological nitrogen use efficiency
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(1) Mohammad Ali Sabeti Amirhandeh, (2) Mehdi Norouzi, (3) Ali Reza Fallah Nosratabad
(1) Guilan Tobacco Research Center, Rasht, Iran; (2) Department of soil science, university of Guilan, Rasht, Iran, (3) Soil and water Research Institute, Karaj, Iran
Mohammad Ali Sabeti Amirhandeh, Guilan Tobacco Research Center, Rasht, Iran
Table 1: Summery of meteorological information at the experimental site in 2009 and 2010 years Climatic properties Month Jan Feb Mar Apr May 2009 Temp. Max. Mean 10.1 10.7 14.6 13.8 22.3 ([degrees]C) Min. Mean 3.8 7.7 8.8 9.4 17.4 Mean monthly 6.8 9.8 11.9 12.1 20.6 Mean Relative humidity % 87.3 85.1 83.2 80.4 77.1 Total Rainfall (mm) 85.7 47.3 25.4 112 33 2010 Temp. Max. Mean 20.1 21.5 22.4 20.4 23.4 ([degrees]C) Min. Mean 7.8 7.4 9.3 10.5 17.9 Mean monthly 9.2 8.8 10.4 13.5 20.4 Mean Relative humidity % 88 87 90 89 83 Total Rainfall (mm) 45.8 84.4 95.8 106.7 51.2 Climatic properties Month Jun Jul Aug Sep Oct 2009 Temp. Max. Mean 26.9 30.7 27.9 27.2 22.7 ([degrees]C) Min. Mean 22.2 24.5 22.1 21.6 17.3 Mean monthly 25.2 28.8 24.6 24.3 20.8 Mean Relative humidity % 75.2 70.4 84.1 80.3 83.3 Total Rainfall (mm) 31.7 4 105.6 110 147.6 2010 Temp. Max. Mean 31.3 33.1 33.4 30.2 26.1 ([degrees]C) Min. Mean 24.5 24.8 24.5 22.3 18.3 Mean monthly 28.3 29.1 29.3 25.6 20.5 Mean Relative humidity % 79 70 69 80 93 Total Rainfall (mm) 19.8 15.6 25.7 18.8 158.4 Climatic properties Month Annual Nov Dec 2009 Temp. Max. Mean 19.1 16.2 20.2 ([degrees]C) Min. Mean 13.1 8.3 14.6 Mean monthly 15.8 10.3 17.6 Mean Relative humidity % 79.5 84.2 80.6 Total Rainfall (mm) 105.7 122 930 2010 Temp. Max. Mean 22.2 21.3 25.4 ([degrees]C) Min. Mean 12.8 11.9 16 Mean monthly 16.4 15.4 18.9 Mean Relative humidity % 80 79 82.2 Total Rainfall (mm) 106.4 9.6 737.9 Table 2: Some physical and chemical characteristics of used soils Texture SP (a) pH ECe (b) (ds.[m.sup.-1]) sand silt clay 56 26 18 28 6.5 0.29 Texture OC (c) Total N Olsen-P (g.[kg.sup.-1]) (g.[kg.sup.-1]) (mg.[kg. sup.-1]) sand silt clay 56 26 18 5.9 0.6 73 Texture Exchangeble K CEC (d) (mg.[kg.sup.-1]) (cmol+.[kg.sup.-1]) sand silt clay 56 26 18 241 12 (a) SP=soil moisture percentage, (b) ECe=Electrical Conductivity, (c) OC=Organic Carbone, (d) CEC=Cations exchangeable capacity. Table 3: Summary of statistics information for quantitative characteristics (height, leaf length, leaf width, leaf number and yield) of tobacco Parameter Plant Leaf Leaf Leaf height length width number cm Maximum 239.0 55.0 33.0 27.0 Median 131.5 45.0 20.0 23.0 Minimum 87.0 34.0 16.0 16.0 Mean 133.1 44.69 20.8 20.8 Range 152.0 21.0 17.0 17.0 SD 19.2 5.1 3.5 1.9 CV (%) 15.8 9.6 13.3 6.8 Parameter Yield (kg.[ha.sup.-1]) Priming Cutter Tip Maximum 467 1460 887 Median 320 550 505 Minimum 200 143 173 Mean 319 620 501 Range 267 1317 714 SD 56 188 86 CV (%) 12.8 25.2 20.1 SD, Standard deviation; CV, coefficient of variation. Table 4: Summary of statistics information for nicotine and reducing sugar contents and nitrogen uptake of tobacco Parameter Nicotine (%) Reducing sugar (%) priming cutter tip priming cutter tip Maximum 3.31 2.89 4.50 18.89 30.75 23.73 Median 1.91 1.88 2.51 7.48 10.51 12.03 Minimum 0.94 0.96 1.57 1.46 2.69 4.63 Mean 1.96 1.85 2.59 8.15 11.64 12.46 Range 2.37 1.93 2.93 17.52 28.06 19.1 SD 0.26 0.2 0.27 3.45 3.03 4.27 CV (%) 6.41 7.51 6.68 14.94 9.83 14.80 Parameter Nitrogen (%) priming cutter tip Maximum 2.67 2.97 3.51 Median 1.90 2.35 2.42 Minimum 1.40 1.61 1.61 Mean 1.93 2.37 2.38 Range 1.27 1.36 1.36 SD 0.24 0.28 0.42 CV (%) 5.13 5.10 5.14 3D, Standard deviation; CV, coefficient of variation. Table 5: Results of ANOVA (significance) and comparison of mean effects of year, nitrogen fertilizer and bacterium inoculation on yield and nitrogen harvest index (NHI) S.O.V df LAI NHI priming cutter tip Replicate 2 * * Year (Y) 1 ** ** ** Nitrogen (N) 3 Bacterium (B) 2 * YxN 3 YxB 2 BxN 6 ** YxBxN 6 Error 46 0.04 11.5 44.8 54.8 CV 11.6 8.2 18.2 20.2 ([cm.sup.2]. % [cm.sup.-2]) Year (Y) 2009 3.23 a 17.3 b 51.9 a 30.7 b 2010 3.54 a 21.2 a 38.9 b 39.9 a Nitrogen (N) 0 2.98 a 20.8 a 43.4 a 35.7 a 15 3.49 a 18.2 a 45.3 a 34.4 a 30 3.54 a 19.0 a 45.6 a 35.4 a 45 3.62 a 19.0 a 45.3 a 35.7 a Bacterium (B) Without 3.35 a 20.6 a 46.5 a 35.2 a inoculation 1 3.67 a 17.9 b 46.5 a 35.6 a 2 3.64 a 19.3 ab 44.1 a 35.1 a * and ** significant at level of 5 and 1%, respectively. Means, in each column, with similar letters are not significantly different at the 5% probability level using Tukey's test. CV, coefficient of variation. Table 6: Results of ANOVA (significance) and comparison of mean effects of year, nitrogen fertilizer and Azotobacter chroococcum on agronomic nitrogen use efficiency (ANUE), nitrogen recovery efficiency (NRE) and physiological nitrogen use efficiency (PNUE). S.O.V df ANUE prim cutter tip Replicate 2 * * Year (Y) 1 ** ** Nitrogen (N) 2 * * Bacterium (B) 2 ** ** YxN 2 YxB 2 * BxN 4 YxBxN 4 Error 34 0.01 0.02 0.03 CV 9.4 12.9 15.7 kg.[kg.sup.-1] Year (Y) 2009 3.57 a 12.62 a 5.17 a 2010 2.27 b 4.97 b 5.17 a Nitrogen (N) 15 4.85 a 15.14 a 7.78 a 30 2.46 b 6.52 b 4.37 a 45 1.70 b 4.73 b 3.37 a Bacterium (B) Without 4.27 a 12.12 a 7.12 a inoculation 1 3.18 ab 10.90 a 4.94 a 2 1.32 b 3.37 b 3.46 a S.O.V df NRE priming cutter tip Replicate 2 ** ** ** Year (Y) 1 ** ** Nitrogen (N) 2 * * Bacterium (B) 2 ** ** YxN 2 YxB 2 ** BxN 4 YxBxN 4 Error 34 0.00 0.02 0.01 CV 4.7 8.9 8.4 kg.[kg.sup.-1] Year (Y) 2009 9.89 a 37.22 a 16.07 a 2010 7.78 b 20.32 b 18.24 a Nitrogen (N) 15 11.63 a 43.23 a 22.47 a 30 7.81 b 23.54 b 15.11 a 45 7.06 b 19.54 b 13.88 a Bacterium (B) Without 11.38 a 35.48 a 20.24 a inoculation 1 8.77 ab 34.99 a 17.18 a 2 6.36 b 15.85 b 14.04 a S.O.V df PNUE priming cutter tip Replicate 2 * Year (Y) 1 ** ** * Nitrogen (N) 2 * * * Bacterium (B) 2 ** ** ** YxN 2 YxB 2 * BxN 4 YxBxN 4 Error 34 0.01 0.01 0.01 CV 6.2 5.7 8.6 kg.[kg.sup.-1] Year (Y) 2009 31.86 a 27.47 a 27.50 a 2010 22.42 b 17.45 b 21.27 b Nitrogen (N) 15 33.50 a 26.06 a 31.92 a 30 25.99 ab 22.01 ab 23.18 ab 45 21.92 b 19.33 b 18.06 b Bacterium (B) Without 33.08 a 28.75 a 33.22 a inoculation 1 30.59 a 25.63 a 22.50 b 2 17.74 b 13.01 b 17.43 b * and ** significant at level of 5 and 1%, respectively. Means, in each column, with similar letters are not significantly different at the 5% probability level using Tukey's test. Table 7: Correlation coefficient between leaf area and nitrogen harvest indices and nitrogen efficiencies with quantitative and qualitative characteristics of tobacco in priming, cutter and tip leaves of tobacco Index Leaf LAI Height Leaf part number LAI Total 1 0.56 ** 0.64 ** NHI Priming -0.09 0.17 0.28 * Cutter -0.08 -0.29 * -0.12 Tip -0.37 ** 0.10 -0.10 ANUE Priming 0.22 0.10 0.20 Cutter 0.34 * 0.09 0.20 Tip 0.00 0.11 0.04 NRE Priming 0.23 0.13 0.20 Cutter 0.35 ** 0.12 0 21 Tip 0.01 0.16 -0.01 PNUE Priming 0.22 0.06 0.10 Cutter 0.24 0.05 0.12 Tip 0.07 0.07 0.06 Index Leaf Leaf Leaf Yield part length width LAI Total 0.84 ** 0.82 ** 0.34 ** NHI Priming 0.07 -0.30 * 0.13 Cutter -0.06 0.04 0.64 ** Tip -0.13 -0.44 ** 0.54 ** ANUE Priming 0.06 0.22 0.64 ** Cutter 0.10 0.35 0.66 ** Tip -0.02 -0.03 0.51 ** NRE Priming 0.10 0.21 0.67 ** Cutter 0.15 0.34* 0.69 ** Tip 0.04 -0.03 0.54 ** PNUE Priming 0.51 0.26 0.39 ** Cutter 0.11 0.26 0.60 ** Tip -0.02 0.06 0.41 ** Index Leaf Nitrogen Nicotine Sugar part LAI Total 0.23 * 0.20 -0.14 NHI Priming 0.07 0.14 0.81 Cutter -0.15 0.20 0.08 Tip 0.22 -0.35 ** -0.38 ** ANUE Priming -0.39 ** -0.39 ** 0.39 ** Cutter -0.46 ** -0.46 ** 0.43 ** Tip -0.39 ** -0.19 0.27 * NRE Priming -0.24 -0.30 * 0.33 * Cutter -0.38 ** -0.25 0.41 ** Tip -0.25 -0.21 0.18 PNUE Priming -0.47 ** -0.47 ** 0.51 ** Cutter -0.58 ** -0.14 0.40 ** Tip -0.61 ** -0.12 0.42 ** * and ** Significance at the 5% and 1% level (df for LAI and NHI=71 and for NUE, NRE and PNUE=53). LAI= leaf area index; NHI= Nitrogen Harvest Index; ANUE = Agronomic nitrogen Use Efficiency; NRE = Nitrogen Recovery Efficiency; PNUE = Physiological Nitrogen Use Efficiency; p = priming; c = cutter; t = tip. Table 8: Regression models of NHI and N use efficiencies No. Model [R.sup.2.adj] 1 LAI = -6.545 + 0.07 L + 0.164 W + 0.98 0.134 Num + 0.002 H 2 [NHI.sub.p] = 0.322 - 0.024 Nic - 0.003W 0.23 3 [NHI.sub.c] = 0.69 + 0.001 Y - 0.003 0.42 H - 0.009 Su 4 [NHI.sub.t] = 0.454 + 0.001 Y - 0.01 0.52 W - 0.07 Nic 5 [ANUE.sub.p] = -1.66 + 0.44 Y-6.2 Nic 0.62 6 [ANUE.sub.c] = 22.8 + 0.028 Y-13.6 Nic 0.59 7 [ANUE.sub.t] = 12.2 + 0.02 Y-7.1 N 0.32 8 [NRE.sub.p] = -3.55 + 0.08 Y-8.8 Nic 0.59 9 [NRE.sub.c] = 66.26 + 0.069 Y-34.7 Nic 0.67 10 [NRE.sub.t] = 9.4+ 0.05 Y-7.12 Nic 0.34 11 [PNUE.sub.p] = 31.06 + 1.19 Su+0.1 Y-27.86 0.49 N+3.5 LAI 12 [PNUE.sub.c] = 22.9 + 0.016 Y - 18.12 N + 0.55 0.28 H 13 [PNUE.sub.t] = 83.67 + 0.03 Y-31.4 N 0.43 [R.sup.2.adj] = Adjusted [r.sup.2] is a measure of goodness of fit in least-squares regression analysis, NHI = Nitrogen Harvest Index; ANUE = Agronomic nitrogen Use Efficiency; NRE = Nitrogen Recovery Efficiency; PNUE = Physiological Nitrogen Use Efficiency; p = priming; c = cutter; t = tip; N = Nitrogen; Y = Yield; Nic = Nicotine; Su = Sugar; LAI = leaf area index; H = plant height; L = leaf length; W = leaf width; Num = leaf number.
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|Title Annotation:||Original Article|
|Author:||Amirhandeh, Mohammad Ali Sabeti; Norouzi, Mehdi; Nosratabad, Ali Reza Fallah|
|Publication:||Advances in Environmental Biology|
|Date:||Jun 1, 2013|
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