Morphological and biochemical variations induced by synergy of salicylic acid and zinc in cockscomb
, , und | 06. Apr. 2021
Article Category: Original Article
Online veröffentlicht: 06. Apr. 2021
Seitenbereich: 79 - 90
Eingereicht: 24. Dez. 2020
Akzeptiert: 01. März 2021
DOI: https://doi.org/10.2478/fhort-2021-0006
Schlüsselwörter
© 2021 Amna Shoaib et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.
The role of zinc (Zn) as a micronutrient for human and plant nutrition is well established. In plant, Zn helps in preventing lodging, better growth, biomass production and high quality of cut-flowers (Shaheen et al., 2015; Khan et al., 2018; Bhatt et al., 2020). Zn participates in all major functions of the plant by acting as a co-factor or structural element in 300 catalytic and non-catalytic proteins and pigment biosynthesis (Pérez et al., 2020). It is involved in stomatal regulation, membrane stability, ion balance, cell wall development, metabolism of carbohydrates, lipids and proteins as well as synthesis of phytohormones (auxins, abscisic acid, gibberellins and cytokinins) (Cabot et al., 2019). Being an essential component of several biomolecules, Zn regulates intracellular signalling pathways and relevant physiological responses in plants by eliminating oxidative stress (Shaheen et al., 2015), by affecting the activity of enzymes, such as catalase (CAT), peroxidase (POX) and polyphenol oxidase (PPO) (Khan et al., 2018). These enzymes are essential for plant growth and development throughout life cycle. For example, the CAT enzyme is integral part of the plant's antioxidative system, contributes in plant defence, aging and senescence (Yang and Poovaiah, 2002). POX enzyme participates in lignification, suberization and cross-linking hydroxyproline-rich proteins (extensins) in the cell wall matrix as well as in control function of redox state in apoplast (Luhová et al., 2006). PPO is nuclear-encoded enzyme involved in pigment biosynthesis, acts against stress and catalyses phenol oxidation (Mastuti et al., 2015). Soil Zn application has been documented already and it stated that Zn application has improved the growth and quality of oriental Lilly by maintaining the activity of antioxidant enzymes and enhancing membrane stability (CAT and POX) (Shaheen et al., 2015). Kolade et al. (2018) findings indicated that Zn fertilisation (50 mg · kg−1) in
Apart from Zn, salicylic acid (SA) contributes a lot in the growth, by acting as an endogenous growth regulator of phenolic nature produced by root cells, by modifying plant performance by modulating key metabolic and physiological processes (Dempsey et al., 2017) that can’t be overlooked. It has been described that SA plays an important role in photosynthesis performance by increasing the stomatal conductance levels, transpiration rates and enzyme activity related to CO2 uptake at the chloroplast level (Janda et al., 2014). Increase in the rate of carbohydrate metabolism of plants lead to more efficient metabolic responses through increased content of soluble sugars and protein (Hasanuzzaman et al., 2017). SA regulates ion uptake by roots, aids in synthesis of kinase protein, which regulate cell division and differentiation, initiate root elongation, stimulate leaves in young shoot and bud behaviours. Hence, it can be concluded that SA positively affects the growth rate and plant productivity and induce flowering in various plant species (Dempsey et al., 2017). Further, SA strengthens vascular cells, delay senescence by regulating the plant water and activity of antioxidants (CAT, POX, etc.) against oxidants generated during senesces (Alaey et al., 2011; Khan et al., 2020). However, the plant responses to exogenous SA vary with species, SA concentrations and mode of application (Miura and Tada, 2014). Pacheco et al. (2013) reported the high flavonoid content and higher biomass production in marigold plants after the consecutive 3 days foliar application of SA (60–140 ppm) before the reproductive stage. Abbas and Ibrahim (2014) suggested spraying of 50 ppm of SA for increasing the growth, yield, and oil ratio indices in black bean. SA (100 ppm) positively and significantly influences number of leaves, spikes and florets spike by accelerating the cell divisions in the apical portion of the sprouts in gladiolus (Pal and Kumar, 2015). The highest rate of vegetative, reproductive and nutrient attributes was obtained due to spraying of 200 ppm SA in thorn apple (Al-Mohammadi and Al-Rawim, 2016) and 100 ppm in Zinnia (Zeb et al., 2017). Marigold received spray of SA (80 ppm and 120 ppm) on aerial parts after 60 days of sowing and exhibited maximum number of leaves, plant height, number of inflorescence, stem diameter, fresh and dry weight of flowers in marigold (Basit et al., 2018). Choudhary et al. (2016) studies revealed that synergism of foliar spraying with Zn (1.0%) and SA (1.0 Mm · L−1) positively increased the growth and flowering parameters in marigold. Ahmad et al. (2018) recommend SA (2 mM) application as foliar at 5th week to achieve the best growth and production of tuberose flowers and bulbs.
Only few literatures are available on physiological responses in ornamental plants after Zn or SA application, but foliar SA and Zn could be used as sustainable approach to ameliorate growth rate, physiological responses and productivity in
The experiment of 12 treatments was conducted in a randomised complete block design having factorial arrangement with replications of three times per treatment. One treatment consisted of 15 plants, as there were 5 plants in each replicate. In the first factor, different concentrations of Zn were applied, while in second factor different concentrations of SA were given and further combinations of Zn and SA were applied under the third factor (Table 1).
Layout of the experiment.
| Factors | Zn (0.0 ppm) | Zn (0.5 ppm) | Zn (1.5 ppm) | Zn (2.0 ppm) |
|---|---|---|---|---|
| SA (0.0 ppm) | T1 | T2 | T3 | T4 |
| SA (50 ppm) | T5 | T6 | T7 | T8 |
| SA (100 ppm) | T9 | T10 | T11 | T12 |
SA, salicylic acid; Zn, zinc.
Field experiment was carried out at the experimental area of 850 sq. ft at Institute of Agricultural Sciences, University of the Punjab, Lahore (31°32′ N latitude and 74°20′ E longitudes), during June–October, 2019. The average temperature during this period was 38 °C, relative humidity was 55% and average rainfall was 48.2 mm.
Total experimental area was 7,600 cm2. To remove hard pans and other plants material, the land was mechanically tilled about 10 cm deep with cultivar and left opened for 3 days for sun solarisation. After ploughing the field, soil was fumigated with 20% formalin solution. Cotton plugs were dipped in a formalin solution and then placed at different positions in soil to disinfect soil. After this, the soil was covered with polythene sheet for 3 weeks. After 21 days, the polythene sheet was removed and soil was mixed well. Then it was left open to remove the formalin residues. Before dividing the field into sub-plots for replicates, the field was irrigated manually. Total experimental area was divided into 36 plots each measuring 120 cm × 120 cm · plot−1. Farm yard manure (FYM) was mixed manually at dose of 1 kg · plot−1. Experimental unit was irrigated as per required at all critical stages of crop. Weeding and hoeing operations were completed according to needs.
Healthy seeds of
Concentration of 0.5 ppm, 1.5 ppm and 2.0 ppm were prepared by dissolving 0.015 g, 0.045 g and 0.06 g of ZnSO4 (Merck) in 3 L of distilled water, respectively. SA (2-hydroxybenzoic acid, Merck) was initially dissolved in 1,000 μL dimethyl sulfoxide to make a volume up to 100 mL, and later 50 ppm and 100 ppm concentration were prepared from the standard solution.
Biochemical attributes like total chlorophyll contents (TCCs), carotenoids (CC), total protein contents, CAT, POX and PPO activities were evaluated following the protocols of Nafisa et al. (2020) from the triplicate samples of leaves collected from each replicates at 60 DST.
TCC and carotenoid contents were determined in ethanolic mixture of leaf extract, obtained after homogenising 0.1 leaf sample in 80% ethanol. Absorbance of supernatant was determined at 470 nm, 645 nm and 663 nm by UV spectrophotometer.
For the estimation of total protein content, leaf sample (0.5 g) was crushed in phosphate buffer (0.1M, pH 7.5) in pre-chilled pestle and mortar in liquid nitrogen followed by centrifugation at 3,000 rpm for 10 min and addition of 1 mL of reagent C [prepared by mixing reagent A (0.2% NaOH and 2% Na2CO3) and reagent B (0.5% CuSO4 in 1% of KNaC4H4O6·4H2O) in 50:1 ratio]. The mixture was shaken for 10 min, then 0.1 mL Folin phenol reagent was added and finally incubated at room temperature for 30 min. The total protein content was estimated at 650 nm by using the standard curve of Bovine serum albumin.
To assess the activity of antioxidant enzymes, 0.1 g of leaf was ground in 0.1 M phosphate buffer, centrifuged at 3,000 rpm for 10 min, and supernatant was used for enzymes activities assays. For CAT, a mixture containing 50 mM phosphate buffer (pH 7.0), 0.3% H2O2 and 100 μL enzyme extract was assessed for absorbance at 240 nm at the intervals of 30 s. POX activity was determined in the reaction mixture contained 2 mL of 0.1 M phosphate buffer (pH 6.8), 1 mL of 0.01 M pyrogallol, 1 mL of 0.05 M H2O2 and 0.5 mL of enzyme extract. The solution was incubated for 5 min at 25°C after which the reaction was terminated with addition of 1 mL of 2.5N H2SO4. The amount of purpurogallin formed was determined by measuring the absorbance at 420 nm. For PPO, the reaction mixture consisted 100 μL enzyme extract and 1.5 mL of 0.1 M sodium phosphate buffer (pH 7.0), the reaction started when 200 μL of 0.01 M catechol was added. The changes in the absorbance were recorded at 30 s intervals for 3 min at 495 nm.
Reproductive indices including number of buds per plant and fresh weight of flower were recorded at 65 and 105 DST, respectively, while growth indices like stem diameter, length, fresh and dry weight of root and shoot were taken at 120 DST.
The statistical analyses were conducted using statistical software Statistics 8.1 at
Table 1 shows various treatments, where three different concentration of Zn and two different concentrations of SA were applied on the plants separately and in combination. A statistically significant difference (
Effect of foliar application of SA and Znon the vegetative and reproductive growth indices in
| DST | Parameters | SA level (ppm) | Zn level (ppm) |
||||
|---|---|---|---|---|---|---|---|
| 0 | 0.5 | 1.5 | 2.5 | Mean | |||
| 120 DST | Stem diameter (mm) | 0 | 23 e | 35 b–d | 38 a–c | 31 b–d | 33 B |
| 50 | 30 c–e | 26 de | 28 de | 31 de | 29 C | ||
| 100 | 34 b–d | 35 b–d | 42 ab | 47 a | 40 A | ||
| Mean | 29 C | 32 BC | 36 AB | 37 A | |||
| Shoot length (cm) | 0 | 29 c | 40 ab | 43 ab | 39 ab | 37 B | |
| 50 | 35 bc | 35 bc | 36 bc | 37 bc | 36 AB | ||
| 100 | 38 bc | 41 ab | 45 a | 46 a | 43 A | ||
| Mean | 34 B | 39 A | 41 A | 42 A | |||
| Shoot fresh weight (g) | 0 | 458 d | 883 ab | 936 ab | 803 bc | 770 B | |
| 50 | 673 d | 536 d | 563 d | 646 cd | 576 C | ||
| 100 | 899 ab | 770 bc | 897 ab | 1023 a | 910 A | ||
| Mean | 638 C | 730 B | 815 AB | 824 A | |||
| Shoot dry weight (g) | 0 | 186 f | 433 cd | 475 bc | 430 cd | 381 B | |
| 50 | 293 ef | 296 e | 320 e | 333 de | 311 C | ||
| 100 | 430 cd | 453 c | 566 b | 687 a | 534 A | ||
| Mean | 304 C | 394 B | 453 A | 483 A | |||
| Root length (cm) | 0 | 19 c | 25 a–c | 26 a–c | 24 a–c | 23 B | |
| 50 | 21 bc | 21 bc | 22 a–c | 22 a–c | 21 B | ||
| 100 | 25 a–c | 26 a–c | 27 ab | 30 a | 27 A | ||
| Mean | 22 A | 24 A | 25 A | 25 A | |||
| Root fresh weight (g) | 0 | 57 c | 77 a–c | 82 a–c | 75 a–c | 72 AB | |
| 50 | 62 bc | 64 bc | 66 a–c | 70 a–c | 66 B | ||
| 100 | 70 a–c | 70 a–c | 85 ab | 93 a | 81 A | ||
| Mean | 62 B | 72 AB | 78 A | 79 A | |||
| Root dry weight (g) | 0 | 37 d | 54 b–d | 59 a–c | 56 a–c | 52 B | |
| 50 | 48 cd | 46 cd | 47 cd | 50 cd | 48 B | ||
| 100 | 59 a–c | 60 a–c | 69 ab | 74 a | 65 A | ||
| Mean | 48 B | 53 AB | 59 A | 58 A | |||
| 65 DST | Number of buds · plant−1 | 0 | 47 e | 61 b–e | 68 bc | 64 b–d | 60 B |
| 50 | 54 c–e | 52 de | 55 c–e | 57 c–e | 54 B | ||
| 100 | 65 b–d | 60 b–e | 73 ab | 80 a | 70 A | ||
| Mean | 55 B | 58 B | 65 A | 68 A | |||
| 105 DST | Flower weight (g) | 0 | 5.83 f | 8.33 c–e | 9.6 bc | 9.3 b–d | 8.25 B |
| 50 | 7.40 ef | 7.41 d–f | 7.46 d–f | 7.83 c–f | 7.52 C | ||
| 100 | 8.56 c–e | 8.51 c–e | 10.50 b | 12.96 a | 10.12 A | ||
| Mean | 7.25 C | 8.06 C | 9.18 B | 10.22 A | |||
Values with same lower case (rows) and upper case (rows and column) show insignificant difference (
DST, days after seedling transplanting; SA, salicylic acid; Zn, zinc.
Root lengths and fresh weights were increased significantly by 30–40% and 50–70%, respectively in T12 followed by T11 as compared with control. However, for the treatments provided with 1.5 ppm and 2.5 ppm of Zn, higher level of SA and their combinations were found optimum for significantly improving the dry biomass by 50–100%. Furthermore, value of uppercase alphabets in rows and column revealed that Zn and SA were optimum for root-related attributes only at their higher levels (Table 2).
Just like vegetative growth assays, mean effect of SA and Zn significantly increased reproductive growth indices at their higher levels. Moreover, interaction of SA + Zn levels boosted the said attributes at higher level of SA (100 ppm) and Zn (2.5 ppm) as well. Lower level of SA (50 ppm) either used separately or with Zn did not improve reproductive indices significantly. Hence, number of buds per plant was significantly less (47) in control, while the highest number (83) was recorded in the plants treated with SA (100 ppm) + Zn (2.5 ppm) followed by 72 buds per plant with SA (100 ppm) + Zn (1.5 ppm). Other treatments that include higher levels of Zn (1.5 and 2.5 ppm), SA (100 ppm) or their combinations exhibited less significant effect in improving number of buds (40%) with respect to the control treatment (Table 2).
Significantly greater flower weight was recorded in T12 followed by T11, which were provided with combined application of higher levels of SA + Zn, and it is observed that flower weight increased significantly by 123% and 80%, respectively over control (5.83 g). Application of 1.5, 2.5 and 0.5 ppm of Zn improved the flower weight significantly by 65, 40 and 43%, respectively, and these treatments were statistically similar. Likewise, the application of SA alone (100 ppm) and in combination with 0.5 ppm of Zn displayed statistically similar increase of 50% in flower weight with respect to control (Table 2).
In the control treatments, the TCC and CC of fresh weight of leaf were 2.52 and 0.44 mg · g−1, respectively. These attributes were significantly improved by 30–50% in the all treatments that are provided with Zn (0.5, 1.5 and 2.5 ppm) and at higher level of SA (100 ppm SA) either separately or in combination. However, lower level of SA (50 ppm) and its combination with Zn levels did not affect the said attributes significantly as compared to the control (Figure 1A and B). Total protein content, as well as activities of POX and PPO were generally enhanced significantly by 20–30% due to applications of different concentrations of Zn or SA applied alone or simultaneously as compared to control (0.43 mg · g−1, 50.14 and 23.59 U · min−1 · mg−1 protein, respectively) (Figure 1C, E and F). The activity of CAT was significantly greater (6.67 U · min−1 · mg−1 protein) in leaves of control treatments as compared to rest of the treatments. The influence of foliar application of Zn or SA alone or in synergism significantly reduced CAT activity by 41–65%. The highest reduction of 65% in the CAT activity was recorded in T3 and T12. The other Zn (0.5 and 2.5 ppm) and SA (100 ppm) levels, when applied alone and in bilateral combination statistically presented similar or same reduction of 51–55% in CAT activity as compared to control. Low dose of SA (50 ppm) also showed significant reduction of 43% when applied separately, while reduction in the CAT activity increased from 43% to 48% when SA (50 ppm) was given with different concentrations of Zn as compared to control (Figure 1D).
Figure 1
(A–F) Effects of foliar application SA and Zn and the physiological attributes in
The PCA score-plot accounted for 89.93% of the total variance and shows a degree of separation among the treatments. The placement of T1 along the negative values of PC1 was due to low measurements of the remaining variables of growth, reproduction and physiology. However, all variables increased with foliar application of SA and Zn as indicated by placement of the remaining treatments. T5–T8, placement along the PC2 revealed a lower value of all variables as compared to other treatments. Moreover, T2–T4, T9 and T10 exhibited intermediate to high values all the traits along the PC1 component. However, the placement of T11 and T12 just opposite to T1 and along the positive side of PC1 exhibited the maximum values of the attributes. The location of morpho-physiological traits on the positive side of PC1 indicated their positive association with each other and with the treatments (T2–T12), while CAT exhibited rather different trend. CAT activity was maximum in T1, and minimum in other treatments (Figure 2).
Figure 2
PCA due to effect of foliar application of SA and Zn on morpho-physiological attributes in
The aggregated data heat-map analysis identified two main clusters. Cluster I corresponding to all treatments (T2, T3, T4, T9, T10, T11 and T12) with higher values of the investigated attributes, whereas T11 and T12 were in same sub-cluster, while T2, T3, T4, T9 and T10 in other sub-cluster. However, cluster II separated T1 from T5, T6, T7 and T8 in sub-clusters based on the difference of morpho-physiological traits (Figure 3).
Figure 3
Heat map visualisation of variables and the treatments due to effect of foliar application of SA and Zn on
In the present study, Zn-fertilised plants (T2–T4) were taller and had higher growth and reproductive indices as compared to the control (T1). The greater benefit was obtained with 1.5 ppm (T3) and 2.5 ppm (T4), where the plants’ height, biomass, stem diameter, number buds and flower weight improved significantly up to 1.5–2.5 folds possibly owing to optimal metabolism obtained by the application of required Zn to plants (García-López et al., 2019) and due to the balancing level of macro and micronutrients (Rahman et al., 2018). The current results were similar to the findings of many previous studies, where the foliar application of Zn improved plant height in flowering plants (Shah et al., 2016; Rehman et al., 2018) because of its potential to induce auxin synthesis, cell division, function in photosynthesis and carbohydrate metabolism (Khan et al., 2018; Awan et al., 2019; Shoaib et al., 2020). The improvements in vegetative growth including stem diameter likely to be responsible for more accumulation of photosynthetic materials and as recorded in current study, photosynthetic pigments improved by 30–50% with application of different Zn levels (0.5–2.5 ppm). Essentiality of Zn in metabolic rate of plant is confirmed, therefore, it is assumed that Zn by facilitating CO2 diffusion to the carboxylation sites in plants through carbonic anhydrase improved chlorophyll content, detoxified reactive oxygen species (ROS) and maintained rubisco structure (García-López et al., 2019). Therefore, observed improvement in plant growth by spraying of plants with 0.5–2.5 ppm of Zn was due to the increase in chlorophyll content, since chlorophyll content is a bioindicator of the photosynthetic efficiency and one of the most important determinants of plant growth. Reports suggested that Zn stimulate bud-out growth by inducing the biosynthesis of hormones involved in the process (Zhu and Kranz, 2012). Moreover increase in number of bud might be associated with the development of adequate storage pool of Zn in stem nodes to redirect Zn effectively to the developing buds for subsequent bud-break and development of new plant organs (Xie et al., 2020).
Foliar application of SA separately was not effective at 50 ppm (T5), while at 100 ppm (T9), the investigated attributes were significantly greater and statistically at part with T2–T4. The present piece of work derives support from the work of earlier researchers, who reported increased growth, physiological and yield responses by exogenous application of SA at higher concentrations (100 and150 ppm) than at the lower ones (Ali and Mahmoud, 2013; Azoz and El-Taher, 2018). Improvement in TCC and CC could be attributed to role of SA in promoting reactions involving photosynthetic pigments, membrane integrity and Photosystem II activity, which may lead to better plant biomass and area (Nazar et al., 2015). Arfan et al. (2007) reported enhanced allocation of N and S to rubisco protein through SA application, and so increase in the availability of CO2 for rubisco protein (key enzyme in photosynthesis catalysing CO2 fixation) might be the reason for positive effect on plant growth and development. Furthermore, phenolic nature and harmonic behaviour of SA has been assigned to play an important role in stimulation in formation of bud and flowers (Zeb et al., 2017).
The combined SA or Zn foliar application significantly enhanced plant vegetative and reproductive growth indices at their higher levels (T10–T12). The synergy of SA (100 ppm) with the highest applied dose of Zn (2.5 ppm) in T12 had resulted in the greatest vegetative and reproductive traits. El-Yazied (2011) recommended that foliar spraying with SA (100 ppm) + Zn (50 ppm) for better nutrient constituentsas well as for better yield and better fruit quality in
A certain amount of ROS is always produced during the whole life of each aerobic organism during stress and development, while its levels are tightly controlled by antioxidant machinery comprising enzymes (e.g. CAT, POX and PPO) (Nafisa et al., 2020). Presently CAT activity was found to be 42–65% lower after Zn, SA and SA + Zn application than control group. The reduction in the CAT activity was greater with higher doses of Zn and with combined application of SA (100 ppm) + Zn, which may indicate good state of plant towards flower production as compared to untreated plants. Alteration in the antioxidant system has been confirmed after application of SA or Zn in different plants through inhibition in the CAT activity and stimulation in POX activity to maintain redox status of the cell (Rao et al., 1997; Khan et al., 2018). Moreover, total protein content, activity of POX and PPO increased by 20–30% with all treatments, might show an active metabolic pool in the leaves because of high photosynthetic activity. Furthermore, high POX and PPO also revealed accumulation of phenolics, lignin and other defence-related secondary compounds in plant (Awan et al., 2019). POX activity accelerates lignin accumulation in cell wall, which not only increases mechanical strength of plant stalk, but also contributes in plant growth, tissue/organ development, resistance against lodging and variety of biotic and abiotic stresses (Pandey et al., 2017). There have been many suggestions regarding influence of PPOs on photosynthesis including its functioning as oxygen buffer reducing lipid peroxidation apart from its role in oxidation of phenols (Vaughn and Duke, 1984).
PCA and heat map are the most frequently utilised multidimensional method to classify treatment, and to identify the key variables and their pattern of correlations (Han et al., 2019; Siddique et al., 2020; Shariatipour et al., 2020). Differences between the treatments are easily evaluated by visualising the distances between the points in biplot, as a greater distance reflects a greater difference and vice versa. PCA and heat map analyses revealed T12 (100 ppm SA + 2.5 ppm Zn) followed by T11 (100 ppm SA + 1.5 ppm Zn) as the optimum treatments for the better growth and reproduction of cockscomb plant. However T2–T4 and T9 exhibited intermediate to high values of the studied traits, while T5–T8 had least influence on the attributes studied. Moreover, note that weak correlation is shown by perpendicular angle between two variables, and high positive/high negative correlation is indicated by nearly parallel (antiparallel) angle between two variables (Han et al., 2019). Generally, all morpho-physiological traits excluding CAT were on the positive side of PCA, and were positively correlated with each other. However, growth attributes (length, biomass, number of buds per plant and flower weight) were clustered very closely (cluster 1: right lower side of biplot), which indicated very strong and positive interrelationship among them. The relationship among total TCC, CC and PPO (cluster 2: upper right side of biplot) was also strong, and these clustered closer to growth attributes as well. Clusters 1 and 2 were moderately and positively correlated with cluster 3 (total protein content and POX). The results showed that greater photosynthetic pigment could modify dynamics of key carbohydrate metabolism in both source to sink organ, resulting in better growth and reproductive indices. Important factors such as stronger stem, bud number and better flower density significantly contributed to greater growth efficiency and better stress resistance in
Morpho-physiological traits of cockscomb are changed by varying the concentrations of SA and Zn sprayed either alone or in synergism. Bilateral combination of SA (100 ppm) and Zn (2.5 ppm) exhibited the highest growth attributes and flower weight. PCA-based biplot and heat map visualisation revealed the positive correlations between all treatments, and majority of the morpho-physiological characteristics of cockscomb are studied.