Evapotranspiration of irrigated and rainfed maize–soybean cropping systems

We have been making year-round measurements of mass and energy exchange in three cropping systems: (a) irrigated continuous maize, (b) irrigated maize–soybean rotation, and (c) rainfed maize–soybean rotation in eastern Nebraska since 2001. In this paper, we present results on evapotranspiration (ET) of these crops for the first 5 years of our study. Growing season ET in the irrigated and rainfed maize averaged 548 and 482 mm, respectively. In irrigated and rainfed soybean, the average growing season ET was 452 and 431 mm, respectively. On average, the maize ET was higher than the soybean ET by 18% for irrigated crops and by 11% for rainfed crops. The mid-season crop coefficient K_c (=ET/ET₀ and ET₀ is the reference ET) for irrigated maize was 1.03 ± 0.07. For rainfed maize, significant dry-down conditions prevailed and mid-season K_c was 0.84 ± 0.20. For irrigated soybean, the mid-season K_c was 0.98 ± 0.02. The mid-season dry down in rainfed soybean years was not severe and the K_c (0.90 ± 0.13) was only slightly lower than the values for the irrigated fields. Non-growing season evaporation ranged from 100 to 172 mm and contributed about 16–28% of the annual ET in irrigated/rainfed maize and 24–26% in irrigated/rainfed soybean. The amount of surface mulch biomass explained 71% of the variability in non-growing season evaporation totals. Water use efficiency (or biomass transpiration efficiency), defined as the ratio of total plant biomass (Y_DM) to growing season transpiration (T) was 5.20 ± 0.34 and 5.22 ± 0.36 g kg^?1, respectively for irrigated and rainfed maize crops. Similarly, the biomass transpiration efficiency for irrigated and rainfed soybean crops was 3.21 ± 0.35 and 2.96 ± 0.30 g kg^?1. Thus, the respective biomass transpiration efficiency of these crops was nearly constant regardless of rainfall and irrigation.     

Nutrient‐solution techniques for evaluation of iron efficiency of soybean 1

Abstract

The iron (Fe) efficiency of soybean [Glycine max (L.) Merr.] genotypes generally has been evaluated in the field on calcareous soil. A nutrient‐solution system has been developed to permit evaluation of Fe efficiency throughout the year. The objectives of this study were to assess the effectiveness of nutrient‐solution tests for evaluating the Fe efficiency of soybean genotypes and to evaluate alternative nutrient‐solution techniques that could minimize the cost of labor and chemicals. Five bicarbonate (HCO₃ ^‐) concentrations and three solution‐change schedules were evaluated in a factorial arrangement. Eight soybean genotypes with a wide range of Fe efficiency were evaluated in each treatment and in replicated field tests on calcereous soil during 3 years. Rank correlation coefficients between mean chlorosis scores of genotypes in nutrient solution and field tests ranged from 0.81 to 0.91 for the three solution‐change schedules and from 0.85 to 0.89 for the five HCO₃ ^‐ concentrations. Replacing the solution every 4 d was not superior to replacing it only at each stage of plant development or not changing the solution throughout the test. A stepwise increase in HCO₃ ^‐ level at each stage of plant development was not superior to utilizing a constant level of HCO₃ ^‐ throughout the test. The most economical evaluation of the Fe efficiency of soybean genotypes in nutrient solution can be achieved with no change in the solution and one or more HCO₃ ^‐ levels that are held constant throughout the test.     

Molecular cloning and biochemical characterization of an endo-β-mannanase gene from soybean for soybean meal improvement

Soybean meal is the most commonly used protein source in animal feeds. Among the undesirable attributes of soybean meal is the high level of β-mannan, which was determined to be detrimental to the growth performance of animals. β-Mannan is a type of hemicellulose in the plant cell wall and can be hydrolyzed by endo-β-mannanase. The goal of this study is to isolate and characterize an endo-β-mannanase gene from soybean that can be used for genetic improvement of soybean meal. From the sequenced soybean genome, 21 putative endo-β-mannanase genes were identified. On the basis of their relatedness to known functional plant endo-β-mannanases, four soybean endo-β-mannanase genes (GmMAN1 to GmMAN4) were chosen for experimental analysis. GmMAN1 and GmMAN4 showed expression in the soybean tissue examined, and their cDNAs without the sequences for signal peptide were cloned and expressed in Escherichia coli to produce recombinant enzymes. Only GmMAN1 showed endo-β-mannanase hydrolase activity. Further gene expression analysis showed that GmMAN1 is specifically expressed in cotyledons of seedlings, suggesting a role of GmMAN1 in degrading mannan-rich food reserves during soybean seedling establishment. Purified recombinant GmMAN1 exhibited an apparent K(m) value of 34.9 mg/mL. The catalytic efficiency (k(cat)/K(m)) of GmMAN1 was determined to be 0.7 mL/(mg·s). GmMAN1 was also shown to be active in hydrolyzing the β-mannan-rich cell wall of soybean seeds.     

Regulation of accumulation of β-subunit of β-conglycinin in soybean seeds by nitrogen

Extract

The storage protein of soybean [Glycine max (L.)] seed mainly consists of glycinin, composed of acidic (38 and 45 kDa) and basic (22 kDa) subunits (Kitamura et al. 1976), and β-conglycinin composed of α′- (75 kDa), α- (72 kDa), and β-(52 kDa) subunits (Thanh and Shibasaki 1978).     

Biofortification of soybean meal: immunological properties of the 27 kDa γ-zein

Legumes, including soybeans ( Glycine max ), are deficient in sulfur-containing amino acids, which are required for the optimal growth of monogastric animals. This deficiency can be overcome by expressing heterologous proteins rich in sulfur-containing amino acids in soybean seeds. A maize 27 kDa γ-zein, a cysteine-rich protein, has been successfully expressed in several crops including soybean, barley, and alfalfa with the intent to biofortify these crops for animal feed. Previous work has shown that the maize 27 kDa zein can withstand digestion by pepsin and elicit an immunogenic response in young pigs. By use of sera from patients who tested positive by ImmunoCAP assay for elevated IgE to maize proteins, specific IgE binding to the 27 kDa γ-zein is demonstrated. Bioinformatic analysis using the full-length and 80 amino acid sliding window FASTA searches identified significant sequence homology of the 27 kDa γ-zein with several known allergens. Immunoblot analysis using human serum that cross-reacts with maize seed proteins also revealed specific IgE-binding to the 27 kDa γ-zein in soybean seed protein extracts containing the 27 kDa zein. This study demonstrates for the first time the allergenicity potential of the 27 kDa γ-zein and the potential that this protein has to limit livestock performance when used in soybeans that serve as a biofortified feed supplement.     

Evaluation of agronomic effectiveness of phosphate rocks for aluminum‐tolerant soybean cultivar

Abstract

The effect of liming on the agronomic effectiveness of three phosphate rocks (PRs) Pesca and Huila from Colombia and Sechura from Peru as compared with TSP was evaluated in a greenhouse experiment for an Al‐tolerant soybean cultivar grown on an acid Ultisol. On both unlimed (pH 4.4) and limed (pH 5.0) soils, the agronomic effectiveness of P sources in terms of increasing seed yield followed the order of TSP > Sechura PR > Huila PR > Pesca PR > check, an order similar to that of solubility of P sources. Liming slightly decreased the effectiveness of Pesca PR, whereas liming had no effect on Huila PR. A significant increase in agronomic effectiveness was observed upon liming for Sechura PR and TSP. Soil‐available P as extracted by the Pi method was closely related to the amount of N fixed by soybean crop that, in turn, was related to the soybean seed yield. Values of relative agronomic effectiveness (RAE) of PRs with respect to TSP were calculated by assuming the check = 0% and TSP = 100%. On unlimed soil, the RAE values of PRs were: Pesca PR = 31%, Huila PR = 42%, Sechura PR = 84%. On the limed soil, the RAE values were: Pesca PR = 8%, Huila PR = 24%, Sechura PR = 66%. It can be concluded that the use of PR with respect to that of TSP for soybean crop is more favorable in the unlimed soil than in the limed soil, provided that the soybean plant is relatively Al‐tolerant.     

Effect of potassium salts on alleviation of lime‐induced chlorosis in soybean 1

Studies were conducted to determine the efficacy of K salts in alleviating lime‐induced chlorosis. Greenhouse studies using a Gibbon silt loam [fine‐silty, mixed (calcareous), mesic Typic Haplaquoll] and a 1: 1 mixture of Gibbon soil and washed sand were conducted with KCl, KNO₃, K₂SO₄, K₂HPO₄, or KHCO₃ applied at rates of 0, 250, and 500 mg K/kg soil. An FeEDDHA treatment was included for comparison. Similar studies were conducted at two field sites known to produce lime‐induced chlorosis. Potassium salts were applied at 0, 20, and 40 g K/m of row. In the greenhouse, plants treated with KCl, KNO₃, and K₂SO₄ on Gibbon soil were less chlorotic than controls or plants treated with K₂HPO₄, or KHCO₃. No K treatment totally alleviated chlorosis except FeEDDHA. Chlorophyll correlated positively with chlorosis rating. No relationship was found between leaf Fe uptake and chlorosis. Plants grown in soil/sand exhibited no chlorosis and had lower Fe uptake than plants grown in Gibbon soil. Thus chlorosis was not due strictly to low soil‐Fe availability or inadequate Fe uptake. Bicarbonate in the soil solutions of both growth media treated with KCl was lower than controls which may explain the reduced chlorosis associated with this treatment.

One field site showed positive effects of K treatments on chlorosis rating, chlorophyll concentration, and seed yield. No treatment was as effective as FeEDDHA in influencing plant growth or yield. Total leaf Fe concentration was unrelated to leaf chlorophyll concentration. Inorganic cation/anion ratios in the plant were from 4.4–8.4 which could cause net H⁺ efflux by the plant and alkalinization of plant tissues. One possibility is that H⁺ efflux solubilizes P in the rhizosphere, which after uptake could immobilize Fe in the plant. Application of KCl, KNO₃, and K₂SO₄ generally lowered HCO₃ content of the upper 15 cm of both soils. High bicarbonate could increase rhizosphere P availability and increase immobilization of Fe in the plant.     

Comparison of three thermostable β-glucosidases for application in the hydrolysis of soybean isoflavone glycosides

A novel thermostable β-glucosidase (Te-BglA) from Thermoanaerobacter ethanolicus JW200 was cloned, characterized and compared for its activity against isoflavone glycosides with two β-glucosidases (Tm-BglA, Tm-BglB) from Thermotoga maritima. Te-BglA exhibited maximum hydrolytic activity toward pNP-β-d-glucopyranoside (pNPG) at 80 °C and pH 7.0, was stable for a pH range of 4.6-7.8 and at 65 °C for 3 h, and had the lowest K(m) for the natural glycoside salicin and the highest relative substrate specificity (k(cat)/K(m))((salicin))/(k(cat)/K(m))((pNPG)) among the three enzymes. It converted isoflavone glycosides, including malonyl glycosides, in soybean flour to their aglycons more efficiently than Tm-BglA and Tm-BglB. After 3 h of incubation at 65 °C, Te-BglA produced complete hydrolysis of four isoflavone glycosides (namely, daidzin, genistin and their malonylated forms), exhibiting higher productivity of genistein and daidzein than the other two β-glucosidases. Our results suggest that Te-BglA is preferable to Tm-BglA and Tm-BglB, but all three enzymes have great potential applications in converting isoflavone glycosides into their aglycons.     

Insensitivity of soybean photosynthesis to ultraviolet‐b radiation under phosphorus deficiency

Soybean [Glycine max (L.) Merr. cv Essex] was grown in sand in a greenhouse under 2 levels of biologically effective ultraviolet‐B radiation (effective daily dose: 0 and 11.5 kJ/m² UV‐BBE and 2 levels of P (6.5 and 52 μM). Plants were grown in each treatment combination up to the fifth trifoliolate stage. UV‐B radiation had no affect on plant growth and net photosynthesis at 6.5 μM P supply but decreased both these parameters when grown in the higher P concentration. Reductions in net photosynthesis were apparently due to direct effects on the photosynthetic machinery, since chlorophyll concentration and stanatal conductance were unaffected by UV‐B radiation. Both UV‐B radiation and reduced P supply increased the level of UV‐B absorbing compounds in leaf tissues and their effects were additive. The reduced sensitivity of P deficient plants to UV‐B radiation may be the result of this increase in UV absorbing compounds and possibly uv protective mechanisms associated with growth inhibition.     

Compositional equivalence of insect-protected glyphosate-tolerant soybean MON 87701 × MON 89788 to conventional soybean extends across different world regions and multiple growing seasons

The soybean product MON 87701 × MON 89788 expresses both the cry1Ac gene derived from Bacillus thuringiensis and the cp4 epsps (5-enolpyruvylshikimate-3-phosphate synthase) gene derived from Agrobacterium sp. strain CP4. Each biotechnology-derived trait confers specific benefits of insect resistance and glyphosate tolerance, respectively. The purpose of this study was to compare the composition of seed and forage from this combined-trait product to those of conventional soybean grown in geographically and climatically distinct regions. Field trials were conducted in the United States during the 2007 growing season, in Argentina during the 2007-2008 growing season, and in the northern and southern soybean regions of Brazil during the 2007-2008 and 2008-2009 growing seasons. Results demonstrated that the compositional equivalence of MON 87701 × MON 89788 to the conventional soybean extended across all regions and growing seasons. Further evaluation of the data showed that natural variation (region and growing season) contributed more to compositional variability in soybean, particularly for such components as isoflavones, fatty acids, and vitamin E, than transgene insertion.     

Ozone tolerances of soybean cultivars and near‐isogenic lines in a fumigation chamber

Ozone (O₃) toxicity is a potential yield‐limiting factor for soybean (Glycine max L. Merr.) in the United States and worldwide. The most economical solution to the problem is to use O₃‐tolerant cultivars. Thirty‐four cultivars and 87 near‐isogenic lines (NILS) of soybean were screened for O₃ tolerance in a fumigation chamber (250 ppb for three hrs). Most tolerant cultivars tested were ‘Cloud’, ‘T‐276’, ‘T263’, and ‘Kindu’. Moderately tolerant cultivars included ‘Davis’, ‘T‐210’, and ‘Elton’. Most sensitive cultivars were ‘Corsoy 79’, ‘Noir’, and ‘Midwest’. The original ‘Clark’ cultivar was not tested, but ‘Clark 63’ tended to be more tolerant than ‘Harosoy’. The aluminum (Al)‐tolerant ‘Perry’ cultivar also tended toward greater O₃ tolerance than the Al‐sensitive ‘Chief’, as observed earlier. Our rankings of ‘Hark’ as moderately sensitive and ‘Davis’ as moderately tolerant are also in agreement with earlier reports. Among NILS, the order of O₃ tolerance was generally Williams>Clark>Harosoy, but differences were also observed within these parental groups. For example, L68–560 was more tolerant than some other NILS of ‘Harosoy’. ‘L76–1988’ appeared more tolerant to O₃ than other NILS of ‘Williams’, but all ‘Williams’ NILS were more tolerant than most NILS of ‘Harosoy’ and ‘Clark’. Ozone‐tolerant and ‐sensitive soybean cultivars or NILS identified in our study may be useful tools in studies on mechanisms of 03 tolerance and differential 03 tolerances in plants and in the development of ameliorative measures.     

Physicochemical properties of β and α'α subunits isolated from soybean β-conglycinin

Soy protein has shown great potential for use in biobased adhesives. β-Conglycinin is a major component of soy protein; it accounts for 30% of the total storage protein in soybean seeds. β-Conglycinin was isolated and purified, and its subunits' (β, α'α) physicochemical and adhesive properties were characterized. Crude β-conglycinin was isolated from soy flour and then purified by the ammonium sulfate precipitation method. The α'α and β subunits were isolated from the purified β-conglycinin by anion exchange chromatography. Yields of α'α subunits and β subunits from 140 g of soy flour were 1.86 g (1.3%) and 0.95 g (0.67%), respectively. The minimum solubility for α'α subunits, β subunits, and β-conglycinin occurred in pH ranges of 4.1-5.4, 3.5-7.0, and 4.8-5.3, respectively. Transmission electron microscopy showed that the β subunits existed as spherical hydrophobic clusters, whereas α'α subunits existed as uniformly discrete particles at pH 5.0. Differential scanning calorimetry showed that β subunits had higher thermal stability than α'α subunits. The pH had a lesser effect on adhesion strength of the β subunits than on that of the α'α subunits. The adhesives made from β subunits also showed greater water resistance than those from α'α subunits and β-conglycinin. Soy protein rich in β subunits is likely a good candidate for developing water-resistant adhesives.     

Composition of transgenic soybean seeds with higher γ-linolenic acid content is equivalent to that of conventional control

γ-Linolenic acid (GLA) has been used as a general nutraceutical for pharmacologic applications, particularly in the treatment of skin conditions such as eczema. Four transgenic soybean lines that produce GLA at high yields (4.21% of total fatty acids, up to 1002-fold) were generated through the stable insertion of the Delta-6-fatty acid desaturase gene isolated from Borago officinalis into the genome of a conventional soybean cultivar. As part of the safety assessment of genetically engineered crops, the transgenic soybean seeds were compared with their parental soybean seeds (nontransgenic) by applying the principle of substantial equivalence. Compositional analyses were conducted by measuring the fatty acids, proximate analysis (moisture, crude protein, crude fat, carbohydrates, TDF, and ash contents), amino acids, lectins, and trypsin inhibitor activity. The present results showed that the specific transgenic cultivar studied was similar to the conventional control.     

Comparative response of corn and soybean to seed‐placed phosphorus over a range of soil test phosphorus

Abstract

The responses of corn and soybean to seed‐placed fertilizer were compared over NaHCO₃‐extractable soil phosphorus (P) levels ranging from 3 to 35 ppm in a two‐year experiment. Early season corn and soybean shoot‐P concentrations were increased with increasing soil test P and were increased with seed‐placed P regardless of soil test P, although the increases were greater for corn than soybean. Corn grain yield increased with increasing soil test P to a plateau level and increased with seed‐placed P regardless of soil test P. A side‐band (5 cm × 5 cm) application of 39 kg P ha^‐1 at a low soil test P increased yield more (P<0.15) than application of 7 kg P ha^‐1 with the seed. A side‐band application of 9 kg P ha^‐1 at a medium soil P test did not increase yield. Soybean yield was increased with increasing soil test P one year out of two, but did not respond to seed‐placed P in either year. The yield response of corn was attributed to the increased P concentration prior to the 6‐leaf stage.     

In vitro assessment of the bioaccessibility of fatty acids and tocopherol from soybean oil body emulsions stabilized with ι-carrageenan

The present investigation aimed to expand the knowledge of the in vitro bioaccessibility of fatty acids and tocopherol from natural soybean oil body emulsions stabilized with different concentrations of ι-carrageenan. Several physicochemical parameters including proteolysis of the interfacial layer, interfacial composition, and microstructure were evaluated with regard to their impact on the bioaccessibility of fatty acids and tocopherol. Results from simulated human digestion in vitro indicated that the bioaccessibility of total fatty acids and tocopherol decreased (62.7-8.3 and 59.7-19.4%, respectively) with the increasing concentration of ι-carrageenan. During the in vitro digestion procedure, ι-carrageenan affected physicochemical properties of the emulsions, thereby controlling the release of fatty acids and tocopherol. These results suggested that soybean oil body emulsions stabilized with ι-carrageenan could provide natural emulsions in foods that were digested at a relatively slow rate, the important physiological consequence of which might be increasing satiety.     

Soil-plant micronutrients dynamics in response to integrated fertilization under wheat–soybean cropping system at Rawalakot,Pakistan

This study provided an insight on improving soil-plant micronutrients availability in response to poultry manure (PM), wheat milling residues (WMR) and urea N (UN) and their integration in wheat–soybean cropping system. The treatments were: control; poultry manure full, PM₁₀₀; wheat milling residues full, WMR₁₀₀; urea N full, UN₁₀₀; PM half and WMR half, PM₅₀+WMR₅₀; UN₅₀+PM₅₀; UN₅₀+WMR₅₀; UN₅₀+PM₂₅+WMR₂₅. All amendments were added at the rate or equivalent to 100 kg total N ha^–1. Results indicated that the integrated treatments increased Cu, Fe, Mn and Zn uptake of wheat by 35.7–103%, 48.4–111.1%, 85.2–267.0% and 33.8–128.2%, respectively over control. In soybean the corresponding increase in micronutrient uptake (Cu, Fe, Mn and Zn) was 18.3–60.3%, 27.5–87.4%, 14.1–54.6% and 13.2–58.0% in integrated treatments. The post-harvest soil analysis indicated 2 to 3-fold increase in micronutrient content with highest values in PM₁₀₀ i.e., 2.66 mg kg^?1 for Cu, 14.41 mg kg^?1 for Fe, 18.58 mg kg^?1 for Mn and 2.44 mg kg^?1 for Zn, respectively. The results showed that the PM either alone or in integrated with WMR and UN can be an effective management strategy for improving micronutrient content of soil–plant.     

Response of soybean to foliar‐applied boron and magnesium and soil‐applied boron 1

Annual plants may partition carbon (C) preferentially to reproductive structures slowing root elongation and subsequent nutrient uptake. Although foliar applications of nitrogen (N), phosphorus (P), potassium (K), and sulfur (S) supplement uptake by roots, soybean [Glycine max (L.) Merr.] yield increases have not been found in most studies. Experiments were designed to determine if foliar applications of boron (B), magnesium (Mg), or B+Mg would increase soybean yield and if soybean would respond to B applied to the soil several weeks prior to planting. Foliar B or Mg applied separately four times during reproductive growth did not affect soybean yield. However, four foliar applications of B+Mg increased soybean yield 12% at Mt. Vernon and 4% at Columbia over a three‐year period. Two foliar applications of B+Mg during the late reproductive stages increased soybean yield 8% over a two‐year period. The yield increase from foliar B+Mg treatment resulted from an increased number of pods on the main stem (18%) and branches (44%). A 2.8 kg/ha B application to soil eight weeks prior to planting increased soybean yield 11% during the first year and 13% the second year but had no effect on soybean yield by the third year after application. When results from the first two years were combined, 2.8 kg/ha B applied to soil increased the number of pods per branch by 17% and the number of branch pods per plant by 39%. Foliar applications of B+Mg increased soybean yield in four of six site‐years in the three‐year experiments at two locations.     

Effect of aluminum on soybean nodulation and nodule activity in a vertical split‐root system

Soybean plants (Glycine max L. cv Santa Rosa) grown hydroponically in nutrient solutions had reduced nodule mass and numbers in the presence of aluminum (Al). Reduced nodule number was attributed mainly to hydrogen (H) ion toxicity, whereas Al had a stronger effect on nodule growth. Using a vertical split‐root system with Al exclusively in the lower (hydroponic) layer also resulted in a significant reduction of nodulation and nodule growth in the surface compartment (vermiculite). This indirect effect could be attributed mainly to Al rather than H. Subsurface Al had no apparent effect on shoot growth or root growth of the upper compartment, but significantly limited root growth in the lower compartment where it was applied. The indirect effect of Al on nodulation could be a reflection of the abnormal root growth in the lower compartment. Split‐root experiments with a high Al soil, however, produced different effects. High Al in soil used exclusively in the lower compartment did not reduce nodule numbers or mass in the upper compartment despite being more harmful than the Al solutions to nodulation and growth of plants when used in a single compartment. Growth of roots in the subsurface compartment was also much less affected by the high soil Al compared with the Al‐containing nutrient solutions. Nodule activity, as estimated by xylem sap ureide levels, was only reduced after direct exposure of nodules to Al. A pronounced increase in the ratio of asparagine/glutamine occurred in all Al treatments where nodulation was reduced, and in some cases, there was an increase in total amino acid concentration of the xylem sap.     

Effect of freeze–thaw cycles on carbon stocks of saline–alkali paddy soil

This study aims to provide basic data to support accurate estimation of carbon stocks and reveal the physicochemical factors that influence the carbon cycle in saline–alkali soils. Soil samples were collected during initial freezing, complete freezing, initial thawing and complete thawing stages. Levels of soil organic carbon (SOC), soil inorganic carbon (SIC), moisture, salinity, pH and available nitrogen were determined, and variations were observed during the freezing and thawing periods. Correlation analysis and regression analysis of carbon contents and physicochemical properties were performed. The results showed that freeze–thaw cycles have significant effects on carbon contents. The SOC content initially decreased in the freezing stage and then increased in the thawing stage. However, the SIC content initially increased in the freezing stage, decreased in the initial thawing stage and finally increased in the complete thawing stage. The migration and transformation of SOC and SIC were observed both temporally and spatially. SOC was positively correlated with available nitrogen, moisture and salinity and negatively correlated with pH; while SIC was negatively correlated with available nitrogen, moisture and salinity and positively correlated with pH. Among the factors evaluated, available nitrogen and salinity exerted the greatest effects on SOC and SIC contents, respectively.