Assessment of Growth and Yield Potential of Different Vegetable Amaranth Type In

Assessment of growth and yield potential of different vegetable amaranth type In

Kleve, Germany

BY Tamanna Tanzin

Matriculation Number: 11520

A Thesis Submitted to the Faculty of Life Sciences, Hochschule Rhein-Waal University of Applied Sciences in Partial Fulfilment of the Requirements for the Degree of Bachelor of Sciences In Sustainable Agriculture

Supervisor Co-supervisor Dr. Katja Kehlenbeck Prof. Dr. Florian Wichern

Kleve, Germany March 2018 Table of Contents

Abstract ...... 5 1. Introduction ...... 6 1.1 Botanical Description ...... 7 1.2 Cultivation and Management of Amaranths ...... 8 1.3 Health Benefits of Amaranths ...... 9 1.4 Justification and Aim of the Study ...... 10 2. Materials and Methods ...... 11 2.1 Field Experiment ...... 11 2.1.1. Location ...... 11 2.1.2 Research Design ...... 12 2.1.3 Variety Selection and Description ...... 14 2.1.4 Management and Cultivation ...... 16 2.1.5 Harvest ...... 16 2.2 Seed Germination Test ...... 18 2.3. Data Collection ...... 19 2.3.1 Continuous Measurements of Field experiment ...... 19 2.3.2 Destructive Measurements of Field Experiment ...... 19 2.3.3 Seed Germination Test ...... 20 2.4 Statistical Analysis ...... 21 3. Results ...... 22 3.1 Field Experiment ...... 22 3.1.1 Plant Height ...... 22 3.1.2 Leaf ...... 25 3.1.3 Stem ...... 29 3.1.4 Root ...... 32 3.1.5 Total Weight of 20 Selected Plants of Each Variety ...... 35 3.1.6 Total Weight of the Fresh Above- Ground Biomass per Plot ...... 36 3.1.7 Dry weight of Leaves, Stems and Roots of 20 Selected Plants of Each Variety ...... 37 3.1.8 Leaf-Stem Ratio ...... 38 3.2 Correlation between Different Variables of Field Experiment ...... 39 3.3 Seed Germination Test ...... 41 4. Discussion ...... 42 5. Conclusions and Recommendations ...... 46 Acknowledgements ...... 48 DECLARATION OF AUTHENTICITY ...... 49 References ...... 50 Appendices ...... 53

Table of Figures

Figure 1: Average temperature and precipitation in Kleve over the year (Meteoblue, 2017) ...... 12 Figure 2: A randomized block design was used for field experiment (A). Here, S1-L8 shows the variety ID of eight selected variety used in this experiment and 'A', 'B' beside the variety ID indicates two replication of each variety. Each sub-plot had four rows in total with 17 cm gap in between rows (B) .... 13 Figure 3: 40cmx40cm frame used to select the core plants of each plot (A), harvested plants of 'Surinamese Spinach' (L7) (B) and total harvested plant of Marog Green (L8) plot after washing the roots (C) ...... 17 Figure 4: The seeds were washed i H₂O₂ ixture ad i distilled water to avoid fungal diseases (A). Later the seeds were placed in petri dishes where each petri dish contains ten seeds of each variety (B). Germinated seeds of leafy variety 'Laal Shak' (L5) at 20 C after three days of placing the seeds in climate cabinet (C) ...... 18 Figure 5: Mean plant height of 20 amaranths plants of each eight different varieties sorted by variety type (stem varieties on top, leafy varieties at the bottom) ...... 23 Figure 6: Boxplot of final mean height (cm) of 20 amaranths plants of each selected varieties sorted by variety type (leafy is coded as L and harvested six weeks after sowing, stem is coded as S and harvested eight weeks after sowing). The H-test were performed separately of each for the four stem and four leafy varieties...... 24 Figure 7: Mean number of leaves of 20 amaranths plants each of eight different varieties sorted by variety type (stem varieties on top, leafy varieties at the bottom) ...... 26 Figure 8: Boxplot of final mean of number of leaves per plant (A) and mean leaf weight in g (B) of 20 amaranths plants of each selected variety sorted by variety type. Here leafy variety is coded as L and harvested six weeks after sowing and stem variety is coded as S and harvested after eight weeks of sowing. The H-Test were performed separately each for the four stem and four leafy varieties...... 27 Figure 9: Boxplot of mean Leaf area (cm²) (A) and mean percentage of leaves from total weight of 20 amaranths plants of each selected varieties sorted by variety type. Here leafy variety is coded as L and

2 harvested six weeks after sowing, stem variety is coded as S and harvested after eight weeks of sowing. The H-tests were performed separately of each for four stem and four leafy varieties...... 28 Figure 10: Boxplot of final mean stem weight in g (A) and stem diameter in mm (B) of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties...... 30 Figure 11: Boxplot of mean percentage of stem from total weight of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties...... 31 Figure 12: Boxplot of mean root length in cm (A) and mean root weigh in g (B) of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties...... 33 Figure 13: Boxplot of mean percentage of roots from total weight of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties...... 34 Figure 14: Boxplot of mean total weight in g of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties ...... 35 Figure 15: Mean dry weight of leaves, stem and roots of 20 amaranths plants of eight selected variety. Here, leafy variety is coded as L and harvested six weeks after sowing and stem variety is coded as S and harvested after eight weeks of sowing and DWL= Dry weight of leaves, DSW= Dry weight of stem and DWR = Dry weight of root. The H-Test were performed separately each for four stem and four leafy varieties...... 37 Figure 16: Boxplot of leaf-stem ratio of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties...... 38 Figure 17: Percentage of germination rate of eight selected varieties at different temperature. Here, stem variety is coded as S and leafy variety is coded as L ...... 41

List of Tables

Table 1: Nutrient elements available in the leaves of five Amaranthus species on the basis of 100g fresh edible weight (Yang & Keding, 2009) ...... 9 Table 2: Species name, origin, variety name, variety Id, variety type, leaf color, stem color and seed color of mostly used-as vegetable of the eight selected variety of Amaranths used in this study...... 15 Table 3: Mean leaf weight (g), stem weight (g), total yield per plot (g/m²) and average total yield per plot in t/ha of eight selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties ...... 36 Table 4: Spearman's Rho correlation showing relationships among yield parameter of Stem varieties ...... 40 Table 5: Spearman's Rho correlation showing relationships among yield parameter of leafy varieties ...... 40

Apendices

Appendix 1:...... 53 Appendix 2:...... 54 Appendix 3:...... 54 Appendix 4:...... 55 Appendix 5:...... 55 Appendix 6:...... 56 Appendix 7:...... 56

Abstract

Amaranthus belongs to the Amaranthaceae family which has approximately 70 species. It is considered as an underutilized plant but some research in the recent years showed that it is a promising plant for future nutrition supply. Amaranths are widely known for its nutritional value and their easy growing habit. This underutilized plant can become a good option for the commercial growers because it is easy to grow, it has high nutritional value and low maintenance cost. Many species of the Amaranthus genus are known as annual weeds, some are used as a vegetable and ornamental plants, some are known as pseudocereals which are mainly used for their grain. The core objective of this study was to assess the growth and yield potential of different vegetable amaranths to select a suitable variety for Kleve, Germany. Eight vegetable amaranths variety (Leafy variety and Stem variety) were grown in the experimental garden of Hochschule Rhein-Waal University. A seed germination test was also conducted at the life science lab to observe the best seed germination rate at different temperatures. Plant height, number of leaves per plant were measured continuously after two weeks of sowing till the harvest week. The fresh and dry weight of leaf, stem and root were measured after final harvest. The statistical analysis of growth and yield performance of stem and leafy varieties were done separately. Leafy varieties showed a high variation in yield and germination rate than stem varieties. Leafy variety Surinamese Spinach (L7) and Marog Green (L8) excelled in all yield parameters whereas Chinese Spinach (L6) was consistent in low growth and yield. Stem variety Danta Bhutan (S2) and Danta Lusina (S3) showed low yield performance. All the varieties showed a high germination rate above 20 C in seed germination test.

1. Introduction

Amaranthus is widely known as amaranth, a short-lived annual plant (National Research Council, 2006). Approximately 60-70 Amaranthus species have recognized so far (Das, 2016). Most of them were known as a weed at the beginning. Some species are still considered as a weed but it has become popular as leafy vegetables, cereal and ornamental plants (Truco & Tranel, 2011). Vegetable amaranths is one of the most popular vegetables in Africa and Asian country for their vitamin, protein and mineral-rich leaves and stems (Das, 2016). According to Das (2016), some popular species of vegetable amaranths are Amaranthus tricolor, Amaranthus blitum, Amaranthus Dubius, Amaranthus cruentus and Amaranthus viridis.

Amaranth is an ancient plant. The domestication of Amaranthus is not determined fully but according to Sauer (1993), the domestication of Amaranthus as a grain crop comes from archeological digs at a cave in Tehaucan, Puebla, Mexico. The seeds of Amaranthus cruentus were dated 6000 years old though he thinks that it first domestication could have appeared earlier in a different location. The known history of Amaranthus cultivation begins with the Aztec civilization of central Mexico. In 1959s, after the arrival of Spanish, they suppressed the Aztec custom and as a result, the use and production of amaranths had been reduced. Grain Amaranths production and use have continued scatteringly in Mexico and Central and South America at least 50 years after the arrival of Spanish. After Spanish invasion, Amaranthus also spread to Europe, Africa and Asia. The seed of amaranth has been used in Central and Latin America since ancient times (Brenner et al. 2010).

According to Early (1992), the seeds were named as Huautli by Aztec and they used to use the seeds in beverage, in sauce, in special kind of tortilla and for various medicinal things. According to National research council (1984), Amranthus tricolor and Amaranthus dubius are famous in India, China, Southeast Asia and South Pacific islands.

1.1 Botanical Description

Amaranthus belongs to the Amaranthaceae family consisting 60-70 species (Das, 2016). Amaranths are easy to grow. Vegetable Amaranth can grow best in hot, humid weather (National Research Council, 1984). Most of the Amaranthus varieties germinate at 20 C or above soil temperature (Myers, 1996). Whitehead et al. (2002) found that, 25 C or higher soil temperature and 28 C- 30 C air temperature are needed for the seed germination. Amaranthus is C4 plant and grows under drought stress, can tolerate unfavorable abiotic conditions including high salinity, acidity or alkalinity which makes this plant uniquely suitable for subsistence farming (Maughan et al. 2011). Amaranth can grow in every type of soil but the well-drained fertile soil is more suitable for a good yield. Soil with high organic matter is ideal for rapid and high yield. The seeds need good fertile soil and adequate moisture to germinate (Amaranthus-Production guideline, 2010). The soil with PH >6 is ideal for high growth and yield (Grubben, 2004a). Amaranth is a broad-leafed annual plant. Mature plant height varies between 0.3m and 2m (van Rensburg et al. 2007) and plants can vary from branched to unbranched, dwarf to tall depending on species, growing condition and growth habit. The amaranth species is an erect or spreading annual with different leaf sizes. Some species have a particular marking on their leaves. Terminal and auxiliary inflorescences can be seen while flowering. The seeds are really small, the colour of the seeds varies from dark brown to shiny black. Some varieties have cream colored seed also (National Research Council, 2006). Amaranth is a widely distributed plant and the new promising food for future because of its easy growing habit and the nutritional value of both grain and leaves (Wu et al. 2000). But the research conducting in improving world food supplies pay little attention to this species. Even though as a neglected plant, the vegetable amaranths is cultivated almost 50 tropical countries (National Research Council, 2006). It is considered that in Africa and Asia, amaranths is one of the most widely eaten vegetables. Amaranths have a high capacity of osmotic adjustment (Liu & Stützel, 2002) and a C4 photosynthetic pathway allows efficient use of CO2 in a large range of temperature and moisture stress environment which consider a major factor in their wide geographical distribution (Stallknecht & Schulz-Schaeffer, 1993). The palatability of amaranths

7 has been rated as similar to spinach in a taste test (Campbell & Abbott, 1982). In Africa and Southeast Asia, Chinese spinach and Amaranthus tricolor are famous as a quality vegetable (National Research Council, 2006). Some varieties are also cultivated and famous in Mexico, Central and South America, in some Carribean areas (Brenner et al. 2010). The plant is very famous and widely used in Taiwan, Thailand, China, Indonesia (National Research Council, 2006).

1.2 Cultivation and Management of Amaranths

The seeds are really small. They need good fine, firm seedbed to grow. Fertile, well-drained, loose soil with high organic matter is the best for high and fast yield (Grubben & Denton, 2004). Adequate soil moisture content is necessary for seed germination and emergence (National Research Council, 2006). Soil PH>6 is necessary for good growth of the plants (Grubben, 2004a). Seeds are often broadcasted or sown in rows with 2–5g/m² seed rate with at least 20cm gap between rows (Achigan-Dako et al. 2014). According to Amaranths production guideline published by agriculture, fishery and forestry department of South Africa, one common disease of amaranth is damping –off fungus which is caused by high soil water content and low soil moisture content. This can also occur for dense planting and insufficient soil aeration. To avoid this problem, using disease-free clean seeds, avoiding over watering and dense planting bed can be helpful. Leaf amaranth often affected by armyworms and curly top viruses. Most of the amaranth varieties are ready for harvesting after four to five weeks of sowing (Achigan-Dako et al. 2014). Most common harvesting method is to pull out the complete plant with roots (Ebert et al. 2011). Plants can be harvested by cutting the main stem also. The main shoot is harvested and side shoots are left to grow in multiple harvests (Achigan-Dako et al. 2014).

1.3 Health Benefits of Amaranths

Amaranthus is rich in vitamins like β-carotene, vitamin B6, vitamin C, riboflavin. They also rich in essential amino acids and dietary minerals like Ca, P, Fe, Mg, K, Cu, Zn, and Mn (Mampholo et al. 2015 ; Venskutonis & Kraujalis, 2013). In Table 1, the nutrient contents of five species of Amaranthus is shown where the difference of nutrient content among the species is listed. 90% vitamin C, 73% vitamin A, 28% calcium and 28% iron of daily nutrient requirement can be obtained from one cup of cooked, boiled and drained amaranth leaves (Achigan-Dako et al. 2014). The daily nutrients requirements for an adult in different region is presented in Appendix 3. An adult in the different region can easily meet their daily requirements of Vitamin A from 100g fresh edible Amaranthus leaves. Other nutrients amount is also very high regarding to daily nutrient intake recommendation for an adult.

Table 1: Nutrient elements available in the leaves of five Amaranthus species on the basis of 100g fresh edible weight (Yang & Keding, 2009)

Species Protein (g) Vitamin A Vitamin C (mg) Ca (mg) Fe (mg) Zn (mg) (mg) Amranthus blitum 3.5 1.7 42 270 3.0 - Amranthus cruentus 3.2 1.8 36 305 3.8 0.7 Amranthus dubius 3.5 3.1 78 582 3.4 1.5 Amaranthus tricolor 3.9 1.8 62 358 2.4 0.8 Amaranthus viridus 4.6 5.7 64 410 8.9 - ***DNIR 59 1.0 100 1000 8.7 9.5

Here, DNIR= Daily nutrient intake recommendation for different age groups in Germany and Switzerland, adapted from Baghurst (2012)

1.4 Justification and Aim of the Study

Amaranths are easy to grow and low maintenance plant. It is considered as a neglected plant and is found most often as a weed. In recent years, some research showing that the plant part of this plant is full of different nutrients and their weather condition adaptability is very diverse. According to some recent research, Amaranths is a new promising plant for future. This plant is not only popular in Africa, widely eaten and cultivated over 50 countries worldwide (National Research Council, 2006). But some popular grain Amaranths still needs some research as well as some vegetable amaranths also (Grubben & Denton, 2004). Vegetable Amaranths are not very much known in Germany. The aim of the study was to test if vegetable Amaranths variety is suitable for cultivation in the environmental conditions in Germany. This study was specifically guided by the following research questions:

 Are there any yield differences among different varieties cultivated in Kleve?  Which variety is the most suitable one for the highest yield in Kleve?  What is the optimum temperature for seed germination for different varieties?

2. Materials and Methods

Eight vegetable amaranths varieties were selected for this study. Eight selected varieties were divided into two groups according to the variety type: leafy variety and stem variety. From these eight selected variety, Danta Drutoraz, Danta Bhutan, Danta Lusina, Danta Ufshi are stem varieties and Laal Shak, Surinamese Spinach , Marog Green, Chinese Spinach are leafy varieties. Seeds of four stem variety (Danta Drutoraz, Danta Bhutan, Danta Lusina, Danta Ufshi ) and one leafy variety (Laal Shak) were collected from Laal Teer Seed Company, Bangladesh. Two leafy variety (Chinese Spinach and Surinamese Spinach) were collected from Templiner-Kraeutergarten, Germany and the seeds of Kenyan leafy variety (Marog Green) were collected from Starke Ayres Company, Kenya. No fertilizer was used during the whole experiment. Seeds were sown on 26th June 2017 on the experiment plot of the greenhouse. All the plants were treated same. The seed germination test was done also by using the same seed. Petri dishes, filter paper and distilled water were used for seed germination test. The seeds were placed in the climate cabinet of Life Science lab for conducting the seed germination test.

2.1 Field Experiment

2.1.1. Location

The experiment plot is located in the garden of greenhouse in Hochschule Rhein-Waal university, Kleve. Kleve is a small city near the dutch-german border which belongs geographically to lower Rhine valley and lower Rhine heights. On an average, the warmest month is July and coolest month is January. The average highest temperature is 24C in Kleve (Figure 1) and 32 C in Bangladesh (Appendix 2) (Meteoblue, 2017). The growing condition is different in two countries. In Bangladesh, the growing season is divided into two part: Rabi and

Kharif. The Rabi season is mainly the winter season starts from October to March and Kharif season is the hot, humid, summer season which starts from April to October. Amaranth (Danta) is a Kharif season vegetable and Laalshak considered as Rabi season vegetable though it can grow throughout the year (BBS, 2016). The agricultural growing season in Germany is from March to October (FAO, 2001)

Figure 1: Average temperature and precipitation in Kleve over the year (Meteoblue, 2017)

2.1.2 Research Design

A randomized block design was used for the experiment plot (Figure 2A). Eight selected varieties were used with two replications. The total plot size was 5.6m in length and 2m in width. This plot was divided into 16 small plots with the size of (70x92) cm each (Figure 2B). There were four rows of each variety in each plot having a 17cm gap in between the rows and 8.5 cm gap in between two new varieties (Figure 2B).

Figure 2: A randomized block design was used for field experiment (A). Here, S1-L8 shows the variety ID of eight selected variety used in this experiment and 'A', 'B' beside the variety ID indicates two replication of each variety. Each sub-plot had four rows in total with 17 cm gap in between rows (B)

2.1.3 Variety Selection and Description

Eight different varieties were selected according to their growing habit and popularity. Vegetable amaranths are popular for its tender leaves and fleshy tender stem in India, Africa and central Africa (National Research Council, 2006). Grain amaranths have got some attention in Germany but vegetable amaranths are still new in Germany. Amaranthus tricolor is a very popular vegetable in Bangladesh (Saha et al. 2013). Most of the varieties used for this study were selected from A. tricolor species. A. tricolor varieties used in this study were Danta Drutoraz, Danta Bhutan, Danta Lusina, Danta Ufshi, Laal Shak, Surinamese Spinach , Marog Green. One A. dubius variety named Chinese Spinach was used also in this study. Amaranthus tricolor and Amaranthus dubius is famous in some tropical regions of India, China, Southeast Asia, and the South Pacific islands (National Research, 1984). Amaranthus tricolor is native in large different regions and it is considered as one of the best-cultivated vegetable amaranth and many varieties of this species are classified according to leaf colour and shape in Southeast Asia (National Research Council, 2006). According to Grubben and Denton (2004), the cultivated type of Amranthus dubius have been exhibited from the weedy ancestor of this species in tropical Asia and then introduced in Africa and Central America by immigrants and is mainly used as a leafy vegetable. Selected eight varieties were divided into two groups according to the variety type which includes four stem variety and four leafy variety. The stem varieties stated as S and leafy varieties as L in this research. More details about the selected Amaranthus varieties are shown in Table 2.

Table 2: Species name, origin, variety name, variety ID, variety type, leaf color, stem color and seed color of the eight selected varieties of amaranths used in this study.

Species Origin Variety Name Variety Type Leaf Stem Seed ID Color Color color

Amaranthus tricolor TAS Danta Drutoraz S1 Stem Green Green Black

Amaranthus tricolor TAS Danta Bhutan S2 Stem Red Green Black

Amaranthus tricolor TAS Danta Lusina S3 Stem Green Green Black

Amaranthus tricolor TAS Danta Ufhsi S4 Stem Green Green Black

Amaranthus tricolor TAS Laal Shak L5 Leafy Red Red Black

Amaranthus dubius TAM Chinese Spinach L6 Leafy Green Green Black

Amaranthus tricolor TAS Surinamese Spinach L7 Leafy Violet Violet Cream

Amaranthus tricolor TAS Marog Green L8 Leafy Green Green Cream

Here, TAS= Tropical Asia and TAM= Tropical America

2.1.4 Management and Cultivation

The plot size was 5.6m in length and 2m wide. The seedbed preparation was done with spike harrow, spade, clod crusher. The soil was loosened by the spade first and leveled the soil with a leveler. The seeds are small and they need a good, firm seedbed. The seeds were sown directly to the soil under 1-1.5 cm depth and then covered with soil gently. Whitehead & Singh (1992) found that sandy loam soil with 6-18% soil moisture is ideal for amaranths. The characteristics of soil in the garden used in this study were analyzed in LUFA lab (Lanwirtschaftkammer, NRW) are presented in Appendix 1. The result of the analysis showed that the soil of experimental plot was sandy loam in texture. The first watering was done immediately after sowing. Then it continued every two days for first four weeks after sowing. The watering was reduced after four weeks and was done once in a week till final harvest. In the rainy season, the irrigation was done based on the soil moisture. The first weeding was done manually after two weeks of sowing. The first soil aeration was done after four weeks of sowing with the help of a spike harrow. The soil aeration was done to reduce the soil compaction which allows the air, water and nutrients to penetrate through the plant roots and helps the root grow deeper and develop a healthy plant. No pest and diseases were found during the field experiment.

2.1.5 Harvest

The harvest was done separately for leafy and stem variety. The growing period for leafy variety is shorter than stem variety. Plants were harvested when they reached their minimum growth period. The harvest of leafy varieties was done on 7th August 2017 and the harvest of stem varieties was done on 21st August 2017. The plants were harvested manually by hand. In each of the plots, a handmade (40x40) cm frame was used to identify the core plants (Figure 3A). Only those plants within the frame were harvested while plants from the outer margin of each sub-

16 plot were excluded from the final harvest to avoid the border effects. Inside the frame, the soil was dug by a spade along the rows. After loosening the soil, each plant was pulled out carefully with roots by hand, collected in bucket separately for each variety and washed thoroughly with water to remove the remaining soil particles from the roots (Figure 3B).

A B C

Figure 3: 40cmx40cm frame used to select the core plants of each plot (A), harvested plants of 'Surinamese Spinach' (L7) (B) and total harvested plant of Marog Green (L8) plot after washing the roots (C)

2.2 Seed Germination Test

The seed germination test was conducted at different temperature in the life science lab of Hochschule Rhein-Waal University. The germination test was started from 5 C and 10 C with three replicates of each variety and continued for 15 C, 20 C, 25 C, 30 C. The seed germination test was done to show the germination capacity of each variety at a different temperature. To avoid any fungal infections, the seeds were washed ito H₂O₂ ixture for seconds (Figure 4A). The H₂O₂ ixture was prepared with l water ad l of % H₂O₂ concentration. After washing the seeds in H₂O₂ ixture, the seeds were placed in distilled water for another 20-30 seconds for five times. Two sterilized filter paper was placed inside each Petri dishes with 10 washed seeds of each variety (Figure 4B). Each variety had 3 replicates with 10 seed in each dish. The first seed germination test with 5 C and 10 C was started on 26th October 2017 and last germination test was started on 6th November 2017 for 25 C. Effect of temperatures on seed germination was observed under continuous darkness at different temperature. Seeds were exposed only to normal light during observations.

A B C

Figure 4: The seeds were washed i H₂O₂ ixture ad i distilled water to avoid fungal diseases (A). Later the seeds were placed in petri dishes where each petri dish contains ten seeds of each variety (B). Germinated seeds of leafy variety 'Laal Shak' (L5) at 20 C after three days of placing the seeds in climate cabinet (C)

2.3. Data Collection

2.3.1 Continuous Measurements of Field experiment

The length of the stem was recorded in centimeter (cm) and the leaves per plant was counted at every week after 14 days of sowing in the experimental plots. Ten plants were randomly selected from the inner rows of each plot for measuring plant height and number of leaves per plant. The height was measured from ground level up to the tip of the growing point. Only unfolded leaves were counted for the total number of leaves per plant. The plant height and leaves per plants were measured continuously once in a week till harvest. The first measurement was taken on 10 July 2017 for all varieties. The last measurement for leafy varieties was taken on 31st July, 2017 and for stem varieties was taken on 14th August, 2017

2.3.2 Destructive Measurements of Field Experiment

Out of the total plants harvested within the (40x40) cm frame, ten plants were randomly selected for final measurement. The fresh weight and dry weight were assessed as a destructive measurement. After washing the roots carefully, the roots, leaves and stems were separated to measure the fresh weight of root, leaf, stem along with stem length, root length and stem diameter. Leaves and roots were detached by a sharp knife from the stem and fresh weight of stem, leaves and roots were measured in gram. Root length were measured in cm and stem diameter was measured at the base of the stem with a slide calipers in mm. Leaf area of green leaves was measured by using LI-3100 leaf area meter (LI-COR, Inc. Nebr.). The data for leaf area was taken two times and later the mean was calculated. The total value of fresh weight was calculated by adding all ten plants fresh weight. After assessing fresh weight, the fresh samples were stored in drying cabin at 80 C for 48 hours for assessing dry weight of stem, leaves and roots. The samples were transferred into desiccators and allowed to cool down at

19 room temperature. The final dry weight of the samples were taken after cooling the samples in a desiccator. Leaf-stem ratio was calculated after harvesting the plant by using following formula:

Fresh Weight of Stem (g)

Leaf-Stem ratio (%) = Fresh Weight of Leaves (g)

2.3.3 Seed Germination Test

Seeds of all selected varieties were placed in climate cabinet under different temperatures. The germination percentage was checked after three, seven and ten days of placing the seeds in climate cabinet and filter paper were moistened with distilled water if needed. Seeds were considered as germinated when the radicle appeared from the seed coat and removed after counting.

2.4 Statistical Analysis

Data of all variables of continuous measurement and destructive measurement were arranged by Microsoft Excel (2010). Normality test was done by performing Shapiro-Wilk test and Q-Q plot with IBM SPSS Statistics for Windows, Version 23.0 (2015) which showed that the data are not normally distributed . A non-parametric Kruskal–Wallis H test and a pairwise comparison test were then performed with IBM SPSS Statistics for Windows, Version 23.0 (2015) on all

20 variables of field experiment to test the significant difference among the yield parameters of tested Amaranthus varieties. The statistical analysis was done separately for leafy and stem varieties. Seed germination data were analyzed by visual assessment of germination curves. Spearmans non-parametric correlation analyses was performed by using IBM SPSS Statistics for Windows, Version 23.0 (2015). Microsoft Excel (2010) and IBM SPSS Statistics for Windows, Version 23.0 (2015) were used for creating boxplots and graphs. Letters above bars denote significant differences between varieties (p < 0.05).

3. Results

3.1 Field Experiment

3.1.1 Plant Height

Plant height was measured continuously starting after two weeks of sowing (Figure 5). Ten plants of each plot A and plot B of all varieties were combined per variety to assess the plant height throughout the field experiment. Growth curves of the stem varieties showed rather similar patterns in plant height while leafy varieties were very different in their growth (Figure 5).

The stem varieties showed small, but significant differences in plant height (Figure 6). For stem variety, the highest plant height was recorded for Danta Bhutan (S2). The leafy varieties showed large significant differences in plant height (Figure 6). In leafy variety, the lowest plant height was recorded for Chinese Spinach (L6) and the highest plant height was recorded for Surinamese Spinach (L7) and Marog Green (L8) on harvest day. The leafy variety Chinese Spinach showed the lowest growth response among all varieties. The lowest plant height was recorded for Chinese Spinach (L6) which is comparatively lower than the highest plant height of other varieties. The plot B of L6 variety was excluded from the final calculation because most of the seedlings died soon after planting. The optimum plant height was noticed for Laal Shak (L5) , flowers appeared with some matured seed until harvest day.

Stem Variety S1 60 S2 S3 50 S4 40 30 20 Height(cm) 10

0 1 2 3 4 5 6 7 8 Weeks after sowing

Leafy Variety

L5 L6 60 L7 L8 50

Height (cm) Height 20

0 1 2 3 4 5 6 Weeks after sowing

Figure 5: Mean plant height of 20 amaranths plants of each eight different varieties sorted by variety type (stem varieties on top, leafy varieties at the bottom)

Figure 6: Boxplot of final mean height (cm) of 20 amaranths plants of each selected varieties sorted by variety type (leafy is coded as L and harvested six weeks after sowing, stem is coded as S and harvested eight weeks after sowing). The H-test were performed separately of each for the four stem and four leafy varieties.

3.1.2 Leaf

Leaves per plants were recorded continuously starting after two weeks of sowing (Figure 7). After two weeks of sowing, the maximum leaves per plant was recorded for Marog Green (L8) and minimum leaves per plant was recorded for Danta Ufshi (S4) among all varieties (Figure 7). In the fourth week of sowing, the highest number of leaves per plant was counted for Marog Green (L8) and lowest number of leaves per plant was counted for Danta Bhutan (S2) and Danta Ufshi (S4).

Stem varieties showed significant differences in number of leaves per plant (Figure 8A). In stem variety, the highest number of leaves per plant was recorded for Danta Lusina (S3) while all other stem varieties showed similarities in number of leaves per plant on harvest day. The leafy varieties also showed significant differences in number of leaves per plant (Figure 8A). In leafy variety, the highest number of leaves per plant was recorded for Surinamese Spinach (L7).

The fresh weight of leaves per plant of stem varieties did not show significant differences (Figure 8B). Leafy varieties showed significant differences in fresh weight of leaves per plant (Figure 8B). In leafy variety, the highest fresh weight of leaves was measured for Surinamese Spinach (L7) which is significantly higher compared to other leafy varieties (Figure 8B).

The four tested stem variety showed no significant differences in leaf area (Figure 9A). The four tested leafy variety showed significant differences in leaf area. The highest leaf area was measured for Surinamese Spinach (L7) and lowest was measured for Chinese Spinach (L6) on harvest day (Figure 9A). Percentage of leaves from total weight showed significance differences in both leafy and stem variety (Figure 9B). Percentage of leaves from total weight was highest for Chinese Spinach (L6) in leafy variety and Danta Drutoraz (S1) in stem variety.

Stem variety

S1 S2 15 S3 S4 12

Number of Leaves per PLant PLant per Leaves of Number 0 1 2 3 4 5 6 7 8 Weeks after sowing

Leafy variety

L5 15 L6 L7 L8 12

3 Number of leaves per Plant per leaves of Number 0 1 2 3 4 5 6 Weeks after sowing

Figure 7: Mean number of leaves of 20 amaranths plants each of eight different varieties sorted by variety type (stem varieties on top, leafy varieties at the bottom)

Figure 8: Boxplot of final mean of number of leaves per plant (A) and mean leaf weight in g (B) of 20 amaranths plants of each selected variety sorted by variety type. Here leafy variety is coded as L and harvested six weeks after sowing and stem variety is coded as S and harvested after eight weeks of sowing. The H-Test were performed separately each for the four stem and four leafy varieties.

Figure 9: Boxplot of mean Leaf area (cm²) (A) and mean percentage of leaves from total weight of 20 amaranths plants of each selected varieties sorted by variety type. Here leafy variety is coded as L and harvested six weeks after sowing, stem variety is coded as S and harvested after eight weeks of sowing. The H-tests were performed separately of each for four stem and four leafy varieties.

3.1.3 Stem

Fresh weight of stem per plant showed significant differences both in stem and leafy variety (Figure 10A). In stem variety, the lowest fresh weight of stem per plant was observed for Danta Drutoraz (S1). In leafy variety, the highest fresh weight of stem per plant was measured for Surinamese Spinach (L7) and the lowest fresh weight of stem per plant was measured for Chinese Spinach (L6) on harvest day (Figure 10A).

Stem varieties did not show any significant differences in stem diameter (Figure 10B). The leafy varieties showed significant differences in stem diameter. Highest stem diameter for leafy variety was recorded for Surinamese Spinach (L7) and lowest was recorded for Chinese Spinach (L6) on harvest day (Figure 10B).

Both leafy and stem varieties showed significant differences in percentage of stem from total fresh weight of 20 plants of each variety (Figure 11). The lowest percentage of stem from total weight was measured for Chinese Spinach (L6) for leafy variety. For stem variety, highest percentage of stem from total weight was measured for Danta Bhutan (S2).

Figure 10: Boxplot of final mean stem weight in g (A) and stem diameter in mm (B) of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties.

Figure 11: Boxplot of mean percentage of stem from total weight of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H- tests were performed separately for each of four stem and four leafy varieties.

3.1.4 Root

Stem varieties did not show any significance differences in root length (Figure 12A). Leafy varieties showed significant differences in root length (Figure 12A). The longest root for leafy variety was measured for Marog Green (L8) and the shortest root was me measured for Laal Shak (L5) on harvest day (Figure 12A).

There were no significant differences found in root weight of stem varieties (Figure 12B). The root weight of leafy varieties showed significant differences (Figure 12B). The highest root weight for leafy variety was recorded for Surinamese Spinach (L7) on harvest day. The root weight for Laal Shak (L5), Chinese Spinach (L6) and Marog Green (L8) showed similar result in root weight which was significantly lower than the root weight of Surinamese Spinach (L7) (Figure 12B).

Stem varieties did not show any significant differences in percentage of root from total fresh weight of 20 plants (Figure 13). The four tested leafy varieties showed significant differences in percentage of roots from total fresh weight of 20 plants (Figure 13). The highest percentage of roots from total fresh weight for leafy variety was observed for Chinese Spinach (L6) (Figure 13).

Figure 12: Boxplot of mean root length in cm (A) and mean root weigh in g (B) of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties.

Figure 13: Boxplot of mean percentage of roots from total weight of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H- tests were performed separately for each of four stem and four leafy varieties.

3.1.5 Total Weight of 20 Selected Plants of Each Variety

Stem varieties did not show any significant differences in total fresh weight of 20 plants of each plot (Figure 14). Leafy varieties showed significant differences in the total fresh weight of 20 plants. In leafy variety, the highest fresh weight of 20 plants was measured for Surinamese Spinach (L7) and lowest fresh weight of 20 plants was measured for Chinese Spinach (L6) (Figure 14)

Figure 14: Boxplot of mean total weight in g of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties

3.1.6 Total Weight of the Fresh Above- Ground Biomass per Plot

Leaves and stems of all the plants harvested inside the (40x40) cm frame were weighed for calculating the total weight of fresh above-ground biomass per hectare. No significant differences were found in the total fresh yield of stem and leafy variety (Appendix 4). The highest fresh yield was observed for Danta Bhutan (S2) in stem variety. The calculated fresh yield for Danta Bhutan was 1.1 t/ha. The highest fresh yield for leafy variety was measured for Green Marog (L8). The calculated fresh yield for Green Marog (L8) was 2.2 t/ha.

Table 3: Mean leaf weight (g), stem weight (g), total yield per plot (g/m²) and average total yield per plot in t/ha of eight selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties

Variety Leaf Weight (g) Stem Total Yield per Calculated Weight (g) plot (g/m²) Total Yield per Plot in t/ha S1 44.3 44.2 88.4 0.6 S2 63.8 104.1 167.8 1.1 S3 59.6 75.4 135.0 0.8 S4 33.4 45.7 79.0 0.5 L5 75.6 110.8 186.4 1.2 L6 28.0 6.81 34.8 0.2 L7 67.4 119.5 187.2 1.2 L8 114.3 235.6 349.8 2.2

3.1.7 Dry weight of Leaves, Stems and Roots of 20 Selected Plants of Each Variety

Stem varieties did not show any significant differences in dry weight of leaves, stems and roots (Figure 15). No significant differences were found also in dry weight of leaves,stems and roots of leafy varities (Figure 15).

80.0 DWL DWS DWR 60.0

40.0

DWL, DWS DWR DWR DWS DWL, 20.0

0.0 S1 S2 S3 S4 L5 L6 L7 L8 Variety

Figure 15: Mean dry weight of leaves, stem and roots of 20 amaranths plants of eight selected variety. Here, leafy variety is coded as L and harvested six weeks after sowing and stem variety is coded as S and harvested after eight weeks of sowing and DWL= Dry weight of leaves, DSW= Dry weight of stem and DWR = Dry weight of root. The H-Test were performed separately each for four stem and four leafy varieties.

3.1.8 Leaf-Stem Ratio

Leaf stem ratio showed significant differences for both stem and leafy varieties (Figure 16). The lowest leaf stem ratio for leafy variety was noticed for Chinese Spinach (L6) while Laal Shak (L5), Surinamese Spinach (L7) and Marog Green (L8) showed similarities. In stem variety, Danta Bhutan (S2) had the highest leaf stem ratio.

Figure 16: Boxplot of leaf-stem ratio of 20 amaranths plants of each selected varieties sorted by variety type. Here, leafy variety is coded as L and harvested six weeks after sowing, stem variety coded as S and harvested after eight weeks of sowing. The H-tests were performed separately for each of four stem and four leafy varieties.

3.2 Correlation between Different Variables of Field Experiment

The Spearmans rho correlation analysis results of selected yield parameters are displayed in Table 5 (Stem variety) and Table 6 (Leafy variety).

In Table 5, the results of correlation analysis of selected yield parameters of stem varieties are summarized. The results of correlation analysis reveals that there was a positively strong significant correlation (r = 0.804) between plant height and stem weight of stem varieties. Positively strong significant correlation (r = 0.792) were also found between number of leaves per plant and leaf weight whereas number of leaves per plant had a positively weak significant correlation (r= 0.391) with root length. There was a very strong positive significant correlation (r = 0.952) between leaf weight and leaf area in stem varieties. Stem diameter and stem weight showed strong positive significant correlation with Root weight (r = 0.895, 0.776). A moderate positive significant correlation (r = 0.474) were found between stem weight and root length. A strong positive significant correlation (r = 0.607) was found between root weight and root length in stem varieties.

In Table 6, the results of correlation analysis of selected yield parameters of leafy varieties are summarized. The results of correlation analysis reveals that there was a very strong positive significant correlation (r =0.913) between plant height and stem weight in leafy varieties. The number of leaves per plant showed strong positive significant correlation with leaf area and stem weight (r = 0.842, 0.840) respectively whereas there was a weak positive significant correlation (r = 0.294) with root length. A very strong positive significant correlation (r = 0.987) was also found between leaf weight and leaf area. Root weight showed a weak positive significant correlation (r = 0.398) with root length in leafy varieties.

Table 4: Spearman's Rho correlation showing relationships among yield parameters of Stem varieties

NL/P LW LA SW SD RW RL Yield Parameter PH .447*** .564*** 624*** .804*** .537*** .524*** .363*** NL/P .792*** .757*** .651*** .695*** .671*** .391*** LW .957*** .798*** .862*** .865*** .523*** LA .813*** .807*** .817*** .548*** SW .776*** .795*** .474*** SD .895*** .557*** RW .607***

Table 5: Spearman's Rho correlation showing relationships among yield parameters of leafy varieties

NL/P LW LA SW SD RW RL Yield Parameter PH .661*** .821*** .867*** .913*** .762*** .725*** .629*** NL/P .831*** .842*** .840*** .822*** .765*** .294*** LW .987*** .931*** .884*** .896*** .515*** LA .957** .889*** .882*** .534*** SW .921*** .891*** .527*** SD .853*** .393*** RW .398***

Here in Table 5 and Table 6, PH = Plant height, NL/P= Number of leaves per plant, LW= Leaf weight, LA= Leaf area, SW= Stem weight, SD= Stem diameter, RW=Root weight, RL=Root length and ***correlation is significant at the 0.01 level

3.3 Seed Germination Test

At 5 C and 10 C none of the tested varieties showed any germination (Figure 17 and Appendix 5). At 15 C, two stem variety S1 and S4 showed low germination rate (< 15 % ) while S2 showed intermediate germination rate ( around 50 %) and S3 and all leafy variety showed high germination rate ( > 70 % ) (Figure 17). At 20 C , S1 had low germination rate ( < 50% ) and germination rate increased for S4 ( > 50 % ) whereas S2 ( > 90 % ) and all leafy variety had high germination rate ( 100 % ). At 25 C, germination rate for S1 ( >80 % ) and S2 ( >70 % ) had increased and all the other variety had maximum germination rate ( 100 % ). At 30 C, S1 (> 90 %) and S4 ( >70 % ) had high germination rate and all the other varieties reached at maximum germination rate ( 100 % ). The ideal minimum temperature for at least 90% germination was therefore 15 C for the varieties S3 and L6, while 20 C for the varieties S2, L5, L7 and L8. Variety S1 needed 30 C for high germination and variety S4 did not show any high germination.

Figure 17: Percentage of germination rate of eight selected varieties at different temperature. Here, stem variety is coded as S and leafy variety is coded as L 41

4. Discussion

The stem and leafy variety showed differences in plant height. In stem variety, Danta Drutoraz (S1) and Danta Ufshi (S4) were comparatively shorter in height than other stem variety. Amaranthus can be harvested at 30 cm to 60 cm height (National Research Council. 2006). In this study, the final plant height ranged from 4.5 cm for Chinese Spinach to 66 cm for Surinamese Spinach on harvest day. The plant height of Surinamese Spinach (L7) was higher than other varieties. Marog Green (L8) was also consistent in higher plant height. Both leaf and stem part used as a vegetable for both varieties. But leafy variety is famous for its leafy part, though the tender stems also used as vegetables and stem variety is mainly famous for its stem part. Laal Shak should be harvested from 25-30 days after sowing to maintain the optimum palatability (Saha et al. 2003). In this study, leafy variety Laal Shak (L5) reached their optimum height till harvest week, which was 42 days after sowing. Some plants were noticed with flowers and grain. Laal Shak normally harvested before flowering because the plants are popular for its leaves and tender stem. Number of leaves per plant is an important measurement for choosing leafy variety. The highest number of leaves per plant (16) was measured for Surinamese Spinach (L7) which is considered as the standard number of leaves per plant for Amranthus tricolor (Miah et al. 2013).

Stem amaranths are mainly popular for its stem part. In this study, the stem amaranth did not reach their optimum growth till harvest week. The average stem diameter is given as approximately 21mm for stem variety under different treatments (Miah et al. 2013). In this study, the highest stem diameter was measured for Danta Bhutan (S2) and Danta Ufshi (S4) was 7.2mm for both varieties which was much lower than the optimum size. Different plant spacing methods influence the growth of stem amaranth. Maintaining a proper spacing is a good criterion for stem amaranth to get the optimum yield. A proper spacing can increase the yield of stem amaranth up to 25% (Bansal et al. 1995). There is different plant spacing recommendation for stem amaranth which ranges between 20 cm to 30 cm (Shanmugavelu,

1989). A field trial was conducted in Florida, the United States to determine the percentage of yield reduction due to plant densities of stem amaranth and they recorded that the percentage of yield reduction can reach 24% with high plant densities (Santos et al. 2003). In this study, there were approximately 25-30 plants in each row and gap between two rows was 17cm. This high plant density could be one reason for the slow growth of stem variety.

According to BBS (2016), in Bangladesh, the yield of Laal Shak was 5 t/ha and the yield of different Danta variety was 7.5 t/ha. In this study, the highest yield of fresh above ground biomass for leafy variety was measured for Marog Green (L8) with 2.2 t/ha and the second highest for Laal Shak (L5) and Surinamse Spinach (L7) which was 1.2 t/ha respectively. Yields of Marog Green (L8) grown under conditions in Kleve can therefore be regarded as intermediate compared to Bangladesh. On the other hand, the total fresh yield of 20 plants per plot was highest for Surinamese Spinach (L7) (Figure 14). In this field experiment, the mean total harvested plant for Marog Green (L8) was 80 per plot whereas the mean total plant for Surinamese Spinach (L7) was 20 plants per plot. Seeds were broadcasted in the plots for sowing and due to the unavailability of seeds, Surinamese Spinach (L7) had comparatively lower number of plants per plot than other varieties. But this variety excelled in all growth parameters than other tested varieties. In this sense, if Surinamese Spinach (L7) had similar number of plants per plots like Marog Green, then the fresh yield could have been assumed approximately 4.8 t/ha which is nearly similar to compare the standard yield of Bangladeshi leafy variety. The highest yield of fresh above ground biomass was measured for Danta Bhutan (S2) with 1.1 t/ha and the second highest was measured for Danta Lusina (S3) 0.9 t/ha for stem variety. Yields of these varieties grown under conditions in Kleve can therefore be regarded as low compared to Bangladesh. On the other hand, the lowest yield of fresh above ground biomass for stem variety was measured for Danta Ufshi (S4) with 0.5 t/ha which comparatively very lower than the standard yield of stem variety in Bangladesh.

Varieties with high yield potential as well as varieties with consistent and reliable yield potential throughout the growing season could be accounted for desirable variety selection. Leaf-stem ratio is an important measurement criterion for variety selection. Both leaf and stems are used as vegetable, hence the variety with higher leaf-stem ratio has more demand (Singh & Whitehead, 1996). According to Campbell & Abbott (1982), the correlation between leaf-stem ratio to yield is negatively correlated and the varieties with highest fresh weight show a tendency of having highest stem weight. In this study, a week positive correlation was found between leaf-stem ratio and yield (Appendix 7), but for 20 plants of each plot, Surinamese Spinach (L7) had the highest fresh weight and stem weight and Chinese Spinach (L6) had lowest fresh weight and stem weight. Chinese Spinach (L6) also had the lowest leaf-stem ratio and lowest yield. This confirms the report of Campbell & Abbott (1982) that A. dubius has lower leaf-stem ratio than A. tricolor.

The average maximum temperature was calculated 24°C during the field study. The optimum air temperature is 28-30°C for maximum seed germination and yield (Whitehead, Carter, & Singh, 2002). Cooler and wetter condition of early growing season affects the growth performance of plants in optimal conditions (Campbell & Abbott, 1982). Plants from the north part of the main plot showed a better result than the plants from south part of the plot. The south part was under shadow during some part of the days due to a tall building in the south, near the plot. In addition, the southern plot part was close to the fence and weed appearance near the fence was higher than in the north part of the plot. A sprinkler irrigation system was used for irrigation during the field experiment. The water from the sprinkler could not reach the south part of the plot. Lack of adequate light , less water availability, more weed stress could be the reason for the slow growth response of the plants from south part of the plot.

Some studies in temperate climate observed that, the Amaranthus varieties which shows highest yield potential in optimum weather condition may also be the most adaptable to wetter

44 and cooler conditions (Campbell & Abbott, 1982). The highest fresh green yield was orserved for Marog Green (L8) and lowest was observed for Chinese Spinach (L6) for leafy variety and for stem variety, the highest fresh green yield was observed for Danta Bhutan (S2) and lowest was observed for Danta Ufshi (S4) in Kleves weather. The leafy variety Chinese Spinach (L6) was consistent in low growth. It is a dwarf variety of amaranthus known as white leaf amaranth, which is cultivated under 21-24 C temperature. This dwarf variety is slow in growth than other amaranthus varieties (Schweig, 2017). Seeds of Chinese Spinach (L6) did not germinate well in the field whereas it showed remarkable germination rate under controlled environment in seed germination test. Low adaptibility of environmental condition compared to other varieties could be one reason for the inability of this variety to germinate at open field experiment. Almost same result was observed for stem varieties also. Stem varieties were also slow in growth in field experiment whereas they showed good response in germination test. Some plants needs more time to germinate even though the climatic conditions are similar required to that amaranths variety (Das, 2016). Amaranths seeds are very small and they often broadcasted in the field for sowing (Ebert, Wu, & Wang, 2011). Though broadcasting method of sowing is easy and not labour intensive but there is high risk which lower the growth of the seedlings due to the competition with weeds, some diseases and pests (Das, 2016). A good firm soil bed with a appropriate planting depth can influence the growth of amaranths. A shallow planting depth may cause decrease the soil-seed contact and increases the risk of the seeds being washed away (Stallknecht & Schulz-Schaeffer, 1993). However, to avoid these problems, starting with seeds in a greenhouse or any other protected structure could be one of the primary ways to maximize the production. Transplanting can be done within two weeks of sowing when plants are about 5-10 cm and late transplanting can affect the yield (Das, 2016). However, the growth response to transplanting has not yet clear, but in some research, it has been stated that growth response to transplanting of some varieties of Amaranthus Tricolor is not satisfactory (Ebert, Wu, & Wang, 2011).

Temperature affects seed germination and seed emergence remarkably. All the varieties failed to show a germination response at 5 C and 10 C. According to Steckel et. al (2004), the seed germination rate response better above 20 C. In this study, all the selected varieties showed approximately 100% germination rate at above 20 C except Danta Drutoraz (S1) and Danta Ufshi (S4). At 20 C, these two varieties showed less germination response where the other varieties reached more than 50% germination rate on the third day of placing seeds. These varieties could not show 100% germination also at 30C where all the other varieties reached the optimum germination rate. The germination rate has a relationship with temperature and germination rate decreases with the decrease of temperature (Das, 2016). According to Cochrane et al. (2011), differences in seed germination rate at different primary temperatures are not exceptional. Steckel et. al (2004) also found that germination rate increases for some amaranths varieties as the temperature increase from 5 C to 30 C. In this study, the germination rate of selected amaranths varieties was increased with the temperature and seeds showed better response above 20 C.

5. Conclusions and Recommendations

The selected stem and leafy varieties showed differences in their growth and seed germination rate. This study showed that leafy variety is in general more suitable than stem variety for the weather condition in Kleve, Germany. Surinamese Spinach (L7) showed highest values in plant height, leaves per plant, leaf area, leaf weight, stem weight, stem diameter than other tested varieties. Leafy variety Chinese Spinach (L6) was consistent in lower growth in field experiment. Marog Green (L8) showed a consistent reliable growth and seed germination

46 response and also had highest yield. Hence, considering the highest growth and yield, the findings of this study may recommend as Marog Green (L8) the suitable variety for Kleve, Germany and considering the consistent growth and germination response, Surinamese Spinach (L7) could also be a good choice for the growers. However, this study was done as a single harvest study. Some further research is necessary to choose the ultimate amaranths variety for Germany. Considering the situation of this study, some further studies in the following areas may be suggested:

 Variety trials with more replication may be performed to observe the differences in growth and yield more broadly.  This study may be carried out in different regions of Germany for observing the regional adaptability.  Plant spacing may be included to compare the plant performance.

Acknowledgements

I would first like to thank my thesis advisor Dr. Katja Kehlenbeck. The door to Dr. Kehlenbecks office was always open whenever I ran into a trouble spot or had a question about my research or writing. She consistently allowed this paper to be my own work, but steered me in the right direction whenever she thought I needed it. Besides my advisor, I would like to thank Michael Hemkemeyer and Christoph Knoblauch for their insightful comments and valuable guidance for my statistical analysis. I would also like to thank Franz-Josef Kuhnigk for his support and help during my field experiment. Last but not the least, I would like to thank my parents and my sister for their continuous encouragement and support throughout the study.

DECLARATION OF AUTHENTICITY

I, Tamanna Tanzin, hereby declare that the work presented here is my own work, and has not been published or submitted for any other degree or qualification. The experimental work is entirely my own work and completed without any aid from external parties and other resources than those listed. Any material from other sources or works done by others has been given due acknowledgement and listed in the reference section. Sentences or parts of sentences quoted literally are marked as quotations; identification of other references with regard to the statement and scope of the work is quoted. The work presented herein has not been published or submitted elsewhere for assessment in the same or a similar form. I will retain a copy of this assignment until after the Board of Examiners has published the results, which I will make available on request

Place, Date Signature

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Appendices

Appendix 1: Soil Characteristics assessed in LUFA lab (Lanwirtschaftkammer, NRW)

Physical Analysis: Type of sample: outdoor Designation: Greenhouse JG 1 Soil type: sand. Loam / loess / loam (sL, uL, L) Cultivation type: Vegetables, Mixed Culture

Chemical Analysis: Test Parameters Unit Test Result pH in CaCl2 6.5 Phosphorus (P2O5) in CAL mg/100g 37 Potassium (K2O) in CAL mg/100g 7 Magnesium (Mg) in CaCl2 mg/100g 5 Humus % 2.4 Entire organ. % 1.4 Carbon (TOC) Total nitrogen (TN) % 0.12

Test parameters / Test Methods used: Potassium (K2O) in CAL, phosphorus (P2O5) in CAL: VDLUFA Method Book Volume I, 1991, A 6.2.1.1 Entire organ. Carbon (TOC), Humus: VDLUFA Method Book Volume I, 2016, A 4.1.3.2 Total nitrogen (TN): DIN ISO 13878 pH in CaCl2: VDLUFA Method Book Volume I, 1991, A 5.1.1 Magnesium (Mg) in CaCl2: VDLUFA Method Book Volume I, 1991, A 6.2.4.

Appendix 2: Average temperatures and precipitation over the years in Bangladesh (Meteoblue, 2017)

Appendix 3: Daily nutrient intake recommendation for different age groups in different areas (Baghurst, 2012)

Nutrients USA/Canada Australia/ Germany/ UK 1991 FAO/WHO 1997-2004 Newzealand Switzerland (19-50 years) RNI 2001 (31-50 years) 2005 2000 (19-50 years) (25-51 years) Protein (g) 58 64 59 45 55 Vitamin A(mg) 1.0 0.9 1.0 0.7 0.6 Vitamin C (mg) 90 45 100 40 45 Calcium(mg) 1000 1000 1000 700 1000 Iron(mg) 10 8 10 8.7 9 Zinc(mg) 15 14 10 9.5 7

Appendix 4: Mean leaf weight and mean stem weight mean of total plants per plot in g. Here, FLW=Fresh weight of leaf and FSW=Fresh weight of stem

400

300

200 FLW FSW

FLW & FSW FSW & FLW 100

0 S1 S2 S3 S4 L5 L6 L7 L8 Variety

Appendix 5: Mean percentage of seed germination of leafy and stem varieties at different temperature.

Type 5°C 10°C 15°C 20°C 25°C 30°C S1 0.0 0.0 10.0 36.7 86.7 93.3 S2 0.0 0.0 56.7 100.0 100.0 100.0 S3 0.0 0.0 93.3 96.7 100.0 100.0 S4 0.0 0.0 3.3 53.3 63.3 76.7 L5 0.0 0.0 83.3 100.0 100.0 100.0 L6 0.0 0.0 93.3 100.0 100.0 100.0 L7 0.0 0.0 73.3 100.0 100.0 100.0 L8 0.0 0.0 86.7 100.0 100.0 100.0

Appendix 6: Total rea and total production of Laal Shak and Danta, 2015-16 (BBS, 2016)

Crop Total Area Total Yield in (ha) Production (T) t/ha Laal Shak 11367 56950 5.0

Danta 11025 82855 7.5

Appendix 7: Correlation analysis of leaf-stem ratio to total fresh yield

6.0

5.0 r = 0.3

4.0

3.0

2.0

1.0

0.0 0 10 20 30 40 50 60