SALINITY TOLERANCE AND NITROGEN USE EFFICIENCY OF QUINOA FOR
EXPANDED PRODUCTION IN TEMPERATE NORTH AMERICA
By
ADAM JOSHUA PETERSON
A thesis submitted in partial fulfillment of the requirements for the degree of
MASTER OF SCIENCE IN CROP SCIENCE
WASHINGTON STATE UNIVERSITY Department of Crop and Soil Science
MAY 2013
To the Faculty of Washington State University:
The members of the Committee appointed to examine the thesis of ADAM JOSHUA PETERSON find it satisfactory and recommend that it be accepted.
______Kevin M. Murphy, Ph.D., Chair
______Joan R. Davenport, Ph.D.
______Kimberly A. Campbell, Ph.D.
ii ACKNOWLEDGEMENT
I’d like to acknowledge those whose support and encouragement has sustained me throughout my time as a graduate student. I owe so much to my parents, Gary and Lisa, who helped cultivate a sense of curiosity in me from a young age. Whether it was allotting me a section of our pasture to grow wheat, or connecting my grain mill to a bicycle, they always found ways to support me, no matter how large or small. I’d also like to thank my sister, Allicia, and my grandmother, Arlene, for their support.
I would like to acknowledge my undergraduate mentors: Martha Rosemeyer,
Donald Morisato, Stephen Bramwell, and Melissa Barker, for guiding an undecided science major into the field of sustainable agriculture. Special thanks to Stephen
Bramwell and Melissa Barker for keeping me in mind for a job managing a quinoa trial at the Evergreen Organic Farm three years ago.
I would like to thank the members of my committee. I owe a huge debt of gratitude to Kevin Murphy, my advisor and mentor. His advice, encouragement, and calm reassurance helped ground me as a graduate student, and he’s truly been a role model for me as a budding plant breeder. He began me on an adventure with this crop that’s taken me from the rolling hills of the Palouse across the planet to the Rift Valley of Africa. I can’t thank him enough.
I would like to thank Joan Davenport and Kim Campbell for their support and expert advice. Joan Davenport helped ignite my interest in soil science and provided me with quick and knowledgeable advice through the challenges I faced in my research. I’m
iii very grateful to Kim Campbell for her assistance in statistical analysis, which was helpful for a complex data set that proved quite the challenge to analyze.
I would like to recognize the help and encouragement I received from teachers, faculty, staff, technicians, and fellow students. Sam Turner, Raymond Kinney, Hannah
Walters, Karen Welch, Kelsey Highet, Dustin Tombleson, Joseph Astorino, Brook
Brouwer, and Edward Olson for their encouragement and help with field and greenhouse work. Thanks to Marc Evans, for his invaluable help with SAS code. Many thanks to
Janet Matanguihan, Max Wood, Brad Jaeckel, and Jacqueline Cruver, for their generous and crucial support. I’d like to acknowledge and thank Tobin Peever for lending me use of the microscopes in his lab, which I used to photograph quinoa crossing techniques.
I’d like to acknowledge WSU BIOAg and the Organic Farming Research
Foundation (OFRF) for their financial support, which made this research possible.
Above all, I’d like to thank and express my deep admiration for the many indigenous peoples of South America who developed and continue to grow quinoa in some of the harshest agricultural environments on the planet. Their perseverance throughout the centuries is reflected in the resilience of this remarkable crop. I’d like to express my sincere hope that the development of quinoa as a crop outside of South
America leads only to their benefit and wellbeing.
iv SALINITY TOLERANCE AND NITROGEN USE EFFICIENCY OF QUINOA FOR
EXPANDED PRODUCTION IN TEMPERATE NORTH AMERICA
Abstract
by Adam Joshua Peterson, M.S. Washington State University May 2013
Chair: Kevin M. Murphy
Quinoa has attracted increasing attention worldwide and in North America due to its high level of mineral nutrition and superior tolerance to marginal agriculture conditions and abiotic stresses. A wide range of challenges and opportunities currently face expanded quinoa production in North America. Heat susceptibility, pre-harvest sprouting, and downy mildew are among the most important of these challenges. In spite of these challenges, varieties with tolerance to high temperatures and resistance to pre- harvest sprouting and downy mildew have been identified. Quinoa’s high level of salinity tolerance will also allow the crop to take advantage of marginal agriculture conditions that limit productivity of other crops.
Two experiments were conducted to explore quinoa’s potential for expanded production in North America. The first experiment examined the relative salinity tolerance of four Chilean lowland varieties to determine their suitability for cultivation on saline soils in North America. All quinoa varieties were grown at 8, 16, and 32 dS m-1
NaCl and Na2SO4 and at a no-salt control. Quinoa demonstrated high levels of salinity tolerance, far exceeding that of barley, a crop generally considered saline tolerant.
v Additionally, variation for salinity tolerance was found among the four quinoa varieties.
On the basis of yield, quinoa was found to better tolerate Na2SO4 than NaCl at equal EC levels.
Previous studies on quinoa and other crops indicate that salinity can significantly impact mineral nutrition of seeds. Our results indicate complex but significant effects from salinity, fertilization level, and variety, and the interaction of these factors, on Ca,
Cu, Fe, Mg, Mn, P, and Zn concentrations in quinoa seed.
The second experiment investigated the response of a wide range of varieties to four levels of a nitrogen-rich organic fertilizer. However, large declines in yield due to high temperatures limited the recovery of useful data on nitrogen use efficiency from the study. Valuable data was gathered on the relative levels of heat tolerance present among the Chilean lowland cultivars currently part of the WSU quinoa program. Field observations indicate that natural selection in 2011 may have increased heat tolerance in the same varieties grown the following year.
vi TABLE OF CONTENTS
Page
ACKNOWLEDGEMENT..…………………………………………………………...iii-iv
ABSTRACT……………………………………………………………………………v-vi
LIST OF TABLES…………………………………………………………………..xii-xii
LIST OF FIGURES.……………………………………………………………………..xv
GLOBAL INTRODUCTION...…………………………………………………………...1
CHAPTER 1: QUINOA CULTIVATION FOR TEMPERATE NORTH AMERICA: CONSIDERATIONS AND AREAS FOR INVESTIGATION…………………………..5
1. INTRODUCTION.……………………………………………………………..6
2. HEAT TOLERANCE…………………………………………………………..9
3. VARIETY SELECTION……………………………………………………...10
4. DROUGHT TOLERANCE…………………………………………………...12
5. WATERLOGGING AND SPROUTING……………………………………..13
6. COLD TOLERANCE…………………………………………………………14
7. SALINITY TOLERANCE……………………………………………………15
8. DISEASE……………………………………………………………………...19
9. INSECTS AND PESTS………………………..……………………………...21
10. WEED CONTROL…………………………………………………………..24
11. FORAGE…………………………………………………………………….25
12. FERTILIZATION……………………………………………………………26
13. PLANTING/SPACING……………………………………………………...29
14. MATURITY AND HARVESTING…………………………………………32
vii
15. SAPONINS…………………………………………………………………..33
16. CONCLUSION………………………………………………………………35
CHAPTER 2: TOLERANCE OF LOWLAND QUINOA CULTIVARS TO SODIUM CHLORIDE AND SODIUM SULFATE SALINITY I: EFFECTS ON YIELD, HEIGHT, AND LEAF GREENNESS…………………...………………………………………….47
1. INTRODUCTION.………. …………………………………………………..48
2. MATERIALS AND METHODS……………………………………………...50
2.1 Experimental design………………………………………………….50
2.2 Statistical analysis …………………………………………………...52
3. RESULTS……………………………………………………………………..53
3.1 Yield………………………………………………………………….53
3.2 Leaf greenness………...……………………………………………..55
3.3 Plant height…………………………………………………………..56
3.4 Correlations…………………………………………………………..56
4. DISCUSSION…………………………………………………………………57
4.1 Yield………………………………………………………………….57
4.2 Leaf greenness………………………………………...……………..58
4.3 Plant height…………………………………………………………..61
4.4 Correlations…………………………………………………………..62
5. CONCLUSION.……………………………………………………………….63
CHAPTER 3: TOLERANCE OF LOWLAND QUINOA CULTIVARS TO SODIUM CHLORIDE AND SODIUM SULFATE SALINITY II: EFFECTS OF SALINITY ON MINERAL NUTRITION OF SEEDS…………………………………………………...85
1. INTRODUCTION…………………………………………………………….86
viii
2. MATERIALS AND METHODS……………………………………………...87
2.1 Experimental design………………………………………………….87
2.2 Statistical analysis……………………………………………………88
3. RESULTS……………………………………………………………..………89
3.1 Ca……………………………………………………………….……89
3.2 Cu……………………………………………………………...……..92
3.3 Fe……………………………………………………………...…...... 95
3.4 Mg…………………………………………………………………....99
3.5 Mn………………………………………………………...………...102
3.6 P…………………………………………………………………….106
3.7 Zn………………………………………………………………...... 110
4. DISCUSSION………………………………………………………………..113
4.1 Ca……………………………………………………………….…..113
4.2 Cu……………………………………………………………...……115
4.3 Fe…………………………………………………………...…….....116
4.4 Mg………………………………………………………….……….117
4.5 Mn………………………………………………………….……….119
4.6 P…………………………………………………………………….120
4.7 Zn…………………………………………………………………...121
5. CONCLUSION………………………………………………………………112
CHAPTER 4: NITROGEN USE EFFICIENCY OF QUINOA UNDER ORGANIC CONDITIONS IN SOUTHEASTERN WASHINGTON……………………………...145
ix 1. INTRODUCTION…………………………………………………………...146
2a. MATERIALS AND METHODS – 2011 QUINOA NUE EXPERIMENT...148
2a.1 Quinoa varieties…………………………………………………...148
2a.2 Experimental design……………………………………………….148
2a.3 Statistical analysis………………………………………………....151
2b. MATERIALS AND METHODS – 2012 MODIFIED QUINOA NUE EXPERIMENT………………………………………………………………....151
2b.1 Experimental design……………………………………………….151
2b.2 Statistical analysis…………………………………………………153
3a. RESULTS – 2011 QUINOA NUE EXPERIMENT………………………..153
3a.1 Yield……………………………………………………………….153
3a.2 Plant height………………………………………………………...154
3a.3 Aphid susceptibility and leaf greenness…………………………...154
3a.4 Days to appearance of flowering bud……………………………...154
3a.5 Days to full senescence………………………………….………...154
3a.6 Correlations………………………………………………………..155
3a.7 Soil samples……………………………………………………….155
3b. RESULTS – 2012 MODIFIED QUINOA NUE EXPERIMENT…….……155
3b.1 Main response variables…………………………………………...155
3b.2 Tissue samples…………………………………………………….156
3b.3 Soil samples……………………………………………………….156
4a. DISCUSSION – 2011 QUINOA NUE EXPERIMENT……………………156
4a.1 Yield……………………………………………………………….156
x
4a.2 Plant height………………………………………………………...158
4a.3 Days to appearance of flowering bud……………………………...159
4a.4 Days to full senescence……………………………………………159
4b. DISCUSSION – 2012 MODIFIED QUINOA NUE EXPERIMENT….…..160
4b.1 Main response variables…………………………………………...160
4b.2 Tissue samples…………………………………………………….161
4b.3 Soil samples……………………………………………………….161
5a. CONCLUSION – 2011 NUE EXPERIMENT…………………………..…161
5b. CONCLUSION – 2012 MODIFIED QUINOA NUE EXPERIMENT….…162
APPENDIX
A. CROSSING METHOD FOR QUINOA USING MANUAL EMASCULATION……………………………………………………………..175
1. Introduction…………………………………………………………..176
2. Materials and Methods……………………………………………….176
3. Results………………………………………………………………..181
GENERAL CONCLUSIONS…………………………………………………………188
xi LIST OF TABLES
CHAPTER 2
1. Analysis of variance with F-values for yield for quinoa and barley ………….67
2. Analysis of variance with F-values for yield, leaf greenness, and plant height for quinoa only…………………………………………………………………...68
3. Analysis of variance with F-value for relative declines in yield under 32 dS m-1 NaCl and Na2SO4……………………………………………………………...... 69
4. Relative declines in yield (g/plant) between the control and 32 dS m-1 NaCl...70
5. Change in yield (g/plant) from low to high fertilization ……………..……….71
6. Pearson correlation coefficients between yield, height, and leaf greenness ….72
7. Pearson correlation coefficients between yield, height, and leaf greenness under NaCl salinity and high fertilization ……………………………………….……..73
8. Pearson correlation coefficients between yield, height, and leaf greenness under NaCl salinity and low fertilization ………………………………………...…….74
9. Pearson correlation coefficients between yield, height, and leaf greenness under Na2SO4 salinity and high fertilization ………………………..………………….75
10. Pearson correlation coefficients between yield, height, and leaf greenness under Na2SO4 and low fertilization………………………………………..……..76
CHAPTER 3
1. Ca concentration (μg Ca g-1) at 0 ds m-1……………………………...……...126
2. Ca concentration (μg Ca g-1) at treatment combinations……….……...….....127
3. Cu concentration (μg Cu g-1) at 0 dS m-1.……………………………………128
4. Cu concentration (μg Cu g-1) at treatment combinations…….……………....129
5. Fe concentration (μg Fe g-1) at 0 dS m-1.………..……...………………..…..130
6. Fe concentration (μg Fe g-1) at treatment combinations…….…….…….…...131
xii
7. Mg concentration (μg Mg g-1) at 0 dS m-1.………………………………..…132
8. Mg concentration (μg Mg g-1) at treatment combinations……….…………..133
9. Mn concentration (μg Mn g-1) at 0 dS m-1………………………………..….134
10. Mn concentration (μg Mn g-1) at treatment combinations………….…...... 135
11. P concentration (μg P g-1) at 0 dS m-1.………….……..………………..…..136
12. P concentration (μg P g-1) at treatment combinations………...... ……..…..137
13. Zn concentration (μg Zn g-1) at 0 dS m-1.………….…...... 138
14. Zn concentration (μg Zn g-1) at treatment combinations…………...... 139
15. Pearson correlation coefficients for response variables for all treatment combinations……………………………………………………………………140
16. Pearson correlation coefficients for response variables, NaCl under high fertilization.……………………………………………………………………..141
17. Pearson correlation coefficients for response variables, NaCl under low fertilization.…………………………………...………………………………...142
18. Pearson correlation coefficients for response variables, Na2SO4 under high fertilization..…………………………………………………………….………143
19. Pearson correlation coefficients for response variables, Na2SO4 under low fertilization....…………………………………………………………………...144
CHAPTER 4
1. List of quinoa varieties included in the 2011 quinoa NUE experiment……...165
2. Varieties in the 2012 modified quinoa NUE experiment…………………….166
3. Analysis of variance with F-values for plant height (PH), leaf greenness (LF), aphid susceptibility (AS), yield, and days to flower bud appearance (DTFB) in the 2011 quinoa NUE experiment.………………………….…………………..….167
4. Height and days to appearance of flowering bud by variety for the 2011 quinoa NUE experiment ………………………………………….………..…………..168
xiii
5. : Analysis of variance with F-values for days to full senescence for 2011 quinoa NUE experiment ………………………………………….………….………...169
6. Correlations between plant height (PH), leaf greenness (LG), aphid susceptibility (AS), yield, and days to flower bud appearance (DFBA) in the 2011 quinoa NUE experiment …………………………………………….……..…..170
7. Nutrient analysis of tissue samples from the 2012 quinoa NUE experiment……………………………………………………………………....171
xiv LIST OF FIGURES
CHAPTER 1
1. Figure 1; Pre-harvest sprouting in quinoa…………………….……………….47
CHAPTER 2
1. Figure 1; Yield of quinoa and barley by salinity treatment…………………...77
2. Figure 2; Yield of quinoa by salinity treatment……………………...... 78
3. Figure 3; Variety x fertilization interaction for yield…………….………...... 79
4. Figure 4; Leaf greenness by variety………………………………...... 80
5. Figure 5; Leaf greenness by fertilization level…….………………...... 81
6. Figure 6; Leaf greenness by salinity treatment……………………...... 82
7. Figure 7; Variety by fertilization interaction for plant height…….………...... 83
8. Figure 8; Fertilization by salinity interaction for plant height..………….……84
CHAPTER 4
1. Figure 1; Yield from the 2011 quinoa NUE experiment …………………....172
2. Figure 2; Weather data for Pullman, WA (2011)...…………………...... 173
3. Figure 3; Weather data for Pullman, WA (2012) ………………………….....174
APPENDIX A
1. Figure 1; 1st Flowering Pattern….…………………...... 183
2. Figure 2; Female flower with protruding bifurcated stigma………………....184
3. Figure 3; Developing inflorescence………………………………...... 185
4. Figure 4; Plant after terminal section of inflorescence is removed……….....186
xv
GLOBAL INTRODUCTION
In the past few decades, interest in quinoa (Chenopodium quinoa Willd.) outside of South
America has greatly increased. Because of quinoa’s nutritional richness and its high level of hardiness in marginal environments, the crop has earned special attention from agronomists and nutritionists alike. The crop has begun to be exported from South America in large quantities.
Programs to cultivate the crop outside of South America have been launched and currently, quinoa has spread from a crop grown only in the Andean highlands and Chilean lowlands of
South America, to having been tested or grown on every continent except Antarctica.
In North America, work on quinoa began in 1983, when a partnership was formed between Colorado State University and Sierra Blanca Associates (Johnson, 1990). A breeding program was launched, and the crop began to be cultivated in the San Luis Valley of Colorado and in parts of Wyoming and New Mexico (Ward, 1994). Quinoa also spread to the Canadian prairies, where it is currently cultivated commercially in Saskatchewan through the Northern
Quinoa Corporation (Alberta Agriculture, Food, and Rural Business, 2005).
As demand for quinoa continues to rise and outpace supply, there has been a surge of interest in North America for expanded domestic quinoa production. Current research on quinoa in North America is largely limited to early research at Colorado State University. However, if quinoa is to expand to new locations in North America, further agronomic testing will be necessary. One of the greatest barriers for quinoa its low level of heat tolerance. Early researchers identified this as limiting factor for its range of cultivation (Johnson, 1990; Oelke et al., 1992; Ward, 1994). Oelke et al. (1992) suggests locations in northern and central Washington state may be suitable for quinoa cultivation, and quinoa was grown commercially in northern
1
Washington state in the 1990s (Schreiber and Ritchie, 1995). Due to its wide range of growing environments and gradients in temperature, annual precipitation, and growing season length,
Washington state will provide a useful environment for testing and improving quinoa’s adaptation to particular pressures. Experiments in this thesis report current work at Washington
State University to reintroduce and expand cultivation of quinoa to the larger region, as well as identify challenges for quinoa cultivation in other parts of the United States and Canada.
Chapter 1 reviews the existing literature on quinoa production, drawing on research conducted at Colorado State University, current and past efforts from quinoa programs in Europe and abroad, and reports from a wide range of South American sources. Potential challenges such as heat susceptibility, pre-harvest sprouting, saponins, disease pressures, and insect pressures are considered. Recommendations for agronomic practices are reviewed.
Quinoa’s high degree of salinity tolerance has also garnered a great deal of attention. Soil salinization is a major and growing problem worldwide. In the United States, 2.2 million hectares are currently classified as saline, largely centered in the western portions of the country and the Upper Great Plains (USDA, 1989, 2011), In Canada, soil salinity is a major problem in the Prairie provinces, affecting 2.19 million hectares in Alberta, Saskatchewan, and Manitoba
(Alberta - Agriculture and Rural Development, 2004). Quinoa’s high salinity tolerance offers a potential solution to this problem. However, studies on the salinity tolerance of Chilean cultivars, which are most suitable to cultivation at temperate latitudes, have been lacking until relatively recently. In chapter 2, a greenhouse experiment testing the relative salt tolerance of quinoa to
NaCl and Na2SO4 is explained. Four Chilean lowland varieties were included and represent a diverse selection from varieties included in initial field trials in 2010.
2
As one of the main nutritional benefits of quinoa is its high level of mineral nutrition, seeds from the experiment described in Chapter 2 were analyzed to determine the effect of salinity, fertilization, and variety on mineral nutrition. Chapter 3 describes the complex effects of these factors on Ca, Cu, Fe, Mg, Mn, P, and Zn concentration
Currently, the only fertilization recommendations for any location for North America are from early work at Colorado State University. In Europe, several experiments examining the nitrogen use efficiency of quinoa indicate that the response of quinoa to fertilization is relatively complex and is dependent on variety and environment. Chapter 4 explains the results of a nitrogen use efficiency experiment conducted in 2011 and 2012. In the first year, 16 varieties were included, representing the best performing varieties grown in initial trials in 2010 and several commercial cultivars. Interference from high temperatures limited collection of useful data on nitrogen uptake. The following year, 14 cultivars were tested. Due an extreme reduction in germination due to close contact of seed with fertilizer and poor seedbed conditions, the experiment was revised to analyze the effects of thinning and plant competition on nutrient uptake.
3
References
Alberta - Agriculture and Rural Development. 2004. Salinity classification, mapping and management in Alberta. Available at http://www1.agric.gov.ab.ca/$department/deptdocs.nsf/all/sag3267#prairies (verified 15 January 2013). Alberta – Agriculture and Rural Development.
Alberta Agriculture, Food and Rural Business. Ag-Entrepreneurship Division. 2005. Quinoa ...: the next cinderella crop for Alberta? Alberta Agriculture, Food and Rural Development, Edmonton, Alberta.
Johnson, D.L. 1990. New grains and pseudograins. p. 122–127. In Janick, J., Simon, J.E. (eds.), Advances in new crops. Timber Press, Portland, OR.
Oelke, E.A., Putnam, D.H., Teynor, T.M., and Oplinger, E.S. 1992. Quinoa. In Alternative Field Crops Manual. University of Wisconsin Cooperative Extension Service, University of Minnesota Extension Service, Center for Alternative Plant & Animal Products.
Schreiber, A., and L. Ritchie. 1995. Washington minor crops. Washington State University, Richland, Wash.
USDA. 1989. The second RCA appraisal: soil, water, and related resources on nonfederal land in the United States. U.S. Government Printing Office, Washington, D.C.
USDA. 2011. 2011 RCA Appraisal. USDA, Washington, D.C.
Ward, S.M. 1994. Developing improved quinoa varieties for Colorado. PhD Dissertation. Colorado State University. Fort Collins, CO.
Wilson, H.D, and J. Manhart. 1993. Crop/weed gene flow: Chenopodium quinoa Willd. and C. berlandieri Moq. Theor Appl Gen. 86: 642–648.
4
CHAPTER ONE
QUINOA CULTIVATION FOR TEMPERATE NORTH AMERICA: CONSIDERATIONS AND AREAS FOR INVESTIGATION
Quinoa Cultivation For Temperate North America: Considerations And Areas For Investigation
Abstract
Quinoa has been commercially cultivated in North America since its introduction in the
1980s. However, quinoa has since largely been limited to high altitude-locations in the Rocky
Mountains and the Canadian Prairies where cool temperatures allow its cultivation. High temperatures remain the largest barrier for its expanded cultivation on the continent. Elsewhere, in regions characterized by a maritime oceanic climate, pre-harvest sprouting and maturity time are the greatest barriers to production. As a relatively new crop, many factors such as pests and disease pressure will likely continue to develop as production of quinoa expands. Additionally, many agronomic factors need to be studied in more detail, informed by initial research conducted in Colorado and by ongoing research in Europe. These will likely vary across the wide range of environments found across the continent. Breeding will play a crucial role in developing varieties uniquely adapted to these environments. Despite these challenges, quinoa's high level of genetic diversity and abiotic stress tolerance provide routes for expanded cultivation and hold potential for its cultivation in areas of marginal agricultural land, particularly in areas affected by soil salinity.
1. Introduction
Quinoa is a pseudograin which originates from the Andean highlands of South America.
Evidence of quinoa cultivation dates back to 5500 BC (Dillehay et al., 2007), and the crop's center of origin is considered to be around Lake Titicaca (National Research Council (U.S.),
6
1989). Quinoa's traditional range of cultivation stretches as far north as Columbia and as far south as southern Chile. As a result of its wide distribution, the crop is adapted to a wide range of environments and forms a diverse range of ecotypes (Risi and Galwey, 1984).
By the time of Spanish arrival in the 16th century, quinoa had reached its maximum range and the crop played a crucial role in Incan agriculture. After suppression of its growth following
Spanish conquest, quinoa cultivation declined. However, it still remained an important crop by indigenous peoples, particularly in areas of marginal ground (Cusack, 1984).
Due to its unique characteristics, quinoa has gained an increasing amount of international attention over the past few decades. The first of these is the crop's superior nutritional characteristics. In contrast to most plant protein sources, quinoa seed protein contains a balanced level of essential amino acids. Quinoa also has higher levels of minerals compared to other grains and contains significant levels of vitamins and essential fatty acids (Schlick and
Bubenheim, 1996; Vega-Gálvez et al., 2010). Additionally, it contains high levels of polyphenols, flavonoids and ecdysteroids which are beneficial for human health (Repo-Carrasco-
Valencia et al., 2010; Kumpun et al., 2011).
Quinoa also exhibits remarkable agronomic characteristics. It is highly tolerant to drought and soil salinity. Additionally, some varieties can tolerate temperatures as low as -8°C for short periods (Jacobsen et al., 2003; Jacobsen et al., 2005). It can tolerate a wide range of soil conditions, subsisting off poor soil fertility, and can withstand pH values ranging from 4.8 to 8.5
(Narrea, 1976; Tapia, 1979; as cited in Risi and Galwey, 1984). Due to its extensive range of cultivation in South America, C. quinoa as a species contains the genetic diversity needed to grow in a wide range of environments outside South America (Jacobsen, 2003).
7
International interest in quinoa began to rise in the late 1970s and 1980s when the first breeding programs outside of South America were begun. In Europe, programs were established in the UK, Denmark, and the Netherlands (Jacobsen, 2003). In North America, efforts were begun in 1983 to grow quinoa in high altitude locations of Colorado through a partnership between Colorado State University and Sierra Blanca Associates (Johnson, 1990). Additionally, private efforts to test and grow quinoa varieties in North America were begun by John Marcille in northern Washington state and Emigdio Ballon in northern New Mexico (Ballon, 1990;
Wilson and Manhart, 1993). Quinoa production expanded outside Colorado to Wyoming and
Northern New Mexico (Ward, 1994). By the late 1980s, efforts to grow quinoa in Canada were underway (National Research Council (U.S.), 1989; Tewari and Boyetchko, 1990; Small, 1999).
Cultivation of quinoa in North America is currently centered in Saskatchewan through the Northern Quinoa Corporation and in the San Luis Valley of Colorado. However, despite efforts to cultivate quinoa in North America and other parts of the world, the majority of worldwide quinoa cultivation takes place in South America. In recent years, there has been a large gap in the supply of quinoa and growing demand for it, leading to high prices and negative social and environmental consequences in the major quinoa producing countries (Jacobsen,
2011; Romero and Shahriari, 2011).
Cultivation of quinoa outside of South America could help ameliorate the current issues associated with rising production and exports of quinoa from its native range. Efforts are increasing to adapt and grow quinoa in various parts of the world (Jacobsen, 2003). Recently, quinoa's potential as a nutritious and resilient crop was recognized by the FAO which declared
2013 as the International Year of Quinoa (FAO, 2013).
8
2. Heat Tolerance
Quinoa production in North America has so far been limited to regions with cool summers, where maximum temperatures do not exceed 35°C. Initial trials in Colorado found that quinoa was not suitable at elevations lower than 2100 m, due to pollen sterility caused by high temperatures (Johnson and Croissant, 1985). Multiple reports exist of high temperatures being an issue in North America, namely in Minnesota (Oelke et al., 1992), Virginia (Bhardwaj et al.,
1996), and Alberta (Alberta Agriculture, Food, and Rural Business, 2005). Oelke et al. (1992) suggest that heat susceptibility would limit the expansion of quinoa cultivation to coastal areas in central California, high altitude locations in central and northern Washington state, and parts of the Canadian prairies. High temperatures have been noted as having a detrimental effect on yield and seed filling in locations outside of North America such as Morocco (Jellen et al., 2005),
Chile (Fuentes and Bhargava, 2011), Greece (Iliadis et al., 2001), and Italy (Pulvento et al.,
2010).
Current work at Washington State University has found maximum summer temperatures to be a major limiting factor for successful seed set in quinoa. While temperatures did not exceed
35°C during anthesis in a 2011 field experiment, high temperatures during the seed fill stage may have caused significant reductions in yield. Many of the inflorescences lacked seeds or contained empty seeds (unpublished data). Despite the large reduction in yield in 2011, some varieties exhibited greater heat tolerance than others. Seed produced in 2011 was replanted in a field experiment the following year. Despite higher temperatures in 2012, higher levels of seed set were observed for many varieties. This suggests that natural selection may have selected for greater heat tolerance among the genetically diverse selections grown.
9
Variation in adaptation to hot, dry conditions has also been observed among Chilean highland varieties grown in a trial performed in the Atacama Desert of northern Chile (Fuentes and Bhargava, 2011). The effect of heat stress on quinoa has been covered in most detail by
Bonifacio (1995), who noted that heat stress can cause reabsorption of endosperm, a phenomenon recognized by Bolivian farmers as "puna”. Bonifacio also observed high temperatures inhibiting anther dehiscence in outdoor plantings in Provo, Utah in 1993, and varietal differences in heat tolerance.
As most areas of temperate North America experience summer temperatures that exceed the 35°C threshold for quinoa (NOAA - National Climatic Data Center, 2011), the development of heat tolerant varieties would greatly expand the area suitable for quinoa production and increase harvest security in the face of damaging heat waves. Despite quinoa's heat susceptibility, closely related Chenopodium spp. inhabit parts of North America that experience intense summer heat (Jellen et al., 2011). Additionally, archaeological evidence indicates the existence of a domesticated chenopod pseudograin, analogous to quinoa, in the prehistoric
Eastern North American agricultural complex (Smith, 1985). This suggests that the development of a heat tolerant chenopod pseudocereal for North America outside of high altitude or cool maritime locations is not an impossibility. Should existing variation for heat tolerance within quinoa prove insufficient, C. berlandieri and C. bushianum to which quinoa can hybridize, may provide a promising source for introgression of genes for heat tolerance.
3. Variety selection
10
Quinoa is traditionally divided into five main ecotypes: Valley, which is grown between
2000-4000 m elevation from Central Peru northwards; Altiplano, grown at 4000 m elevation around Lake Titicaca; Salar, grown at 4000 m around salt flats in southern Bolivia; Subtropical, from the Yungas region of Bolivia; and Sea level, from temperate latitudes in southern Chile
(Tapia et al., 1980 as cited in Risi and Galwey, 1984). The Sea level ecotype, also referred to as
Chilean lowland ecotype, is recognized as the best adapted to temperate latitudes and high summer temperatures. After initial trials, Chilean cultivars were chosen to form the basis of breeding programs in Europe and in Colorado (Johnson, 1990; Jacobsen, 1999). Chilean lowland varieties have shown to be the best adapted to cultivation in Washington state (unpublished data), and only Chilean and southern Bolivian varieties were noted to set seed in Colorado
(Johnson, 1990). Bertero (2003) concluded that Chilean lowland and Altiplano cultivars, characterized by low photoperiod sensitivity and short basic vegetative period, are most suited for cool, temperate latitudes.
There are reports of non-Chilean varieties successfully producing yields at high latitude locations in Europe. A wide range of Peruvian and Bolivian varieties produced seed in a field trial in Finland, at two locations above 60°N (Carmen, 1984). Likewise, Valley and Altiplano cultivars produced significant yields in England, as did Bolivian and Peruvian cultivars in
Denmark. However, non-Chilean cultivars generally exhibit greater times to maturity (Risi and
Galwey, 1991a; Jacobsen and Stolen, 1993; Jacobsen, 1997). Christiansen et al. (2010) found that photoperiod induced a stay green reaction in Real, a Bolivian cultivar.
11
Non-Chilean cultivars largely failed to set seed in field trials in Washington state
(unpublished data). Bertero (2003) also noted failure of these cultivars to reach maturity in
Argentina.
In an experiment with an Altiplano cultivar, long daylength and high temperatures were found to have a detrimental effect on seed fill. The two factors alone had small but significant effects, but when 16 hour daylength was combined with 28°C temperature, seed size decreased by 73% (Bertero et al., 1999). This relatively low threshold for high temperatures, combined with the influence of long summer daylength indicates that non-Chilean varieties likely have little place for cultivation in temperate regions of North America.
4. Drought Tolerance
Quinoa is considered to have remarkable drought tolerance. It has been reported to grow with as little as 200 mm in annual precipitation in pure sand (Aguilar and Jacobsen, 2003).
Yields exceeding 1000 kg ha-1 have been reported with as little as 50 mm irrigation in the
Atacama Desert of Northern Chile. However, yields are much improved in arid regions under irrigation (Martínez et al., 2009). Initial research in Colorado found that quinoa yielded best with
208 mm of combined irrigation and rainfall on a sandy loam soil (Flynn, 1990). Later recommendations to growers in the San Luis Valley were 25-38 cm of combined irrigation and rainfall (Johnson and Croissant, 1985). The addition of organic matter has been shown to be useful in increasing quinoa yield under arid conditions (Martínez et al., 2009).
The effect of drought on yield varies depending on the stage of plant development. Geerts et al. (2006) found that drought occurring in early growing stages improved overall water use
12
efficiency. When drought occurred during the pre-flowering stage up until the dough stage, significant decreases in yield were seen. Jensen et al. (2000) also found decreases in yield when drought was applied during flowering and seed fill. Additionally, yield increases were seen when simulated drought occurred during the vegetative stage.
However, contrasting responses have been reported. Razzaghi et al. (2012) found that yield did not significantly decrease when simulated drought was applied during the seed filling stage. Darwinkel and Stølen (1997) reported greater drought tolerance in later growth stages.
Jacobsen and Stolen (1993) note that in Denmark, the greatest impact from drought occurs during the vegetative stage.
5. Waterlogging and Sprouting
Quinoa has been reported to grow in regions of high rainfall. For instance, a quinoa variety was collected from a location in Chiloé Island in Chile (Wilson, 1981), an island characterized by high levels of annual precipitation (1400-2000 mm) and temperate rainforest
(Vera, 2006). Pre-harvest sprouting can be an issue when rainfall coincides with seed maturity.
This was seen during quinoa trials held in 2010 in Olympia, Washington. Here, a rare heavy rainfall event in late summer caused heavy sprouting in a majority of the 44 varieties (Figure 1).
However, a few accessions proved resistant to this, including PI 614880 (unpublished data,
2010), the aforementioned variety collected from Chiloé Island. This accession has been shown to exhibit seed dormancy which confers pre-harvest sprouting resistance (Ceccato et al., 2011).
Pre-harvest sprouting has also proven to be a problem in the Netherlands. Quinoa varieties in the
Dutch quinoa breeding program were successfully screened for pre-harvest sprouting resistance
13
by testing the dormancy of seeds relative to the date of ripeness. Variability was found within the accessions and this trait was successfully selected upon in their breeding program (Mastebroek and Limburg, 1997).
In a greenhouse experiment, Gonzales et al. (2009) noted the negative impact of waterlogging, which was more severe than that of drought. Unnecessary irrigation of seedlings can cause stunting and damping off. During later growth stages, irrigation of quinoa caused increases in vegetative growth but no increase in seed production (Oelke et al., 1992).
Waterlogging had a deleterious impact on quinoa in a trial in the UK, with differential responses from varieties being noted (Risi and Galwey, 1989).
6. Cold Tolerance
Reports on quinoa's tolerance to frost in the field vary. Risi and Galwey (1984) reviewed reports of quinoa's frost tolerance in South America, with some sources indicating little or no frost resistance and others indicating high levels of frost resistance.
Experiments under controlled conditions have shown superior frost tolerance in an
Altiplano type variety compared to a Valley type variety. Frost tolerance varies with genetic background, but it is also linked with quinoa's ability to increase proline and soluble sugars with exposure to cooler temperatures (Jacobsen et al., 2007). Growth stage also affects tolerance to frost. Exposure after the initiation of flowering proved far more damaging than exposure during earlier growth stages. A 66% yield reduction was seen when plants in anthesis were exposed to -
4°C. Seedlings at the two-leaf growth stage only saw a 9% reduction in yield to the same
14
exposure. Humidity also interacts with the impact of frost on quinoa, with drier conditions proving more damaging (Jacobsen et al., 2005).
A few reports exist of frost tolerance in Chilean lowland cultivars. In Colorado, reports indicate that quinoa could withstand light frosts of -1°C to 0°C. In accordance with Jacobsen's findings, a heavy frost -4.4°C during flowering caused losses exceeding 70% (Johnson and
Croissant, 1985). Oelke et al. (1992) suggested that temperatures below -2°C during flowering would cause significant losses. However, once seeds are in the soft dough stage, frost resistance increases and plants are reported to withstand temperatures down to -7°C. In England, Risi and
Galwey (1984) found that several Chilean cultivars tolerated a number of frosts in spring, one of which reached -5° C. Darwinkel and Stølen (1997) reported varietal differences in frost tolerance and noted -3°C as the threshold for quinoa.
7. Salinity Tolerance
Soil salinity is a significant agricultural problem for large parts of temperate North
America. In the United States, there are 2.2 million hectares of saline agricultural land, with a further 30.8 million hectares of agricultural land under threat of salinization (USDA, 2011). The majority of these saline affected soils are confined to arid and semi-arid regions of the West
(USDA, 1989). In Canada, salinity is a significant problem in the Prairie provinces of Alberta,
Saskatchewan, and Manitoba, affecting an estimated 2.19 million hectares (Alberta - Agriculture and Rural Development, 2004).
15
Quinoa is generally recognized as one of the most saline tolerant crops known (Jacobsen,
2007). In its native range, quinoa is cultivated in areas with highly saline soils. For instance,
Salares varieties are grown around salt flats in Southern Bolivia (Risi and Galwey, 1984).
Quinoa has exhibited a remarkable level of salinity tolerance under controlled conditions compared to other cultivated crops. Koyro and Eisa (2007) and Hariadi et al. (2011) demonstrated the ability of quinoa to survive and produce seed under 500 mmol NaCl, an equivalent concentration to seawater. Jacobsen et al. (2003) found that the variety Kanckolla was able to germinate at 57 mS/cm, a level exceeding that of seawater.
The salt tolerance of quinoa has been demonstrated to vary significantly within and between ecotypes. In an analysis of Peruvian quinoa germplasm, Gómez-Pando et al. (2010) found significant variation for saline tolerance at both the seedling and adult stage. Tolerance at the seedling stage was not necessarily connected with tolerance during the adult stage (Jacobsen et al., 1999b). Adolf et al. (2012) investigated the response of Danish, Bolivian, and Peruvian varieties to salinity and noted a large range in physiological responses and relative tolerance.
Despite the complex variation exhibited, Danish varieties of Chilean lowland background were among the less saline tolerant cultivars, while the Real type quinoa varieties from Southern
Bolivia exhibited greater levels of salinity tolerance.
Investigation of salinity tolerance within Chilean lowland germplasm will prove important, as these varieties are most adapted to the conditions found in areas of North America affected by soil salinity. Many salt affected areas in western North America experience high summer heat temperatures characterized by a continental climate. For some areas, such as the
San Joaquin Valley of California, mild climate may allow early planting and avoidance of high
16
temperatures. However, in regions with a short growing season but high summer temperatures, breeding cultivars with greater heat tolerance will be necessary. Characterizing the level of salinity tolerance in Chilean lowland cultivars, which have the greatest heat tolerance, will prove key in developing quinoa as a successful halophytic crop for these areas.
Several studies have focused on salinity tolerance in Chilean lowland material, both under controlled conditions and in the field. Two germination studies conducted with Chilean germplasm have noted significant geographical trends in adaptation. Delatorre-Herrera and Pinto
(2009) found higher tolerance among a Chilean highland variety compared to a Chilean lowland variety and connected this to adaptation to the saline soils found in the highlands of northern
Chile. Ruiz-Carrasco et al. (2011) tested varieties within the range of the Chilean lowland ecotype, comparing varieties from central Chile with a variety from southern Chile. The southern
Chilean variety was found to have lower levels of saline tolerance compared to the central
Chilean varieties. This difference was linked to the gradient of decreasing exposure to salinity and increasing precipitation that runs from north to south in Chile.
Further investigation to confirm a geographical gradient of saline tolerance among
Chilean lowland accessions could prove fruitful in locating the most saline tolerant Chilean lowland germplasm. Additionally, given the increasing temperatures that are also found at more northern and lower latitudes in Chile (Dirección Meteorológica de Chile; Ruiz-Carrasco et al.,
2011) greater heat tolerance may be found in conjunction with greater salinity tolerance.
A field study performed in Southern Italy found that a Danish quinoa cultivar had no significant difference in yield when irrigated with saline water of 22 dS m-1, mixed to approximate 1:1 ratio of seawater to freshwater, compared to a fresh water control (Pulvento et
17
al., 2012). For comparison, barley, considered a saline tolerant grain, sees yield decreases when yield exceeds 8 dS m-1 (Maas, 1986); however, a lower threshold of 6 dS m-1 has been reported by Royo et al (2000). When quinoa was grown on a saline-sodic soil in Greece with an EC of 6.5 dS m-1, Karyotis et al. (2003) saw poor establishment, which the authors linked to high pH, high
Na+, and poor soil physical characteristics due to soil sodicity. Cultivars responded differently when grown on a non-saline soil, indicating varietal differences in tolerance. Overall, yield was much decreased, with the best performing variety under saline-sodic conditions yielding 1.27 t ha-1, while the best performing cultivar under non-saline conditions yielded 2.3 t ha-1. The added effects of sodicity to salinity likely explains the large differences in yield response compared to the findings by Pulvento et al (2012). As quinoa emergence is reduced by soil crusting, the tendency of sodic soils to crust may be a barrier for quinoa's potential to grow on these soils.
Additionally, the higher levels of sodium characteristic of soil sodicity may have had an impact.
A greater effect by sodicity is backed by an early experiment comparing quinoa's response to saline, saline-sodic, and sodic soils. Sodicity was found to have a detrimental impact on quinoa biomass yield compared to salinity, though differences in sodicity tolerance were seen between the two varieties examined (Torres, 1955).
Sodium chloride is the salt most commonly investigated for crop salinity tolerance, but other areas in North America experience problems with other salt types. For instance, sulfate salts are predominant in North Dakota (Keller et al., 1986), which contains the largest area of saline and saline-sodic soils of any state in the United States (USDA, 1989). In the Yakima
Valley of Washington state, both sodium sulfate and sodium chloride are predominant problem salts (Smith et al., 1958)
18
Differential response to salt type has been noted in several crop species. At high concentrations, sodium sulfate had a greater detrimental effect on sorghum than sodium chloride
(Boursier and Läuchli, 1990). Chenopodium rubrum, a species which inhabits sodium sulfate affected soils in Western Canada, saw a more detrimental impact on growth at high concentrations of sodium sulfate than sodium chloride (Warne et al., 1990).
Quinoa has been reported to grow in soils affected by chloride and sulfate salts
(Delatorre-Herrera, 2003). However, no studies contrasting the tolerance of quinoa to sulfate and chloride salts have been conducted. All studies have either used NaCl or a mixture of salts in determining salinity tolerance. Searching for and characterizing differences tolerance based on salt type could allow for more precise use of quinoa as a salt tolerant crop for saline affected areas.
8. Disease
Due to isolation, quinoa in North America has escaped many of the disease pressures it faces in its native range. However, at least one major pathogen of quinoa, downy mildew
(Peronospora variabilis), has been reported in quinoa grown in Canada (Tewari and Boyetchko,
1990). Downy mildew has also been identified growing on field trials in Washington state
(unpublished data) and in Pennsylvania (Testen et al., 2012). Downy mildew infecting quinoa has been shown to be closely related to strains that infect Chenopodium album, a common weed in North America. Choi et al. (2010) suggest the possibility that C. album may be a potential inoculum reservoir for quinoa. More investigation is needed on the potential of C. album and other closely related Chenopodium spp. to host strains of P. variabilis to which C. quinoa is
19
susceptible. As Choi et al. (2010) demonstrated, downy mildew exhibits a high level of host specificity. No threat is posed from strains of downy mildew that infect the related crops spinach
(Spinacia oleracea L.) and beets (Beta vulagris L.), as these strains belong to Peronospora farinosa, a separate species from Peronospora variabilis.
Variation for resistance to downy mildew exists within quinoa strains. Chilean lowland varieties have been reported to contain more resistance to the pathogen than other ecotypes
(Fuentes et al., 2008). However, reports from Denmark of P. variabilis infecting Dutch and
Danish cultivars indicate that problematic levels of susceptibility to P. variabilis do exist among varieties with a Chilean lowland background (Danielson et al., 2002).
The close relative C. berlandieri has been noted to have high resistance to downy mildew
(Jellen et al., 2011). The domesticated subspecies C. berlandieri sbsp. nuttalliae has been used to introgress downy mildew resistance into quinoa (Bonifacio, 2004). In an experiment in India conducted on downy mildew resistance, accessions of C. berlandieri sbsp. nuttalliae and the
North American species C. bushianum exhibited no symptoms of infection. Diverse responses, ranging from lack of infection to high susceptibility, were also seen from a wide range of quinoa cultivars, leading the authors to postulate the existence of multiple pathotypes (Kumar et al.,
2006). Reports from Dutch breeding efforts have found a source of downy mildew resistance that is dominantly inherited (Mastebroek and Van Loo, 2000). The reported diversity for downy mildew resistance should allow for the breeding of resistant cultivars. As C. berlandieri sbsp. nuttalliae and C. bushianum have both been successfully crossed with quinoa, these species represent another source for downy mildew resistance (Wilson, 1980)
20
Danielsen and Munk (2004) developed a standardized method – the three-leaf technique - to determine the severity of downy mildew infection in terms of yield reduction. They also noted the preference of downy mildew for cool, moist conditions. The Salares type cultivar Utusaya was found to be highly susceptible to downy mildew. However, in its usual range of cultivation,
Utusaya escapes infection due to the dry climate. Escape from downy mildew infection should be possible in western regions of the United States characterized by dry summers. This is supported by reports of reduced infection in the Europe under dry summer conditions as compared to more humid summers (Jacobsen, 1999). Areas characterized by wet conditions and high humidity during the growing season may find downy mildew to be a more significant problem. As downy mildew in quinoa has been shown to be transmissible by seed, care must be taken when introducing quinoa in a new area to prevent introducing the pathogen (Danielsen et al., 2004).
One other report of quinoa disease in North American exists. In a fall quinoa variety trial in Southern California, Sclerotium rolfsii caused damping off of quinoa seedlings, as well as seed rot (Beckman, 1980).
9. Insects and Pests
Many insect pests have been reported for quinoa in its native range in South America.
The most damaging of these pests is the quinoa moth (Eurysacca melanocampta, E. quinoae;
Rasmussen et al., 2003). Quinoa grown in North America has escaped the pests found in its native range through isolation.
21
As quinoa continues to be grown in North America, it is likely that native pest species will extend their host range to include quinoa. This has occurred in Europe, where two pests that normally feed on C. album, Scrobipalpa atriplicella and Cassida nebulosa, have begun to attack quinoa (Sigsgaard et al., 2008).
Oelke et al. (1992) reports that insect pressure was not determined to be a significant factor for Colorado grown quinoa. However, a study of pest pressures in Colorado found a wide range of insect pests on quinoa several years after the introduction of quinoa. Many of the reported pests were also pests of C. album and Beta vulgaris. Some particular pests were found to lower yields and cause major losses in some locations. The major seedling infecting insects were found to be Melanotrichus coagulatus (Uhler) and the false cinch bug, Nysius raphanus
Howard. Beet armyworm, Spodoptera exigua (Hübner), caused large scale defoliation at one location. Another problematic foliar feeder was the boat gall aphid Hayhurstia atriplicis (L.).
Lygus spp. were noted as problematic seed feeding pests and sugarbeet root aphid (Pemphigus populivenae Fitch) caused yield declines (Cranshaw et al., 1990). Oelke et al. (1992) records sugarbeet root aphid as a significant quinoa pest, saying that cracked soil provided a point of entry. Additionally, flea beetles and aphids were reported on quinoa grown in Minnesota.
Darwinkel and Stølen (1997) note some overlap in the pests of beets and quinoa in
Europe. Beet flea beetles (Chaetocnema concinna and C. tibialis) and beet carion beetles
(Aclypea opaca) were found to feed on quinoa grown in sandy soil, though no data is given on the relative seriousness of these pests. Black bean beetle (Aphis fabae) was also reported as a pest likely causing decreases in yield.
22
In Washington state, aphids and Lygus sp. have been the major pest at test plots throughout the state since trials began in 2010. Harvesting aphid infested plants results in hardened honeydew particles which prove considerably difficult to clean from harvested quinoa seed. Lygus sp. have been observed to take shelter in more compact inflorescences (unpublished data).
Various control methods for quinoa pests in South America have been recommended, although much research remains to be done for development of proper IPM strategies. Current pest control strategies that are most transferable to North American conditions are crop rotation, intercropping, and biocontrols. Varietal resistance for pests, such as for quinoa moth, has been observed (Rasmussen et al., 2003).
As pest pressures change and develop with expanding quinoa production in North
America, strategies for pest control will likewise change. Given the current pest pressures seen, some recommendations can be made. In Colorado, Cranshaw et al. (1990) noted syrphid larvae,
Diaeretiella rapae, and the convergent lady bird beetle (Hippodamia convergens) as predators of
H. atriplicis. Variations between sites in aphid pressures were also seen. The aforementioned species and other aphid parasitoids should be investigated as potential biocontrol agents.
Additionally, the factors that determine the severity of aphid infestation require further study.
As quinoa and other Chenopodium species share common pests, studying the potential of
Chenopodium spp. weeds as alternate hosts is warranted. Close rotations of quinoa with related crops such as beets, should be carefully monitored to ensure that pests common to both crops do not spread.
23
10. Weed Control
Quinoa's slow growth after germination makes weeds particularly challenging. Related
Chenopodium spp. are often a major pressure, due to their similarities to quinoa in early growth habit and appearance. Common lambsquarter was noted as a major weed pressure in southern
Colorado, along with pigweed (Amaranthus sp.), kochia (Bassia sp.), and sunflower (Helianthus sp.; Oelke et al., 1992). In Colorado, recommendations for growers were to plant in areas of low weed pressure and to cultivate for weed management (Johnson and Croissant, 1985). Early planting was also found to be key. Tilling the soil for planting after irrigating was found to be effective, particularly with controlling Chenopodium spp. (Oelke et al., 1992). Late planting of quinoa in England led to complete loss to heavy weed pressure (Risi and Galwey, 1991b), confirming the importance of early establishment of quinoa. Grasses were found to cause major yield reductions in quinoa. Johnson and Ward (1993) saw reductions in yield from 1,822 kg ha-1 to 640 kg ha-1 from pressure from grassy weeds.
Jacobsen et al. (2010) compared the efficacy of inter-row hoeing to harrowing for weed control under Danish conditions. They found that inter-row hoeing, when combined with a row spacing of 50 cm, was most effective at reducing weed pressures. Harrowing had a significant but less pronounced impact. Although both methods had a significant decrease on plant stands during one of the two years the study was conducted, yield significantly increased by both methods. Of most interest is that Chenopodium album was the predominant weed species, indicating these methods were effective in controlling weedy Chenopodium spp.
24
Observations in a study by Risi and Galwey (1991) of ten quinoa varieties in England noted distinct differences in the negative impact of weed pressure on plant height. This seems to indicate that some quinoa varieties may have greater ability to compete with weeds.
Cross-pollination of C. quinoa with weedy C. berlandieri has been confirmed at a farm in
Northern Washington state (Wilson and Manhart, 1993). Oelke et al. (1992) reports putative hybrids between quinoa and common lambsquarter in Colorado (species unspecified). Such pollination could have consequences in terms of maintaining purity of quinoa varieties. Control of C. berlandieri will prove critical in preventing undesired cross-pollination.
11. Forage
Quinoa has attracted attention as a forage crop. Carlsson et al. (1984) investigated quinoa in southern Sweden and concluded it was a promising crop for the production of green LPC
(liquid protein concentrate). Dry matter production was found to increase with high levels of fertilization, up to 470 kg N ha-1.
In tests in the Netherlands, quinoa performed poorly as a forage crop in comparison with grass and clover (Schooten and Pinxterhuis, 2003). Early reports on quinoa's forage potential present a contrasting picture. Quinoa biomass had high crude protein and low fiber when harvested near the flowering period and was competitive with alfalfa and grass when compared on a cost basis. Quinoa silage was found to be an acceptable method of storing quinoa fodder long term. Increased nitrogen applications were recommended for forage quinoa, with 100 kg N ha-1 applied at sowing followed by an additional 200 kg N ha-1 applied five weeks later
(Darwinkel and Stølen, 1997).
25
12. Fertilization
Fertilizer recommendations for quinoa in Colorado were 135 kg N ha-1 based on results from early trials (Johnson and Croissant, 1985). However, Oelke et al. (1992) notes that fertilization recommendations were increased to 170-200 kg N ha-1 after more extensive research.
Nitrogen exceeding these levels lead to lodging and delayed maturity. Under Danish conditions, yield increases were seen at N applications of 160 kg N ha-1 compared to the lower levels of 120,
80, and 40 kg N ha-1. However, the response of yield under higher levels was minimal. At 40 kg
N ha-1, yield was only 24.1% lower than at 160 kg N ha-1 (Jacobsen et al., 1994). Trials in
Denmark and the Netherlands resulted in differences in nitrogen response across locations and years, although general recommendations of 100-150 kg N ha-1 were given. Too little nitrogen resulted in poor yields, while nitrogen in excess of 150 kg N ha-1 was found to have little benefit
(Darwinkel and Stølen, 1997).
Trials in southern Germany found quinoa quite responsive to increased nitrogen fertilization. Yields at no fertilization produced 1790 kg ha-1, while 120 kg N ha-1 caused yields to almost double to 3495 kg ha-1. Nitrogen utilization efficiency, a measure of grain yield in proportion to aboveground plant nitrogen, was not found to change under increasing fertilization, though a significant difference was seen between the two varieties tested (Schulte auf’m Erley et al., 2005). In a greenhouse experiment conducted by Thanapornpoonpong (2004), nitrogen utilization efficiency showed significant decreases under increasing N, contrasting the results found by Schulte auf'm Erley et al (2005). This difference is notable in that one of the varieties tested – Faro – was common to both experiments, indicating contrasting nitrogen dynamics due to environment. Again, there was a significant difference between varieties, though in nitrogen
26
uptake efficiency, which measures the proportion of above ground plant nitrogen in proportion to available soil nitrogen supply.
The large differences seen at different locations for nitrogen dynamics, as well as significant varietal differences, pose several challenges and opportunities. Varietal differences in response to nitrogen introduce the possibility of breeding for increased nitrogen use efficiency.
Quinoa has historically thrived in marginal areas and varieties that are adapted to low soil nitrogen might prove valuable. Traditional cultivation of quinoa on the Peruvian Altiplano generally involves no fertilization. In crop rotations, quinoa follows potatoes and scavenges remaining nutrients in the soil (Aguilar and Jacobsen, 2003).
As most demand is for organic quinoa, a look at the response of quinoa varieties to fertilization and management under organic conditions is necessary. Quinoa cultivars developed under university or private breeding programs may represent selection under mineral fertilization application, while traditional quinoa landraces represent selection under traditional agricultural practices. Selection of cultivars under organic and conventional conditions has been shown to result in cultivars adapted to those respective conditions, as has been shown in wheat (Murphy et al., 2007). Properly evaluating differences between cultivars and determining their nitrogen use efficiency under organic conditions will be useful both for calculating NUE and for informing future breeding efforts. Currently, little data exists on quinoa's response to organic fertilization.
Bilalis et al. (2012) reported yield increases of 6% with compost application and of 10% with cow manure application.
Seed composition is influenced by fertilization. In the aforementioned study, saponin levels increased under organic fertilization as compared to a non-fertilized control. Johnson and
27
Ward (1993) found that protein levels are responsive to nitrogen applications, increasing by
0.1% per kg of ammonium nitrate.
Soil texture is reported to have a major impact on nitrogen uptake and nitrogen use efficiency. In a lysimeter experiment where 120 kg N ha-1 was applied, nitrogen uptake was 134 kg N ha-1 under a sandy clay loam but only 77 kg N ha-1 under a sandy soil, leading to differing yields of 3.3 Mg ha-1 and 2.3 Mg ha-1 respectively (Razzaghi et al., 2012).
Most studies have applied nitrogen in one large application, although several studies report splitting applications between planting and a later stage (Aguilar and Jacobsen, 2003;
Schulte auf’m Erley et al., 2005; Pulvento et al., 2010). However, there is no published research investigating the effect of split fertilizer applications with quinoa. One study conducted on wheat found differential response to the number of fertilizer applications, with higher grain yield under three split applications as opposed to two split applications (Abdin et al., 1996). Darwinkel and
Stølen (1997) suggest splitting fertilizer applications exceeding 150 kg N ha-1 to prevent excessive salt exposure.
There are contradictory reports for quinoa's response of phosphorus fertilization. In Chile, yield increased with 100 kg P ha-1 and 200 kg P ha-1 when accompanied by nitrogen fertilization
(Delatorre-Herrera, 2003). Recommendations from South America indicate that quinoa responds well to 80 kg N ha-1 and 80 kg P ha-1 (Mujica, 1977, as cited in Aguilar and Jacobsen, 2003).
However, Gandarillas (1982, as cited in Johnson and Ward, 1993) found no yield response to
-1 phosphorus or potassium application. In Colorado, phosphorus applications of 34 kg P2O5 ha failed to produce an effect on yield (Oelke et al., 1992). Darwinkel and Stølen (1997) reported
-1 requirements of 70 kg P2O5 ha for quinoa prior to seed filling and note that existing levels of
28
phosphorus in many agricultural soils are likely sufficient. They also note a fairly large requirement for potassium, with uptake of 400 kg K ha-1 , and recommend application of 100-200 kg K ha-1 (actual). Data is largely absent on the micronutrient requirements of quinoa. However,
Darwinkel and Stølen (1997) report that manganese is a critical micronutrient for quinoa.
13. Planting/Spacing
Row spacing for quinoa varies widely. In South America, a variety of planting techniques are used. When quinoa is planted in rows, spacing ranges from 0.4 and 0.8 m. Wide ranges of optimal sowing density are reported (Tapia et al., 1980, as cited in Risi and Galwey, 1984).
Recommendations for Colorado are based on a plant density of 320,000 plants ha-1 and a sowing density of 0.6-0.8 kg ha-1. No set row spacing is recommended, though spacing greater than 36 cm is beneficial (Johnson and Croissant, 1985). In experiments in England, highest yields were seen for the Chilean variety Baer under the closest tested row spacing of 20 cm seeded at 20 kg seed ha-1. Increasing density caused a decrease in the amount of branching on plants and also resulted in earlier maturity in the field (Risi and Galwey, 1991b).
Jacobsen et al. (1994) investigated row spacings of 50, 25, and 12.5 cm and found no significant differences between them. The widest spacing of 50 cm was combined with mechanical hoeing to control weeds and plants under this spacing appeared healthiest, while plants at the 25 and 12.5 cm row spacings were able to crowd out weeds. Additionally, optimal plant density was calculated to be 327 plants m-2 with a standard deviation of 220 plants m-2, leading Jacobsen et al. (1994) to conclude that yield could remain stable over a wide range of plant densities. Based on later experiments in Denmark and the Netherlands, Darwinkel and
29
Stølen (1997) confirm the stability of yield across a wide range of row spacings, ranging from
12.5 to 75 cm, and plant densities ranging from 30 to 250 plants m-2. Based on harvesting considerations and weed competition, a seeding rate of 6 kg ha-1 with target plant densities of
100 to 150 plants m-1 were recommended for quinoa cultivation in northern Europe.
The large range of plant densities, row spacing, and seeding rates found in these studies indicate the large influence that location and weed pressure, as well as cultivar characteristics, have on yield. Given the wide range of conditions found across temperate North America, further research will be necessary to find the proper spacing and seeding rates for new locations.
Planting depth is also variable depending on soil type and conditions. Oelke et al (1992) recommends planting at a depth of 1.3 to 2.5 cm in Colorado. Excessively deep sowings were noted to lead to poor emergence due to waterlogging, while too shallow of plantings left seeds vulnerable to soil drying. In Europe, high germination required adequate contact of seeds with soil at the proper moisture level (Darwinkel and Stølen, 1997). Aufhammer et al. (1994) found that soil crusting has shown a highly detrimental effect on seedling emergence. They stressed the importance of a light-textured seedbed for successful germination. Quinoa germination rates were found to drop steeply at depths greater than 2 cm. At shallower depths, greater emergence was seen in a sandy soil than a loamy clay soil.
Reports from Europe confirm the need for a fine-textured even seedbed, and indicate that heavy clay soils can cause problems with early establishment of the root system. Plant depths of
1 to 2 cm were recommended (Darwinkel and Stølen, 1997)
Temperature has a strong effect on successful germination and reports on optimum temperature vary. In Denmark, seed is usually planted when the soil temperature is 7-8°C. An
30
experiment with a Danish cultivar “Olav” of Chilean lowland origin shows maximum germination between 15-20°C. However, high rates of germination were seen for a wide range of temperatures, including 8°C, the lowest temperature tested (Jacobsen and Bach, 1998).
Recommended soil temperatures for planting for Colorado were 7-10°C (Johnson and Croissant,
1985). For England, reports indicate soil temperatures of 5-8°C are suitable for planting quinoa in the spring (Galwey, 1989). Darwinkel and Stølen (1997) recommend soil temperatures exceeding 10°C for planting and notes that lower temperatures inhibit proper germination and establishment.
In a germination experiment with the Danish cultivar Olav, temperatures of 6°C inhibited germination, resulting in only 25% of germination found at 20°C. Additionally, at low temperatures, the date of harvest became a significant factor. For seed that had been harvested early, 0% germination was seen, increasing to a maximum of 45% with later harvest time
(Jacobsen et al., 1999a)
Christiansen et al. (1999) investigated five Peruvian quinoa lines for their ability to germinate at low temperatures. In contrast to the results reported by Jacobsen, high germination was seen at temperatures as low as 2°C, indicating that variation exists in quinoa for minimum germination temperature.
For many areas, sufficiently high soil temperatures will be determining factor for planting date. As Jacobsen's temperature germination study investigated only one cultivar with a Chilean lowland background, a more detailed look at a wide range of Chilean lowland cultivars for their ability to germinate at lower temperatures could allow for earlier planting. If this trait is not successfully located, this trait could potentially be brought in from non-Chilean cultivars. Earlier
31
planting would mean earlier establishment, enhanced competitiveness with weeds, and early harvest time.
14. Maturity and Harvesting
Days to maturity is a critical factor for the successful growth of quinoa. The period varies widely. In Colorado, maturation time ranged from 90-125 days (Johnson and Croissant, 1985). In
Greece, maturation time was shown to vary from 110-116 days compared to the 150 days to maturity noted in Denmark. Within the same location, maturity time has been shown to vary widely from year to year (Gesinksi, 2000; Jacobsen, 2003). The wide range of maturity time to location and environmental conditions emphasizes the importance of trialing quinoa in new locations.
Quinoa can be harvested by combine. However, adjustments are needed for the small seed size and large stems. In Colorado, quinoa harvest recommendations are similar to sorghum.
Combining does not result in a totally clean product and further processing is necessary after combining. Recommendations include the use of a fanning mill and gravity separator (Oelke et al., 1992). Darwinkel and Stølen (1997) also repeat the necessity of cleaning after combining, although they note that the percentage of chaff can be kept under 5% if done under optimum field conditions with a properly adjusted combine.
Jacobsen et al. (1994) compared the efficacy of swathing compared to combining and found no statistically significant differences between the two methods. Swathing did allow for harvest earlier in the season than combining. This may lend it an advantage in situations where early maturity is important.
32
In Colorado, steady yields of 1340 kg ha-1 were reported over a three-year period
(Johnson and Croissant, 1990). With proper management, however, this can increase to over
2000 kg ha-1 (Oelke et al., 1992). In Saskatchewan, yield is reported to range from 840-1400 kg ha-1, although complete losses and yields exceeding 2200 kg ha-1 can occur (Alberta Agriculture,
Food, and Rural Business, 2005).
Reported yields in Europe exceed those of Colorado. Quinoa yields in Denmark typically range from 2,000-3,000 kg ha-1 (Jacobsen et al., 2010). Under two field experiments in England, the variety Baer yielded 5.14 t ha-1 (Risi and Galwey, 1991a). In Southern Italy, variety Regalona
Baer yielded 3.42 t ha-1 and 3.00 t ha-1 over two years (Pulvento et al., 2010). In Chile, the variety Baer is reported to produce yields of 3000 kg ha-1 under field conditions and 6500 kg ha-1 under experimental conditions (Baer, as cited in Delatorre-Herrera, 2003).
This wide variation in yield indicates the large effect of location and cultivar on the performance of quinoa. As yield figures from only two locations in temperate North America are available, variety testing will help illuminate yield potential for new quinoa growing areas.
15. Saponins
Many varieties of quinoa contain saponins in the pericarp of the seed. Saponins give quinoa a bitter taste and make it unpalatable. Additionally, saponins have potential negative health effects and have been shown to be disruptive to intestinal membranes in rats (Gee et al.,
1993). For quinoa to be marketable, saponins must be removed after harvest. This is generally done through abrasion or by washing the saponin from the seed (Johnson and Ward, 1993).
Alternatively, brushing has been used as a saponin removal method (Darwinkel and Stølen,
33
1997). Removal of the saponin through abrasion can result in losses of minerals such as calcium
(Konishi et al., 2004).
While industrial uses for quinoa saponin have been proposed (Jacobsen, 2003), the presence of saponins currently remains an obstacle rather than an opportunity. The required infrastructure for removal of saponins remains a challenge for small-scale quinoa growers.
Commercial production of quinoa in Colorado only began once machinery to remove saponin was obtained (Johnson, 1990).
Saponin free varieties of quinoa exist, though they are not found in Chilean lowland material. The Dutch quinoa program successfully introgressed the saponin-free trait into improved varieties (Limburg and Mastebroek, 1997). The presence of saponin is both qualitatively and quantitatively controlled. Its presence is governed by a single gene, the recessive allele of which results in seeds lacking saponin in the pericarp (Ward, 2001). When the dominant phenotype is expressed, saponin levels are then governed quantitatively. Efforts to breed low saponin varieties quantitatively have resulted in little success (Ward, 2000). Future efforts to develop saponin-free varieties will likely rely on generating homozygous recessive individuals for the qualitative trait. However, this comes with the potential drawback of saponin production being restored through cross-pollination from other varieties.
While the removal of saponin remains an obstacle for quinoa production, saponin free varieties have drawbacks. Reports from South America and from Europe indicate that saponin free varieties can suffer grain losses, some quite severe, due to feeding by birds (Risi and
Galwey, 1991a; Rasmussen et al., 2003; Darwinkel and Stølen, 1997). Losses to birds may also
34
be a problem with saponin containing varieties, as rain has been reported to wash away saponins
(Oelke et al., 1992).
Drought and salinity can have significant effects on saponin levels. Irrigation with saline water resulted in a 30% increase in saponins compared to freshwater irrigation. At the lowest irrigation level, saponins levels decreased 42% in comparison to the full irrigation control
(Gómez-Caravaca et al., 2012).
16. Conclusion
Expanded production of quinoa in temperate areas of North America holds many challenges and opportunities. Despite the main obstacle of heat susceptibility, breeding efforts utilizing within-species diversity may result in cultivars with improved heat tolerance. Should that prove insufficient, the North American species Chenopodium berlandieri lies within quinoa's secondary gene pool and may prove to be a promising source for introgression of heat tolerance genes. Existing genetic diversity has been identified that should allow for challenges such as downy mildew, saponins, and pre-harvest sprouting to be overcome. Many unknowns remain for disease and pest pressures due to quinoa's novelty to North America and its isolation from pest pressures found in its native range of cultivation. Quinoa's high level of abiotic stress tolerance gives it a distinct advantage on areas of marginal agricultural land, specifically in areas affected by salinity. Interest and demand for domestically produced quinoa remains high, and with further research and breeding efforts, there is considerable potential for expanded quinoa production in North America.
35
References
Abdin, M.Z., K.C. Bansal, and Y.P. Abrol. 1996. Effect of split nitrogen application on growth and yield of wheat (T. aestivum L.) genotypes with different N-assimilation potential. J. Agron. Crop Sci. 176(2): 83–90.
Adolf, V.I., S. Shabala, M.N. Andersen, F. Razzaghi, and S.E. Jacobsen. 2012. Varietal differences of quinoa’s tolerance to saline conditions. Plant Soil. 357: 1–13.
Aguilar, P.C., and S.E. Jacobsen. 2003. Cultivation of quinoa on the Peruvian altiplano. Food Reviews International. 19: 31–41.
Alberta - Agriculture and Rural Development. 2004. Salinity Classification, Mapping and Management in Alberta. Available at http://www1.agric.gov.ab.ca/$department/deptdocs.nsf/all/sag3267#prairies (verified 15 January 2013). Alberta – Agriculture and Rural Development.
Alberta Agriculture, Food and Rural Business. Ag-Entrepreneurship Division. 2005. Quinoa ...: the next cinderella crop for Alberta? Alberta Agriculture, Food and Rural Development, Edmonton, Alberta.
Aufhammer, W., H. Kaul, M. Kruse, J. Lee, and D. Schwesig. 1994. Effects of sowing depth and soil conditions on seedling emergence of amaranth and quinoa. Eur. J. Agron. 3: 205– 210.
Baer Von, E. Unpublished data. 1999.
Ballon, Emigdio. 1990. Low-input agriculture in the Southwestern United States. General Technical Report RM-GTR-198. Rocky Mountain Forest and Range Experiment Station, Fort Collins, CO.
Beckman, P.M. 1980. Seed rot and damping-off of Chenopodium quinoa caused by Sclerotium rolfsii. Plant Dis. 64: 497.
Benlhabib, O., M. Atifi, E.N. Jellen, and S. Jacobsen. 2004. The introduction of a new Peruvian crop “quinoa” to a rural community in Morocco. In 8 ESA Congress, 2004, Copenhagen, Denmark : [European agriculture in a global context] : book of proceedings. 11-15 July 2004. Samfundslitteratur, Frederiksberg, Denmark. 881-884.
Bertero, H.D. 2003. Response of developmental processes to temperature and photoperiod in quinoa (Chenopodium quinoa Willd.). Food Rev Intern. 19: 87–97.
Bhardwaj, H.L., A. Hankins, T. Mebrahtu, J. Mullins, M. Rangappa, O. Abaye, and G.E. Welbaum. 1996. Alternative crops research in Virginia. p. 87–96. In Janick, J. (ed.), Progress in new crops. ASHS Press, Alexandria, VA.
36
Bilalis, D., I. Kakabouki, A. Karkanis, I. Travlos, V. Triantafyllidis, and D. Hela. 2012. Seed and saponin production of organic quinoa (Chenopodium quinoa Willd.) for different tillage and fertilization. Not Bot. Horti Agrobot. Cluj-Napoca. 40: 42–46.
Bonifacio, Alejandro. 1995. Interspecific and Integeneric Hybridization in Chenopod Species. MS Thesis. Brigham Young University. Provo, Utah.
Bonifacio, Alejandro. 2004. Genetic variation in cultivated and wild chenopodium species for quinoa breeding. PhD Dissertation. Brigham Young University. Provo, Utah.
Boursier, P., and A. Läuchli. 1990. Growth responses and mineral nutrient relations of salt- stressed sorghum. Crop Sc. 30: 1226–1233.
Carlsson, R., P. Hanczakowski, and T. Kaptur. 1984. The quality of the green fraction of leaf protein concentrate from Chenopodium quinoa Willd. grown at different levels of fertilizer nitrogen. Anim. Feed. Sci. Tech. 11(4): 239–245.
Carmen, M.L. 1984. Acclimatization of quinoa (Chenopodium quinoa Willd.) and canihua (Chenopodium pallidicaule Allen) to Finland. Annal. Agric. Fenniae. 23: 135–144.
Ceccato, D.V., H. Daniel Bertero, and D. Batlla. 2011. Environmental control of dormancy in quinoa (Chenopodium quinoa) seeds: two potential genetic resources for pre-harvest sprouting tolerance. Seed Science Research. 21(2): 133.
Choi, Y.-J., S. Danielsen, M. Lübeck, S.-B. Hong, R. Delhey, and H.-D. Shin. 2010. Morphological and molecular characterization of the causal agent of downy mildew on quinoa (Chenopodium quinoa). Mycopath. 169: 403–412.
Christiansen, J.L., S.-E. Jacobsen, and S.T. Jørgensen. 2010. Photoperiodic effect on flowering and seed development in quinoa (Chenopodium quinoa Willd.). Acta Agriculturae Scandinavica, Section B - Soil & Plant Sci. 60: 539–544.
Christiansen, J.L., E.N. Ruiz-Tapia, B. Jornsgard, and S.-E. Jacobsen. 1999. Fast seed germination of quinoa (Chenopodium quinoa) at low temperature. In Proceedings of COST 814-Workshop: Alternative Crops for Sustainable Agriculture, 13-15 June 1999, Turku, Finland: 220–225.
Cranshaw, W.S., B.C. Kondratieff, and T. Qian. 1990. Insects Associated with Quinoa, Chenopodium quinoa, in Colorado. J. Kans. Entomol. Soc. 63: 195–199.
Cusack, D. 1984. Quinua: Grain of the Incas. Ecol. 14: 21–31.
Danielsen, S., V.H. Mercado, T. Ames, and L. Munk. 2004. Seed transmission of downy mildew (Peronospora farinosa f.sp. chenopodii) in quinoa and effect of relative humidity on seedling infection. Seed Sci Tech. 32: 91–98.
37
Danielsen, S., and L. Munk. 2004. Evaluation of disease assessment methods in quinoa for their ability to predict yield loss caused by downy mildew. Crop Prot. 23: 219–228.
Danielson, S., S.-E. Jacobsen, and J. Hockenhull. 2002. First report of downy mildew of quinoa caused by Peronospora farinosa f. sp. chenopodii in Denmark. Plant Dis. 86: 1175.
Darwinkel, A., and O. Stølen. 1997. Understanding the quinoa crop: Guidelines for growing in temperate regions of N.W. Europe. European Commission, Brussels.
Delatorre-Herrera, J. 2003. Current use of quinoa in Chile. Food Rev Intern. 19: 155–165.
Delatorre-Herrera, J., and M. Pinto. 2009. Importance of ionic and osmotic components of salt stress on the germination of four quinua (Chenopodium quinoa Willd.) selections. Chilean Journal of Agricultural Research. 69(4): 477–485.
Dillehay, T.D., J. Rossen, T.C. Andres, and D.E. Williams. 2007. Preceramic adoption of peanut, squash, and cotton in northern Peru. Science. 316(5833): 1890–1893.
Dirección Meteorológica de Chile. Anuarios Climatológicos 1920-2009. Dirección Meteorológica de Chile. Available at http://164.77.222.61/climatologia/php/menuAnuarios.php (verified 23 March 2013).
FAO. 2013. International Year of Quinoa IYQ-2013. Available at http://www.rlc.fao.org/en/about-fao/iyq-2012/ (accessed 11 March 2013; verified 11 March 2013). Food and Agriculture Organization of the United Nations. Regional Office for Latin America and the Caribbean. Vitacura, Santiago, Chile.
Flynn, R.P. 1990. MS Thesis. Colorado State University. Fort Collins, CO.
Fuentes, F., and A. Bhargava. 2011. Morphological analysis of quinoa germplasm grown under lowland desert conditions. J of Agron Crop Sci. 197: 124–134.
Fuentes, F.F., E.A. Martinez, P.V. Hinrichsen, E.N. Jellen, and P.J. Maughan. 2008. Assessment of genetic diversity patterns in Chilean quinoa (Chenopodium quinoa Willd.) germplasm using multiplex fluorescent microsatellite markers. Conserv Gen. 10: 369–377.
Galwey, N.W. 1989. Exploited plants - Quinoa. Biologist. 36: 267–274.
Gandarillas, H. 1982. Quinoa production. IBTA-CIID. (Trans. by Sierra-Blanca Assoc., Denver, CO. 1985.)
Gee, J.M., K.R. Price, C.L. Ridout, G.M. Wortley, R.F. Hurrell, and I.T. Johnson. 1993. Saponins of quinoa (Chenopodium quinoa): Effects of processing on their abundance in quinoa products and their biological effects on intestinal mucosal tissue. J Food Sci Agr. 63: 201–209.
38
Geerts, S., R.S. Mamani, M. Garcia, and D. Raes. 2006. Response of quinoa (Chenopodium quinoa Willd.) to differential drought stress in the Bolivian Altiplano: Towards a deficit irrigation strategy within a water scarce region. In Proceedings of the 1st International Symposium on Land and Water Management for Sustainable Irrigated Agriculture, 4-8 April, Adana, Turkey.
Gesinksi, K. 2000. Potential for Chenopodium quinoa acclimitisation in Poland. In Abstracts/Proceedings of COST 814 Conference, Crop Development for Cool and Wet Regions of Europe, 10-13 May, 2000, Pordenone, Italy: 547–552.
Gómez-Caravaca, A.M., G. Iafelice, A. Lavini, C. Pulvento, M.F. Caboni, and E. Marconi. 2012. Phenolic compounds and saponins in quinoa samples (Chenopodium quinoa Willd.) grown under different saline and nonsaline irrigation regimens. Journal of Agr Food Chem. 60: 4620–4627.
Gómez-Pando, L.R., R. Álvarez-Castro, and A. Eguiluz-de la Barra. 2010. SHORT COMMUNICATION: Effect of salt stress on Peruvian germplasm of Chenopodium quinoa Willd.: A promising crop. J Agr Crop Sci. 196: 391–396.
González, Juan A., M. Gallardo, M. Hilal, M. Rosa, and F.E. Prado. 2009. Physiological responses of quinoa (Chenopodium quinoa Willd.) to drought and waterlogging stresses: dry matter partitioning. Bot Studies. 50: 35–42.
Hariadi, Y., K. Marandon, Y. Tian, S.E. Jacobsen, and S. Shabala. 2011. Ionic and osmotic relations in quinoa (Chenopodium quinoa Willd.) plants grown at various salinity levels. J. Expt. Bot. 62: 185–193.
Hugh D. Wilson. 1980. Artifical hybridization among species of Chenopodium sect. Chenopodium. Syst Bot. 5: 253–263.
Hugh D. Wilson. 1978. Chenopodium quinoa Willd.: Variation and relationships in southern South America. Nat. Geog. Soc. Res. Rep. 19: 711–721.
Iliadis, C., T. Karyotis, S.E. Jacobsen, S.E. Jacobsen, and Z. Portillo. 2001. Adaptation of quinoa under xerothermic conditions and cultivation for biomass and fibre production. Memorias, Primer Taller Internacional sobre Quinua—Recursos Geneticos y Sistemas de Producción, 10-14 May, 1999, Lima, Peru: 371–378.
Jacobsen, S.E. 1997. Adaptation of quinoa (Chenopodium quinoa) to Northern European agriculture: studies on developmental pattern. Euphytica. 96: 41–48.
Jacobsen, S.E. 1999. Potential for quinoa (Chenopodium quinoa Willd.) for cool and wet regions of Europe. In Proceedings of COST 814-Workshop: Alternative Crops for Sustainable Agriculture, 13-15 June 1999, Turku, Finland: 87–99.
39
Jacobsen, S.E. 2003. The worldwide potential for quinoa (Chenopodium quinoa Willd.). Food Rev. Intern. 19: 167–177.
Jacobsen, S.E. 2007. Quinoa’s world potential. p. 109–122. In Ochatt, S., Jain, S.M. (eds.), Breeding of neglected and under-utilized crops, spices and herbs. Science Publishers. Enfield, NH.
Jacobsen, S.E. 2011. The situation for quinoa and its production in southern Bolivia: from economic success to environmental disaster. J Ag Crop Sci. 197: 390–399.
Jacobsen, S.E., and A.P. Bach. 1998. The influence of temperature on seed germination rate in quinoa (Chenopodium quinoa Willd.). Seed Sci and Tech. 26: 515–524.
Jacobsen, S.E., J.L. Christiansen, and J. Rasmussen. 2010. Weed harrowing and inter-row hoeing in organic grown quinoa (Chenopodium quinoa Willd.). Outlook on Ag. 39: 223–227.
Jacobsen, S.E., I. Joergensen, and O. Stolen. 1994. Cultivation of quinoa (Chenopodium quinoa) under temperate climatic conditions in Denmark. J Agr Sci. 122(1): 47.
Jacobsen, S.E., B. Jornsgard, J.L. Christiansen, and O. Stolen. 1999a. Effect of harvest time, drying technique, temperature and light on the germination of quinoa (Chenopodium quinoa). Seed Sci and Tech. 27: 937–944.
Jacobsen, S.E., C. Monteros, J.L. Christiansen, L.A. Bravo, L.J. Corcuera, and A. Mujica. 2005. Plant responses of quinoa (Chenopodium quinoa Willd.) to frost at various phenological stages. Eur. J. Agron. 22: 131–139.
Jacobsen, S.E., C. Monteros, L.J. Corcuera, L.A. Bravo, J.L. Christiansen, and A. Mujica. 2007. Frost resistance mechanisms in quinoa (Chenopodium quinoa Willd.). European journal of agronomy. 26(4): 471–475.
Jacobsen, S.E., A. Mujica, and C.R. Jensen. 2003. The resistance of quinoa (Chenopodium quinoa Willd.) to adverse abiotic factors. Food Rev. Intern. 19: 99–109.
Jacobsen, S.E., H. Quispe, and A. Mujica. 1999b. Quinoa: an alternative crop for saline soils in the Andes. In Scientists and farmer–partners in research for the 21st century. CIP Program Report 1999-2000: 403–408.
Jacobsen, S.E., and O. Stolen. 1993. Quinoa - Morphology, phenology and prospects for its production as a new crop in Europe. Eur. J. Agron. 2: 19–29.
Jellen, E.N., B.A. Kolano, M.C. Sederberg, A. Bonifacio, and P.J. Maughan. 2011. Chenopodium. p. 35–61. In Kole, C. (ed.), Wild Crop Relatives: Genomic and Breeding Resources. Springer. Berlin, Heidelberg.
Jensen, C.R., S.E. Jacobsen, M.N. Andersen, N. Núñez, S.D. Andersen, L. Rasmussen, and V.O.
40
Mogensen. 2000. Leaf gas exchange and water relation characteristics of field quinoa Chenopodium quinoa Willd.) during soil drying. Eur. J. Agron.13: 11–25.
Johnson, D.L. 1990. New Grains and Pseudograins. p. 122–127. In Janick, J., Simon, J.E. (eds.), Advances in new crops. Timber Press, Portland, OR.
Johnson, D.L., and Croissant, R.L. 1990. Alternate crop production and marketing in Colorado. Technical Bulletin LTB90-3. Coop Extension, Colorado State University, Fort Collins, CO.
Johnson, D.L., and Croissant, R.L. 1985. Quinoa production in Colorado. Service-In-Action No. 112. Colorado State University Coop Ext, Fort Collins, CO.
Johnson, D.L., and Ward, Sarah. 1993. Quinoa. p. 219–221. In Janick, J., Simon, J.E. (eds.), New crops. Wiley, New York.
Karyotis, T.H., C. Iliadis, C.H. Noulas, and T.H. Mitsibonas. 2003. Preliminary research on seed production and nutrient content for certain quinoa varieties in a saline–sodic soil. J Agron Crop Sci. 189: 402–408.
Keller, L.P., G.J. McCarthy, and J.L. Richardson. 1986. Mineralogy and stability of soil evaporites in North Dakota. Soil Sci. Soc. Am. J. 50: 1069–1071.
Konishi, Y., S. Hirano, H. Tsuboi, and M. Wada. 2004. Distribution of minerals in quinoa (Chenopodium quinoa Willd.) seeds. Biosc. Biotech Biochem. 68: 231–234.
Koyro, H.-W., and S.S. Eisa. 2007. Effect of salinity on composition, viability and germination of seeds of Chenopodium quinoa Willd. Plant Soil 302: 79–90.
Kumar, A., A. Bhargava, S. Shukla, H.B. Singh, and D. Ohri. 2006. Screening of exotic Chenopodium quinoa accessions for downy mildew resistance under mid-eastern conditions of India. Crop Prot. 25: 879–889.
Kumpun, S., A. Maria, S. Crouzet, N. Evrard-Todeschi, J.-P. Girault, and R. Lafont. 2011. Ecdysteroids from Chenopodium quinoa Willd., an ancient Andean crop of high nutritional value. Food Chem. 125: 1226–1234.
Limburg, H., and H.D. Mastebroek. 1997. Breeding high yielding lines of Chenopodium quinoa Willd. with saponin free seed. 103-114. In Stolen, O., K. Bruhn, K. Pithan & J. Hill (Eds.). Proc COST 814 Workshop on Small Grain Cereals and Pseudo-cereals, Copenhagen, Denmark, 22-24 February 1996.
Maas, E.V. 1986. Salt tolerance of plants. Applied Ag. Res.1: 12–26.
Martínez, E.A., E. Veas, C. Jorquera, R. San Martín, and P. Jara. 2009. Re-Introduction of quínoa
41
into arid Chile: Cultivation of two lowland races under extremely low irrigation. J Agron Crop Sci. 195: 1–10.
Mastebroek, H.D., and H. Limburg. 1997. Breeding harvest security in Chenopodium quinoa. 79-86. In Stolen, O., K. Bruhn, K. Pithan & J. Hill (Eds.). Proceedings of the COST 814 Workshop on Small Grain Cereals and Pseudo-cereals, Copenhagen, Denmark, 22-24 February 1996.
Mastebroek, H.D., and R. van Loo. 2000. Breeding of Quinoa - state of the art. 491–496. In: Abstracts/Proceedings of COST 814 Conference, Crop Development for Cool and Wet Regions of Europe, Pordenone, Italy, 10-13 May, 2000.
Mujica, A. 1977. Tecnología del cultivo de la quinua. Fondo Simón Bolivar. Ministerio de Alimentación. Zona Agraria XII. IICA, UNTA 1977. Puno, Perú.
Murphy, K.M., K.G. Campbell, S.R. Lyon, and S.S. Jones. 2007. Evidence of varietal adaptation to organic farming systems. Field Crops Res. 102: 172–177.
Narrea R., A. 1976. Cultivo de la Quinua. Bol. No. 5. Dirección General de Producción, Ministerio de Alimentación, Lima, Peru.
National Research Council (U.S.). Advisory Committee on Technology Innovation. 1989. Lost crops of the Incas : little-known plants of the Andes with promise for worldwide cultivation. National Academy Press, Washington, D.C.
NOAA - National Climatic Data Center. 2011. Annual mean extreme maximum temperature. NCDC: CLIMAPS. Available at http://hurricane.ncdc.noaa.gov/climaps/tmp13a13.pdf (verified 23 March 2013).
Oelke, E.A., D.H. Putnam, T.M. Teynor, and E.S. Oplinger. 1992. Quinoa. In Alternative Field Crops Manual. University of Wisconsin Cooperative Extension Service, University of Minnesota Extension Service, Center for Alternative Plant & Animal Products.
Pulvento, C., M. Riccardi, A. Lavini, R. d’ Andria, G. Iafelice, and E. Marconi. 2010. Field Trial Evaluation of two Chenopodium quinoa genotypes grown under rain-fed conditions in a typical Mediterranean environment in south Italy. J Agron Crop Sci. 196: 407–411.
Pulvento, C., M. Riccardi, A. Lavini, G. Iafelice, E. Marconi, and R. d’ Andria. 2012. Yield and quality characteristics of quinoa grown in open field under different saline and non-saline irrigation regimes. J Agron Crop Sci. 198: 254–263.
Rasmussen, C., A. Lagnaoui, and P. Esbjerg. 2003. Advances in the knowledge of quinoa pests. Food Rev Intern. 19: 61–75.
Razzaghi, F., F. Plauborg, S.-E. Jacobsen, C.R. Jensen, and M.N. Andersen. 2012. Effect of
42
nitrogen and water availability of three soil types on yield, radiation use efficiency and evapotranspiration in field-grown quinoa. Ag Water Mgmt. 109: 20–29.
Repo-Carrasco, R., C. Espinoza, and S.E. Jacobsen. 2003. Nutritional value and use of the Andean crops quinoa (Chenopodium quinoa) and kañiwa (Chenopodium pallidicaule). Food Rev Intern. 19: 179–189.
Repo-Carrasco-Valencia, R., J.K. Hellström, J.M. Pihlava, and P.H. Mattila. 2010. Flavonoids and other phenolic compounds in Andean indigenous grains: Quinoa (Chenopodium quinoa), kañiwa (Chenopodium pallidicaule) and kiwicha (Amaranthus caudatus). Food Chem. 120: 128–133.
Risi, J.C., and N.W. Galwey, N.W. 1984. The Chenopodium grains of the Andes - Inca crops for modern agriculture. Adv. Appl. Biol.10: 145–216.
Risi, J.C., and N.W. Galwey. 1989. The pattern of genetic diversity in the Andean grain crop quinoa (Chenopodium quinoa Willd). I. Associations between characteristics. Euphytica. 41: 147–162.
Risi, J.C., and N.W. Galwey. 1991a. Genotype x environment interaction in the Andean grain crop quinoa (Chenopodium quinoa) in temperate environments. Plant breed. 107: 141– 147.
Risi, J.C., and N.W. Galwey. 1991b. Effects of sowing date and sowing rate on plant development and grain yield of quinoa (Chenopodium quinoa) in a temperate environment. J. Agric. Sci. 117: 325.
Romero, S., and S. Shahriari. 2011. A food’s global success creates a quandary at home. The New York Times. 20 Mar. 6.
Royo, A., R. Aragüés, E. Playán, and R. Ortiz. 2000. Salinity–Grain yield response functions of barley cultivars assessed with a drip-injection irrigation system. Soil Sci. Soc. Am. J. 64: 359.
Ruiz-Carrasco, K., F. Antognoni, A.K. Coulibaly, S. Lizardi, A. Covarrubias, E.A. Martínez, M.A. Molina-Montenegro, S. Biondi, and A. Zurita-Silva. 2011. Variation in salinity tolerance of four lowland genotypes of quinoa (Chenopodium quinoa Willd.) as assessed by growth, physiological traits, and sodium transporter gene expression. Plant Physiol. Biochem. 49: 1333–1341.
Schlick, G., and D.L. Bubenheim. 1996. Quinoa: Candidate crop for NASA’s controlled ecological life support systems. p. 632–640. In Janick, J. (ed.). Progress in new crops. ASHS Press, Arlington, VA.
Schulte auf’m Erley, G., H.-P. Kaul, M. Kruse, and W. Aufhammer. 2005. Yield and nitrogen
43
utilization efficiency of the pseudocereals amaranth, quinoa, and buckwheat under differing nitrogen fertilization.Eur. J. Agron. 22: 95–100.
Sigsgaard, L., S.E. Jacobsen, and J.L. Christiansen. 2008. Quinoa, Chenopodium quinoa, provides a new host for native herbivores in Northern Europe: Case studies of the moth, Scrobipalpa atriplicella, and the Tortoise Beetle, Cassida nebulosa.J Insect Sci. 8: 1–4.
Small, E. 1999. New crops for Canadian agriculture. p. 15–52. In Janick, J. (ed.), Perspectives on new crops and new uses. ASHS Press, Alexandria, VA.
Smith, B.D. 1985. Chenopodium berlandieri ssp. jonesianum: Evidence For a Hopewellian Domesticate From Ash Cave, Ohio. Southeastern Archaeology. 4: 107–133.
Smith, L.H., C.H. Dwyer, F.K. Nunns, G.M. Schafer, H. Olsen, and D.W. Klauss. 1958. Soil survey, Yakima County, Washington. United States Department of Agriculture, Washington, D.C.
Tapia, M.E. 1979. Historia y distribución geográfica. In M.E. Tapia (ed.). “Quinoa y Kañiwa. Cultivos Andinos” Serie Libros y Materiales Educativos No. 49. pp. 11-15. Instituto Interamericano de Ciencias Agrícolas, Bogota, Colombia.
Tapia, M.E., S. Mujica, and A. Canahua. 1980. Origen distribución geográfica y sistemas de producción en quinua. In “Primera Reunion sobre Genética y tomejoramiento de la Quinua.” pp. A1-A8. Universidad Nacional Técnica del Altiplano, Instituto Boliviano de Tecnología Agropecuaria, Instituto Interamericano de Ciencias Agrícolas, Centro de Investigación Internacional para el Desarrollo, Puno, Peru.
Testen, A.L., J.M. McKemy, and P.A. Backman. 2012. First report of quinoa downy mildew caused by Peronospora variabilis in the United States. Plant Dis. 96: 146–146.
Tewari, J.P., and S.M. Boyetchko. 1990. Occurrence of Peronospora farinosa f. sp. chenopodii on quinoa in Canada. Can Plant Dis Surv. 70: 127–128.
Thanapornpoonpong, S. 2004. Effect of nitrogen fertilizer on nitrogen assimilation and seed quality of amaranth (Amaranthus spp.) and quinoa (Chenopodium quinoa Willd.). PhD Dissertation. Georg-August University of Göttingen. Göttingen, Germany.
Torres, Julian Donoso. 1955. Comparative study of resistance of alfalfa, brome grass and quinoa seedlings to synthetic saline soils, saline alkali and alkali soils. M.S. Thesis. Cornell University. Ithaca, NY.
USDA. 1989. The second RCA appraisal: soil, water, and related resources on nonfederal land in the United States. U.S. Government Printing Office, Washington, D.C.
44
USDA. 2011. 2011 RCA Appraisal. USDA, Washington, D.C. van Schooten, H.A., and J.B. Pinxterhuis. 2003. Quinoa as an alternative forage crop in organic dairy farming. 445-448. In Kirilov, A., Todorov, N., Katerov, I. (eds.), Optimal forage systems for animal production and the environment : proceedings of the 12th symposium of the European Grassland Federation, Pleven, Bulgaria, 26-28 May 2003. British Grassland Society.
Vega-Gálvez, A., M. Miranda, J. Vergara, E. Uribe, L. Puente, and E.A. Martínez. 2010. Nutrition facts and functional potential of quinoa (Chenopodium quinoa willd.), an ancient Andean grain: a review. J Sci Food and Agric. 90: 2541–2547.
Vera, R.R. 2006. Chile. Country/Pasture/Forage Resource Profiles. Available at http://www.fao.org/ag/AGP/AGPC/doc/Counprof/Chile/cile.htm.
Ward, S.M. 1994. Developing improved quinoa varieties for Colorado. PhD Dissertation. Colorado State University. Fort Collins, CO.
Ward, S.M. 2000. Response to selection for reduced grain saponin content in quinoa (Chenopodium quinoa Willd.). Field Crops Research. 68(2): 157–163.
Ward, S.M. 2001. A recessive allele inhibiting saponin synthesis in two lines of Bolivian quinoa (Chenopodium quinoa Willd.). J Heredity. 92: 83–86.
Warne, P., R.D. Guy, L. Rollins, and D.M. Reid. 1990. The effects of sodium sulphate and sodium chloride on growth, morphology, photosynthesis, and water use efficiency of Chenopodium rubrum. Can J. Bot. 68: 999.
Wilson, H.D, and J. Manhart. 1993. Crop/weed gene flow: Chenopodium quinoa Willd. and C. berlandieri Moq. Theor Appl Gen. 86: 642–648.
45
Figure 1: Pre-harvest sprouting in quinoa.
46
CHAPTER TWO
TOLERANCE OF LOWLAND QUINOA CULTIVARS TO SODIUM CHLORIDE AND SODIUM SULFATE SALINITY I: EFFECTS ON YIELD, HEIGHT, AND LEAF GREENNESS
Tolerance Of Lowland Quinoa Cultivars To Sodium Chloride And Sodium Sulfate Salinity I: Effects On Yield, Height, And Leaf Greenness
Abstract
As soil salinity increases as a major agriculture problem worldwide, halophytic crops such as quinoa may provide a route of adaptation to these marginal soil conditions. Until recently, little research had been conducted on the relative levels of salinity tolerance of Chilean lowland cultivars, which are best adapted to temperate latitudes and high summer temperatures characteristic of many saline affected regions. In this study, four Chilean lowland cultivars and the salt-tolerant barley cultivar “Albacete” were exposed to a no-salt control and 8, 16, and 32 dS
-1 m of NaCl and Na2SO4 combined with two levels of fertilization. Yield, plant height, and leaf greenness were measured. Quinoa exhibited greater tolerance to sodium sulfate applications than to sodium chloride, and demonstrated greater saline tolerance than barley at 32 dS m-1 NaCl and
Na2SO4. Additionally, high fertilization had a detrimental effect on yield, with the notable exception of variety UDEC-1 which increased in yield. Significant levels of variation were found in salinity tolerance to NaCl and Na2SO4 within the four quinoa cultivars.
1. Introduction
Soil salinity is a growing problem worldwide, affecting 20-50% of irrigated arable land
(Pitman and Läuchli, 2004). In the United States, 2.2 million hectares are affected by salinity and sodicity, with a further 30.8 million hectares at risk (USDA, 2011). Quinoa has gathered much attention in recent years for its high level of salinity tolerance. It has been shown to produce seed under salinity levels as high as seawater and showed no reduction in yield when irrigated with a
48
1:1 mix of freshwater and approximated seawater (Koyro and Eisa, 2007; Hariadi et al., 2011;
Pulvento et al., 2012). All previous studies with quinoa have focused on quinoa’s response to various strengths of sodium chloride or to seawater or mixed salt solutions. However, many areas affected by salinity, such as the Upper Midwest, deal predominantly with sulfate salts (Keller et al., 1986). Some species, such as sorghum (Sorghum bicolor L. Moench) and Chenopodium rubrum, have shown greater sensitivity at high concentrations to sulfate salinity as compared to chloride salinity at isosmotic solutions (Weimberg et al., 1984; Boursier and Läuchli, 1990;
Warne et al., 1990). Other species show no difference, or are more tolerant of sulfate salinity at isosmotic or equal EC levels (Cramer and Spurr, 1986; Manchanda and Sharma, 1989). Grattan and Grieve (1999) note a trend of greater sulfate tolerance than chloride tolerance for many plants. The mechanisms causing differences in tolerance to chloride salts and sulfate salts are not well understood (Pitman and Läuchli, 2004). Characterizing differences in quinoa’s response to chloride and sulfate salinity could allow for a more precise use of the crop in agricultural land affected by soil salinity.
Before this study was conducted, there had been little investigation into the salinity tolerance of Chilean lowland cultivars. This group of cultivars is most adapted to cultivation at temperate latitudes (Bertero, 2003). Chilean lowland cultivars are also the most tolerant varieties of quinoa to heat stress, which is a major limiting factor for quinoa. Most saline affected soils are located in the western United States (Ghassemi et al., 1995), in locations that experience high summer temperatures. Therefore, investigating the relative salinity tolerance of Chilean lowland varieties will be key in developing quinoa as a successful halophytic crop for such areas.
49
The objectives of this experiment were to 1) characterize the relative salinity tolerance of a range of Chilean cultivars, 2) determine the differences in tolerance of quinoa to NaCl versus
Na2SO4 salinity and 3) determine if fertility level had any interaction with salinity type or concentration.
2. Materials and Methods
2.1 Experimental design
A greenhouse study with a split-split-plot design was conducted. Factors included variety/species (four quinoa varieties and one barley variety); fertility, at two levels; and seven salinity levels. Three subsamples, each representing a single plant, were grown for each treatment combination in each of the three replications. The experiment was conducted twice in two separate greenhouse rooms. The first run was planted September 10, 2011 and the second on
October 7, 2011.
The four quinoa varieties, Colorado 407D (PI 596293), UDEC-1 (PI 634923), Baer (PI
634918), and QQ065 (PI 614880) were of Chilean origin or background. UDEC-1, Baer, and
QQ065 originated from Chile and were chosen based on their latitude of origin within the country. Based on their GRIN passport data, UDEC-1 (34.63° N), Baer (38.70° N), and QQ065
(42.50° N) represent the northernmost, middle, and southernmost points of distribution of
Chilean lowland cultivars. This range of latitudes spans environmental gradients such as rainfall and increased salinity exposure (Ruiz-Carrasco et al., 2011). Colorado 407D, hereafter referred to as CO407D, is a variety released by Colorado State University and was bred from Chilean varieties. The barley variety, Albacete (PI 467780), is a Spanish cultivar that has had its salinity tolerance previously characterized and is considered relatively salt tolerant (Royo et al., 2000).
50
Seed from the four quinoa varieties was grown at the WSU Organic Farm the previous year, and the seed from Baer represents a single plant selection. The original seed was obtained from North
Central Regional Plant Introduction Station (Ames, IA). Seed for Albacete was provided by the
National Small Grains Collection (Aberdeen, ID).
Fertility was supplied by a mixture of alfalfa meal (3:1:3), monoammonium phosphate
(11:52:0), and feather meal (12:0:0). These were mixed to provide a per pot level of 1 g N, 0.29 g P, and 0.29 g K (P and K actual) at the low fertilization level, and 3 g N, 0.86 g P, and 0.86 g K
(P and K actual) at the high fertilization level. The relative levels of fertility were set to match a previous greenhouse experiment on quinoa’s response to fertilization (unpublished data).
Salinity levels included a no-salt control, 8 dS m-1 NaCl, 16 dS m-1 NaCl, 32 dS m-1
-1 -1 -1 NaCl, 8 dS m Na2SO4, 16 dS m Na2SO4, and 32 dS m Na2SO4. The tap water used to water pots had a measured EC of ~0.3 dS m-1, and EC rates were adjusted upward to compensate. For simplicity, the no-salt control will hereafter be referred to as 0 dS m-1.
Sunshine Mix #1 from Sun Gro Horticulture (Bellevue, WA) was used and added to pots in equal volumes. Seeds were planted and watered with tap water. After germination, salinity was applied over two periods. Saline solution was mixed in a large container, and its strength was measured with a HI 8633 handheld salinity meter (Hannah Instruments, Woonsocket, RI), which was calibrated to the temperature of the tap water. A predetermined volume of saline solution, measured to fully saturate the potting soil, was added in halves at 20 days after planting
(dap) and at 33 dap. By 20 dap, seedlings had become established. Plants were thinned to a single plant per pot at 27 dap.
51
Before the first addition of saline water, plastic trays were placed underneath the pots to
-1 control leaching and prevent the loss of salt. For pots under the 32 dS m Na2SO4 treatment, saline water was added in one addition. This caused overflow of saline water from the trays and loss of salt from the pot-tray system. Therefore, these pots were discarded. Throughout the duration of the experiment, plants were watered so that standing water was present in the trays.
This was done to ensure leached salts would not dry and accumulate in the trays. As water levels in the trays were variable and salts accumulated at the high water mark, the trays and pots were rinsed at 44-45 dap and the resulting water was added to the pots.
Plant heights were measured for all quinoa plants 70 dap. Heights were averaged from all surviving subsamples for each treatment combination. Leaf greenness was measured at 73 dap.
Three measurements were taken with a Konica Minolta SPAD-502 Plus chlorophyll meter
(Minolta Camera Co., Ltd., Osaka, Japan) on leaves from the top third of the plant. These three values were averaged to obtain an average value for each plant, and then all surviving subsamples were averaged to obtain an average measure of leaf greenness for that replicate of the given treatment combination.
Plants were harvested at maturity. Seeds were stripped by hand from the inflorescences and the resulting material was threshed in a wheat head thresher manufactured by Precision
Machine Company (Lincoln, NE). The threshed material was cleaned in a Clipper Office Tester
(Seedburo, Des Plaines, IL). Yields were averaged across surviving subsamples for each treatment combination.
2.2 Statistical Analysis
52
Data was analyzed using PROC MIXED in SAS (SAS Institute, Cary NC). Experiment repeats were treated as a random factor. Replication was nested within experiment repeat and was also included as a random factor. Variety/species, salinity, and fertilization were treated as fixed factors. Appropriate error terms were used. Percent yield declines were arcsine transformed to restore normality. Pearson correlation coefficients between height, leaf greenness, and yield were determined via PROC CORR (SAS Institute, Cary NC) using the means generated from
PROC MIXED.
3. Results
3.1 Yield
Salinity had a highly significant effect on yield. A highly significant interaction between variety and fertility complicated interpretation of these factors (Tables 1 & 2).
When salinity was examined across genotype, several trends appeared (Figure 1). The control was not found to significantly differ in yield from 8 dS m-1 NaCl, 16 dS m-1 NaCl, 8 dS
-1 -1 -1 m Na2SO4, and 16 dS m Na2SO4. At the highest concentration of 32 dS m , both NaCl and
-1 Na2SO4 resulted in a significant decline in yield. The decline for 32 dS m NaCl was higher than that of 32 dS/m Na2SO4.
As that analysis included both barley and quinoa, a separate analysis was run excluding barley yield data in order to obtain means applicable to quinoa (Figure 2). Several changes occurred under this modified analysis. Sodium sulfate was found to have no statistically significant effect on yield at any concentration when compared with the control. In contrast, sodium chloride had a significant detrimental impact on yield. Significant decreases in yield
53
were seen starting at 16 dS m-1 NaCl, where yield decreased 14.8%. At the highest concentration of 32 dS m-1 NaCl, yield decreased 45.4%.
No significant interaction was found between variety and salinity. This contrasts with what would be expected if quinoa and barley differed in salinity tolerance. However, the analysis that was run measured absolute declines as opposed to relative declines. Quinoa yields exceeded that of barley on a per plant basis. An equal gram per plant decrease in yield for both species could have resulted in total losses for barley but only partial yield decreases for quinoa.
Therefore, percent declines in yield from the no-salt control are more relevant when comparing the effects of salinity. Additionally, quinoa varieties exhibited large differences in yield under the no-salt control. Therefore, a given decrease in grams of yield per plant may represent a larger percent decrease for one variety than another. This makes relative declines also more relevant when comparing salinity tolerance between quinoa varieties.
To determine relative changes in yield under salinity, the means for variety at each salinity level were compared. Due to the decreased statistical power, only differences between
-1 the control and 32 dS m NaCl and Na2SO4 were statistically significant. A separate PROC
-1 MIXED analysis was run on the percent decline in yield at 32 dS m NaCl and Na2SO4 from the control. Significant variety effects were found (Table 3). This allowed determination of the statistical significance of relative decreases between varieties at these levels. Under 32 dS m-1
NaCl, Albacete, the barley cultivar, exhibited the greatest decrease by far at -98.0%. A significant range in responses was seen for the four quinoa cultivars. UDEC-1 and Baer had the lowest decreases of 43.7% and 49.3% respectively. CO407D declined by over half (-65.4%) but
54
did not differ significantly from all other quinoa varieties in yield decline. QQ065 was the most sensitive to NaCl salinity, decreasing in yield by 73.7% (Table 4).
-1 Under 32 dS m Na2SO4, fewer statistically significant differences appeared between
-1 varieties. This was partly due to the reduced statistical power as the 32 dS m Na2SO4 treatment had been discarded in one of the two experiment runs. Barley had the greatest decline at -82.4%, which was greater decline than all quinoa varieties except QQ065, which declined 51.9%.
UDEC-1 had the lowest decline (-10.8%) although this was only statistically different from
QQ065 and Albacete (Table 4). When relative decreases were compared between 32 dS m-1
NaCl and Na2SO4, 32 dS m-1 NaCl resulted in larger decreases for all varieties/species
(p<0.0009).
The highly significant interaction between variety and fertility was due to a difference in response to increased fertilization (Figure 3). Albacete, the barley cultivar, and quinoa varieties
CO407D, Baer, and QQ065 all showed significant decreases in yield under high fertilization compared to low fertilization. However, quinoa variety UDEC-1 increased in yield under high fertilization (Table 5; Figure 3).
3.2 Leaf greenness
Leaf greenness content was found to differ significantly by variety (p<0.0001), salinity
(p<0.0001), and fertility (p<0.0001) (Table 2). UDEC-1 was highest in leaf greenness, followed by Baer. CO407D and QQ065 ranked lowest in leaf greenness and did not differ significantly from each other (Figure 4).
55
High fertilization caused a noted decrease in leaf greenness (Figure 5). Leaf greenness under salinity decreased at all levels in comparison to the control. However, this varied depending on salt type. With NaCl, the decrease in leaf greenness increased under higher EC level. However, the impact of Na2SO4 did not vary with concentration and appeared steady across EC levels (Figure 6).
3.3 Plant height
Significant interactions between fertility and variety (p=0.018) and between salinity and fertility (p=0.004) complicated interpretation of plant height (Table 2). Despite these interactions, plants under high fertilization exhibited decreases in plant height.
A crossover interaction occurred between fertilizer and variety (Figure 7). Under low fertilization, CO407D (93 cm) and UDEC-1 (91 cm) were tallest, followed by Baer (83 cm) which ranked intermediate, and QQ065 (72 cm) which ranked shortest. UDEC-1 remained the tallest under high fertilization, while all other varieties did not differ significantly. Additionally, it had the lowest relative decline in height, by 31.6%.
The effect of salt type was influenced by fertilization. At low fertilization, height decreased for both salts as salinity increased. Under high fertilization, the trend of decreasing height held for NaCl. However, under Na2SO4 with high fertilization, height did not differ between salinity levels (Figure 8).
3.4 Correlations
56
Moderately strong correlations were found between all factors (Table 6). The three correlations were moderately strong and highly significant (p<0.0001). When correlations were run for individual combinations of salinity and fertility treatments, some particular patterns appeared. For both salts under high fertilization, yield, plant height, and leaf greenness were positively correlated, with the exception of leaf greenness and yield under Na2SO4, which exhibited a marginally significant correlation (p=0.053) (Tables 7 & 9).
Under low fertilization, only height and yield were significantly correlated with each other for NaCl (r=0.83) and for Na2SO4 (r=0.90) (Tables 8 & 10). In contrast to the situation under high fertilization, leaf greenness was not significantly correlated with either height or yield.
4. Discussion
4.1 Yield
When yields of both quinoa and barley were analyzed together, NaCl was found to have a much greater impact on yield than Na2SO4 in terms of both absolute yields and relative yields.
When only yields of the quinoa varieties were analyzed, yields under all levels of Na2SO4 were not significantly different from the control. This indicates that at equal EC levels, quinoa’s tolerance to Na2SO4 exceeds that for NaCl.
Sodium chloride began to show a detrimental impact on quinoa yield starting at 16 dS m-1
NaCl. This indicates that the salinity threshold after which quinoa yield declines lies somewhere between 8 dS m-1 and 16 dS m-1 NaCl. This contrasts with other findings, where two quinoa cultivars saw an increase in yield at 15 dS m-1 NaCl compared to the control, after which
57
decreases in yield occurred (Jacobsen et al., 1999). These cultivars originated from the Andean highlands, which might suggest lower salinity tolerance among Chilean lowland cultivars. Adolf et al. (2012) tested salinity tolerance of a wide range of quinoa germplasm under greenhouse conditions. Reactions to salinity were complex and varied widely, but non-Chilean cultivars saw the lowest decreases in biomass. Danish varieties tended to rank among those most impacted by salinity, although the authors noted that the higher relative decreases seen in Danish varieties should be considered in light of their higher absolute yields of biomass.
In contrast, Morales (2011) found no differences in plant height or weight with a Chilean lowland cultivar, KU-2, and Chipaya, a variety originating from the southern Altiplano. This further demonstrates that, at least in terms of height and biomass weight, there is an overlap in salinity tolerance between Chilean varieties and varieties from the Altiplano.
Differences in the relative yield declines for the Chilean lowland varieties largely matched with latitude of origin. The northernmost variety, UDEC-1, along with Baer saw the lowest declines in yield under the highest NaCl level. QQ065, the southernmost variety, declined the most. The only difference among quinoa varieties at 32 dS m-1 Na2SO4 was between UDEC-
1 and QQ065, with QQ065 declining more. These results supports the hypothesis of Ruiz-
Carrasco et al. (2011) and confirms the results of their study. Seedlings of Chilean varieties selected across the same environmental gradient in Chile were found to differ in root to shoot ratios and concentrations of compatible solutes, with the variety of southernmost origin demonstrating maladapted characteristics for salinity stress.
Only limited comparisons could be made for differences in salt tolerance between quinoa
-1 and barley, as only yield at 32 dS m NaCl and Na2SO4 was significantly different from the
58
control. Yield declines for barley under 32 dS m-1 NaCl were almost complete at 98.0%. This
-1 decline was lower than that of the four quinoa varieties. Under 32 dS m Na2SO4, the relative decline in yield for barley (-82.4%) was lower than the declines for all quinoa varieties except
QQ065 (-51.9%). However, the barley yield decline might have been exaggerated due to the effects of calcium deficiency, caused by the high level of Na+ present under 32 dS m-1 NaCl. This has been reported for corn (Maas and Grieve, 1987) and for barley under high levels of sodium sulfate (Curtin et al., 1993). Royo et al (2000) calculated a 50% yield reduction for Albacete at
15.3 dS m-1, far below what was seen for quinoa. This confirms that quinoa is superior to barley in salinity tolerance.
Quinoa appeared to escape calcium deficiency. There were no signs of calcium
+ deficiency and no decreases in yield under Na2SO4, which had a greater concentration of Na than NaCl, along with high levels of sulfate, which can decrease the activity of calcium in solution (Manchanda and Sharma, 1989; Curtin et al., 1993). Therefore, quinoa appears to respond similarly to results seen by Curtin et al. (1993) for Kochia scoparia and may be similarly efficient in calcium uptake under high sodium conditions.
A negative effect in yield was seen for most varieties under high fertilization, along with the decrease in leaf greenness content and plant height. Lower rates of germination and stunting were also seen in pots receiving high fertilization, indicating that high levels of fertilization may have a detrimental effect on quinoa. In a field trial grown in 2012, described in Chapter 4, seed was mixed with fertilizer and planted together, which resulted in near-total lack of germination.
More research should be conducted to determine the causal mechanism of this sensitivity. One potential explanation for this observation is ammonium toxicity. Members of Chenopodiaeceae
59
are noted for their relative susceptibility to ammonium. Ammonium toxicity has been shown to cause yield decreases in many plants and can sometimes result in plant mortality (Britto and
Kronzucker, 2002)
UDEC-1 exhibited a peculiar response to high fertilization. In contrast to all other quinoa varieties, which saw significant decreases in yield, UDEC-1 saw a large increase in yield
(31.8%), concomitant with a sharp decrease in height (-31.6%). It appears that this variety was impacted negatively by high fertilization, as reflected in the decrease in height, but that it also benefited from high fertilization as seen in the yield increase.
4.2 Leaf greenness
Leaf greenness decreased under high fertilization. Based on the correlation of leaf greenness with yield and plant height under high fertilization, it appears to be a stress indicator.
Decreases in leaf greenness have been seen in other crops due to stress, and has been proposed as an indicator of stress tolerance. For instance, SPAD values were found to be correlated with salinity tolerance, heat resistance, and spot blotch resistance in wheat (El-Hendawy et al., 2007;
Rosyara et al., 2010). SPAD values have been proposed as a method to detect drought tolerance in peanut (Arunyanark et al., 2008).
Varietal differences in leaf greenness deserve further explanation. For the three Chilean cultivars, leaf greenness seemed to follow latitude of origin. UDEC-1, the northernmost, was highest in leaf greenness, followed by Baer which was intermediate, and finally QQ065, the southernmost which was lowest. CO407D ranked lowest in leaf greenness along with QQ065.
60
However, when all four varieties were included in a field experiment, described in Chapter 4, no significant differences were detected in leaf greenness.
Salinity induced stress was also visible in leaf greenness. All salinity treatments resulted in a decrease in leaf greenness relative to the control. Under NaCl salinity, leaf greenness progressively decreased with increasing EC level, although for Na2SO4 this did not change. This appears to match results found for yield that indicates greater tolerance to Na2SO4 than NaCl.
4.3 Plant height
Stress from salinity treatment and high fertilization was readily visible in reductions in plant height. There are multiple reports of decreased plant height in quinoa due to salinity stress
(Jacobsen et al., 1999; Wilson et al., 2002; Morales, 2009; Orsini et al., 2011). Exceptions have been reported, however. Adolf et al. (2012) tested a diverse range of quinoa varieties and found that some varieties decreased in height under high salinity while others did not significantly change. Gómez-Pando et al. (2010) found that two of fifteen tested Peruvian accessions increased in height at 30 dS m-1, while the remaining thirteen accessions decreased in height.
A few interactions between salinity and plant height were observed. Despite a decrease in height for all varieties under high fertilization, UDEC-1 seemed particularly resilient and was the tallest variety under high fertilization, and experienced the smallest decline across fertilization levels. High fertilization interacted with Na2SO4 salinity and appeared to have a stabilizing effect on the effect of increasing Na2SO4 concentrations on height. Under low fertilization, height decreased with increasing Na2SO4 concentration.
61
Due to the interaction with fertilization, finding baseline differences in plant height is complicated. However, observations under low fertilization are likely more indicative of field conditions, given the negative impact from high fertilization. Under low fertilization, varietal differences in height appeared to match with latitude of geographical origin, as was seen with leaf greenness. Among the Chilean cultivars, UDEC-1, the northernmost, was the tallest. Baer, which is from a central latitude, was intermediate in height, followed by QQ065, the southernmost variety and shortest. Due to the small number of cultivars included in this study, no certain conclusions can be drawn between these results and geographical influences on varieties.
One speculative possibility for the observed height differences is the effect of annual precipitation on rooting depth. Risi and Galwey (1984), in their review of South American reports, note an association between rooting depth and plant height. This, taken in light of gradient in annual precipitation reported by Ruiz-Carrasco et al. (2011), may explain why
UDEC-1 was tallest. Rooting depth has been linked to greater drought tolerance in wheat and a wide range of zoysiagrasses (Narayan and Misra, 1991; Marcum et al., 1995). If taller height among northern varieties is linked to deeper rooting depth, that could allow for greater interception of water in the soil profile in the drier environment at the more northern location.
4.4 Correlations
The significant correlations among yield, plant height, and leaf greenness make sense in light of the corresponding decreases in all three variables due to the stressors of salinity and high fertilization.
When correlations were run separately for each combination of salt type and fertilization
62
level, an interesting change occurred. Only height was correlated significantly with yield under low fertilization. With high fertilization, leaf greenness was highly correlated with both height and yield. As low fertilization is likely more reflective of field conditions than high fertilization, it appears that leaf greenness may not be a satisfactory measure of salinity tolerance for quinoa.
Height, however, may be a promising factor for selecting saline tolerant varieties in the field.
Considering that height measurements were taken just as flowering had begun, this strong correlation could allow for the identification and crossing of more saline tolerant varieties in the same season.
5. Conclusion
Chilean lowland cultivars demonstrated a high level of salinity tolerance, greatly exceeding that of other crops considered to be salt tolerant, such as barley. Differences in salinity tolerance among the varieties appeared to sort along latitude of geographic origin in Chile, while the American bred cultivar CO407D ranked intermediate in salinity tolerance. Stress from high fertilization is a significant problem for quinoa, although there appeared to be contrasting reactions in yield to high fertilization and this phenomenon deserves further investigation. Salt type had a large impact on quinoa, which showed significantly higher tolerance to sodium sulfate than to sodium chloride.
63
References
Adolf, V.I., S. Shabala, M.N. Andersen, F. Razzaghi, and S.E. Jacobsen. 2012. Varietal differences of quinoa’s tolerance to saline conditions. Plant Soil. 357: 117-129.
Arunyanark, A., S. Jogloy, C. Akkasaeng, N. Vorasoot, T. Kesmala, R.C. Nageswara Rao, G.C. Wright, and A. Patanothai. 2008. Chlorophyll stability is an indicator of drought tolerance in peanut. J Agr Crop Sci. 194: 113–125.
Bertero, H.D. 2003. Response of developmental processes to temperature and photoperiod in quinoa (Chenopodium quinoa Willd.). Food Rev Int. 19: 87–97.
Boursier, P., and A. Läuchli. 1990. Growth responses and mineral nutrient relations of salt- stressed sorghum. Crop Sci. 30: 1226–1233.
Britto, D.T., and H.J. Kronzucker. 2002. NH4 toxicity in higher plants: a critical review. J. Plant Physiol. 159: 567–584.
Cramer, G.R., and A.R. Spurr. 1986. Responses of lettuce to salinity. I. Effects of NaCl and Na2SO4 on growth. J. Plant Nutr. 9: 115–130.
Curtin, D., H. Steppuhn, and F. Selles. 1993. Plant responses to sulfate and chloride salinity: growth and ionic relations. Soil Sci. Soc. Am. J. 57: 1304–1310.
El-Hendawy, S.E., Y. Hu, and U. Schmidhalter. 2007. Assessing the suitability of various physiological traits to screen wheat genotypes for salt tolerance. Journal of Integrative Plant Biol. 49: 1352–1360.
Ghassemi, F., Jakeman, A.J. and Nix, H.A. 1995. Salinisation of land and water resources: Human causes, extent, management and case studies. UNSW Press: Sydney, Australia and CAB International: Wallingford, UK.
Gómez-Pando, L.R., R. Álvarez-Castro, and A. Eguiluz-de la Barra. 2010. SHORT COMMUNICATION: Effect of salt stress on Peruvian germplasm of Chenopodium quinoa Willd.: A promising crop. J Agr Crop Sci. 196: 391–396.
Grattan, S.R., and C.M. Grieve. 1999. Mineral nutrient acquisition and response by plants Grown in saline environments. p. 203–229. In Handbook of plant and crop stress. 2nd ed., rev. and expanded. Books in soils, plants, and the environment. M. Dekker, New York.
Grattan, S.R., and C.M. Grieve. 1999. Salinity-mineral nutrient relations in horticultural crops. Sci. Hortic. 78: 127–157.
64
Hariadi, Y., K. Marandon, Y. Tian, S.E. Jacobsen, and S. Shabala. 2011. Ionic and osmotic relations in quinoa (Chenopodium quinoa Willd.) plants grown at various salinity levels. J. Exp. Bot. 62: 185–193.
Jacobsen, S.E., H. Quispe, A. Mujica, and others. 1999. Quinoa: an alternative crop for saline soils in the Andes. Scientists and farmer–partners in research for the 21st century. CIP Program Report 2000: 403–408.
Keller, L.P., McCarthy, G.J., and J.L. Richardson. 1986. Mineralogy and stability of soil evaporites in North Dakota. Soil Sci. Soc. Am. J. 50: 1069–1071.
Koyro, H.-W., and S.S. Eisa. 2007. Effect of salinity on composition, viability and germination of seeds of Chenopodium quinoa Willd. Plant Soil 302: 79–90.
Maas, E.V., and C.M. Grieve. 1987. Sodium-induced calcium deficiency in salt-stressed corn. Plant Cell Environ. 10: 559-564.
Manchanda, H.R., and S.K. Sharma. 1989. Tolerance of chloride and sulphate salinity in chickpea (Cicer arietinum). J. Agric. Sci. (Cambridge). 113: 407–410.
Marcum, K.B., M.C. Engelke, S.J. Morton, and R.H. White. 1995. Rooting characteristics and associated drought resistance of zoysiagrasses. Agron. J. 87: 534.
Morales, A.J. 2009. Physiological assessment of Chenopodium quinoa to salt stress. MS Thesis. Brigham Young University. Provo, Utah.
Morales, J.A. 2011. Physiological responses of Chenopodium quinoa to salt stress. International J. Plant Physiol. Biochem. 3: 219-232.
Narayan, D., and R.D. Misra. 1991. Root growth and productivity of wheat cultivars under different soil moisture conditions. Int J Ecol Environ Sci. 17: 19–26.
Orsini, F., M. Accorsi, G. Gianquinto, G. Dinelli, F. Antognoni, K.B.R. Carrasco, E.A. Martinez, M. Alnayef, I. Marotti, and S. Bosi. 2011. Beyond the ionic and osmotic response to salinity in Chenopodium quinoa: functional elements of successful halophytism. Funct Plant Biol. 38: 818–831.
Pitman, M., and A. Läuchli. 2004. Global impact of salinity and agricultural ecosystems. p 3-20. In Läuchli, A., Lüttge, U. (eds.), Salinity: Environment-Plants-Molecules.
Pulvento, C., M. Riccardi, A. Lavini, G. Iafelice, E. Marconi, and R. d’ Andria. 2012. Yield and quality characteristics of quinoa grown in open field under different saline and non-saline irrigation regimes. J Agron Crop Sci. 198: 254–263.
Risi, J.C., and Galwey, N.W. 1984. The Chenopodium grains of the Andes - Inca crops for modern agriculture. Adv. Appl. Biol. 10: 145–216.
65
Rosyara, U.R., S. Subedi, E. Duveiller, and R.C. Sharma. 2010. Photochemical efficiency and SPAD value as indirect selection criteria for combined selection of spot blotch and terminal heat stress in wheat. J. Phytopathol. 158: 813–821.
Royo, A., R. Aragüés, E. Playán, and R. Ortiz. 2000. Salinity–grain yield response functions of barley cultivars assessed with a drip-injection irrigation system. Soil Sci. Soc. Am. J. 64: 359.
Ruiz-Carrasco, K., F. Antognoni, A.K. Coulibaly, S. Lizardi, A. Covarrubias, E.A. Martínez, M.A. Molina-Montenegro, S. Biondi, and A. Zurita-Silva. 2011. Variation in salinity tolerance of four lowland genotypes of quinoa (Chenopodium quinoa Willd.) as assessed by growth, physiological traits, and sodium transporter gene expression. Plant Physiol. Biochem. 49: 1333–1341.
USDA. 2011. 2011 RCA Appraisal. USDA. Washington, D.C.
Warne, P., R.D. Guy, L. Rollins, and D.M. Reid. 1990. The effects of sodium sulphate and sodium chloride on growth, morphology, photosynthesis, and water use efficiency of Chenopodium rubrum. Can J. Bot. 68: 999.
Weimberg, R., H.R. Lerner, and A. Poljakoff-Mayber. 1984. Changes in growth and water- soluble solute concentrations in Sorghum bicolor stressed with sodium and potassium salts. Physiol Plantarum 62: 472–480.
Wilson, C., Read, J.J., and Abo-Kassem, E. 2002. Effect of mixed-salt salinity on growth and ion relations of a quinoa and a wheat variety. J. Plant Nutr. 25: 2689–2704.
66
Table 1: Analysis of variance with F-values for yield for quinoa and barley.
Effect DF Yield variety 4 64.67*** salinity 6 16.85*** fertility 1 22.48*** salinity*variety 24 0.52 fertility*variety 4 12.28*** salinity*fertility 6 0.81 salinity*fertility*variety 24 0.85 ***=p<0.001
67
Table 2: Analysis of variance with F-values for yield, leaf greenness, and plant height for quinoa only.
Effect DF Yield1 Leaf Plant greenness height variety 3 32.27*** 17.88*** 14.5*** salinity 6 12.44*** 16.35*** 15.54*** fertility 1 11.17** 21.23*** 281.64*** salinity*variety 18 0.3 0.77 0.42 fertility*variety 3 14.18*** 0.69 3.43* salinity*fertility 6 1.4 1.2 3.34** salinity*fertility*variety 18 0.42 0.86 0.59 ***=p<0.001, **=p<0.01, *=p<0.05
68
Table 3: Analysis of variance with F-value for relative declines in yield under 32 dS m-1 NaCl and Na2SO4.
Effect DF Yield Declines variety 4 16.63*** salinity 1 12.82*** fertility 1 30.58*** salinity*variety 4 0.14 fertility*variety 4 1.28 salinity*fertility 1 0.01 salinity*fertility*variety 4 1.27 ***=p<0.001
69
Table 4: Relative declines in yield (g/plant) between the control and 32 dS m-1 NaCl.
-1 -1 Variety/Species 32 dS m NaCl 32 dS m Na2SO4 Percent Percent decrease* decrease* Albacete (Barley) -98.0% C -82.4% D CO407D -65.4% AB -24.5% AB UDEC-1 -43.7% A -10.8% A Baer -49.3% A -11.6% AB QQ065 -73.7% B -51.9% BCD LSDs are from arcsine transformed data *Percent decreases were back transformed from arcsine transformed data
70
Table 5: Change in yield (g/plant) from low to high fertilization.
Variety Low Fertilization High Fertilization % change Albacete 7.33 A 3.40 B -53.7% CO407D 16.21 A 10.95 B -32.4% UDEC-1 13.97 B 18.41 A +31.8% Baer 15.41 A 11.81 B -23.4% QQ065 10.07 A 6.76 B -32.9% LSDs significant at p<0.05, significant within variety
71
Table 6: Pearson correlation coefficients between yield, height, and leaf greenness.
yield leaf greenness leaf 0.62*** greenness height 0.60*** 0.64*** ***=p<0.001
72
Table 7: Pearson correlation coefficients between yield, height, and leaf greenness under NaCl salinity and high fertilization.
yield leaf greenness leaf greenness 0.74** height 0.90*** 0.82** ***=p<0.001, **=p<0.01
73
Table 8: Pearson correlation coefficients between yield, height, and leaf greenness under NaCl salinity and low fertilization.
yield leaf greenness leaf greenness 0.38 ns height 0.83*** 0.47 ns ***=p<0.001, ns=not significant
74
Table 9: Pearson correlation coefficients between yield, height, and leaf greenness under Na2SO4 salinity and high fertilization.
yield leaf greenness leaf greenness 0.57† height 0.76** 0.90*** ***=p<0.001, **=p<0.01, †=p<0.07
75
Table 10: Pearson correlation coefficients between yield, height, and leaf greenness under Na2SO4 and low fertilization.
yield leaf greenness leaf greenness 0.17 ns height 0.70** 0.46 ns **=p<0.01, ns=not significant
76
Figure 1: Yield (g/plant) of quinoa and barley by salinity treatment.
16
A 14 AB A ABC
12 BC
C
10
8 D
6 yield yield (g/plant)
4
2
0 Control 8 dS/m 16 dS/m 32 dS/m 8 dS/m 16 dS/m 32 dS/m NaCl NaCl NaCl Na2SO4 Na2SO4 Na2SO4
LSDs significant at p<0.05
77
Figure 2: Yield of quinoa (g/plant) by salinity treatment.
18
16 A AB AB 14 ABC C BC
12
10 D 8
yield (g/plant) yield 6
4
2
0 Control 8 dS/m 16 dS/m 32 dS/m 8 dS/m 16 dS/m 32 dS/m NaCl NaCl NaCl Na2SO4 Na2SO4 Na2SO4
LSDs significant at p<0.05
78
Figure 3: Variety x fertilization interaction for yield (g/plant).
20 18
16
14 Albacete 12 CO407D 10 UDEC-1 8
6 Baer yield (g/plant) yield 4 QQ065 2 0 High Fertilization Low Fertilization
79
Figure 4: Leaf greenness by variety.
50 A 49 48 B 47 46 45 C
SPAD level SPAD 44 C 43 42 41 CO407D UDEC-1 Baer QQ065
LSDs significant at p<0.05
80
Figure 5: Leaf greenness by fertilization level.
48 A 47
46
45 B
SPAD Value SPAD 44
43
42 High Fertilization Low Fertilization
LSDs significant at p<0.05
81
Figure 6: Leaf greenness by salinity treatment.
52 A 50 B B 48 B B 46 44 C 42 D SPAD Value SPAD 40 38 36 Control 8 dS/m 16 dS/m 32 dS/m 8 dS/m 16 dS/m 32 dS/m NaCl NaCl NaCl Na2SO4 Na2SO4 Na2SO4
LSDs significant at p<0.05
82
Figure 7: Variety by fertilization interaction in plant height.
100 A A 90 B
80
C 70 D
60 height (cm) height E E 50 E
40
30 CO407D UDEC-1 Baer QQ065
High Fertilization Low Fertilization
LSDs significant at p<0.05
83
Figure 8: Fertilization by salinity interaction for plant height.
110 A 100 A A 90 B B 80 BC 70 CD D 60 DE DE DE DE
height (cm) height E 50
40 F
30
20 Control 8 dS/m 16 dS/m 32 dS/m 8 dS/m 16 dS/m 32 dS/m NaCl NaCl NaCl Na2SO4 Na2SO4 Na2SO4 High Fertilization Low Fertilization
LSDs significant at p<0.05
84
CHAPTER THREE
TOLERANCE OF LOWLAND QUINOA CULTIVARS TO SODIUM CHLORIDE AND SODIUM SULFATE SALINITY II: EFFECTS OF SALINITY ON MINERAL NUTRITION OF SEEDS
Tolerance Of Lowland Quinoa Cultivars To Sodium Chloride And Sodium Sulfate Salinity Ii: Effects Of Salinity On Mineral Nutrition Of Seeds
Abstract
Quinoa (Chenopodium quinoa Willd.) has attracted increasing attention as a crop for saline affected areas due to its high level of salinity tolerance. One of the nutritional advantages of quinoa is its high level of mineral nutrition; however, previous studies have found negative and positive effects of salinity on the concentration of minerals in quinoa. This objective of this study was to examine the effect of two salts at three strengths on the Ca, Cu, Fe, Mg, Mn, P, and
Zn concentration of the seed of four Chilean lowland quinoa varieties. Two fertilization levels were included to investigate potential interactions of salinity and soil nitrogen. A highly complex set of responses occurred and salinity, fertilization, and variety were found to significantly interact. Under high fertilization, general decreases were seen in Ca, Cu, and Mn. Few general trends for salinity appeared, but NaCl application caused increases in Cu and Zn and decreases in
Mg. Na2SO4 caused significant decreases in Ca at all EC levels. These results underscore the large and complex effect that soil conditions have on the mineral concentration of quinoa seeds and the importance of environment in determining the levels of mineral nutrition in quinoa.
1. Introduction
Despite the well-documented effects of soil salinity on yield and nutrient uptake, there are relatively few studies examining the impact of soil salinity on grain and seed nutrition. Two studies have examined the effects of salinity and saline-sodic soils on mineral nutrients in quinoa
(Karyotis et al., 2003; Koyro and Eisa, 2007). However, no studies have looked at the effects of
86
salinity from different salts on a range of micronutrients in quinoa. As one of the positive qualities of quinoa is its high level of minerals, any changes in mineral concentration due to soil salinity will be important to characterize.
Additionally, quinoa has a wide reported range of mineral concentration, which is attributed to both genetic factors and differences in environment and growing conditions (Schlick and Bubenheim, 1996; Vega-Gálvez et al., 2010). Schlick and Bubenheim (1996) suggest that mineral concentration of the seeds could be influenced considerably by nutrient availability.
Given the range of interactions with nutrient uptake that have been reported for salinity (Grattan and Grieve, 1999), its reasonable to expect that salinity might result in differences in seed mineral concentration.
The objectives of this experiment were to 1) determine the influence of NaCl and Na2SO4 application and application strength on mineral nutrition in quinoa seeds, 2) determine the effect of high and low fertilization on mineral nutrition and 3) characterize the varietal differences in mineral nutrition for the range of Chilean lowland varieties tested.
2. Materials and Methods
2.1 Experimental Design
This experiment was conducted on seeds produced in the experiment described in chapter
2. For each treatment combination in the experiment, seed samples were combined across replications for each experiment run. This resulted in two samples, one from each experiment run. Combined samples were sent to the Grand Forks Human Nutrition Research Center (Grand
Forks, ND) for mineral analysis. Each combined sample was split into three subsamples.
87
Subsamples were individually ground to a powder (60 s) in a laboratory grinder (Janke &
Kunkel, Model 20S3; Staufen, Germany) with a stainless steel chamber and blade.
Approximately 0.6 g of each subsample was weighed and placed in triplicate into separate Pyrex beakers. Watchglasses were placed on the beakers and the samples were ashed in a muffle furnace at 200°C for 2 h and at 490°C for 12 h. The ash was dissolved in 10 mL of concentrated nitric acid (J.T. Baker Instra-Analyzed; Phillipsburg, NJ) and heated on a hotplate at 120°C, refluxing for 2 h. Three milliliters of 30% hydrogen peroxide (J.T. Baker; Phillipsburg, NJ) were slowly added to each beaker. Samples were allowed to dry and then were ashed again in a muffle furnace following the procedure outlined above. The resulting white ash was dissolved in 2 mL of 6 M HCl (J.T. Baker Instra-Analyzed; Phillipsburg, NJ) with heating and subsequently diluted to 10 mL with deionized water. Samples were analyzed simultaneously for Ca, Cu, Fe, Mg, Mn,
P, and Zn by inductively coupled argon plasma (ICAP) techniques by using a PerkinElmer 3300 instrument (Waltham, MA). Four durum wheat standards (National Institute of Standards and
Technology, Gaithersburg, MD) and four acid blanks were analyzed with each batch of samples.
2.2. Statistical Analysis
Data was analyzed with the PROC MIXED procedure in SAS (SAS Institute, Cary NC).
Each subsample was treated as a replication. Experiment repeat and replication were treated as random factors. Variety, salinity, and fertilization were treated as fixed factors. Appropriate error terms were used. Pearson correlation coefficients for nutrients, yield, plant height, and leaf greenness were determined via PROC CORR in SAS (SAS Institute, Cary NC) using the means generated from PROC MIXED.
88
3. Results
A highly complex set of reactions occurred for all nutrients. There were significant three- way interactions between variety, salinity, and fertilization for all nutrients tested. This necessitated an in-depth analysis of all treatment combinations for all nutrients tested.
3.1 Ca
Ca at 0 dS m-1
At the no-salt control (0 dS m-1 NaCl), there were significant varietal differences in Ca.
This was seen within fertilizer treatments, though a greater range in Ca was seen under higher fertilization (508.9-328.1 μg Ca g-1) than with low fertilization (476.7-409.3 μg Ca g-1).
Additionally, variety influenced response of Ca to fertilization level. Ca increased in CO407D by
24.3% with higher fertilization while in UDEC-1 and QQ065, it decreased by 27.2% and 31.2% respectively. Baer had no significant changes in Ca between fertilization levels. At low fertilization, varietal differences were fewer. Only CO407D was significantly different, having the lowest level of Ca (Table 1).
NaCl
Changes across fertilization
At all strengths of NaCl, Ca was higher under low fertilization than under high fertilization. Ca increased 34.1% under 8 dS m-1 (p<0.0001), 15.4% under 16 dS m-1 (p<0.0001) and 10.8% under 32 dS m-1 (p=0.002).
Response to Increased Salinity
89
Under high fertilization, reaction differed by variety. Ca did not change in UDEC-1 and
QQ065 across NaCl concentrations. However, at 8 dS m-1, Ca in UDEC-1 increased 16.4% compared to the control and in QQ065, Ca increased 19.0%. QQ065 also increased 18.8% at 32 dS m-1. CO407D and Baer both had different responses. At 8 dS m-1, Ca in CO407D decreased
20.9% compared to the control while Ca in Baer decreased 19.6%. Both varieties did not differ in Ca concentration from the control at 16 dS m-1, though at 32 dS m-1, Ca declined by 33.6% for
CO407D and 18.3% for Baer.
Under low fertilization, Ca responses to salinity were more consistent among varieties.
Ca at 8 dS m-1 was 14.1% higher than the control (p<0.0001). At 32 dS m-1, the opposite reaction occurred and Ca decreased by 11.0% (p<0.0001). At 16 dS m-1, only CO407D was significantly different than the control, rising 21.7% (Table 2).
Na2SO4
Changes across fertilization
-1 The results from Na2SO4 treatments were less clear. Only Baer at 8 dS m and QQ065 at
16 dS m-1 differed in Ca across fertilization levels. In both cases, low fertilization resulted in higher levels of Ca. For Baer, Ca increased 21.7% under low fertilization at 8 dS m-1. For QQ065 at 16 dS m-1, Ca increased 34.9% under low fertilization.
Response to Increased Salinity
All levels of Na2SO4 caused a decrease in Ca relative to the control (p<0.0001). Under low fertilization, Ca decreased significantly with higher Na2SO4 concentration. Under 8, 16, and
32 dS m-1, Ca dropped 21.4%, 28.3%, and 47.8% respectively compared to the control
90
(p<0.0001). The application of 32 dS m-1 had the largest negative decrease of any salinity treatment under both fertilization levels (p<0.0001).
Under high fertilization, the situation was more complicated. Ca concentration was higher at 8 and 16 dS m-1 than at 32 dS m-1 (p<0.0001). Only CO407D and QQ065 differed between 8 and 16 dS m-1, declining by 13.8% and 7.9% respectively at 16 dS m-1. Significant varietal differences were seen under 32 dS m-1. Baer (293.7 μg Ca g-1) along with UDEC-1
(266.8 μg Ca g-1) ranked highest in Ca. CO407D (199.1 μg Ca g-1) and QQ065 (202.7 μg Ca g-1) together ranked lowest in Ca, although QQ065 did not differ significantly from UDEC-1 (Table
2).
NaCl vs. Na2SO4
When compared at equal EC levels, NaCl treatments resulted in higher Ca than Na2SO4
-1 - treatments (p<0.0001). Na2SO4 was 23.4% lower at 8 dS m than NaCl (347.8 vs. 454.3 μg Ca g
1), 27.4% lower at 16 dS m-1 NaCl (317.9 vs. 437.7 μg Ca g-1), and 37.8% lower at 32 dS m-1
(239.3 vs. 384.9 μg Ca g-1).
Variety Effects
The high number of interactions prevented determination of general varietal effects.
CO407D was the most variable in Ca level under both fertilization treatments, ranging from
(508.9-199.1 μg Ca g-1) under high fertilization and (556.5-234.6 μg Ca g-1) under low fertilization.
Correlations
Ca had a weak correlation with plant height (r=0.31) (Table 15). The strength of this
91
correlation rose under NaCl application (r=0.80) and Na2SO4 application (r=0.76) under low fertilization, but disappeared for both salts under high fertilization (Tables 16-19).
Ca was strongly correlated with Mg under NaCl salinity under high fertilization (r=0.65) and low fertilization (r=0.79) (Tables 16 & 17). Under NaCl salinity with high fertilization, there was a moderately strong correlation with Mn (r=0.61) (Table 16). Under Na2SO4 salinity, Ca was negatively correlated with Cu under high fertilization (r=-0.75) and positively correlated with Fe under low fertilization (r=0.66) (Table 18 & 19).
3.2 Cu
Cu at 0 dS m-1
At the control treatment, low fertilization (5.58 μg Cu g-1) resulted in 40.4% higher Cu than high fertilization (3.97 μg Cu g-1) across all varieties (p<0.0001) (Table 3). QQ065 had the highest Cu for any variety across all fertilization and salinity levels, including the control
(p<0.0001).
Under high fertilization, CO407D and UDEC-1 ranked intermediate below QQ065 with no significant differences in Cu between the two varieties. Baer was lowest in Cu. Under low fertilization, UDEC-1 ranked second behind QQ065. CO407D was overall lowest, though neither of these differed significantly from Baer (Table 3).
NaCl
Changes across fertilization
92
For all NaCl concentrations, Cu concentration was lower under high fertilization than at low fertilization (p<0.0001). Under high fertilization, decreases in Cu of 26.8%, 27.1%, and
17.0% were seen at 8, 16, and 32 dS m-1 respectively.
Response to increased salinity
Looking at effects on variety as salinity increased, a clear trend emerged. Across varieties and both fertilizer levels, all NaCl concentrations resulted in significantly higher Cu concentrations than the control (p<0.0001).
Under high fertilization, 32 dS m-1 resulted in significantly higher Cu concentration than 8 and 16 dS m-1 (p<0.0001). Only CO407D varied in Cu between 8 and 16 dS m-1 at high fertilization, with 11.6% higher Cu under 16 dS m-1. In contrast, 16 dS m-1 was the highest under low fertilization when compared to 8 and 32 dS m-1 (p=0.006). Cu concentration did not vary between 8 and 32 dS m-1 for any varieties.
Na2SO4
Changes across fertilization
Cu concentration was lower under high fertilization than low fertilization for both 8 and 16 dS m-1 (p<0.0001). The most dramatic decrease was seen with the variety Baer. At 8 dS m-1, Cu decreased 40.5% under high fertilization, and at 16 dS m-1, a 38.9% decrease occurred. At 32 dS m-1, Cu decreased a more modest 19.0% under high fertilization.
For the other three varieties under 32 dS m-1, only UDEC-1 changed significantly across fertilization, decreasing 15.8% under high fertilization.
93
Response to Increasing Salinity
As salinity levels increased, varieties either exhibited increased Cu compared to the control or they did not differ from the control. No combination of Na2SO4 salinity and fertilization led to a decrease in Cu concentration.
Under high fertilization, all varieties increased in Cu concentration at 16 dS m-1 (p<0.0001) and 32 dS m-1 (p<0.0001) relative to the control. Cu rose 21.3% at 16 dS m-1 and 44.6% at 32 dS m-1 compared to the control. At 8 dS m-1, CO407D did not change relative to the control.
However, UDEC-1 (16.0%), Baer (21.7%) and QQO65 (13.4%) all rose significantly (Table 4).
Low fertilization resulted in a larger range of responses by variety. Cu concentration of
CO407D rose 18.8% at 16 dS m-1 and 17.2% at 32 dS m-1 compared to the control. UDEC-1 had no significant differences in Cu for any Na2SO4 concentrations compared to the control. In contrast, Cu in Baer rose at 8 dS m-1 (16.1%), 16 dS m-1 (15.9%), and 32 dS m-1 (17.1%) relative control, although increasing concentration did not have a differing impact. QQ065 only increased in Cu concentration at 32 dS m-1 (14.5%) relative to the control (Table 4).
NaCl vs. Na2SO4
In general, plants grown under NaCl had higher Cu concentration than the equivalent EC
-1 -1 level of Na2SO4. This was highly significant for 8 dS m by 17.8% (p<0.0001) and for 16 dS m by 13.5% (p<0.0001) under high fertilization. This was also seen for 8 dS m-1 by 24.05%
(p<0.0001), 16 dS m-1 by 22.3% (p<0.0001), and 32 dS m-1 by 17.6% (p<0.0001) under low fertilization.
94
Variety Effects
QQ065 stood out as having the highest Cu concentration across all treatment combinations.
Correlations
A moderately strong and highly significant negative correlation was seen between Cu and yield (r=-0.50). Several strong correlations were seen between Cu and other nutrients. A highly significant and moderately strong correlation was found between Cu and Zn (r=0.73) (Table 15).
Under NaCl, two correlations appeared. With high fertilization, a strong negative correlation appeared between Cu and Fe (r=-0.81) and with low fertilization, a positive correlation appeared between Cu and P (r=0.82) (Tables 16 & 17). Two correlations also appeared under Na2SO4 salinity. Cu had a strong negative correlation with height (r=-0.80) under low fertilization. A strong negative correlation appeared between Cu and Ca (r=-0.75) under high fertilization (Table
18 & 19)
3.3 Fe
Fe at 0 dS m-1
Fe levels at the control differed by variety. Under high fertilization, CO407D had the highest value of 69.84 μg Fe g-1 while UDEC-1 had the lowest at 54.51 μg Fe g-1. Baer ranked intermediate, while QQ065 was not significantly different from Baer or CO407D.
Under low fertilization, UDEC-1 had the highest Fe concentration of 72.20 μg Fe g-1,
95
representing a 32.5% increase from the level of Fe under high fertilization. All other varieties did not significantly differ from each other. Overall, UDEC-1 had the highest range in Fe concentration (54.51-72.20 μg Fe g-1). It was also the only variety to show a significant difference in Fe concentration between the two fertilization treatments (Table 5).
NaCl
Changes across fertilization
At 8 dS m-1, all varieties had a statistically higher Fe concentration under low fertilization than under high fertilization (p=0.023). However, this difference was relatively low. Fe concentration under low fertilization was only 4.6% higher. At 16 dS m-1, only Baer had was different, with 13.0% higher Fe concentration under high fertilization compared to low fertilization. This stands in contrast to the reaction seen at 32 dS m-1. At this level, Baer and
UDEC-1 both had 15.7% and 16.1% higher Fe under high fertilization, respectively. The opposite trend occurred in CO407D, with 13.3% lower Fe under high fertilization.
Response to increased salinity
Varietal responses to increasing salinity were complex. Under high fertilization, CO407D decreased 9.2% at 16 dS m-1 and 18.5% at 32 dS m-1 in comparison to the control. This was also seen with QQ065, where all NaCl treatments resulted in a decrease in Fe compared to the control. Fe decreased 21.4% at 8 dS m-1, 18.0% at 16 dS m-1, and 33.9% at 32 dS m-1 compared to the control.
In contrast, UDEC-1 showed increases in Fe for 8 dS m-1 , 16 dS m-1 , and 32 dS m-1 of
17.1%, 23.7%, and 23.2% respectively. This level of increase did not differ among the different
96
NaCl concentrations. Baer had no differences across the control, 8, and 16 dS m-1. However, at
32 dS m-1, there was a 12.9% increase in Fe concentration compared to the control (Table 6).
Under low fertilization, increased NaCl salinity caused decreasing Fe concentration in
UDEC-1, QQ065, and Baer. UDEC-1 saw a decreases at 8 dS m-1 (-9.0%), 16 dS m-1 (-14.1%), and 32 dS m-1 (-19.9%). QQ065 also decreased at all levels, by 11.0% at 8 dS m-1, 24.0% at 16 dS m-1, and by 24.4% at 32 dS m-1. Baer decreased in Fe only at 16 dS m-1 by 16.1%, but
CO407D did not change at any level (Table 6).
Na2SO4
Changes across fertilization
At 8 dS m-1, only Baer and QQ065 showed different Fe concentrations between fertilization treatments. Baer had higher Fe under low fertilization at 94.00 μg Fe g-1, the highest
Fe concentration found under any treatment combination. This was 39.4% higher than Fe for
Baer under high fertilization at the same salinity level. QQ065, in contrast, had 14.5% greater Fe under high fertilization compared to low fertilization. Neither CO407D nor UDEC-1 demonstrated any change in Fe over the two fertilization levels.
At 16 dS m-1, Baer had 15.8% higher Fe concentration under low fertilization than high fertilization. QQ065 was 23.2% higher in Fe under high fertilization than low fertilization.
UDEC-1 had 11.0% greater Fe concentration at low fertilization than high fertilization. No significant effect was seen for CO407D.
Under 32 dS m-1, only Baer had a significant difference between fertilization levels, with
11.0% higher Fe concentration under low fertilization than high fertilization.
97
Response to increased salinity
Trends in Fe concentration were seen for varieties under increasing Na2SO4 salinity. With high fertilization, only increases or no changes in Fe concentration were seen compared to the control. Under high fertilization, Fe increased for UDEC-1 (25.4%) and QQ065 (26.8%) at 8 dS m-1. UDEC-1, Baer, and QQ065 increased at 16 dS m-1 by 20.0%, 16.0%, and 25.7% respectively. Only UDEC-1 responded at 32 dS m-1, with Fe increasing by 18.37% (Table 6).
Under 8 dS m-1 at low fertilization, Fe increased for Baer and QQ065 in comparison to the control, by 49.8% and 10.9% respectively. Only Baer changed in Fe at 16 dS m-1, increasing by 35.8% relative to the control. The only decreases in Fe occurred at 32 dS m-1. Fe concentration of UDEC-1 and QQ065 decreased by 23.2% and 13.6% respectively (Table 6).
NaCl vs. Na2SO4
-1 Both salt types could be directly compared at 8 dS m across both fertilization levels.
-1 -1 -1 NaCl at 8 dS m NaCl resulted in 15.5% lower Fe (62.72 µg Fe g ) than 8 dS m Na2SO4 (74.24
µg Fe g-1).
-1 Under low fertilization, NaCl was again lower than Na2SO4 at 16 dS m . Application of
-1 -1 -1 16 dS m NaCl resulted in 20.8% lower Fe (58.34 µg Fe g ) than at 16 dS m Na2SO4 (73.66 µg
Fe g-1).
Variety Effects
QQ065 was significantly lower than the other varieties when it was treated with NaCl
(p<0.0001). Under Na2SO4 salinity at high fertilization, it saw a significant elevation in Fe
-1 concentration at 8 and 16 dS m Na2SO4 compared to the other varieties, and exhibited
98
significantly higher concentrations compared to other treatment combinations. Under low fertilization, this trend disappeared.
Under Na2SO4 salinity under low fertilization, Baer saw a pronounced and significant
-1 increase in Fe concentration at all levels of Na2SO4. Only at 32 dS m Na2SO4 does another variety match Baer in Fe concentration. Most notably, the highest Fe concentration for any
-1 -1 treatment combination, 94.00 µg Fe g , was seen with Baer at 8 dS m Na2SO4 under low fertilization.
Correlations
Fe had strong correlations with several other nutrients at particular combinations of salinity and fertilization. A moderately strong negative correlation was seen between Fe and Zn with NaCl under low fertilization (r=-0.68) (Table 17). Fe also had a negative correlation with
Cu with NaCl, but under high fertilization (r=-0.81) (Table 16). A positive correlation between
Fe and Mn (r=0.58) was seen with NaCl under low fertilization (Table 17). Under Na2SO4 salinity with low fertilization, Fe was positive correlated with Mg (r=0.77) and Ca (r=0.66)
(Table 19).
Although correlations between Fe and yield, plant height, and leaf greenness were quite low, there was a strong positive correlation between Fe and yield with NaCl salinity under low fertilization (r=0.71) (Table 17).
3.4 Mg
Mg at 0 dS m-1
Significant differences were seen between varieties within both fertilizer treatments.
99
Under higher fertilization, QQ065 stood as significantly lower than the rest, while the other three were not statistically significantly different. Under low fertilization, Baer was significantly the highest, while QQ065 was again the lowest along with CO407D (Table 22). High fertilization caused a 7.3% decrease in Mg compared to low fertilization at the control (p<0.0001) (Table 7).
NaCl
Changes across fertilization
For all EC levels of NaCl, higher Mg resulted from low fertilization concentration than with high fertilization (p<0.0001). This increase was 9.9% at 8 dS m-1, 12.9% at 16 dS m-1, and
14.4% at 32 dS m-1.
Response to increased salinity
Under high fertilization, Mg at 8 dS m-1 had no significant differences from Mg at the control. Mg decreased 8.9% from 2370 μg Mg g-1 at the control to 2159 μg Mg g-1 at 16 dS m-1
(p<0.0001). At 32 dS m-1, Mg decreased 27.4% for CO407D, 17.6% for UDEC-1, and 17.3% for
Baer. QQ065 did not differ significantly in Mg at 32 dS m-1 (Table 8).
Similar responses were seen with low fertilization. However, QQ065 again was an exception, with no significant differences in Mg between the control and all NaCl levels. For 8 dS m-1 and 16 dS m-1, only Baer decreased significantly in Mg compared to the control with decreases of 8.8% and 15.7%, respectively. At 32 dS m-1, CO407D, UDEC-1 and Baer all decreased significantly compared to their respective controls, with decreases of 8.2%, 9.8%, and
21.7% (Table 8).
100
Na2SO4
Changes across fertilization
As with NaCl, Mg under Na2SO4 treatments was generally higher under lower fertilization. Higher Mg was seen under low fertilization than under high fertilization for 8 dS m-
1 (2510 vs. 2399 μg Mg g-1) and 16 dS m-1 (2536 vs. 2397 μg Mg g-1) (p<0.0001). At 32 dS m-1 , only varieties UDEC-1 and Baer were significantly higher in Mg under low fertilization, by
13.6% and 16.4% respectively.
Response to increased salinity
Compared to the control under low fertilization, Na2SO4 only had significant but relatively small effects for CO407D and Baer. For CO407D, Mg decreased 7.8% at 32 dS m-1.
Baer decreased 4.9% at 16 dS m-1 and 5.8% at 32 dS m-1 (Table 8).
Under high fertilization, mixed responses appeared. Mg decreased significantly at 32 dS m-1 for CO407D (6.7%), UDEC-1 (11.3%), and Baer (6.5%), compared to the control. QQ065, in contrast, increased in Mg by 7.2% at 16 dS m-1 compared to the control (Table 8).
Na2SO4 vs. NaCl
Few direct comparisons could be made between NaCl and Na2SO4. However, under high
-1 -1 fertilization, the two salts could be compared at 8 dS m and 16 dS m . Na2SO4 treatment resulted in 5.1% higher Mg at 8 dS m-1 (p=0.0004) and 11.0% higher Mg at 16 dS m-1
(p<0.0001) than NaCl treatment.
101
Variety Effects
CO407D was found to be particularly variable in its Mg concentration under high fertilization. It had the highest Mg concentration at control (2458 μg Mg g-1) as well as the lowest Mg level (1785 μg Mg g-1), which was seen at 32 dS m-1 NaCl.
Under low fertilization, Baer proved to be the most variable with Mg concentration ranging from 2790 μg Mg g-1 at the control to 2186 μg Mg g-1 at 32 dS m-1 NaCl. It was also the variety highest in Mg under Na2SO4 salinity combined with low fertilization.
Correlations
Mg had a moderately weak correlation with yield (r=0.46). Slightly stronger correlations were seen with leaf greenness (r=0.55) and height (r=0.66). Mg had a moderately strong correlation with phosphorus (r=0.60) (Table 15). Ca and Mg were correlated with NaCl salinity, under both high fertilization (r=0.65) and low fertilization (r=0.79) (Tables 16 & 17).
3.5. Mn
Mn at 0 dS m-1
UDEC-1 had the highest Mn concentration of the four varieties under both fertilization levels (p<0.0001). Only for Baer did Mn concentration change with fertilization, which decreased 35.0% under high fertilization. Under low fertilization, CO407D, Baer, and QQ065 were not significantly different. When fertilization was increased, significant differences appeared for all three. CO407D ranked second highest behind UDEC-1, QQ065 third, and Baer ranked lowest (Table 9).
102
NaCl
Changes across fertilization
When salinity was increased to 8 dS m-1, Mn concentration for all varieties was affected by fertilization. UDEC-1 was higher in Mn under high fertilization, while Mn in the other varieties decreased under high fertilization.
At 16 and 32 dS m-1, a simple pattern arose. At these levels, Mn was higher under low fertilization for all varieties (p<0.0001).
Response to increased salinity
When salinity increased, there was a complex range of responses for varieties under high fertilization. In contrast, a more uniform response was seen as salinity increased under low fertilization.
Under high fertilization, Baer and QQ065 both showed no significant differences in Mn among the NaCl levels. This held when these levels were compared to the control. The sole exception was for Baer at 16 dS m-1 where Mn was 21.4% higher than the control. Compared to the control, Mn declined significantly at 32 dS m-1 for CO407D and UDEC-1, with respective declines of 20.7% and 14.6%. Significant increases of 14.1% for CO407D at 16 dS m-1 and
14.2% for UDEC-1 at 8 dS m-1 were observed (Table 10).
Due to the lack of confounding interactions, 16 and 32 dS m-1 under low fertilization were compared to the control. Mn at 16 dS m-1 (65.65 µg Mn g-1) and Mn concentration at 32 dS m-1 (64.03 µg Mn g-1) were 17.4% and 14.5% higher than the control (55.93 µg Mn g-1),
103
respectively. There were no significant differences between 16 and 32 dS m-1 for any varieties.
At 8 dS m-1, all varieties but UDEC-1 had significant increases from the control. CO407D increased by 31.9%, Baer by 21.3%, and QQ065 by 14.9% (Table 10).
Na2SO4
Changes across fertilization
Na2SO4 induced a more complex range of responses than NaCl. For all varieties at 8 dS m-1, Mn concentration changed in response to fertilization. UDEC-1 and QQ065 increased
17.4% and 24.5% respectively in Mn concentration under high fertilization, while a decrease was seen in CO407D (12.1%) and Baer (20.8%). Increased salinity moderated the response in Mn concentration to fertilizer. At 16 dS m-1, only Baer exhibited a significant change over fertilization, with Mn decreasing 15.1% under higher fertilization. Baer also decreased in Mn at
32 dS m-1 by a larger margin of 23.3%. At this salinity under high fertilization, CO407D increased in Mn by 19.2% from 78.47 µg Mn g-1 under low fertilization to 93.57 µg Mn g-1, the highest level of Mn for any treatment combination.
Response to increasing salinity
Increasing Na2SO4 salinity resulted in differing patterns of Mn concentration in the four varieties. CO407D and Baer exhibited similar trends, with increasing salinity generally resulting in higher Mn concentration.
Under high fertilization, CO407D exhibited no significant differences at 8 dS m-1.
However, a 22.1% increase in Mn relative to the control occurred at 16 dS m-1. A significantly larger increase of 64.4% occurred when salinity was further increased to 32 dS m-1. For CO407D
104
under low fertilization, significant increases relative to the control were seen at all Na2SO4 levels. The levels of increase for 8 dS m-1 (21.2%) and 16 dS m-1 (25.1%) did not differ significantly. However, at 32 dS m-1, there was a significantly higher increase in Mn of 42.9%.
Baer exhibited significant increases in Mn for all Na2SO4 levels under high fertilization.
While the levels of increase at 8 dS m-1 (35.9%) and 16 dS m-1 (52.3%) were not significantly different, there was a significantly larger increase at 32 dS m-1 (83.8%). When Baer was under low fertilization, no changes were seen at 8 dS m-1. However, Mn increased 16.5% at 16 dS m-1.
This increase grew further to 55.6% at 32 dS m-1.
For both UDEC-1 and QQ065, few changes in Mn occurred with the application of
-1 Na2SO4. At 8 dS m under high fertilization, Mn of UDEC-1 increased 17.6%. The only significant change with QQ065 was a 24.6% decrease in Mn at 8 dS m-1 under low fertilization
(Table 10).
Several main effects for Na2SO4 concentration were interpretable, despite the presence of interactions. Under low fertilization, Mn concentration was 26.5% higher at 32 dS m-1 (70.76 µg
Mn g-1) than under the control (55.93 µg Mn g-1) (p<0.0001). Under high fertilization, Mn was
13.0% higher at 8 dS m-1 (56.57 µg Mn g-1) (p=0.0002), 20.6% higher at 16 dS m-1 (60.42 µg Mn g-1) (p<0.0001), and 38.9% higher at 32 dS m-1 (69.55 µg Mn g-1) than at the control (50.08 µg
Mn g-1) (p<0.0001).
NaCl vs. Na2SO4
-1 NaCl and Na2SO4 could be directly compared at 8 dS m under low fertilization. NaCl
-1 -1 -1 at 8 dS m (64.54 µg Mn g ) resulted in a 9.3% decrease in Mn compared to 8 dS m Na2SO4
105
(71.19 µg Mn g-1) (p<0.0001). Under high fertilization, NaCl and Na2SO4 could be compared at
-1 -1 -1 16 and 32 dS m . NaCl was 16.9% lower than Na2SO4 at 16 dS m (60.98 vs. 73.36 µg Mn g ) and 6.0% lower at 32 dS m-1 (59.71 vs. 63.52 µg Mn g-1).
Correlations
Mn had significant but weak correlations with yield, leaf greenness, and plant height.
When correlation analysis was limited to NaCl salinity under high fertilization, strong correlations appeared between Mn and yield (r=0.61) and Mn and height (r=0.74) (Table 16).
A few correlations appeared between Mn and other nutrients at particular combinations of salinity and fertilization. Mn and P had moderately strong negative correlations with NaCl under low fertilization (r=-0.59) and with Na2SO4 under high fertilization (r=-0.62) (Tables 17 &
18). Ca and Mn were correlated with NaCl under high fertilization (r=0.61) (Table 16) and Fe and Mn were correlated with NaCl under low fertilization (r=0.58) (Table 17).
3.6 P
P at 0 dS m-1
The most distinct varietal difference was seen with Baer, which ranked the highest in P across both fertilization levels. CO407D and UDEC-1 ranked intermediate in P concentration and did not change significantly over fertilization levels. QQ065 had similar P levels to CO407D and UDEC-1 under low fertilization. However, P concentration of QQ065 dropped significantly under high fertilization compared to UDEC-1 and CO407D under high fertilization and by 5.2% compared to QQ065 under low fertilization (Table 11).
106
NaCl
Changes across fertilization
The response of varieties to fertilizer remained similar to the control at 8 and 16 dS m-1
NaCl. At these NaCl levels, only QQ065 showed changes in P across fertilizer levels, decreasing under high fertilization by 8.5% at 8 dS m-1 and by 11.6% at 16 dS m-1. However, at 32 dS m-1,
QQ065 did not differ between fertilization levels. At this level, CO407D and Baer did exhibit differences between fertilization level. CO407D had 5.1% lower P under high fertilization. Baer exhibited the opposite reaction, with 4.1% higher P concentration under high fertilization.
Response to increasing salinity
Different trends were seen by variety as NaCl salinity increased. CO407D, UDEC-1, and
Baer all saw decreases in P concentration with increasing salinity under high fertilization.
QQ065 exhibited a different trend under high fertilization. P concentration held steady across the control and at 8 and 16 dS m-1. At 32 dS m-1, a significant increase in P concentration occurred, with an 8.1% increase compared to the control (Table 12).
Under low fertilization at 32 dS m-1, P declined by 5.5%, 6.7%, and 11.7% for CO407D,
UDEC-1, and Baer, respectively. For CO407D, P at 8 and 16 dS m-1 was not significantly different than the control. UDEC-1 had a more complex response with 4.1% lower P at 8 dS m-1, but no significant difference at 16 dS m-1. Baer had the most notable decline, with a 6.5% decline in P at 8 dS m-1 and a 10.9% decline at 16 dS m-1. QQ065 was lowest in P under the control. All
NaCl concentrations elicited significant increases (4.1-6.5%) in P for QQ065, however these increases were not significantly different from each other (Table 12).
107
Despite the high number of interactions, several direct comparisons between the salinity treatments could be made. P concentration at 16 dS m-1 was lower than that at the control
(p<0.0001) and at 8 dS m-1 (p=0.001). P under 16 dS m-1 (5054 µg P g-1) was 5.0% lower than P at the control (5318 µg P g-1).
Na2SO4
Changes across fertilization
At 8 dS m-1, P concentration was 2.2% higher under high fertilization (5356 µg P g-1) than low fertilization (5242 µg P g-1) for all four varieties (p=0.029). No changes occurred at 16 and 32 dS m-1, except for Baer at 32 dS m-1 which increased 8.7% under low fertilization.
Response to increased salinity
Trends by variety differed under Na2SO4 salinity. Under high fertilization, UDEC-1 and
QQ065 saw no significant differences across all Na2SO4 concentrations. When compared to the control, UDEC-1 had lower P concentration at 16 dS m-1 (-4.1%) and 32 dS m-1 (-7.8%) and
QQ065 had significantly higher P concentration at 8 dS m-1 (+5.0%) (Table 12).
P concentration remained stable under low fertilization. Similar to under high fertilization, QQ065 and UDEC-1 had no differences among Na2SO4 concentrations. For QQ065,
-1 none of the Na2SO4 treatments were significantly different than the control. Only at 8 dS m was
UDEC-1 different than the control. P concentration decreased by a minimal 2.8%.
For C0407D, only at 32 dS m-1 was P concentration significantly different than the control, with a 6.9% decrease.
P concentration of Baer increased or decreased depending on the salinity concentration. At 16 dS
108
m-1, there was no significant decrease in P concentration compared to the control. However, at 8 dS m-1, P concentration was 4.6% higher and at 32 dS m-1 it was 5.2% lower (Table 12).
Due to interactions, only one comparison between the means of Na2SO4 levels could be made. P was significantly higher at 16 dS m-1 (5270 µg P g-1) than at 32 dS m-1 (5064 µg P g-1)
(p=0.002).
NaCl vs. Na2SO4
Due to interactions, few comparisons could be made for equal levels of NaCl and
-1 Na2SO4. However, a comparison could be made at 16 dS m under high fertilization. P under 16
-1 -1 -1 -1 dS m Na2SO4 (5270 µg P g ) was 4.3% higher than P at 16 dS m NaCl (5054 µg P g )
(p<0.0001).
Variety
-1 -1 The overall highest P value of 5872 µg P g was found with Baer at 8 dS m Na2SO4 under high fertilization. Baer also had the widest range of P values (5872-5083 µg P g-1). When compared across all fertility and salinity levels, Baer had 8.6% higher P than CO407D and 8.3% higher P than UDEC-1 (p<0.0001). Baer also exceeded QQ065 in P under high fertilization,
-1 except at the highest salinity treatments of 32 dS m NaCl and Na2SO4 where no significant differences were seen between the varieties. Under low fertilization, QQ065 had significantly higher P than Baer under all NaCl levels. In the control and Na2SO4 treatments, this trend reversed and P in Baer was significantly higher than QQ065.
Correlations
P had a moderately strong positive correlation with Mg (r=0.60). A very weak negative
109
correlation occurred between P and Mn (r=-0.27) (Table 15), but this increased with NaCl under low fertilization (r=-0.59) and with Na2SO4 under high fertilization (r=-0.62) (Tables 17 & 18).
3.7 Zn
Zn at 0 dS m-1
At the control, fertilization affected the Zn levels of two varieties. CO407D and Baer increased 22.8% and 10.4% respectively in Zn under high fertilization, but there was no change for UDEC-1 or QQ065. Under high fertilization, CO407D and QQ065 ranked highest while
UDEC-1 and Baer had the least Zn. Similarly under low fertilization, two groups formed.
However, QQ065 and UDEC-1 had the highest Zn, while Baer and CO407D had the least Zn
(Table 13).
NaCl
Changes across fertilization
Responses under NaCl salinity were complex, and few consistent trends emerged. At 8 dS m-1, fertilizer had a notable effect on two varieties, UDEC-1 and QQ065, which did not display a response in Zn concentration to fertilizer when under the control. UDEC-1 increased in
Zn concentration by 6.8% under high fertilization. Fertilization had the opposite effect on
QQ065. Low fertilization resulted in a relative 16.9% increase in Zn.
Under 16 dS m-1, QQ065 again showed higher Zn under low fertilization. At this salinity,
Zn concentration increased 7.1%. CO407D was the only other variety to respond differently across fertilizer levels at this NaCl level. It reacted oppositely than the control and Zn decreased under high fertilization, dropping 7.8%.
110
At 32 dS m-1, QQ065 showed an increase in Zn under low fertilization. It increased
13.3% as fertilization level decreased. While UDEC-1 did not exhibit an increase in Zn with high fertilization at 16 dS m-1 as it did at 8 dS m-1, this reaction reappeared at 32 dS m-1. Zn increased
11.7% under high fertilization.
Response to increasing salinity
Examining varieties across increasing NaCl revealed a consistent trend of increasing Zn.
Under high fertilization, Zn levels at 8 dS m-1 (51.25 µg Zn g-1) and 32 dS m-1 (55.94 µg Zn g-1) were significantly higher than the control (46.27 µg Zn g-1) (p<0.0001). Zn under 32 dS m-1 was
9.2% higher than Zn at 8 dS m-1 (p<0.0001). At 16 dS m-1, UDEC-1, Baer, and QQ065 were higher in Zn than the control, by 24.8%, 20.4%, and 20.5% respectively. CO407D did not differ in Zn at 16 dS m-1 than the control, but it was lower in Zn than at 32 dS m-1 (Table 14).
Zn was found to increase at all NaCl levels under low fertilization. Compared to the control, it rose 24.3% at 8 dS m-1 (53.43 µg Zn g-1), 30.5% at 16 dS m-1 (56.11 µg Zn g-1), and
31.7% at 32 dS m-1 (56.62 µg Zn g-1) compared to the control (42.99 µg Zn g-1)(p<0.0001).
Similar to under high fertilization, Zn at 8 dS m-1 and 32 dS m-1 could be directly compared due to the lack of interactions. Zn at 32 dS m-1 was found to be 6.0% greater than Zn 8 dS m-1
(p<0.0001) (Table 14).
Na2SO4
Changes across fertilization
Some small changes occurred when fertilization was applied. This was most notable under 8 dS m-1, where Zn concentration increased by 7.0% for CO407D, by 9.1% for UDEC-1,
111
and by 11.2% for QQ065 under high fertilization. At 16 dS m-1, only CO407D increased in Zn under high fertilization, by 12.3%. Baer, which did not change across fertilizer levels at 8 dS m-1, exhibited the opposite trend and decreased by 8.3%. The only varietal change induced by different fertilizer level at 32 dS m-1 was QQ065, which increased by 8.6%.
Response to increasing salinity
A more complex range of responses was generated with increasing Na2SO4 than with
NaCl. Under high fertilization, Zn at 16 dS m-1 (49.45 µg Zn g-1) was found to have increased
6.9% compared to the control (46.27 µg Zn g-1) (p<0.0001). Responses from varieties at 8 and 32 dS m-1 were mixed. CO407D decreased 8.3% in Zn concentration at 8 dS m-1 and 10.2% at 32 dS m-1 as compared to the control. UDEC-1 and QQ065 both rose in Zn at 8 and 32 dS m-1. The most notable increase was for QQ065 at 32 dS m-1, which saw a 29.2% increase to 63.71 µg Zn g-1, one of the highest Zn concentrations detected. Baer did not differ from the control in Zn at 8 dS m-1, but increased 11.0% at 32 dS m-1 (Table 14).
Under low fertilization, several main effects could be compared due to the lack of interactions between varieties. Zn concentration at 16 dS m-1 was found to be 11.8% higher than the control, increasing from 42.99 µg Zn g-1 to 48.06 µg Zn g-1 (p<0.0001). An even greater increase of 17.4% occurred at 32 dS m-1, with Zn rising to 50.47 µg Zn g-1 (p<0.0001).
Zn levels at 8 dS m-1 could be compared directly to both 16 dS m-1 and 32 dS m-1 and were lower by 5.9% and 10.3% respectively (p<0.0001). When Zn at 8 dS m-1 was compared to the control, only Baer showed a significant difference, rising 18.4% (Table 14).
112
NaCl vs. Na2SO4
-1 -1 -1 Zn was 12.3% higher at 8 dS m NaCl (52.34 µg Zn g ) than at 8 dS m Na2SO4 (46.59
-1 µg Zn g ) (p<0.0001). Interactions complicated the comparison between NaCl and Na2SO4 at 16 dS m-1 and 32 dS m-1 across fertilization levels, but the two salts could be compared at these strengths under low fertilization.
At both 16 dS m-1 and 32 dS m-1 under low fertilization, NaCl resulted in higher Zn than
-1 -1 Na2SO4 (p<0.0001). The increase was highest at 16 dS m , where Zn under NaCl (56.11 dS m )
-1 -1 -1 was 16.7% greater than under Na2SO4 (48.06 µg Zn g ). NaCl at 32 dS m (56.62 µg Zn g )
-1 was 12.2% higher than Zn under 32 Na2SO4 (50.47 µg Zn g ).
Variety Effects
QQ065 was found to have the highest Zn under low fertilization (p<0.0001) at 56.80 µg
Zn g-1. UDEC-1 ranked second highest in Zn at 50.31 µg Zn g-1 (p<0.0001).
Correlations
Zn had negative correlations with yield (r=-0.57), leaf greenness (r=-0.52), and height
(r=-0.39) (Table 15). The strength of the correlation between Zn and yield increased considerably under low fertilization for NaCl (r=-0.91) and Na2SO4 (r=-0.82) (Tables 17 & 19).
The correlation between Zn and plant height also increased considerably under low fertilization for NaCl (r=-0.65) and Na2SO4 (r=-0.88) (Tables 17 & 19).
4. Discussion
4.1 Ca
113
Ca and Na have an antagonistic relationship in the plant and soil. Increased sodium can result in decreased Ca uptake and concentration in plant tissues (Grattan and Grieve, 1999;
Cramer, 2002). Karyotis et al. (2003) saw a significant decrease in average seed Ca for quinoa varieties grown on a saline-sodic soil. In contrast, Koyro and Eisa (2007) found higher Ca in seeds under high NaCl treatment, though Ca was found to decrease in exterior parts of the seed. No changes in Ca concentration were found for wheat grown under saline conditions (Labanauskas et al., 1978).
Under NaCl treatment, the effects were rather complicated and no universal decrease from the control was seen. High fertilization complicated the response of varieties to NaCl. Under low fertilization, some interesting observations were made. Ca was found to be higher at the lowest salinity level, while lowest at the highest salinity level. As low fertilization is more likely to be observed under typical field conditions, this could be considered to be a more relevant response to soil salinity.
Na2SO4 resulted in decreased Ca at all levels when compared to the control. One contributing factor may be due to the difference in molar concentration at equal electrical conductivity levels between NaCl and Na2SO4. Based on preliminary tests determining the quantity
+ of salt to add for a given EC level, a greater amount of Na could be expected from Na2SO4 than
NaCl at equal EC (unpublished data). This greater Na+ concentration may be the cause of the
-1 reduced Ca, particularly as 32 dS m Na2SO4 resulted in the lowest Ca of any salinity treatment.
This could explain the greater level of Ca under NaCl compared to Na2SO4 at equivalent EC levels. However, it would not explain why Ca decreased at 8 dS m-1 Na2SO4, but either showed no change or increased at 16 dS m-1 NaCl. Different effects from the anions Cl- and SO42- may play a
114
role in seed Ca, but these effects are difficult to determine due to the different levels of Na associated with NaCl and Na2SO4.
2+ Na2SO4 application may have also resulted in reduced activity of Ca due to interaction
2- with SO4 . This was seen in an experiment with barley, which exhibited Ca deficiency under sulfate salinity (Curtin et al., 1993).
High fertilization had a significant impact on Ca uptake, generally causing a decrease in seed Ca. The strong correlations between Ca and plant height for both salts under low fertilization could be explained by the common impact of salinity on both these factors. Under low fertilization, salinity was the main determinant of height, in contrast to under high fertilization, where fertilizer related stress was the main factor limiting height. Under both fertilization levels, Ca saw a decrease with greater salinity, which reflects the antagonistic relationship between Na+ and Ca2+.
4.2 Cu
The only report on the effects of soil salinity on seed Cu in quinoa is by Karyotis et al.
(2003), who found that saline-sodic soils did not have a significant impact on average seed Cu levels. Reports for Cu concentration in wheat and in common beans have recorded no change in
Cu for plants grown under saline conditions (Labanauskas et al., 1978; Carbonell-Barrachina et al., 1998).
In this experiment, the response of Cu to the various treatments was more easily interpreted due to relatively low number of interactions. All salinity treatments resulted in either increases or no significant change in Cu from the control. NaCl at all concentrations was related to increases in seed Cu. For Na2SO4, there were either no changes or increases in seed Cu, with
115
increases usually occurring at higher Na2SO4 concentrations. There was a greater increase in
-1 -1 seed Cu with NaCl than Na2SO4 at 8, 16, and 32 dS m under low fertilization and at 16 dS m under high fertilization. High seed Cu was found with low fertilization for the control and all
-1 salinity levels except 32 dS m Na2SO4. High nitrogen supply can induce Cu deficiency by interfering with retranslocation of Cu within the plant (Marschner, 1995). If retranslocation of
Cu was inhibited by high fertilization, less Cu would be expected to have reached the seeds, which would explain the observed results.
The negative correlation between Cu and yield should be considered with some caution.
QQ065, the variety highest in Cu, was also the lowest yielding. Due to the small number of varieties tested, this likely skewed this correlation upwards. However, lower Cu concentration under higher yields might be related to the effect of dilution. An increase in yield would result in a larger sink for the limited amount of Cu present in the plant, resulting in lower overall Cu seed concentration. Further investigation with a wider range of varieties is warranted to determine the cause of this correlation.
4.3 Fe
In a review of the effects of plant salinity on plant shoot Fe concentration, Grattan and
Grieve (1999) found contrasting reports. No changes in the average Fe concentration of quinoa seed were seen for plants grown on a saline-sodic soil (Karyotis et al., 2003). Fe was found to increase in the common bean in response to salinity, although Fe concentration of wheat grain did not change significantly (Labanauskas et al., 1978; Carbonell-Barrachina et al., 1998).
116
The response of Fe was relatively complex and few discernable trends appeared. QQ065 was lowest in Fe under NaCl salinity, indicating that Fe concentration of this variety was particularly impacted. Fertilization level played a large role in changing the varietal response to salinity. A few distinct varietal trends appeared under particular combinations of salinity and fertility.
Baer showed a relatively strong reaction to low and moderate Na2SO4 salinity under low
-1 fertilization. Baer was higher in Fe than all other varieties at 8 and 16 dS m Na2SO4 under low fertilization, and at 8 dS m-1, it had the highest Fe concentration of any treatment combination at almost 50% greater than the control. QQ065 also had a strong increase in Fe with Na2SO4 salinity, but under high fertilization.
4.4 Mg
Many studies which examine the effects of salinity on Mg focus on the Ca2+/Mg2+ ratio, as high Ca levels can affect Mg uptake (Grattan and Grieve, 1999). However, in this study, Ca was provided by the potting soil and was constant between salinity treatments. More relevant to this study would be the results found by Bernstein et al. (1974), where increased salinity resulted in either no change or a decrease in Mg in plant tissue, depending largely on species. Karyotis et al. (2003) saw lower average seed Mg for quinoa varieties cultivated on a saline-sodic soil. In contrast, Koyro and Eisa, (2007) found that salinity increased overall Mg in the seed increased at high salinity. Concentration was found to drop in the pericarp, while Mg remained concentrated in the seed embryo.
117
In this study, general declines in seed Mg were seen with NaCl salinity. Under Na2SO4 salinity, the situation was more complex, with either no change in seed Mg or decreases occurring, mainly under higher Na2SO4 levels. Only QQ065 showed an increase in Mg under
-1 Na2SO4 salinity, at 16 dS m under high fertilization. As Mg uptake is vulnerable to interference from other cations, one potential explanation for reduced Mg may be an excess of Na+ from high salinity levels (Marschner, 1995).
For almost all salinity levels, with the sole exception of 32 Na2SO4 for CO407D and
+ QQ065, high fertilization reduced Mg levels. High concentrations of NH4 can interfere with uptake of Mg2+ (Kurvitis and Kirkby, 1980 as cited in Marschner, 1995). Only speculative
+ statements can be made here, however, as NH4 was not measured in this experiment.
CO407D showed a higher level of variability in Mg for salinity treatments under high fertilization. In contrast, Baer was quite variable in Mg over salinity treatments, but under low fertilization.
The positive correlations between Mg and yield, plant height, and leaf greenness are likely due to two factors. First, other cations can inhibit uptake of Mg2+ (Marschner, 1995), and the high amount of Na+ present in the saline solutions may have reduced Mg uptake. The simultaneous reduction in yield, plant height, leaf greenness, and Mg under salinity would have been responsible for the positive correlation. However, calcium which is also taken up as a divalent cation vulnerable to competition from Na+, was positively correlated with these factors only under low fertilization. However, Mg was found to generally decline in seeds under high fertilization. Leaf greenness and plant height also showed declines under high fertilization. This
118
would explain why Mg exhibited positive correlations with these factors under both high and low fertilization.
4.5 Mn
Complicated and contrasting reports exist for the effect of salinity on Mn uptake. Despite this complexity, it is reported that divalent cations usually have a positive effect of Mn in plant tissue while monovalent cations have the opposite effect (Grattan and Grieve, 1999).
A complex set of results were seen in Mn concentration. Despite the presence of both increases and decreases in Mn and that monovalent Na+ was the predominant cation, increases in
Mn due to salinity were most common. However, it is important to note that the studies reviewed by Grattan and Greieve (1999) examined Mn in plant tissue and not concentration in seeds.
Karyotis et al. (2003) provides the only study looking at response of Mn in quinoa seed, reporting that quinoa grown on a saline-sodic soil resulted in significantly lower average Mn in seeds. However, this was in a saline-sodic soil and may not reflect observations on a non-sodic saline soil. Labanauskas et al. (1978) found increased Mn in wheat grain in response to salinity.
In contrast, Carbonell-Barrachina et al. (1998) found no change in Mn concentration of seeds of the common bean in response to salinity application.
In this experiment, NaCl application under low fertilization resulted in increased Mn for all varieties and levels, except for UDEC-1 at 8 dS m-1 NaCl. This stimulatory effect disappeared under high fertilization, and mixed responses were seen with NaCl under high fertilization. In contrast, increased Mn was seen with Na2SO4 compared to the control, but only under high
-1 fertilization. Under low fertilization, a stimulatory effect appeared at 32 dS m Na2SO4, but at
119
lower concentrations, differences due to variety were predominant. This demonstrates that NaCl and Na2SO4 induced different responses that depend strongly on the level of fertilization.
Of particular note is relatively high Mn concentration of variety CO407D under the
-1 combination of 32 dS m Na2SO4 and high fertilization. Mn concentration at this level was significantly higher than any other treatment combination, though it should be noted that Mn at
-1 32 dS m Na2SO4 under low fertilization was also relatively high.
4.6 P
Karyotis et al. (2003) found no significant difference in the average seed P for a range of quinoa cultivars grown on a saline-sodic soil. Koyro and Eisa (2007) noted that increasing salinity caused an overall increase of P in the seed. However, this differed spatially in the seed, with lower P in the pericarp and elevated P in the cotyledon.
Salinity reactions with P are notably variable and complex, with a large effect from varietal and environmental influences. Of note for this experiment, chloride and sulfate ions have an inhibitory effect on P uptake (Grattan and Grieve, 1999). Findings in this experiment and in others for P concentration in seeds should be taken cautiously, particularly when predicting performance in field conditions.
In this study, varietal differences appeared in response to NaCl salinity. CO407D,
UDEC-1, and Baer either had no change in P or decreases in P, particularly at higher NaCl levels. QQ065 had a contrasting response and saw Zn increases under low fertilization, while it both increased and decreased under high fertilization. However, under Na2SO4 salinity, mixed responses were observed across varieties. Under low fertilization, P either stayed the same or
120
decreased compared to the control.
Baer was notable for its large range of P concentration and relatively high P concentration across treatment combinations. The exception was for NaCl salinity combination with low fertilization, where QQ065 was highest in P.
4.7 Zn
Contrasting responses have been reported for the effect of salinity on Zn in plant shoots
(Grattan and Grieve, 1999). Zn concentration increased for wheat seed from plants grown in saline conditions (Labanauskas et al., 1978). Karyotis et al. (2003) found a decrease in average seed Zn concentration for quinoa varieties grown on a saline-sodic soil.
In this study, increased Zn was found with NaCl compared to the no-salt control, with the
-1 exception of 16 dS m NaCl under high fertility. No change was found in seed Zn with Na2SO4 treatments.
When the two salts were compared, NaCl was generally associated with higher Zn than
-1 -1 Na2SO4. This held for 8 dS m under both fertilization levels, and for 16 and 32 dS m under low fertilization. This confirms findings by Gupta and Gupta (1984), where Zn in plant shoots was higher when salinity was increased using Cl versus SO4 anions.
A few distinct varietal trends appeared across salinity levels. Under low fertilization,
QQ065 was highest in Zn, followed by UDEC-1 which ranked second highest in Zn. As low fertilization is more representative of proper field conditions, QQ065 might be expected to produce seed with higher Zn concentration.
The high positive correlation between Zn and Cu could be attributable to the high amount
121
of Cu and Zn in variety QQ065, which likely skewed the correlation between these two nutrients upwards. This correlation and the negative correlation between Zn and yield might also be due to the effects of dilution. If greater yields resulted in a larger sink for a limited amount of plant Zn and Cu, then lower concentrations in the seeds could be expected.
5. Conclusion
The effects of salinity application, variety, and fertilization were highly complex and interactions were found between all factors for every nutrient analyzed. This matches the general consensus that the effects of salinity on plant nutrition are highly complex and dependent on factors including but not limited to species, variety, salt type, salt strength, and plant tissue type
(Grattan and Grieve, 1999; De Pascale et al., 2005; Martínez-Ballesta et al., 2010). Due to this complexity, only speculation can be made about the underlying mechanisms responsible for the observed responses.
Several large trends came to light, despite the complex set of responses that occurred.
General decreases in Mg and increases in Cu and Zn were found with NaCl treatments.
Differences were seen between the two salt types. Higher seed Ca, Cu, and Zn were found with
NaCl than Na2SO4 at equal EC levels. In the only varietal difference that was consistent across all combinations of salinity and fertility, QQ065 was found to be high in seed Cu.
Fertilization level had a general trend in effect for Ca, Cu, and Mn, with high fertilization resulting in lower nutrient concentration. The results of nutrient changes under high fertilization may be of limited agronomic relevance, as high fertilization and the accompanying plant stress would not be reflective of typical field conditions.
122
From these findings, it appears that mineral concentration of quinoa seed is directly affected by salinity type and strength, soil fertility, and varietal differences, and that the interactions among these factors are particularly complex. Some changes were quite pronounced, with significant changes in the level of seed ineral nutrition of quinoa. These changes in mineral nutrition should be considered if quinoa is developed as a nutrient-rich crop for cultivation in saline affected areas.
123
References
Bernstein, L., L.E. Francois, and R.A. Clark. 1974. Interactive effects of salinity and fertility on yields of grains and vegetables. Agron. J. 66: 412–421.
Carbonell-Barrachina, A.A., F. Burlo, and J. Mataix. 1998. Response of bean micronutrient nutrition to arsenic and salinity. J. Plant Nutr. 21: 1287.
Cramer, G.R. 2002. Sodium-calcium interactions under salinity stress. p. 205–227. In Läuchli, A., Lüttge, U. (eds.), Salinity: Environment - Plants - Molecules.
Curtin, D., H. Steppuhn, and F. Selles. 1993. Plant responses to sulfate and chloride salinity: growth and ionic relations. Soil Sci. Soc. Am. J. 57: 1304–1310.
De Pascale, Stefania, Maggio, Albino, and Barbieri, Giancarlo. 2005. Soil salinization affects growth, yield and mineral composition of cauliflower and broccoli. Eur. J. Agron. 23: 254–264.
Grattan, S.R., and C.M. Grieve. 1999. Mineral nutrient acquisition and response by plants grown in saline environments. p. 203–229. In Handbook of plant and crop stress. 2nd ed., rev. and expanded. Books in soils, plants, and the environment. M. Dekker, New York.
Grattan, S.R., and C.M. Grieve. 1999. Salinity-mineral nutrient relations in horticultural crops. Sci. Hortic. 78: 127–157.
Gupta, M.K., and Gupta, S.P. 1984. Effect of zinc sources and levels on growth and Zn nutrition of soybeam (Glycine max. L.) in the presence of chloride and sulphate salinity. Plant Soil. 81: 299–304.
Karyotis, T.H., C. Iliadis, C.H. Noulas, and T.H. Mitsibonas. 2003. Preliminary research on seed production and nutrient content for certain quinoa varieties in a saline–sodic soil. J Agron Crop Sci. 189: 402–408.
Koyro, H.-W., and S.S. Eisa. 2007. Effect of salinity on composition, viability and germination of seeds of Chenopodium quinoa Willd. Plant Soil. 302: 79–90.
Kurvits, A., and E.A. Kirkby. 1980. The uptake of nutrients by sunflower plants (Helianthus annum) growing in a continuous flowing culture system, supplied with nitrate or ammonium as nitrogen source. Z. Pflanzenernaehr. Bodenkd. 143: 140–149.
Labanauskas, C.K., F.T. Bingham, and A. Cerda. 1978. Free and protein amino acids, and nutrient concentrations in wheat grain as affected by phosphorus nutrition at various salinity levels. Plant Soil. 49: 581–593.
Marschner, H. 1995. Mineral nutrition of higher plants. Academic Press, London; San Diego.
124
Martínez-Ballesta, M.C., R. Dominguez-Perles, D.A. Moreno, B. Muries, C. Alcaraz-López, E. Bastías, C. García-Viguera, and M. Carvajal. 2010. Minerals in plant food: effect of agricultural practices and role in human health. A review. Agron. Sustain. Dev. 30: 295– 309.
Schlick, G., and D.L. Bubenheim. 1996. Quinoa: Candidate crop for NASA’s controlled ecological life support systems. p. 632–640. In Janick, J. (ed.), Progress in new crops. ASHS Press, Arlington, VA.
Vega-Gálvez, A., M. Miranda, J. Vergara, E. Uribe, L. Puente, and E.A. Martínez. 2010. Nutrition facts and functional potential of quinoa (Chenopodium quinoa Willd.), an ancient Andean grain: a review. J Sci Food and Agric. 90: 2541–2547.
125
Table 1: Ca concentration (μg Ca g-1) at 0 dS m-1. Comparisons across fertilization and across variety.
Across fertilization Across variety Variety High fert Low fert High fert Low fert CO407D 508.9 A 409.3 B 508.9 A 409.3 B
UDEC-1 345.6 B 474.6 A 345.6 B 474.6 A Baer 444.7 A 464.5 A 444.7 A 464.5 A QQ065 328.1 B 476.7 A 328.1 B 476.7 A
LSDs significant at p<0.05
126
Table 2: Ca concentration (μg Ca g-1) at treatment combinations.
CO407D UDEC-1 Baer QQ065 Salinity High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 508.9 409.3 345.6 474.6 444.7 464.5 328.1 476.7 A C BC AB A B BC AB 8 dS m-1 402.6 556.5 402.1 505.9 357.4 521.9 390.4 497.4 NaCl B A A A C A A A 16 dS m-1 463.4 498.2 388.6 507.6 409.9 436.1 363.3 434.2 NaCl A B AB A AB BCD AB BC 32 dS m-1 338.2 374.4 369.6 452.0 363.2 367.7 389.6 424.7 NaCl CD CDE AB B BC EFG A C 8 dS m-1 380.8 343.9 342.3 355.8 319.8 389.1 305.2 345.5 Na2SO4 BC DE BC CDE CD DEF C DEFG 16 dS m-1 328.1 332.9 315.3 307.0 345.4 335.8 246.5 332.4 Na2SO4 D E CD FG CD GHI D EFG 32 dS m-1 199.1 234.6 266.8 227.0 293.65 257.8 202.7 232.6 Na2SO4 E F D H D J D H
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
127
Table 3: Cu concentration (μg Cu g-1) at 0 dS m-1. Comparisons across fertilization and across variety.
Across fertilization Across variety
Variety High fert Low fert High fert Low fert CO407D 3.75 B 4.76 A 3.75 D 4.76 C UDEC-1 3.51 B 5.68 A 3.51 D 5.68 B
Baer 2.96 B 5.21 A 2.96 E 5.21 BC QQ065 5.67 B 6.66 A 5.67 B 6.66 A
LSDs significant at p<0.05
128
Table 4: Cu concentration (μg Cu g-1) at treatment combinations.
CO407D UDEC-1 Baer QQ065 Salinity High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 3.75 C 4.76 E 3.51 E 5.68 BC 2.96 D 5.21 C 5.67 E 6.66 C
8 dS m-1 4.55 B 6.44 B 5.16 B 6.61 A 4.01 BC 6.38 AB 7.31 C 9.28 A NaCl 16 dS m-1 5.08 A 6.98 A 4.79 BC 7.07 A 4.52 AB 6.63 A 7.48 BC 9.33 A NaCl 32 dS m-1 5.32 A 6.58 AB 5.70 A 6.76 A 4.83 A 6.24 AB 8.04 AB 9.20 A NaCl 8 dS m-1 3.74 C 5.04 DE 4.075 D 5.53 BC 3.60 C 6.05 B 6.43 D 6.53 C Na2SO4 16 dS m-1 4.99 AB 5.66 C 4.31 CD 5.92 B 3.69 C 6.04 B 6.28 D 6.92 C Na2SO4 32 dS m-1 5.55 A 5.58 CD 4.36 CD 5.18 C 4.94 A 6.10 AB 8.12 A 7.62 B Na2SO4
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
129
Table 5: Fe concentration (μg Fe g-1) at 0 dS m-1. Comparisons across fertilization and across variety.
Across fertilization Across variety
Variety High fert Low fert High fert Low fert CO407D 69.84 A 69.84 A 69.84 A 69.84 B UDEC-1 54.51 B 54.51 A 54.51 C 54.51 A
Baer 63.43 A 63.43 A 63.43 B 63.43 B
QQ065 65.35 A 65.35 A 65.35 AB 65.35 B
LSDs significant at p<0.05
130
Table 6: Fe concentration (μg Fe g-1) at treatment combinations.
CO407D UDEC-1 Baer QQ065 Salinity High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 69.84 64.61 54.51 72.20 63.43 62.76 65.35 65.28 AB AB B A BC C B B 8 dS m-1 65.51 68.47 63.86 65.68 62.88 65.93 51.35 58.07 NaCl BC A A B BC C C C 16 dS m-1 63.41 69.05 67.43 62.04 59.48 52.65 53.60 49.62 NaCl C A A BC C D C D 32 dS m-1 56.90 65.59 67.15 57.83 71.60 61.87 43.20 49.36 NaCl D AB A C A C D D 8 dS m-1 66.12 70.46 68.37 72.30 67.41 94.00 82.87 72.36 Na2SO4 ABC A A A AB A A A 16 dS m-1 72.32 70.16 65.39 72.56 73.57 85.23 82.17 66.70 Na2SO4 A A A A A B A AB 32 dS m-1 65.00 59.53 64.52 55.46 60.45 67.13 64.09 56.43 Na2SO4 ABC B A C BC C B CD
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
131
Table 7: Mg concentration (μg Mg g-1) at 0 dS m-1. Comparisons across fertilization and across variety.
Across fertilization Across variety Variety High fert Low fert High fert Low fert
CO407D 2458 A 2483 A 2458 A 2483 BC UDEC-1 2385 B 2554 A 2385 A 2554 B Baer 2415 B 2790 A 2415 A 2790 A
2220 B 2397 A 2220 B 2397 C QQ065
LSDs significant at p<0.05
132
Table 8: Mg concentration (μg Mg g-1) at treatment combinations.
CO407D UDEC-1 Baer QQ065 Salinity High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 2458 2483 2385 2554 2415 2790 2220 2397 A AB A A AB A BC AB 8 dS m-1 2336 2596 2307 2442 2305 2546 2181 2447 NaCl AB A A A BC C C AB 16 dS m-1 2320 2447 2137 2433 2141 2353 2037 2513 NaCl B B B A D D D A 32 dS m-1 1785 2280 1965 2305 1996 2186 2329 2467 NaCl C C C B E E ABC AB 8 dS m-1 2361 2481 2396 2465 2508 2721 2330 2372 Na2SO4 AB AB A A A AB AB B 16 dS m-1 2407 2521 2391 2517 2408 2654 2381 2454 Na2SO4 AB AB A A ABC BC A AB 32 dS m-1 2292 2288 2115 2403 2259 2628 2256 2433 Na2SO4 B C BC AB CD BC ABC AB
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
133
Table 9: Mn concentration (μg Mn g-1) at 0 dS m-1.
Across fertilization Across variety Variety High fert Low fert High fert Low fert CO407D 56.93 A 54.93 A 56.93 ABC 54.93 BC
UDEC-1 60.82 A 63.23 A 60.82 AB 63.23 A
Baer 34.70 B 53.42 A 34.70 E 53.42 CD QQ065 47.85 A 52.15 A 47.85 D 52.15 CD
LSDs significant at p<0.05
134
Table 10: Mn concentration (μg Mn g-1) at treatment combinations. Comparisons across fertilization and across variety.
CO407D UDEC-1 Baer QQ065 Salinity High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 56.93 54.93 60.82 63.23 34.70 53.42 47.85 52.15 D C BC B E D AB BC 8 dS m-1 62.42 72.48 69.47 62.40 40.53 64.77 52.21 59.91 NaCl CD AB A B CDE B AB A 16 dS m-1 64.96 72.39 58.36 72.21 42.12 61.35 50.01 56.65 NaCl BC AB CD A CD BC AB AB 32 dS m-1 45.12 66.00 51.95 73.59 40.43 57.06 44.33 59.47 NaCl E B D A DE CD B A 8 dS m-1 58.56 66.59 71.55 60.95 47.16 59.51 48.99 39.34 Na2SO4 CD B A B BC BCD AB D 16 dS m-1 69.53 68.70 66.71 62.45 52.84 62.25 52.60 47.89 Na2SO4 B B AB B B BC A C 32 dS m-1 93.57 78.47 65.34 68.82 63.80 83.14 55.49 52.61 Na2SO4 A A ABC AB A A A ABC
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
135
Table 11: P concentration (μg P g-1) at 0 dS m-1. Comparisons across fertilization and across variety.
Across fertilization Across variety
Variety High fert Low fert High fert Low fert CO407D 5337 A 5174 A 5337 B 5174 BC UDEC-1 5298 A 5231 A 5298 B 5231 BC
Baer 5601 A 5757 A 5601 A 5757 A
QQ065 5035 B 5310 A 5035 C 5310 B
LSDs significant at p<0.05
136
Table 12: P concentration (μg P g-1) at treatment combinations.
CO407D UDEC-1 Baer QQ065 Salinity High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 5337 5174 5298 5231 5601 5757 5035 5310 A A A A B A CD B 8 dS m-1 5062 5189 5145 5016 5548 5385 5172 5654 NaCl BC A AB BC BC C BCD A 16 dS m-1 4993 5012 4977 5084 5271 5127 4975 5630 NaCl C ABC BCD ABC D D D A 32 dS m-1 4636 4887 4796 4880 5289 5083 5443 5528 NaCl D BC D C D D A A 8 dS m-1 5149 5105 5117 5023 5872 5617 5286 5222 Na2SO4 ABC A ABC BC A AB AB B 16 dS m-1 5241 5046 5080 5147 5536 5469 5222 5201 Na2SO4 AB AB BC AB BC BC ABC B 32 dS m-1 4966 4790 4886 5085 5308 5768 5096 5118 Na2SO4 C C CD ABC CD A BCD B
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
137
Table 13: Zn concentration (μg Zn g-1) at 0 dS m-1. Comparisons across fertilization and across variety.
Across fertilization Across variety Variety High fert Low fert High fert Low fert CO407D 49.70 A 40.49 B 49.70 A 40.49 EF
UDEC-1 44.23 A 45.77 A 44.23 CD 45.77 BC Baer 41.82 A 37.88 B 41.82 DE 37.88 F QQ065 49.32 A 47.84 A 49.32 A 47.84 AB
LSDs significant at p<0.05
138
Table 14: Zn concentration (μg Zn g-1) at treatment combinations.
Salinity CO407D UDEC-1 Baer QQ065 High Low High Low High Low High Low Fert. Fert. Fert. Fert. Fert. Fert. Fert. Fert. Control 49.70 40.49 44.23 45.77 41.82 37.88 49.32 47.84 BC E D D C E E C 8 dS m-1 49.84 50.37 55.00 51.51 45.63 48.09 54.52 63.73 NaCl BC B B BC B BC CD A 16 dS m-1 49.00 53.15 55.18 57.36 50.37 50.30 59.42 63.61 NaCl C A B A A B B A 32 dS m-1 52.71 53.31 60.16 53.86 53.00 53.70 57.88 65.60 NaCl A A A B A A BC A 8 dS m-1 45.56 42.59 49.51 45.37 43.42 44.84 53.40 48.05 Na2SO4 D DE C D BC D D C 16 dS m-1 51.99 46.29 50.80 49.13 42.85 46.72 52.15 50.12 Na2SO4 AB C C C C CD D C 32 dS m-1 44.64 45.59 49.27 49.17 46.43 48.48 63.71 58.65 Na2SO4 D CD C C B BC A B
LSDs significant at p<0.05, comparisons down columns for each combination of variety and fertilization level
139
Table 15: Pearson correlation coefficients for response variables for all treatment combinations.
yield leaf height Ca Cu Fe Mg Mn P greenness leaf 0.62*** greenness height 0.60*** 0.64*** Ca 0.06 ns -0.05 ns 0.31* Cu -0.50*** -0.25† 0.10 ns 0.14 ns Fe 0.36** 0.33* 0.30* -0.10 ns -0.30* Mg 0.46*** 0.55*** 0.66*** 0.24 ns 0.08 ns 0.32* Mn 0.42** 0.21 ns 0.43** -0.08 ns 0.06 ns 0.05 ns 0.29* P 0.02 ns 0.28* 0.05 ns 0.17 ns 0.02 ns 0.10 ns 0.60*** -0.27* Zn -0.57*** -0.52*** -0.39** 0.06 ns 0.73*** -0.40** -0.36** -0.06 ns -0.16 ns ***=p<0.001, **=p<0.01, *=p<0.05, †=p<0.07, ns=not significant
140
Table 16: Pearson correlation coefficients for response variables, NaCl under high fertilization.
yield leaf height Ca Cu Fe Mg Mn P greenness leaf 0.74** greenness height 0.90*** 0.82** Ca 0.41 ns -0.09 ns 0.37 ns Cu -0.57† -0.38 ns -0.40 ns -0.09 ns Fe 0.56† 0.51 ns 0.39 ns 0.05 ns -0.81** Mg 0.41 ns 0.15 ns 0.42 ns 0.65* -0.04 ns -0.13 ns Mn 0.61* 0.43 ns 0.74** 0.61* -0.09 ns 0.25 ns 0.42 ns P 0.02 ns 0.03 ns -0.06 ns 0.11 ns -0.01 ns -0.19 ns 0.64* -0.35 ns Zn -0.24 ns -0.07 ns -0.23 ns -0.26 ns 0.71** -0.30 ns -0.37 ns 0.06 ns -0.36 ns ***=p<0.001, **=p<0.01, *=p<0.05, †=p<0.07, ns=not significant
141
Table 17: Pearson correlation coefficients for response variables, NaCl under low fertilization.
yield leaf height Ca Cu Fe Mg Mn P greenness leaf 0.38 ns greenness height 0.83*** 0.47 ns Ca 0.41 ns 0.47 ns 0.80** Cu -0.84*** -0.08 ns -0.51 ns -0.08 ns Fe 0.71** -0.02 ns 0.68 ns 0.43 ns -0.69 ns Mg 0.03 ns 0.21 ns 0.46 ns 0.79** 0.31 ns 0.07 ns Mn 0.40 ns -0.15 ns 0.53 ns 0.51 ns -0.48 ns 0.58* 0.16 ns P -0.56† 0.11 ns -0.27 ns 0.14 ns 0.82** -0.55† 0.54 ns -0.59* Zn -0.91*** -0.17 ns -0.65* -0.24 ns 0.94*** -0.68* 0.10 ns -0.44 ns 0.67* ***=p<0.001, **=p<0.01, *=p<0.05, †=p<0.07, ns=not significant
142
Table 18: Pearson correlation coefficients for response variables, Na2SO4 under high fertilization.
yield leaf height Ca Cu Fe Mg Mn P greenness leaf 0.57† greenness height 0.76** 0.90*** Ca 0.53 ns -0.14 ns 0.05 ns Cu -0.75** -0.37 ns -0.47 ns -0.75** Fe -0.10 ns -0.03 ns -0.04 ns 0.10 ns 0.24 ns Mg 0.29 ns 0.12 ns 0.28 ns 0.51 ns -0.35 ns 0.34 ns Mn 0.28 ns 0.35 ns 0.50 ns -0.32 ns -0.06 ns -0.44 ns -0.28 ns P -0.04 ns 0.07 ns 0.00 ns 0.39 ns -0.32 ns 0.22 ns 0.71** -0.62* Zn -0.44 ns -0.45 ns -0.44 ns -0.46 ns 0.83*** 0.13 ns -0.30 ns -0.17 ns -0.38 ns ***=p<0.001, **=p<0.01, *=p<0.05, †=p<0.07, ns=not significant
143
Table 19: Pearson correlation coefficients for response variables, Na2SO4 under low fertilization.
yield leaf height Ca Cu Fe Mg Mn P greenness leaf 0.17 ns greenness height 0.70** 0.46 ns Ca 0.44 ns 0.32 ns 0.76** Cu -0.77** -0.33 ns -0.80** -0.29 ns Fe 0.36 ns 0.37 ns 0.34 ns 0.66* -0.01 ns Mg 0.31 ns 0.61* 0.24 ns 0.41 ns 0.02 ns 0.77** Mn 0.44 ns 0.08 ns 0.02 ns -0.46 ns -0.43 ns -0.15 ns 0.15 ns P 0.01 ns 0.50 ns 0.00 ns 0.21 ns 0.21 ns 0.57* 0.84*** 0.07 ns Zn -0.82*** -0.21 ns -0.88*** -0.65* 0.85*** -0.37 ns -0.16 ns -0.20 ns 0.01 ns ***=p<0.001, **=p<0.01, *=p<0.05, ns=not significant
144
CHAPTER FOUR
NITROGEN USE EFFICIENCY OF QUINOA UNDER ORGANIC CONDITIONS IN SOUTHEASTERN WASHINGTON
Nitrogen Use Efficiency Of Quinoa Under Organic Conditions In Southeastern Washington
Abstract
As quinoa cultivation expands into new areas in North America, it will be useful to characterize its response to fertilization in novel environments. Existing research indicates that quinoa’s nitrogen dynamics vary depending on environment and variety. The objective of this study was to determine the responses of a diverse range of 16 quinoa varieties to fertilization under organic cultivation in Washington state. In 2011, a field experiment was conducted with a high-nitrogen organic fertilizer applied at 0, 50, 100, and 150 kg N ha-1. In 2012, a modified version of the experiment was conducted due to low germination. Five plots with adequate germination were split into two halves, one thinned and one not thinned.
Several interfering environmental factors prevented the determination of nitrogen dynamics in both years. In 2011, large reductions in seed set appear to have been caused by high temperatures later in the growing season. However, significant differences in heat tolerance among varieties were potentially revealed. Variation in valuable traits such as days to full senescence was observed as well. In 2012, yield and leaf greenness were not different between thinned and unthinned treatments, but a marginally significant difference in leaf nitrogen indicated greater nitrogen uptake for plants in thinned plot halves.
1. Introduction
Quinoa cultivation in North America has so far been limited to Colorado and other high- altitude locations in the American West, as well as the Canadian Prairies. Early research at
146
Colorado State University generated fertilization recommendations of 135 kg N ha-1, later revised upwards to 170-200 kg N ha-1 (Johnson and Croissant, 1990; Oelke et al., 1992).
However, it remains to be seen if those recommendations hold for other regions as quinoa cultivation expands in North America and new varieties are developed from different germplasm.
Differing reports exist for quinoa's response to fertilization. In Denmark, yield response of quinoa at 40 kg N ha-1 was 24.1% lower than that at 160 kg N ha-1. Increased nitrogen provided decreasing benefits, with only 12.0% and 2.7% decreases in yield at 80 and 120 kg N ha-1 compared to 160 kg N ha-1 (Jacobsen et al., 1994). In a field trial in southern Germany, Schulte auf’m Erley et al. (2005) found that yield nearly doubled from 1790 kg ha-1 under the non- fertilized control to 3495 kg ha-1 under 120 kg N ha-1. In the same experiment, significant differences in nitrogen uptake efficiency (g yield g-1 plant N) were found between the cultivars
Faro and Cochabamba. Increasing fertilization did not impact nitrogen uptake efficiency. This contrasts with the results of a greenhouse study by Thanapornpoonpong (2004) who found decreasing nitrogen uptake efficiency with greater nitrogen application. Additionally, differences in nitrogen use efficiency (g yield g-1 N) were found between the two cultivars used in this study.
Soil texture also appears to play a role in the nitrogen uptake dynamics of quinoa. Razzaghi et al.
(2012) found nitrogen use efficiency was highest under soil types with greater percentages of clay.
The complicated picture of nitrogen responses and varietal differences indicates the need for further investigation of quinoa's nitrogen dynamics in novel environments with a wide range of germplasm.
147
Many who are attracted to quinoa's nutritional and health benefits are predominately interested in organically grown quinoa. Quinoa has also drawn a great amount of interest from organic farmers, particular due to its reputation as a low-input, hardy crop. However, only one study has so far been reported for quinoa grown under organic conditions (Bilalis et al., 2012).
Nitrogen dynamics in organic agriculture differ from those in conventional production, with greater dependence on biological factors such as mineralization of organic matter (Davis and
Abbott, 2006). By testing quinoa's response to fertilization under organic conditions, the goal of this experiment was to collect information on the quinoa varieties currently part of the WSU quinoa program and to determine their suitability for cultivation under organic cultivation in
Washington state.
2a. Materials and Methods – 2011 Quinoa NUE Experiment
2a.1 Quinoa varieties
Sixteen quinoa varieties were included in the experiment (Table 1). These represent the best performing varieties from initial field trials in 2010. Several commercially available varieties in the US and Canada were also included.
2a.2 Experimental design
A split-plot experiment was conducted at the WSU Organic Farm in Pullman, WA
(latitude 46.73° N, longitude 117.13° W) on a Palouse silt loam soil (fine-silty, mixed, superactive, mesic Pachic Ultic Haploxerolls). The climate is characterized by hot, dry summers and cold, wet winters, with an average annual precipitation of 540 mm.
148
The two factors in this experiment were variety and fertilization. There were four total fertilization levels. This included a non-fertilized control and three levels of Perfect Blend 7-2-2
(Perfect Blend Biotic Fertilizers, Bellevue WA) applied at 50, 100, and 150 kg N ha-1. Soil samples were taken from each of the blocks on June 22nd to determine existing nitrogen supply in the field. Ten samples were taken randomly within each block and combined into a single sample.
The site was 15.8 m x 83.5 m and was split into three blocks along its width, perpendicular to the downward slope along the length of the field. Quinoa variety constituted the whole plot, while fertilizer level constituted the subplot. Plots were 1.2 m x 3.0 m and had four rows spaced 0.25 m apart. Furrows were made by tractor. Fertilizer was broadcast by hand into the rows on June 2nd.
The site was planted by hand in early June. The seeding rate was 5 g per plot (3.7 m2).
The first block was planted by hand June 3-4, the second June 4-5, and the third June 9-10.
Plants were thinned in mid-July to within-row spacing of 10 cm.
The appearance of flower buds, equivalent to phenological stage 2 as described by
Jacobsen and Stolen (1993), was recorded for each variety. Leaf greenness was measured on
August 30, 2011 using a Konica Minolta SPAD-502 Plus chlorophyll meter (Minolta
Camera Co., Ltd., Osaka, Japan). A minimum of ten healthy leaves was measured from the top quarter of the plant at the base of the inflorescence and the values obtained were averaged. At the time leaf greenness was measured, the majority of plants were at phenological stage 13 as described by Jacobsen and Stolen (1993). Aphid susceptibility was measured on August 11,
2011. Percent plot coverage was taken at this time in order to adjust yield measurements, as
149
many plots did not have full germination. Plant height was recorded during senescence. A minimum of ten plants were randomly chosen from each of the plots and the height measured from the soil level to the top of each plant. These values were averaged to obtain an average height value for each plot.
Plots were harvested on October 1, 2011 with a 1999 Wintersteiger Nursery Master Elite plot combine. Due to the high level of extraneous material that was harvested, the seed was further threshed with a wheat head thresher manufactured by Precision Machine Company
(Lincoln, NE) and with a custom built belt thresher located at Western Regional Plant
Introduction Station (Pullman, WA). Seed was cleaned in a Clipper Office Tester (Seedburo, Des
Plaines IL). Yield was adjusted according to the percentage of plot coverage. Yield was recorded as 0 g for plots with <20% plot emergence to avoid unrealistically inflating yield values. Plots with low emergence had low yield and often contained stunted plants.
Days to complete senescence, corresponding to phonological stage 21 as described by
Jacobsen and Stolen (1993), were measured for the varieties. Seed for all varieties matured, although some varieties were not completely dry before harvest. Only maturity dates for varieties that had completely senescence by the harvest date were included in the final analysis.
Soil samples were collected on October 3, 2011. For each block, one sample 15 cm deep was taken from each of sixteen the plots receiving 150 kg N/ha. All sixteen samples, one taken from each variety, were combined into one sample. The same was conducted for the control plots receiving no fertilizer. Soil samples were sent to the Analytical Sciences Laboratory at the
University of Idaho (Moscow, Idaho) for analysis. All soil samples were analyzed for NO3-N,
NH4-N, P, K, Cu, Fe, Mn, Zn, organic matter, and pH.
150
2a.3 Statistical Analysis
PROC MIXED in SAS (SAS Institute, Cary NC) was used to analyze data. Block and variety by block interaction were treated as random factors. Variety and fertilization were treated as fixed factors. Appropriate error terms were used. PROC UNIVARIATE in SAS (SAS Institute,
Cary NC) was used to assess normality. Pearson correlation coefficients were generated using
PROC CORR in SAS (SAS Institute, Cary NC) using the least square means generated from
PROC MIXED.
2b. Materials and Methods - 2012 Modified Quinoa NUE Experiment
2b.1 Experimental design
The same experiment was planted on May 16, 2012 at Boyd Farm in Pullman, WA
(latitude 46.75 N, longitude 117.08 W) on a Palouse silt loam soil (fine-silty, mixed, superactive, mesic Pachic Ultic Haploxerolls). A second site was planted on May 17, 2012 at the WSU
Organic Farm in Pullman, WA (latitude 46.73 N, longitude 117.13 W) on a Palouse silt loam soil.
The only modification from the previous year was the variety selection (Table 2). Several varieties had produced insufficient seed the previous year for replanting and were replaced with new varieties.
At both sites, germination was greatly impaired or completely reduced for plots that received fertilizer. This was due to the method of planting in which seed and fertilizer was planted together using a mechanical seeder. Only the control plots receiving no fertilization had adequate levels of emergence at Boyd Farm. However, this varied due to poor soil conditions
151
around planting time. At the WSU Organic Farm site, emergence was low for all plots and the experiment was abandoned.
A modified approach was taken at the Boyd site to retrieve useful data on nutrient competition. Five plots that had high levels of emergence and even plot coverage were divided into two plot halves. One plot half was chosen at random to be thinned and the other plot half was left unthinned. The five plots were treated as replicates and the experiment was treated as an
RCBD. As different varieties were in each of the replications, the aim of the experiment shifted towards finding effects of nutrient competition in quinoa as a whole and not on variety specific effects.
On July 12, 2012, plants within one half of the plot, randomly chosen, were thinned to approximately 10 cm within-row spacing. The other plot half was left unthinned. Soil samples were taken on July 2, 2012, with ten samples taken from 0-30 cm and combined and another ten samples taken from 30-61 cm and combined.
On July 31, 2012, leaf tissue samples were taken from the upper quarter of the plant, just below the inflorescence. At this stage, most plants were at phenological stage 13 as described by
Jacobsen and Stolon (1993). Three leaves were taken from 10 plants each from each plot half.
These were dried at 60˚C for 72 hours. A second set of tissue samples was taken on August 28,
2012, when the plants were in the seed fill stage, corresponding roughly with phenological stages
16-17 (Jacobsen and Stolen, 1993).
Plant height was measured on a minimum of ten plants chosen randomly per plot. These heights were averaged to determine plant height. Maturity date was recorded for each plot half.
152
The plots were hand harvested on September 26, 2012 and threshed in a Vogel thresher. Seed was cleaned in a Clipper Office Tester (Des Plaines, IL).
Due to the lack of rain into late autumn and the difficulty of sampling the hard soil, final soil samples could not be taken until late in the season. On November 7, 2012, adequate rain had allowed samples to be taken 0-30 cm. Beyond 30 cm, the ground was still dry and further sampling was not possible.
All soil samples were analyzed by a Lachat flow injection analyzer (Lachat Instruments;
Loveland, CO) for NO3-N and NH4-N. Analysis was conducted at the Lachat Service Center in the Department of Crop and Soil Sciences at Washington State University (Pullman, WA). Tissue samples were analyzed for N, P, K, Ca, Mg, Mn, Zn, Fe, B, Na, and Al at Brookside Labs (New
Bremen, OH).
2b.2 Statistical Analysis
PROC MIXED in SAS (SAS Institute, Cary NC) was run to analyze data on yield, height, and days to maturity, along with tissue and soil sample data. Yield data lacked normality and was log transformed before analysis. Replication was treated as a random factor and thinning and sampling date was treated as fixed factors. Appropriate error terms were used. R
FOURNormality was tested via the PROC UNIVARIATE procedure in SAS (SAS Institute, Cary
NC). Due to a lack of normality, yield data was log transformed.
3a. Results – 2011 NUE Experiment
3a.1 Yield
153
No differences were found in yield across fertilization treatment (p=0.780) (Table 3).
Differences were seen among varieties (p=0.0002). Yield varied from a high of 25.80 g per plot with QQ74, to a near complete lack of seed production at 0.16 g per plot with QQ065 (Figure 1).
Of the 16 varieties, 9 yielded less than 2 g per plot. The remaining varieties, QQ74, Colorado
407D, Kaslaea, Cherry Vanilla, Red Head, BBR, and Baer did not differ significantly in yield.
3a.2 Plant height
Plant height did not vary due to fertilization level (p=0.400), but there were significant differences among varieties (p=0.039). Height ranged from 108 cm to 81 cm (Table 4).
3a.3 Aphid susceptibility and leaf greenness
Fertilization had no effect on either the level of aphid susceptibility (p=0.992) or leaf greenness (p=0.628).
3a.4 Days to appearance of flowering bud
Variety had a highly significant difference on days to first appearance of the flowering bud (p<0.0001). The earliest variety was Black-seeded, at 30 days, and the latest was QQ63, at
36 days (Table 4).
3a.5 Days to full senescence
For varieties that had completely senesced by the time of harvest, there was a marginally significant difference in maturity times (p=0.061) (Table 5). QQ63 had a longer time to full
154
senescence than the rest (p=0.05), at 111 days to maturity. Baer showed no significant differences from any of the other varieties, while varieties UDEC-1, Kaslaea, Brightest Brilliant
Rainbow, Cherry Vanilla, QQ065, Red Head, and UDEC-3 all ranked earliest.
3a.6 Correlations
Several weak but statistically significant correlations were found (Table 6). Plant height was positively correlated with leaf greenness (r=0.33) and yield (r=0.41). Days to flowering bud appearance was negatively correlated with both plant height (r=-0.37) and yield (r=-0.42). Days to full senescence was positively correlated with plant height (r=0.34), though this correlation only holds for the subset of varieties that fully senesced by harvest.
3a.7 Soil samples
There were no significant differences in the measured soil nutrients or soil characteristics between the non-fertilized control (0 kg N/ha) and the highest fertilization treatment (150 kg
N/ha). When measurements at the end of the season were compared to the initial samples, only
Mn and Zn levels were different, being higher later in the season regardless of fertilizer application.
3b. Results – 2012 Modified Quinoa NUE Experiment
3b.1 Main response variables
Yield, plant height, leaf greenness, and days to full senescence did not differ significantly between thinning treatments. Only aphid susceptibility was different between the two plots
155
(p=0.015). Thinned plots (5.3/10) were rated more susceptible to aphids than unthinned plots
(2.7/10).
3b.2 Tissue samples
Of the nutrients tested, N, P, Cu, and Zn were significantly lower when measured later in the season. Mg, Ca, Fe, Mn, and Al were significantly higher later in the season (Table 7). No significant differences were seen with K, S, and B between dates.
Thinning had no significant effect on the biologically significant nutrients tested. A marginally significant effect (p=0.064) was seen in nitrogen concentration. Nitrogen was higher under the thinned plots (2.25%) compared to unthinned plots (1.77%).
3b.3 Soil samples
There were no significant differences in NO3-N or NH4-N between the thinned and unthinned plot half. When the thinned and unthinned plots were compared with the initial samples in spring, no significant changes in NH4-N were detected. However, NO3-N increased significantly. Thinned plots sampled on November 7th had NO3-N of 6.60 ppm compared to 2.17 ppm in spring (p=0.0002). For unthinned plots, NO3-N was 8.258 ppm compared to 2.17 ppm in spring (p<0.0001).
4a. Discussion – 2011 NUE Experiment
4a.1 Yield
156
Very low yields were seen for all varieties, likely due to the effects of heat stress from high temperatures occurring later in the growing season (Figure 2). According to field observations, most plants were full of empty seeds and few plants exhibited high levels of successful seed set.
In early field trials in Colorado, temperatures exceeding 35ºC (95ºF) were found to cause pollen sterility (Johnson and Croissant, 1990). Reports from Morocco, Greece, and Northern
Chile have also found heat to be a significant impacting factor for yield (Iliadis et al., 2001;
Benlhabib et al., 2004; Fuentes and Bhargava, 2011). Heat later in the season has also been found to affect seed fill. Bonifacio (1995) noted that high temperatures can cause the reabsorption of endosperm (likely referring to the perisperm which contains the main starch reserves of the seed, as quinoa lacks an endosperm of appreciable size) in developing seeds. High maximum temperatures exceeding 30ºC occurred later in the growing season during the seed fill stages.
This likely contributed to the empty seeds observed on many of the plants.
Fertilizer was not found to have an effect on yield. Quinoa has been found to respond the fertilization at levels similar to those used in the current experiment (Johnson and Croissant,
1990; Jacobsen et al., 1994; Schulte auf’m Erley et al., 2005). The most likely scenario is that the effects of heat stress acted as a confounding factor.
The significant effect of variety on yield was likely attributable to differential heat tolerance among varieties. When varieties matched up with their location or background of origin from GRIN passport data, a trend appeared. Among the group of cultivars that yielded more than 2 g per plot, most originated from locations characterized by high summer temperatures. QQ74 and Baer originate from the inland Chilean Central Valley, which
157
experiences the highest temperatures within the geographic distribution of Chilean lowland varieties (Dirección Meteorológica de Chile). Red Head, Cherry Vanilla, and Brightest Brilliant
Rainbow were bred in the Willamette Valley of Oregon, which is known to experience temperatures exceeding that of the reported tolerance threshold for quinoa.
4a.2 Plant height
Fertilization appeared to have no significant impact on plant height, although the lack of height differences across fertilization could also be due to the confounding effects of heat stress.
Oelke et al. (1992) noted that high temperatures can cause vegetative dormancy. If heat stress was the main limiting factor for growth, no increase in height would be expected under increased fertilization.
Moderately weak correlations were found between plant height and yield (r=0.41) and with plant height and leaf greenness (r=0.33). This may reflect the correlated levels of stress in plants with different levels of heat tolerance. For instance, taller varieties might be less stunted and yield better due to superior heat tolerance. Another possibility is that taller plant height may provide adaptation to heat stress. Risi and Galwey (1984) quote South American sources which report that root length is proportional to plant height. Deeper rooting depth may allow for greater access to water deeper in the soil profile, which may allow the plants to better cope with the effects of heat stress. Another potential supporting piece of evidence is a significant but weak positive correlation between plant height and days to full senescence (r=0.34). If shorter varieties had less access to water in the soil profile due to their shorter rooting depth, this may have resulted in earlier senescence.
158
4a.3 Days to appearance of flowering bud
The differences in time to flowering bud appearance were consistent with results by Risi and Galwey (1991) where a wide range of cultivars of several ecotypes were found to vary for days to anthesis and days to maturity. Jacobsen (1998) also reported diversity in timing for phenological stages among a group of mostly Chilean lowland cultivars. Early time to the initiation of flowering could prove an important trait for quinoa as it would allow for temporal separation between flowering and the onset of high temperatures. In Morocco, early flowering quinoa varieties were able to survive a 47ºC heat wave through temporal avoidance (Benlhabib et al., 2004). In our trials, early flowering seemed to have a beneficial impact. Increased days to flowering bud appearance was negatively correlated with yield (r=-0.42) and plant height (r=-
0.37). Temperatures during 2011 only exceeded the 35ºC threshold reported by Johnson and
Croissant (1990) in late August, well past the flowering stage. If early initiation of flowering was linked to earlier seed fill, which heat has been reported to affect, then this correlation could be explained by escape of plants to high temperatures (>35ºC) occurring later in the season (Figure
2).
4a.4 Days to full senescence
Results for this trait are limited to early maturing varieties, as full senescence was not reached by all varieties at harvest time. Plants of some varieties remained partly green, despite full maturation of the seed by harvest. Despite the differences that were found, a majority of the cultivars grouped together as the earliest to fully senesce. No cultivars ranked as significantly
159
early, which will be crucial in breeding for areas with short growing seasons or limiting environmental pressures. In the Dutch quinoa breeding program, maturity and flowering time were found to significantly vary and be governed largely by additive variation (Mastebroek et al.,
2002). If maturity time could be selected upon to generate earlier maturing cultivars, this would reduce the current risk of pre-harvesting sprouting due to quinoa’s relatively long maturity times or of early frost in areas with short growing seasons.
4b. Discussion - 2012 Modified Quinoa NUE Experiment
4b.1 Main response variables
The lack of significant differences for all but aphid susceptibility may be attributable to the range of varieties included. The significant effect of thinning on aphid susceptibility matched observations in the field, where thinned plot halves fared much worse than those left unthinned.
This could partly be explained by the increased exposure of thinned plants. Unthinned plants were more densely packed in rows and helped to shade each other, while plants in the thinned rows may have experienced increase environmental stress, making them more susceptible to pest pressures. This confirms with observations in the 2011 experiment, where isolated plants were more susceptible to aphids.
Despite the lack of difference between thinning treatments for yield, heat stress likely resulted in greatly reduced seed set. Temperatures in 2012 were higher than in 2011 (Figures 2 &
3). However, for several varieties not included in the experiment due to lack of sufficient plot coverage, significantly higher levels of seed set were observed among plants. This suggests that natural selection acted on varieties grown in 2011 and selected for greater heat tolerance.
160
4b.2 Tissue samples
The significant changes in nutrient concentration across sampling dates were consistent with the remobilization of nutrients from the vegetative portion of the plants to the seeds as they developed. Nitrogen, P, and Cu were lower in leaves at the later sampling date, matching observations in other crops (Marschner, 1995). The increase in Ca, Mg, and Mn, along with the decrease in Cu corresponds with observations in senescing soybean leaves (Wood et al., 1986).
Thinning appeared to have little effect on nutrient status, although there was a marginally significant effect on nitrogen concentration (p=0.064). The higher level of nitrogen in leaves of thinned plants indicates reduced competition for nitrogen in the thinned plot halves.
4b.3 Soil samples
The significant increase in NO3-N seen at the later sampling date is attributable to oxidation of organic matter in the soil. The lack of differences in NO3-N between the thinned and unthinned plot halves may be due to the volatility of NO3 and the wide gap between harvest and the later sampling date, rather than the lack of differences in nitrogen uptake.
5a. Conclusion – 2011 NUE Experiment
High temperatures were likely the main limiting factor for cultivation of quinoa in our location in 2011. Due to the confounding effects of heat, little information was obtained on quinoa's response to fertilization. However, important information was collected on the relative levels of heat tolerance among quinoa varieties. For many varieties, heat tolerance appeared linked to their place of origin. Variance in growing season length and time to initiation of
161
flowering was also found for many of the varieties. Adjusting days to maturity and days to different phonological stages will prove useful in areas with short growing seasons or where early flowering may prevent avoidance of heat during crucial periods.
5b. Conclusion - 2012 Modified Quinoa NUE Experiment
The negative effect of fertilizer when applied in contact with the seed appears particularly severe with quinoa and underscores the importance of distancing seed from fertilization applications either spatially or temporally, applied after germination. Additionally, a fine seedbed is important for proper germination and establishment. Inclement weather near the planting date caused soil clodding in the field, disrupting germination in plots that had received no fertilization. The modified Quinoa NUE experiment revealed increased aphid pressure due to the effects of thinning, indicating that thick stands provide a protective effect for plants. Despite the lack of yield differences between thinning treatments, high heat late in 2012 resulted in reduced yields in many of the plot halves. However, several varieties not included in the modified Quinoa
NUE experiment had much improved seed set in plants compared to the previous year. This suggests that natural selection has acted on the genetic diversity present in many of the quinoa accessions and that heat tolerance has improved.
162
References
Benlhabib, O., M. Atifi, E.N. Jellen, and S. Jacobsen. 2004. The introduction of a new Peruvian crop “quinoa” to a rural community in Morocco. In 8 ESA Congress, 2004, Copenhagen, Denmark : [European agriculture in a global context] : book of proceedings. 11-15 July 2004. Samfundslitteratur. 881-884.
Bilalis, D., I. Kakabouki, A. Karkanis, I. Travlos, V. Triantafyllidis, and D. Hela. 2012. Seed and saponin production of organic quinoa (Chenopodium quinoa Willd.) for different tillage and fertilization. Not Bot. Horti Agrobot. 40: 42–46.
Bonifacio, Alejandro. 1995. Interspecific and intergeneric hybridization in Chenopod species. MS Thesis. Brigham Young University. Provo, Utah.
Davis, J., and L. Abbott. 2006. Soil fertility in organic farming systems. p. 25–51. In Kristiansen, P., A. Taji, and J. Reganold (eds). Organic Agriculture: A Global Perspective. Comstock Publishing Associates, Ithaca, N.Y.
Dirección Meteorológica de Chile. Anuarios Climatológicos 1920-2009. Dirección Meteorológica de Chile. Available at http://164.77.222.61/climatologia/php/menuAnuarios.php (verified 23 March 2013).
Fuentes, F., and A. Bhargava. 2011. Morphological analysis of quinoa germplasm grown under lowland desert conditions. J Agron Crop Sci. 197: 124–134.
Iliadis, C., T. Karyotis, S.E. Jacobsen, S.E. Jacobsen, and Z. Portillo. 2001. Adaptation of quinoa under xerothermic conditions and cultivation for biomass and fibre production. Memorias, Primer Taller Internacional sobre Quinua—Recursos Geneticos y Sistemas de Producción: 371–378.
Jacobsen, S.E. 1998. Developmental stability of quinoa under European conditions. Ind Crops Products. 7: 169–174.
Jacobsen, S.E., I. Jørgensen, and O. Stølen. 1994. Cultivation of quinoa (Chenopodium quinoa) under temperate climatic conditions in Denmark. J. Agric. Sci. (Cambridge). 122: 47-52.
Jacobsen, S.E., and O. Stolen. 1993. Quinoa - morphology, phenology and prospects for its production as a new crop in Europe. Eur. J. Agron. 2: 19–29.
Johnson, D.L., and Croissant, R.L. 1990. Alternate crop production and marketing in Colorado. Technical Bulletin LTB90-3. Cooperative Extension, Colorado State University, Fort Collins, Colorado.
Marschner, H. 1995. Mineral nutrition of higher plants. Academic Press, London; San Diego.
163
Mastebroek, H.D., E.N. Van Loo, and O. Dolstra. 2002. Combining ability for seed yield traits of Chenopodium quinoa breeding lines. Euphytica. 125: 427–432.
Oelke, E.A., D.H. Putnam, T.M. Teynor, and E.S. Oplinger. 1992. Quinoa. In Alternative Field Crops Manual. University of Wisconsin Cooperative Extension Service, University of Minnesota Extension Service, Center for Alternative Plant & Animal Products.
Razzaghi, F., F. Plauborg, S.-E. Jacobsen, C.R. Jensen, and M.N. Andersen. 2012. Effect of nitrogen and water availability of three soil types on yield, radiation use efficiency and evapotranspiration in field-grown quinoa. Agric Water Management. 109: 20–29.
Risi, J.C., and N.W. Galwey. 1984. The Chenopodium grains of the Andes - Inca crops for modern agriculture. Advances in applied biology. 10: 145–216.
Risi, J.C, and N.W. Galwey. 1991. Genotype x environment interaction in the Andean grain crop quinoa (Chenopodium quinoa) in temperate environments. Plant Breeding. 107: 141–147.
Schulte auf’m Erley, G., H.-P. Kaul, M. Kruse, and W. Aufhammer. 2005. Yield and nitrogen utilization efficiency of the pseudocereals amaranth, quinoa, and buckwheat under differing nitrogen fertilization. Eur. J. Agron. 22: 95–100.
Thanapornpoonpong, S. 2004. Effect of nitrogen fertilizer on nitrogen assimilation and seed quality of amaranth (Amaranthus spp.) and quinoa (Chenopodium quinoa Willd). PhD Dissertation. Georg-August-University of Göttingen. Göttingen, Germany.
Wood, L.J., B.J. Murray, Y. Okatan, and L.D. Nooden. 1986. Effect of petiole phloem disruption on starch and mineral distribution in senescing soybean leaves. Am. J. Bot. 73: 1377.
164
Table 1: List of quinoa varieties included in the 2011 quinoa NUE experiment.
Variety Origin UDEC-3 (PI 634925) Llico, Chile1 QQ065 (PI 614880) Chiloé, Chile1 Colorado 407D (PI 596293) Colorado, US1 Kaslaea (Ames 13745) New Mexico, US1 1ESP (Ames 13730) New Mexico, US1 Isluga (Ames 13743) New Mexico, US1 Baer (PI 634918) Bio-Bio, Chile1 UDEC-1 (PI 634923) Bucalemu, Chile1 Brightest Brilliant Rainbow Philomath, OR2 Cherry Vanilla Philomath, OR2 Red Head Philomath, OR2 Multi-Hued Kelowna, BC3 UDEC-4 (PI 634922) Llico, Chile1 QQ74 (PI 614886) Maule, Chile1 QQ63 (PI 614887) Bio-Bio, Chile1 Black-seeded Mosca, Colorado4 1 – from NPGRIS 2 – from Wild Garden Seed 3 – from Sunshine Farm 4 – from White Mountain Farm
165
Table 2: Varieties in the 2012 modified quinoa NUE experiment.
Replication Variety Origin 1 Oro de Valle Philomath, OR 2 Biobio* Willits, CA (seed from Boistfort, WA) 3 Isluga (Ames 13743) New Mexico, US 4 Titicaca* Denmark 5 Oro de Valle Philomath, OR * new varieties added in 2012
166
Table 3: Analysis of variance with F-values for plant height (PH), leaf greenness (LF), aphid susceptibility (AS), yield, and days to flower bud appearance (DTFB) in the 2011 quinoa NUE experiment.
Effect DF PH LG AS Yield DTFB Variety 15 2.11* 1.6 0.77 4.91*** 5.83*** Fertilization 3 0.99 0.58 0.03 0.36 0.52 Variety*Fertilization 45 0.89 0.84 1.02 0.95 1.01 ***=p<0.001, *=p<0.05
167
Table 4: Height and days to appearance of flowering bud by variety for the 2011 quinoa NUE experiment.
Variety Plant height Days to appearance (cm) of flowering bud 1ESP 101.9 AB 32.9 CDEFGH
Baer 92.0 BCD 31.8 FGHI
Black- 96.8 BCD 30.0 I seeded Brightest 105.6 AB 31.4 FGHI Brilliant Rainbow Cherry 102.4 AB 32.5 EFGH Vanilla Colorado 96.5 BCD 33.3 BCDEF 407D Isluga 102.9 ABC 31.4 FGHI
Kaslaea 97.7 BCD 32.8 DEFGH
Multi-Hued 82.8 CD 33.0 CDEFG
QQ065 81.0 D 35.2 AB
QQ63 91.7 BCD 35.6 A
QQ74 96.2 BCD 31.0 HI
Red Head 118.0 A 31.3 GHI
UDEC-1 90.8 BCD 34.5 ABCD
UDEC-3 105.3 AB 34.8 ABC
UDEC-4 89.4 BCD 34.3 ABCDE
LSDs significant at p<0.05
168
Table 5: Analysis of variance with F-values for days to full senescence for 2011 quinoa NUE experiment.
Effect DF F-value Days to full senescence Variety 8 2.43†
Fertilization 3 1.40
Variety*Fertilization 24 0.88
†=p<0.07
169
Table 6: Correlations between plant height (PH), leaf greenness (LG), aphid susceptibility (AS), yield, and days to flower bud appearance (DFBA) in the 2011 quinoa NUE experiment.
PH LG AS Yield DFBA Leaf greenness 0.332* Aphid susceptibility -0.169 ns 0.229† Yield 0.413** 0.137 ns 0.038 ns Days to flower bud -0.374* -0.229† 0.020 ns -0.422** appearance Days to complete 0.344* 0.078 ns -0.308† -0.269 ns 0.260 ns senescence **=p<0.001, *=p<0.05, †=p<0.07
170
Table 7: Nutrient analysis of tissue samples from the 2012 quinoa NUE experiment.
%N %P Cu (ppm) Zn (ppm) %Mg %Ca Fe (ppm) Mn (ppm) 7/31/12 2.488 0.2869 8.89 21.65 1.2323 1.588 94.18 450.7 8/28/12 1.768 0.1436 6.79 15.59 2.372 2.711 118.7 710.3 P-value <0.0001 <0.0001 0.0006 0.0006 <0.0001 <0.0001 0.0005 0.0003 % change -28.9% -49.9% -23.6% -28.0% 92.5% 70.7% 26.0% 57.6%
171
Figure 1: Yield from the 2011 quinoa NUE experiment.
30 A 25
A 20 A
15 AB AB
g yield per plotper yield g 10 ABC ABC 5 BCD BCD BCD CD CDE DE DE DE E 0
LSDs significant at p<0.05; Yield data has been back transformed from natural log transformation
172
Figure 2: Weather data for Pullman, WA (2011).
40
35
30
25
20
15
10
5
0
-5
2011-06-08 2011-07-28 2011-06-03 2011-06-13 2011-06-18 2011-06-23 2011-06-28 2011-07-03 2011-07-08 2011-07-13 2011-07-18 2011-07-23 2011-08-02 2011-08-07 2011-08-12 2011-08-17 2011-08-22 2011-08-27 2011-09-01 2011-09-06 2011-09-11 2011-09-16 2011-09-21 2011-09-26 2011-10-01 Total Precip (mm) Avg Air Temp (°C) Max Air Temp (°C) Min Air Temp (°C)
*Data provided by AgWeatherNet, Prosser, WA (accessible at http://weather.wsu.edu/awn.php)
173
Figure 3: Weather data for Pullman, WA (2012).
40
35
30
25
20
15
10
5
0
-5
-10
2012-06-09 2012-09-01 2012-05-16 2012-05-22 2012-05-28 2012-06-03 2012-06-15 2012-06-21 2012-06-27 2012-07-03 2012-07-09 2012-07-15 2012-07-21 2012-07-27 2012-08-02 2012-08-08 2012-08-14 2012-08-20 2012-08-26 2012-09-07 2012-09-13 2012-09-19 2012-09-25 Total Precip (mm) Avg Air Temp (°C) Max Air Temp (°C) Min Air Temp (°C)
Data provided by AgWeatherNet, Prosser, WA (accessible at http://weather.wsu.edu/awn.php)
174
APPENDIX
Appendix A.
Crossing Method For Quinoa Using Manual Emasculation
1. Introduction
Manually crossing quinoa varieties via emasculation has generally been considered a difficult process. Due to the small size of the flowers and fragility of the inflorescence, the plants can easily become damaged during the process.
Crossing is most easily done when one of the parents contains a dominant morphological marker, such as purple stem color or axil pigmentation. F1 plants can easily be determined in such cases (Jacobsen and Stolen, 1993). However, in the case that both parents lack any differentiating trait, crossing becomes increasingly more difficult. This makes a method of manual emasculation with consistently high success crucially important.
Chilean lowland cultivars lack differences for many of the monogenic morphological traits used to determine successful F1 progeny. As these varieties form the bulk of the WSU breeding program, a method was developed to manually emasculate quinoa plants and was tested.
Several crossing techniques used in South America were reported by (Risi and Galwey,
1984) However, based on observations of flowering patterns in the greenhouse, a different but similar approach was developed that was found to be more suitable. It should be noted that the range of Chilean varieties crossed with this method had glomerulate inflorescence as opposed to an amaranthiform inflorescence.
2. Materials and Methods
176
Planting/Observation
In the initial tests, three sets of three male plants were grown, spaced one week apart.
Seeds were planted near the edge of the pots. In Week 1, three male individuals were planted. In the second week, three male individuals were planted along with the three female individuals. In the final week, three male individuals were grown. The wide spacing in planting dates allows for adequate leeway in having an available pollen donor. Delays in development or stress to the female during manual emasculation can result in large gaps in pollen productivity and stigma receptivity. From the tests, it appeared that manual emasculation delayed the female plant by at least a week. A useful modification might be to plant a set of the male and of the female plants in the first week, followed by two male sets over the next two subsequent weeks.
As the male and female reached the point of crossing, the pots were rotated so that both plants remained upright. This allowed both the inflorescence of the male and female plants to be in the same isolating bag while remaining upright.
As the plants began to form floral buds, cellulose tubing was placed over the female plants. One end was secured with paper clips, while the bottom end was secured around the stem with string. This prevented pollination from other varieties. The plants were observed closely each day. The flower bud was clipped once the flower bud grew in size to where developing glomeruli (flower clusters) were visible beneath it.
Not removing the terminal flower bud would result in a panicle with hundreds of densely packed flowers, making adequate emasculation prohibitively difficult. Instead, the small glomeruli at the base of the plant were allowed to grow and develop. These are smaller and easier to manage. Three to four of these glomeruli were allowed to grow per plant.
177
Flowering patterns
Quinoa is gynomonoecious, having both hermaphroditic and female flowers. Three generally flowering patterns were observed. These were usually specific within variety, though variation within variety and even between flower clusters on the same plant was observed.
1st Flowering Pattern
In the first flowering pattern, the terminal flower of the glomerulus is hermaphroditic.
This flower was larger than the other smaller flowers that flanked it on the cluster (Photos 1 &
2). These smaller flowers were invariably female. To make crosses, the terminal floret simply needed to be emasculated or removed. With either approach, the risk of selfing was eliminated.
Careful monitoring of the female side flowers is advised, as with time, they may actually be immature hermaphroditic flowers.
2nd Flowering Pattern
In the second flowering pattern, the terminal flower and some of the flanking flowers were hermaphroditic. The rest of the flanking flowers were female. Checking all flanking side flowers and emasculating as necessary is needed to eliminate the potential for selfing. If a small number of side flowers were hermaphroditic, they could be removed. However, removal of too many flowers is not advised as the whole glomerulus can desiccate as a result of excessive injury.
178
3rd Flowering Pattern
The third flowering pattern is most difficult to deal with. With this pattern, all or most of the flowers in the side flower cluster are hermaphroditic. The large number of flowers in a given side flower cluster makes emasculating each flower prohibitive. Additionally, the excessive handling could result in damaging the flower cluster. If this pattern is not seen on all side flower clusters produced by an individual, it’s advised to remove flower clusters with this pattern and focus on those with the 1st or 2nd flowering pattern. Varieties that exhibit this pattern are best utilized as males.
If a cross needs to be made with a variety solely exhibiting this flowering pattern, the glomerulus can be trimmed back as close to the plant’s stem as possible, leaving a small number of flowers that are then emasculated. Clipping the glomerulus close to the plant’s main stem ensures that injury is limited to the stem running central through the glomerulus.
However, the remaining flowers that were emasculated will soon be flanked by new flowers. These flowers are exceedingly small, but produce anthers. However, given their small size, lack of anthesis, and the temporal gap in maturity with the older, larger emasculated flowers, the risk of self-pollination is reduced.
While emasculating with this particular flowering type is difficult, it can be done. Fewer seeds will be produced due to the small number of emasculated flowers that can be pollinated and produce seed. Therefore, the number of plants as females and/or the number of side flower clusters retained per plant might be adjusted upward.
179
Emasculation
The emasculation process was conducted under a compound microscope at 80x magnification. This provided significant magnification of the whole flower, though reduced magnifications would work satisfactorily. The cellulose bag was removed from the inflorescence and the plant tipped on its side, so the flower cluster could be examined under the compound microscope.
The flower cluster was carefully held between the forefinger and thumb to secure it. Once in the field of view, the tweezers were grasped with the other hand and emasculation of the target flowers began.
The sepals of quinoa flowers curl around and protect the developing stamens and the central pistil. The tweezers need to carefully lift each sepal and remove the anther located beneath it. Bending the tip of the tweezers can aid in this. Rough handling of the sepals can result in excess stress to the flower and cause it to abort. While the usual number of stamens is 5, flowers with 6, 7, and 8 stamens have been seen. Care must be taken not to remove the central pistil which can be knocked loose by the tweezers. The bifurcated stigma can also be damaged.
Once removed, the process continues to the remaining flowers.
Once complete, a male plant that has recently entered anthesis is chosen. Both the inflorescence of the female and male are placed in the cellulose bag together. As anthesis occurs over a number of days, this ensures that pollen will be produced over the period that the female plant will be receptive. To aid in the flow of pollen and increase seed set, the bag can be shaken as the anthers dehisce to help distribute pollen to the female flowers. Protrusion of the stigma may be observed in the female at this point.
180
If the female is adequately pollinated, seed production will begin. If it does not, the female and emasculated flowers will continue to exhibit protruded stigmas and fail to set seed.
3. Results
As of this writing, F1s from the first set of experimental crosses have begun to set seed.
Based on dominant morphological characteristics from the male, which was used to determine relative success of this crossing method, there appears to be a high rate of success. For two varieties, success appears to be at least 80%. However, for one variety that exhibited the third flowering type and that had difficulties in crossing, success was reduced. However, a successful cross with this variety was still maintained.
Due to the high rate of success, it appears that this will be a valuable method to cross quinoa in the future, especially where dominant visible qualitative traits exist. Even if success is low for a particular variety, increasing the percent of successful F1 progeny would allow identification at the F2 generation when segregation of quantitative traits would allow for easy identification.
Other Crossing Techniques
Bonifacio (1995) describes in detail a crossing method devised to cross quinoa varieties and to make interspecific crosses. This report was found after the technique described here was developed and differs in some ways, but it also is reported to result in a high degree of success.
Another report, titled “Gynomonoecy in Chenopodium quinoa (Chenopodiaceae): variation in inflorescence and floral types in some accessions”, discovered after this technique
181
was developed, describes the complex set of flowering patterns found in quinoa in far greater detail than described here (Bhargava et al., 2007). For the purposes of making crosses, however, the current set of classification and approach should suffice.
182
Figure 1: 1st Flowering Pattern. Side flower cluster exhibiting flowering pattern 1. Sepals were removed on the relatively larger terminal flower, making anthers and stigma visible. Sepals were also removed from a flanking female flower halfway along the side flower cluster, making the pistil visible.
183
Figure 2: Female flower with protruding bifurcated stigma. A flanking female flower with a protruding stigma. Numerous pigmented epidermal bladder cells are visible on the surface of the flower.
184
Figure 3: Developing inflorescence. Inflorescence at the stage the terminal section should be removed.
185
Figure 4: Plant after terminal section of inflorescence is removed. Three small glomeruli are visible below where the terminal section of the inflorescence was removed.
186
References
Bhargava, A., S. Shukla, and D. Ohri. 2007. Gynomonoecy in Chenopodium quinoa (Chenopodiaceae): variation in inflorescence and floral types in some accessions. Biologia. 62: 19–23.
Bonifacio, Alejandro. 1995. Interspecific and integeneric hybridization in Chenopod species. MS Thesis. Brigham Young University. Provo, Utah.
Jacobsen, S.-E., and O. Stolen. 1993. Quinoa - morphology, phenology and prospects for its production as a new crop in Europe. Eur. J. Agron. 2: 19–29.
Risi, J.C., and N.W. Galwey. 1984. The Chenopodium Grains of the Andes - Inca Crops for Modern Agriculture. Advances in applied biology. 10: 145–216.
187
GENERAL CONCLUSIONS
Quinoa holds many challenges and opportunities for expanded cultivation in North
America. Heat tolerance currently limits quinoa cultivation to high altitude, northern, and coastal locations while pre-harvesting sprouting pressures pose a threat in cool, coastal regions of the continent characterized by an oceanic climate. Downy mildew poses a threat in these regions, as well as locations in Central and Eastern parts of the continent characterized by humid summers.
Despite these challenges, sources of adaptation have been identified. Even though little information on nitrogen dynamics was found in field experiments held in 2011 and 2012, varieties with greater heat tolerance were identified and natural selection has appeared to improve this tolerance. Pre-harvest sprouting tolerance was located during a heavy rainfall event in trials in 2010, and downy mildew resistance has been reported within quinoa and for related species. With the right goals specific to the area of adaptation and enough the right support, superior varieties of quinoa can be developed that will greatly increase the possible range of quinoa cultivation.
Quinoa, due its high level of agronomic resiliency and stress tolerance, also holds some unique opportunities. This was demonstrated in the greenhouse salinity experiment, where the
Chilean lowland cultivars of quinoa that form the basis of current WSU breeding efforts were found to have high levels of salinity tolerance. Soil salinity is a significant problem worldwide, and quinoa’s ability to tolerate extremely high levels of salinity may prove useful in cultivating soils that were formerly abandoned. Additionally, the large difference seen in quinoa’s tolerance to sodium chloride and sodium sulfate indicates that quinoa may be better able to tolerate land
188
affected by sulfate salinity, such as areas in the Upper Midwest of the United States and the
Canadian Prairies. Despite being more greatly affected by sodium chloride salinity, quinoa also exhibited high levels of tolerance to sodium chloride.
Quinoa produced under saline conditions can also expect to have altered levels of mineral nutrients. This should be observed carefully, as the levels of mineral nutrition may be enhanced or decreased depending on soil conditions and the nutrient in question. Many factors are at play as well, and variety played a significant role in the response of mineral nutrition to salinity.
As demand for quinoa rises and outstrips supply from South America, increasing domestic production will be important in helping to balance the current situation, which has led to unfortunate social and economic consequences in the Andean countries from which quinoa is exported. Quinoa’s unique combination of being both a staple food crop and a halophyte holds great opportunities for its cultivation on marginal land that was formerly abandoned due to high soil salinity. Further research and field trials, combined with breeding and selection for increased heat tolerance will be necessary to fully realize its potential in this area.
189