Field experiments were conducted in the 2016/17 and 2017/18 cropping seasons in Presidente Bernardes, São Paulo, Brazil, 22º07'32" S and 51º23'20" W, 475 m asl. The soil is a typic Rhodustult, sandy loam (USDA 2010). The climate is tropical with dry winters and wet, hot summers. Soil samples were taken at the depths of 0 to 20 cm and 20 to 40 cm on May 05, 2016, for chemical analysis as in Raij et al. (2001). Selected chemical and particle size distribution of the soil are in Table 1. Lime (45% of CaO and 4.9% of MgO) was broadcast on the soil surface at 1,400 kg ha− 1 in September 2016 and 2,000 kg ha− 1 in October 2017. Rainfall, maximum and minimum temperatures, and vapor pressure deficit during the experiment are shown in Fig. 1.
Table 1
Selected chemical properties and particle size distribution of the soil
depth | pH (CaCl2) | SOM | P (Resin) | S | Al3+ | H + Al | K+ | Ca2+ | Mg2+ | | CEC |
cm | | g dm− 3 | mg dm− 3 | | mmolc dm− 3 |
0–20 | 4.7 | 13.6 | 2.8 | 4.0 | 0.6 | 17.6 | 0.8 | 7.3 | 5.1 | | 30.7 |
20–40 | 4.8 | 11.6 | 2.0 | 3.3 | 2.3 | 18.6 | 0.8 | 6.5 | 4.4 | | 30.3 |
| B | Cu | Fe | Mn | Zn | m | BS | sand | Silt | clay | |
| ------------------mg dm− 3------------------- | ---%--- | --------g kg− 1--------- | |
0–20 | 0.34 | 2.0 | 26.0 | 0.9 | 0.4 | 4.3 | 42.6 | 848 | 36 | 116 | |
20–40 | 0.41 | 1.4 | 31.5 | 1.0 | 0.3 | 16.4 | 38.6 | 841 | 23 | 137 | |
The treatments were an early (FM 913GLT) and a late (FM 983GLT) cotton cultivar and six K fertilization schemes (Table 2). Treatments were repeated in the same plots in both seasons. The experimental design was a 2 x 6 factorial in complete randomized blocks with five replicates. Each plot had four 7.0-m cotton rows spaced 0.8 m from each other. The two outer rows and 1.0 at the end of each row were considered borders and not included in evaluations.
Table 2
Scheme of cotton K fertilization
Treatments | Ruzigrass (RZ) | K rate | Time of K fertilization |
0K | no | 0 | - |
116K | no | 116 kg ha− 1 | 58 kg ha− 1applied 30 DAE and 58 kg ha− 1 at 45 DAE |
0K + RZ | yes | 0 | - |
116K on RZ | yes | 116 kg ha− 1 | applied on RZ vegetative stage (90 DAE) |
58K on RZ + 58K on C | yes | 116 kg ha− 1 | 58 kg ha− 1applied on RZ vegetative stage and 58 kg ha− 1 on cotton 30 DAE |
116K on C + RZ | yes | 116 kg ha− 1 | 58 kg ha− 1 30 DAE and 58 kg ha− 1 45 DAE |
DAE: days after emergence |
Table 3
Eigenvalues of all variables analyzed of FM 913GLT and FM 983GLT under different K management and the use of ruzigrass
Total variance explained (%) | FM 913GLT | FM 983GLT |
LAI 30 | − 0.6417 | -0.0285 |
LAI 60 | -0.4406 | -0.3578 |
LAI 90 | -0.8885 | -0.7625 |
LAI 120 | -0.9217 | -0.7661 |
Gs F1 | -0.4848 | -0.0170 |
Gs F7 | -0.0224 | -0.5170 |
Gs C2 | -0.6766 | -0.2506 |
SAI | -0.7899 | -0.8911 |
SPS | -0.8047 | -0.6714 |
SUSY | -0.7150 | -0.8713 |
Cellulose | -0.6132 | -0.5821 |
Ruzigrass was sown on May 05, 2016, and June 06, 2017 using 14 kg ha− 1 pure viable seeds. Potassium fertilizer was applied to the grass in the corresponding treatments early September each year, using potassium chloride. Ruzigrass was desiccated on November 1, 2016 and November 6, 2017, using glyphosate (1.92 g ha− 1 a.i.). Ruzigrass averaged 5.6 Mg ha− 1 of dry biomass. Cotton was sown on December 9, 2016, and November 23, 2017. Phosphorus was applied at 56 kg ha− 1 and N at 30 kg ha− 1 as mono ammonium phosphate at cotton planting. The rate of 140 kg ha− 1 of N was side dressed and equally split 35 and 45 days after plant emergence (DAE), using ammonium sulphate and urea, respectively. Boric acid was sprayed at 2.0 kg ha− 1 split into 4 weekly applications from the first flower. Weed, pests, and diseases were controlled according to standard farm practices in São Paulo State. Seven days before harvest plants were defoliated with Tidiazurom (60 g ha− 1 a.i.) + Diuron (30 g ha− 1 a.i). Cotton was handpicked 141 and 132 DAE in 2016/17 and 2017/18, respectively.
Leaf area index 1st and 2nd season
Leaf area index was measured at 30, 60, 90, and 120 days after emergence using a ceptometer (Accupar LP-80 – Decagon Devices) in three sub-samples per plot in both seasons.
Enzyme extraction and assay – 2nd season
At mid flowering (F6/F7 stage – Marur and Ruano 2001), twelve plants per plot had their first position flowers tagged, and 24 days later (24 DAA – days after anthesis) four bolls and their leaves were collected. At 48 DAA only bolls were collected since leaves were absent. This material was ground to a fine powder in liquid nitrogen. Leaf samples (0.5 fresh weight) and 3.5 mL of extraction buffer (50 mM Hepes-KOH, pH 7.5, 10 mM MgCl2, 2 mM EDTA, 5 mM DTT, 2% (w/v) PVP) were ground into a homogenate in an ice bath (Grof et al. 2007). The samples were centrifuged subsequently at 12,000 xg for 20 minutes at 4 ºC. The supernatant was gradually added with Dowex 1x4 and then centrifuged at 12,000 xg for 10 minutes at 4 ºC. The supernatant was collected and the Sucrose Phosphate Synthase (SPS - EC 2.4.1.14), Sucrose Synthase (SuSy - EC 2.4.1.13) and Soluble Acid Invertase (SAI - EC 3.2.7.26) were analyzed. All procedures were done at 4 ºC. Protein measurement was performed as in Bradford (1976), using bovine serum albumin as standard.
SPS activity was determined as in Huber and Huber (1991). A 50 µL enzyme solution alicote was added to 50 µL solution contained (100 mM Hepes-NaOH buffer, 50 mM MgCl2, 20 mM UDPG, 5 mM fructose 6-phosphate, and 17.5 mM glucose 6-phosphate. The reaction was started by the addition of extract and incubated at 25°C for 10 min. After stopping the reaction with 100 µL of 5 M KOH and 10 min of heating at 100°C, followed by 1 mL of 0.14% (w/v) anthrone in 80% (v/v) H2SO4, was added before 40 min of incubation at 40°C (King et al. 1997). SUC-6-P (and SUC) content was determined by comparing the A628 to that of a standard curve (0-200 nmol of SUC).
SuSy activity (synthetic direction) was determined by replacing fructose 6-phosphate with fructose in the same way as SPS activity. The unit of enzyme activity was expressed as µmol µmol Suc min–1 mg–1 prot. The SAI activity was determined as in King et al (1997). Shortly, 0.2 mL enzyme solution was added into 0.8 mL reaction solution (pH 4.8 0.1M Na2HPO4–0.1 M sodium citrate, 0.1 M sucrose), and reacted at 37°C for 30min.
Determination of cellulose in cotton fiber – 2nd season
Fruit samples were taken from the first position (P1) of the 10th sympodial branch 24 and 48 DAA and dried in a forced-air oven at 65 ºC for 96 hours. Determinations were conducted in three replications, according to Van Soest et al. (1991). The acid-detergent fiber (ADF) content was determined using an acid detergent (trimethylammonium bromide, standardized sulfuric acid. After examining the ADF content, the cellulose content was assayed in cotton samples using a standardized solution of sulfuric acid.Statistical analysis
After testing for homogeneity and normality, data were submitted to ANOVA. The experiment was arranged in a complete randomized block design in a 2 x 6 factorial scheme (cultivars x K management) and five replicates. The data were analyzed with three-way (cultivar, K management and year) analysis of variance and Tukey test (P < 0.05) was used to compare treatment means. When the differences between cultivars were not significant, their averages were presented and discussed.
Multivariate analysis was performed via principal component analysis (PCA) to verify the grouping of the different responses to K fertilizer management and the use of ruzigrass in the second year (2017/2018). Considering that the measurement units differed between variables, the data were log-transformed to reduce the effect of the numeric scale (McGarigal et al. 2000). The ordination graphic (with two major components) was demarcated by two axes designated as the first (PC1) and second (PC2) principal components.