EFFECT OF GIBBERELLIC ACID 4 AND 7 AND 6-BENZYL ADENINE ON GROWTH, YIELD AND QUALITY OF SUGARCANE IN KAKAMEGA COUNTY, KENYA ANDREW ADODI ACHOLA (Bsc. Agribusiness Management) AI44/OL/KKA/24082/2014 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE MASTER OF SCIENCE (AGRONOMY) IN THE SCHOOL OF AGRICULTURE AND ENTERPRISE DEVELOPMENT OF KENYATTA UNIVERSITY SEPTEMBER, 2020 ii DECLARATION This thesis is my original work and has not been presented for a degree in any other University. Signature………………………………………. Date…………………………. NAME: ANDREW ADODI ACHOLA REG NO: A144/OL/KKA/24082/2014 SUPERVISORS We confirm that the work reported in this thesis was carried out by the candidate under our supervision and has been submitted with our approval as university supervisors. Signature…………………………………………. Date……………………………… Dr. Nicholas Kibet Korir Department of Agricultural Science and Technology Kenyatta University Signature…………………………………………. Date……………………………… Dr. Joseph P. Gweyi Onyango Department of Agricultural Science and Technology Kenyatta University Signature…………………………………………. Date……………………………… Dr. George Omoto Department of Crop Development Sugar Research Institute iii DEDICATION I dedicate this work to my late father whose belief in the pursuit of academic excellence helped strengthen my resolve. To my loving wife for her fervent prayers and encouragement. To my children for their moral support and to my mother for instilling in me the spirit of hard work. May God bless you all! iv ACKNOWLEDGEMENTS The successful completion of any work is without doubt a team effort. I want to acknowledge with appreciation the versatile team that brought this research work to fruition. Special appreciation goes to my team of Supervisors: Dr. Nicholas Kibet Korir, Dr. Joseph Gweyi Onyango and Dr. George Omoto for their tireless guidance, insight and deft execution of this research. I wish to thank the management of Butali Sugar Mills Limited for facilitating my research. Specifically, I appreciate the contribution of Mr. Jayanti Patel, Mr. Sanjay Patel, Mr. Pratap Keshwala, Mr. Daniel Kiyondi, Mr. Maurice Lukano and Mr. Karthik Gairaj. I am grateful to the team in Cane Development section of Butali Sugar Mills Limited led by Mr. Ravi Kumar for untiring support and encouragement. Mr. Samson Onyango Auma deserves special mention for discharging his duties as Research Assistant with noteworthy determination. My special gratitude goes to Mr. Kailash M. Bhadania of Greenlife Africa for availing chemicals for this research; my brothers Mr. Andrew Acholla Jnr and Mr. Patrick Achollah for their support. I must acknowledge my mother, Mrs. Mary Achieng Achola for her encouragement. My sincere gratitude goes to my loving wife Mrs. Lucy Achola and my children: Wendy Achieng’, Hedley Achola, Perry Adhiambo, Ivy Akoth, Garry Achola and Rainey Achola for their moral support. I am forever grateful to God for giving me the ability to think, visualize and peep into my destiny. v TABLE OF CONTENTS DECLARATION ........................................................................................................ ii DEDICATION ........................................................................................................... iii ACKNOWLEDGEMENTS ....................................................................................... iv TABLE OF CONTENTS ............................................................................................ v LIST OF TABLES .................................................................................................. viii LIST OF FIGURES ................................................................................................... ix LIST OF APPENDICES ............................................................................................. x ABBREVIATIONS AND ACRONYMS ................................................................... xi ABSTRACT .............................................................................................................. xiv CHAPTER ONE: INTRODUCTION......................................................................... 1 1.1 Background information ...................................................................................... 1 1.2 Problem statement ................................................................................................ 6 1.3 Objectives ............................................................................................................ 6 1.3.1 General objective ...........................................................................................6 1.3.2 Specific objectives .........................................................................................7 1.4 Research hypotheses ............................................................................................ 7 1.5 Justification of the study ...................................................................................... 7 1.6 Conceptual framework ......................................................................................... 9 CHAPTER TWO: LITERATURE REVIEW .......................................................... 10 2.1 Overview of sugarcane production ..................................................................... 10 2.2 Challenges of sugarcane production ................................................................... 13 vi 2.3 Effect of plant growth regulators on crop growth ............................................... 18 2.4 Role of gibberellins and cytokinins in sugarcane growth, yield and sucrose........ 26 2.5 Cost-benefit analysis .......................................................................................... 31 CHAPTER THREE: MATERIALS AND METHODS ........................................... 33 3.1 Study area .......................................................................................................... 33 3.2 Research design and treatments .......................................................................... 34 3.3 Soil sampling and analysis ................................................................................. 35 3.4 Planting materials .............................................................................................. 36 3.5 Land preparation and planting ............................................................................ 37 3.6 Field management .............................................................................................. 37 3.7 Data collection ................................................................................................... 38 3.7.1 Effect of levels of GA 4+7 and 6-BA on growth of selected sugarcane varieties ................................................................................................................................ 38 3.7.2 Effect of GA 4+7 and BA on yield and sucrose of selected sugarcane varieties ................................................................................................................................ 38 3.7.3 Cost-benefit analysis of use of gibberellic acid 4 and 7 and 6-benzyl adenine .. 39 3.8 Data analysis ...................................................................................................... 41 CHAPTER FOUR: RESULTS AND DISCUSSION ............................................... 43 4.1 Germination ....................................................................................................... 43 4.2 Tillering ............................................................................................................. 45 4.3 Stem diameter .................................................................................................... 50 4.4 Plant height ........................................................................................................ 53 4.5 Internode length ................................................................................................. 56 vii 4.6 Number of leaves ............................................................................................... 59 4.7 Yield .................................................................................................................. 62 4.8 Pol% .................................................................................................................. 65 4.9 Profitability of use of GA 4+7 and 6-BA at 3 L/Ha ............................................ 67 CHAPTER FIVE: CONCLUSIONS AND RECOMMENDATIONS ..................... 69 5.1 Conclusions ....................................................................................................... 69 5.2 Recommendations .............................................................................................. 70 REFERENCES .......................................................................................................... 71 APPENDICES ........................................................................................................... 88 viii LIST OF TABLES Table 3.1: Physio-chemical status of soil at Chegulo ................................................... 35 Table 3.2: Treatment levels and labels ......................................................................... 37 Table 3.3: Extra cost per hectare .................................................................................. 39 Table 3.4: Yields at 0 and 3 litres per hectare of GA 4+7 and 6-BA ............................. 40 Table 3.5: Extra yields at 3 litres per hectare of GA 4+7 and 6-BA .............................. 40 Table 3.6: Discounted value of extra yield ................................................................... 41 Table: 4.1: Number of tillers as affected by variety and GA 4+7 and 6-BA.................. 47 Table 4.2: Growth rate of shoots as affected by variety and GA 4+7 and 6-BA ............ 49 Table 4.3: Stem diameter in centimeters as affected by variety and GA 4+7 and 6-BA 51 Table 4.4: Growth rate of diameter as affected by variety and GA 4+7 and 6-BA ........ 52 Table 4.5: Plant height in centimeters as affected by sugarcane varieties and GA 4+7 and 6-BA .................................................................................................. 54 Table 4.6: Growth rate of height as affected by variety and GA 4+7 and 6-BA ............ 55 Table 4.7: Internode length in centimeters as affected by variety and GA4+7 and 6-BA .................................................................................................................... 57 Table 4.8: Growth rate of internodes as affected by variety and GA4+7 and 6BA ........ 58 Table 4.9: Number of leaves as affected by variety and GA4+7 and 6-BA levels ......... 59 Table 4.10: Growth rate of leaves as affected by variety and GA 4+7 and 6-BA .......... 61 Table 4.11: Yield as affected by variety and GA 4+7 and 6-BA ................................... 63 Table 4.12: Yield due to interaction effect between varieties and GA 4+7 and 6-BA ... 64 Table 4.13: Pol % as affected by variety and GA 4+7 and 6-BA .................................. 66 Table 4.14: Benefit-Cost Ratio .................................................................................... 67 ix LIST OF FIGURES Figure 1.1: Conceptual framework ................................................................................9 Figure 2.1: Sugarcane stool showing primary and secondary shoots ............................ 27 Figure 3.1: Map of Kakamega County showing areas of cane development (source: GOK, 2013) .............................................................................................. 33 Figure 3.2: Map of Malava Sub-County showing Chegulo study site ........................... 34 Figure 4.1: Effect of GA 4+7 and 6-BA on germination of sugarcane .......................... 44 Figure 4.2: Effect of GA 4+7 and 6-BA on tillering of sugarcane ................................ 46 Figure 4.3: Benefit-Cost Ratio across varieties ............................................................ 68 x LIST OF APPENDICES Appendix 1: Definition of Terms………………………………….………………….88 Appendix 2: List of marketed plant growth regulators…..............…………………...90 Appendix 3: Salient features of 5 sugarcane varieties grown in Kakamega County....91 Appendix 4: Cane establishment and management practices.......................................92 xi ABBREVIATIONS AND ACRONYMS AFLP: Amplified Fragment Length Polymorphism AMP: Antimicrobial peptides ANOVA: Analysis of variance BA: Benzyl adenine BAP: Benzyl amino urine CBA: Cost Benefit Analysis Ck: Cytokinin CO421: Coimbatore 421 variety from India CO945: Coimbatore 945 variety from India COMESA: Common Market for Eastern and Southern Africa D8484: Demerara 8484 variety from Guyana DAP: Days after planting DNA: Deoxyribonucleic Acid EAC: East African Community EAK 73-335: East African Kenyan variety 73-335 EC: European Commission EU: European Union FAO: Food and Agriculture Organization FAOSTAT: Food and Agriculture Organization Corporate Statistical Database xii GA: Gibberellic Acids HA: Hectare IAA: Indole 3-acetic acid IBR: Institute for Biotechnology Research ICSB: International Consortium for Sugarcane Biotechnology IISR: International Institute for Scientific Research IPT: Isopentenyl transferase ISSR: Inter Simple Sequence Repeats KEN 82-472: Kenyan variety 82-472 from Mtwapa KEN 82-493: Kenyan variety 82-493 from Mtwapa KEN 83-737: Kenyan variety 83-737 from Mtwapa KESREF: Kenya Sugar Research Foundation KSB: Kenya Sugar Board KSI: Kenya Sugar Industry LPT: Lipopolysaccharide transport LSD: Least Significant Difference MAP: Months after planting NDO: NDO certified seed of oat NPK: Nitrogen Phosphorus and Potassium OC: Organic Carbon PGR: Plant growth regulators xiii pH: Potential Hydrogen RAPD: Random Amplified Polymorphic DNA RCBD: Randomized Complete Block Design SADC: South African Development Committee SAS: Statistical Analysis System SCAR: Sequence Characterized Amplified Region SD: Sugar Directorate SONY: South Nyanza SRI: Sugar Research Institute SSR: Simple Sequence Repeat STMS: Sequence Tagged Microsatellite TRAP: Target Region Amplification Polymorphism UPO: UPO certified seed of oat USDI: Unique Systems Design Inc WTO: World Trade Organization ZOG: Zeatin O-glucosyltransferase ZOX: Zeatin O-xylosyltrasferase xiv ABSTRACT Declining sugarcane productivity in Kenya has attracted a plethora of interventions such as optimal fertilizer regimes, improved seedcane quality and use of plant growth regulators (PGRs). Although application of PGRs in leading sugarcane producing countries like China and India has registered success, sugarcane farmers in Kenya are yet to exploit this technology. Globalization and liberation of world sugar industry now subjects the local millers and farmers to stiff competition. The local sugar millers have responded by seeking to adopt payment model based on yield and sucrose content. To address this problem, a study was conducted to determine the effect of different levels of gibberellic acid 4 and 7 and 6-benzyl adenine on growth, yield and sucrose content of five selected sugarcane varieties (CO 421, KEN 83-737, D8484, CO 945 and EAK 73- 335). The study was carried out at the Butali Sugar Company Research and Demonstration farm in Chegulo, Kakamega County-Kenya. The trial was laid out in a Randomized Complete Block Design in 5 by 5 factorial arrangement. The treatments consisted of gibberellic acid 4 and 7 and 6-benzyl adenine at 0 1, 2, 3 and 4 litres per hectare and the five sugarcane varieties. All treatments were replicated three times. Data on germination, tillering, height, girth, length of internodes and leaf number was collected monthly for six months for two seasons. Data on yield was collected by harvesting all the sugarcane from each plot at the 12th month after planting and recording the stalk biomass. Pol% was was generated by extracting juice from 10 randomly selected stalks per plot to obtain a homogenized, composite sample before adding lead sub-acetate clarifier and measuring the filtrate using a polarimeter before multiplying the recorded value by the corresponding pol factor. All the data was subjected to ANOVA using SAS 9.1 software; and means separated using Tukey’s test (P≤0.05). Finally, cost-benefit analysis was computed for all varieties under study. It was observed that use of gibberellic acid 4 and 7 and 6-benzyl adenine led to significant and or linear increasing growth, yield and quality (sucrose content) of sugarcane. It was also observed that the varieties differed in their response to application of the gibberellic acid 4 and 7 and 6-benzyl adenine. In yield, D8484 grown with GA4+7 and 6-BA at 4 litres per hectare recorded the best performance at 75.35 and 75.23 tons/ha in Seasons 1 and 2 respectively. EAK 73-335 recorded superiority in sucrose accumulation with pol% of 14.70% and 14.69 % in seasons 1 and 2 respectively when treated with GA 4+7 and 6-BA at 4 litres per hectare. There was no significant difference at 4 litres per hectare in all parameters under study. Application of gibberellic acid 4 and 7 and 6-benzyl adenine increased growth, yield and quality of the sugarcane varieties with D8484 recording the best overall performance and the highest net positive benefit-cost ratio at 3.7. Gibberellic acid 4 and 7 and 6-benzyl adenine increased yield by 11-22% and sucrose content by 3-6%. Therefore, it is recommended that variety D8484 and EAK 73-335 be grown and gibberellic acid 4 and 7 and 6-benzyl adenine at 3 litres per hectare be incorporated in the production of sugarcane in Kakamega County and areas with similar agro ecological conditions. In addition, there is need for evaluation of response of more cane varieties and in other agro ecologies. Further study is recommended on performance of ratoon crop previously applied with gibberrellic acid 4 and 7 and 6-benzyl adenine. 1 CHAPTER ONE: INTRODUCTION 1.1 Background information Sugarcane, Saccharum officinarum (Gramineae) is a cash crop of global and regional importance providing over 70% of the world’s sugar requirement. It is cultivated in more than 23.8 million hectares in tropical and subtropical regions of the world, producing up to 1.5 billion metric tons of crushable stems (Paiva et al., 2004; Han and Wu, 2004). It has served as a source of sugar since hundreds of years and is an important renewable biofuel source (Pandey et al., 2000). Sugarcane is generally used to produce sugar, accounting for almost two thirds of the world’s production and has lately gained increased attention because of ethanol which is derived from cane. Sugarcane bagasse is largely used for energy cogeneration at the mill or paper production (Sangnark and Noomhorm, 2004). Today, sugarcane is grown in over 110 countries. In 2019, 1,683 million metric tons was harvested from estimated 26.2 million hectares globally which amounts to 22.4% of the total world agricultural production by weight (FAOSTAT 2019). About 50 percent of sugarcane production occurs in Brazil and India. Brazil has the highest area under cane at 10.1 million hectares with annual sugarcane production of 670.78 million metric tons, India 4.7 million hectares with 347.87, China at 123.46, Thailand at 96.5 and Pakistan at 58.39. Australia has the highest productivity at 85.1 tons per hectare. (FAOSTAT, 2014). Out of the total sugar production, approximately 70% comes from sugarcane and 30% from sugar beet. (FAO, 2019). 2 In sub-Saharan Africa, the largest sugarcane growing countries are South Africa, Mozambique and Cameroon. Generally, Africa contributes 5% of the world’s sugarcane production with 1.5% coming from East Africa and 0.2% from Kenya, whose annual sugar production is 591,658 metric tons against domestic consumption of 826,000 metric tons (Sugar Directorate, 2019). In Kenya, sugarcane is grown on about 206,809 hectares mainly in Kakamega, Bungoma, Busia, Kisumu, Migori, Kwale, Narok and Siaya counties. Sugarcane productivity in Kenya has declined over the years. The national average cane yield is 57.6 tons per hectare, which is far below existing potential (Sugar Directorate cane census records of 2019). Sugarcane is ranked 4th after horticulture, tea and coffee and contributes about 15% of the Agricultural gross domestic product. An estimated 250,000 small scale farmers depend directly on sugarcane with 25% of the population depending on the sugar industry either directly or indirectly (FAOSTAT 2019). All sugar cane species interbreed, and the major commercial cultivars are complex hybrids. Under field conditions germination starts from 7 to 10 days and usually lasts for about 30-35 days after planting (DAP) (Seabra and Macedo 2011). Germination denotes activation and subsequent sprouting of the eye bud which is influenced by both external as well as internal factors. The external factors are the soil moisture, soil temperature and aeration. The internal factors include the bud health, sett moisture, sett reducing sugar content and sett nutrient status (Rademacher, 2015). 3 Tillering starts from around 40 and may last up to 180 days. It is a physiological process of repeated underground branching from compact nodal joints of the primary shoot, providing the crop with appropriate number of stalks required for a good yield (Subhashisa et al., 2016). Various factors as variety, light, temperature, soil moisture and fertilizer practices influence tillering. Early formed tillers give rise to thicker and heavier stalks. Late formed tillers either die or remain short or immature. Cultivation practices such as spacing, time of fertigation, water availability and weed control influence tillering. Inducing good tillering is important to build adequate population (Ullah et al., 2012). Grand growth phase starts from 120 DAP and lasts up to 270 days in a 12-month crop. During the early period of this phase tiller stabilization takes place. Out of the total tillers produced only 40-50% survives by 150 days to form millable cane. This is one of the most important phases of the crop where in the actual cane formation and elongation and thus yield build up takes place. Under favourable conditions stalks grow rapidly almost 2-3 internodes per month (Shirzad et al., 2012). Ripening and maturation phase in a twelve-month crop lasts for about three months starting from 270-360 days. Sugar synthesis and rapid accumulation of sugar takes place during this phase and vegetative growth is reduced. As ripening advances, simple sugars (monosaccharide viz., fructose and glucose) are converted into cane sugar (sucrose, a disaccharide). Cane ripening proceeds from bottom to the top and hence bottom portion contains more sugars than the top portions (Sobol et al., 2014). 4 Ample sunshine, clear skies cool nights and warm days (i.e., more diurnal variation in temperature) and dry weather are highly conducive for ripening. Sugarcane area and productivity differ widely from country to country. Abiotic and biotic stresses are the primary cause of crop loss worldwide, reducing average yields for most major crop plants by more than 50% (Nishiyama et al., 2011). Such declines have been attributed to biotic and abiotic constraints such as pests and diseases, low soil fertility, drought, salinity (Idrees et al., 2004). Sugarcane is grown in a range of climates from hot dry environment near sea level to cool and moist environments at high elevations (Plaut et al., 2000). Efforts to improve sugarcane production through application of plant growth regulators have been suggested. Plant growth regulators are essential in plant growth and in mitigating the effects of drought (Khan et al., 2003). They are mainly involved in initial growth processes such as replication of chromosomes, synthesis of deoxyribonucleic acids and nuclear protein and thus influence plant growth (Shiranzi et al., 2005). Therefore, application of plant growth regulators such as gibberellic acids 4 and 7 may be useful in enhancing sugarcane yields. Benzyladenine plant growth regulator plays an important role in regulation of many physiological responses. It is involved in almost all growth and developmental processes of plant organs (Frankenberger, 2002). 5 Previous studies show that gibberellic acid 4 and 7 induces tillering in most crops. Tillering is a desirable characteristic in sugarcane as it increases yields (Gomez and Brewer 2008). The species Saccharum officinarum has the simplest tillering model that can be roughly represented by one primary shoot, three secondary shoots and ten tertiary shoots: showing determinate tillering (Hussein et al., 2007). Thus, tillering is a primordial characteristic of sugarcane: the main sink of the product of photosynthesis are the stalks formed from the growth of tillers and therefore the profitability of the crop depends primarily on the tillers produced that will dictate the final number of harvestable stalks (Vishwakarma et al., 2008). Although the higher the number, the thinner the diameter of stalks, the final result is a higher volume and thus higher productivity (Nishumura et al., 2004). Today, there are very many plant growth regulators known to increase cell division by stimulating the production of proteins needed for growth and development. To assess economic efficiency of use of gibberellic acid 4 and 7 and 6-benzyl adenine on sugarcane, economic evaluation of costs and benefits is preferred. Cost benefit analysis (CBA) has been used to inform the right decisions while employing limited resources in the best way possible in order to achieve value for money. In many respects, the scope of CBA is much broader compared to other forms of economic evaluation since it converts all costs and benefits into monetary value (Perraillon, 2016). The current study investigated the effect of gibberellic acids 4 and 7 and 6-benzyl adenine on germination, growth, yield and sucrose content of sugarcane. 6 1.2 Problem statement Sugarcane farmers in Kenya have to contend with new payment formula based on tonnage and sucrose content; this despite declining yields and sucrose content as a result of biotic and abiotic stresses. According to Sugar Directorate annual report of 2019, average yield was 57.6 tons of cane per hectare while pol% was 12.8. This was low compared to other countries like Australia with 85.1 tons of cane per hectare and pol% 17.2 and South Africa with 76 tons per hectare and pol% of 15.4. Plant growth regulators modulate plant growth and development and mediate responses to both biotic and abiotic stresses. Promalin (a mixture of two naturally occurring plant growth regulators: gibberellic acid 4 and 7 and 6-benzyl adenine) has been used in leading sugarcane producing countries like Brazil and India with a lot of success. Therefore, there exists a need to induce growth parameters like germination, length of internodes, height, girth, tillering and number of leaves to promote yield and sucrose. The current research study focused on the best level of GA 4+7 and 6-BA for optimum sugarcane and sugar yields in Kakamega County. The varieties selected for this study were CO 945, CO 421, D8484, KEN 83-737 and EAK 73-335; considered dominant in Kakamega County. 1.3 Objectives 1.3.1 General objective To determine and evaluate the effect of gibberellic acid 4 and 7 and 6-benzyl adenine on growth, yield and sucrose content of selected sugarcane varieties. 7 1.3.2 Specific objectives i. To determine the effect of different levels of gibberellic acid 4 and 7 and 6-benzyl adenine on growth of selected sugarcane varieties. ii. To determine the effect of different levels of gibberellic acid 4 and 7 and 6-benzyl adenine on yield and sucrose content of selected sugarcane varieties. iii. To evaluate cost and benefit due to use of gibberellic acid 4 and 7 and 6-benzyl adenine at the recommended level. 1.4 Research hypotheses i. There is no statistically significant effect of different levels of gibberellic acid 4 and 7 and 6-benzyl adenine on growth of sugarcane varieties. ii. Gibberellic acid 4 and 7 and 6-benzyl adenine does not significantly affect sugarcane yield and sucrose content and thus cannot be effectively used to achieve optimum yields. iii. Use of gibberellic acid 4 and 7 and 6-benzyl adenine results into more cost than benefit and thus should not be recommended for sugarcane farmers. 1.5 Justification of the study Sugarcane productivity and profitability in Kenya is on the steady decline despite high cost of production. Globalization and liberalization have introduced new dynamism making it necessary to address competitiveness at every level. Farmers and millers are facing competition from countries with high sugarcane productivity but low-cost sugar. 8 It is against this backdrop that millers are in the process of adopting payment formula based on tonnage and sucrose content. In response, farmers are looking for appropriate technologies including but not limited to use of plant growth regulators to increase yields and sucrose content. Therefore, this study could not have come at a better time. Plant growth regulators have been widely used in leading sugarcane producing countries to improve productivity (Yoder, 2001). Exogenous application of promalin was reported to increase productivity in lettuce and celery (Apiumgraveolens), (Thomas and Van Staden, 2000). In an empirical search for compounds which might stimulate early growth of sugarcane plant, it was found that gibberellic acids 4 and 7 and 6-benzyl adenine caused considerable stimulation of germination and growth at later stages resulting in increased production of both cane and sugar (Riefler et al., 2006). The study sought to generate specific information on optimal use of gibberellic acid 4 and 7 and 6-benzyl adenine to increase growth, yield and sucrose content of selected sugarcane varieties in Kakamega County and similar agro ecologies. Information on use and effect of gibberellic acid 4 and 7 and 6-benzyl adenine would then be availed to the benefit farmers and for sustainability of the industry. 9 1.6 Conceptual framework Figure 1.1: Conceptual framework 10 CHAPTER TWO: LITERATURE REVIEW 2.1 Overview of sugarcane production Sugarcane Saccharum officinarum (Grammineae) is cultivated in the tropical countries and provides over 70% of the world’s sugar requirements (Cox et al., 2000). The main product of sugarcane is sucrose, constituting about 10% of the crop (Tecson-Mendoza, 2000). Sucrose is a highly valued food and sweetener but also serves as a preservative for other foods. The biggest world sugar producers are Brazil with 20.3 million metric tons, India with 19.9 million metric tons and the European Union with 15.5 million metric tons (FAO, 2013). Kenya produces only about 0.6 million metric tons. However, it is a major cash crop in Kenya, ranked fourth in terms of its contribution to Agriculture Gross Domestic Product (AGDP) after horticulture, tea and coffee (FAO, 2013). It supports about 6 million people in the country directly or indirectly and contributes to Kenya’s economic growth (FAO, 2013). The Industry’s contribution to AGDP is about 15% (KSI, 2009). The sector supports more than 250,000 small holder farmers supplying about 90% of sugarcane processed while about 10% comes from nucleus estates (KSI, 2009; KSB, 2010). At least 25% of the Country’s population depends directly or indirectly on the sugar industry for their livelihoods. Many households are able to pay school fees and put up decent houses’ courtesy of sugarcane crop. Farmers earn their income directly from sugarcane proceeds while farm workers, transporters and cane cutters are paid for services (KSB, 2010). 11 Another category of service provider include agro-chemical suppliers, extension officers and sugar researchers who immensely contribute to the improvement of sugarcane production and derive their income as a result (KSI, 2009). Financial institutions like Agricultural Finance Corporation have also emerged and developed suitable loan facilities for sugarcane production (KSB, 2010). Economic growth and development in the rural areas have been spurred by sugarcane crop. Roads and electricity infrastructure and the emergence of rural markets are some of the strong indicators of positive impact (Economic survey, 2001). Apart from supporting beverage industry, confectionary industry, pharmaceutical industry, alcohol industry and animal feed industry, sugarcane production supports domestic sugar consumption considerably (FAOSTAT, 2014). Imported sugar only serves to close the gap created by deficit. To maintain quality supplies of sugar and attain self-sufficiency local sugar factories must be supported to mill fresh cane from the growers (KSB, 2010). Sugarcane was introduced in Kenya in 1902 (Osoro, 1997). The first sugar cane factory was set up at Miwani in Kisumu County in 1922 and later at Ramisi in Kwale County in 1927 (Osoro, 1997). Muhoroni Sugar followed in 1966, Chemelil Sugar in 1968, Mumias Sugar in 1973, Nzoia Sugar in 1978, SONY Sugar and Kabras in 1979, Kibos Sugar in 2007, Butali Sugar in 2008, Ndhiwa Sugar in 2010 (EU, 2012). Trans Mara Sugar followed in 2012, Olepito Sugar in 2016 and Busia Sugar in 2019 (SD, 2019). Today, Kenya has 12 Sugar companies among them 5 are state-owned and 7 privately-owned. 12 The government involvement in the sugar sub-sector particularly after independence was due to five main policy considerations: first, there was need to ensure self-sufficiency with exportable surplus in sugar production (Keya, 1995). Second, sugar production was regarded as an essential import substitution strategy to save the country the much-needed foreign exchange (Keya, 1995). Third, sugarcane growing was seen as a tool for social development. It provided employment opportunities and wealth in the Kenyan rural areas (Keya, 1995). Fourth, sugarcane growing was viewed as an agent for infrastructural and rural development (Keya, 1995). Fifth, at the time of independence; the domestic Kenyan market was served by imported sugar. In order to protect this domestic market, local sugarcane production was a viable alternative (Keya, 1995).Deliberate policy measures were thus put in place to attain this goal among them establishment of Kenya Sugar Research Foundation to undertake research and extension in the sugar industry (Keya, 1995). Although sugar factories have grown in number from 1922 to date, Kenya continues to be sugar deficit country attracting importation from other countries and regions. Only 4 out of 12 licensed mills continue to operate below their installed capacity due to inadequate raw material and factory inefficiencies (SD, 2018). During the period between 2016 and 2017, 475,670 metric tons of sugar was produced against a domestic demand of 718,396 metric tons, leaving a deficit of more than 200,000 metric tons (SD, 2018). This is a clear indication that there is need to increase sugarcane production to meet the demand. To meet the high sugar needs in Kenya, there is need to 13 increase production by 5.2% annually (SD, 2018). However, land for sugarcane cultivation in the high potential areas is becoming increasingly scarce. Therefore, future gains in sugar production must come from increased productivity on already planted land and exploitation of other marginal areas (SD, 2018). It is against this backdrop that a research was conducted in Kakamega County to assess the role of plant growth regulators on sugarcane yields. Global interest in sugarcane has increased significantly in recent years due to its economic impact on sustainable energy production. Globally, fuel technology research and development are a shift from hydrocarbon to carbohydrate (Singh, 2003). Biomass was the principal source of energy from the dawn of human civilization until the middle of the 19th century. Biomass is essentially a form of solar energy and renewable, unlike the fossil fuels that are the main causes of the environmental problems (Lakshmanan et al., 2005). Therefore, importance of bio-energy-generating crops such as sugarcane is increasing rapidly and is likely to play an important role in environmental and economic challenges of fossil fuel usage (Lakshmanan et al., 2005). In Kenya, biofuel from bagasse has been released to the national grid as seen in Mumias Sugar, Butali Sugar and Kibos Sugar (SD, 2018). 2.2 Challenges of sugarcane production Economic liberalization and global trade de-regulation present challenges to the sugar industry sub-sector of the Kenyan economy (KSI, 2009). Multi-lateral and regional trade treaties, especially those associated with COMESA, EAC, SADC and WTO, have 14 facilitated the importation of sugar into Kenya at minimal or zero tariffs from producer member states (Wolfgang and Owegi, 2012). Imported sugar in most cases is heavily subsidized by its source government thus adversely affecting marketability of locally produced sugar, which because of its high production cost relative to imported sugar, cannot compete favorably with foreign sugar in the domestic and foreign markets (KSB, 2010). Although quotas for imported sugar to address the shortfall have been set over the years, the Kenyan government has been unable to enforce these quotas, resulting in annual gluts of sugar supply in the local market (KSI, 2009). Usually, the import figures have no relationship with the shortfall in consumption. It can only be assumed that consumption figures are either exaggerated are that substantial imports are not recorded (KSI, 2009). Large imported stocks have worsened the cash flow problems of domestic sugar companies and the sugar industry as a whole. Farmers have to contend with low sugarcane price and delayed payment as a result. This presents real challenge in sugarcane production (KSI, 2009). Comparative analysis of the cost of sugarcane production between Kenya and other COMESA and EAC Countries reveal high input costs, punitive taxation regime and middlemen activities (Odek et al., 2003). The ten lowest cost sugarcane producers of the world are Australia, Brazil, Columbia, Guatemala, Fiji, Malawi, Swaziland, Thailand, Zambia and Zimbabwe. 15 The total cost of sugarcane production in Swaziland per ton is USD 168.6; while the total average cost including overheads is USD 265.5 (Odek et al., 2003). In Kenya, the cost of producing sugarcane is USD 420 per ton; while in Sudan, the Kenana average cost of production is US 230 per ton. In Sudan, Kenana sugar sells at USD 345 per ton while Kenyan Sugar is sold at an average of USD 430-USD 600 per ton; while the world market sugar sells at USD130-170 (Odek et al., 2003). The rising cost of farm inputs like seedcane, fertilizers, herbicides, transport and labour have presented real challenge to struggling farmers (Wawire et al., 1999). Land held under African tenure system has promoted sub-division to uneconomic land sizes (Wawire et al., 1999). Besides, most small holder farmers pursue survival rather than growth and graduation. Thus, cost of production is usually high and unsustainable (Wawire et al., 2001). Another challenge facing sugarcane production in Kenya is decline in sugarcane productivity. Farm level efficiency has been in constant decline for the last 10 years. While area under cane has increased steadily from 122,580 Ha in 2010 to 198,000 Ha in 2018, average cane yield stands at 57.6 tons per hectare from 60.52 tons per hectare in 2010 (FAOSTAT, 2018). Poor cane husbandry, declining soil fertility status, poor sugarcane nutrition, overdependence on natural rainfall and poor weeds management have contributed to the undesirable decline (FAOSTAT, 2018). Inadequate research and extension has seen farmers stick to very old technologies in terms of seed cane varieties and other agronomic practices. 16 As a matter of fact, research and development into more productive varieties and methods of cane husbandry has not been given much attention, particularly in this era of globalization and intense competition (KSB, 2010). Factories have left this important aspect of cane development in the hands of Sugar Research Institute, which due to lack of resources has not made much progress in the promotion of new varieties (KSB, 2010). The sucrose content of cane grown by Kenyan farmers is much lower than that found in sugar exporting countries such as Sudan and Brazil (FAO, 2013). Cane Testing Unit is a new development in the Kenya sugar industry and is largely viewed as a game changer. If fully implemented, farmers will be paid based on sucrose (SD, 2018). Poor access to affordable credit has frustrated farmers efforts to purchase the necessary farm inputs and make improvements to their farms. The result is that farmers have scaled down their acreage sugarcane or abandoned the crop altogether (Okwach, 2009). Stakeholder and shareholder institutions such as factories and grower cooperatives have been slow to take the initiative and provide farmers with the badly needed credit (EU, 2012). With changing circumstances, sugarcane farmers are seeking varieties which seem to address their various socio-economic and agro-ecological constraints. Early efforts to identify improved sugarcane varieties in Kenya involved importing and testing varieties for adaptation to local conditions (KESREF, 2002). The major commercial varieties currently grown are of Indian and South African origin, and include CO 421, CO 1148, CO 331, CO 945, CO 617 and N 14 (KESREF, 2002). 17 The first organized sugarcane breeding program was established in the mid-1960s to serve the then East African Community (KESREF, 2002).The expansion of sugarcane cultivation into diverse agro climatic conditions over the past 40 to 50 years has increased the demand for new improved varieties (KSB, 2003). The objective of the current variety improvement program is to develop high yielding, pest and disease resistant varieties that are adapted to the cane growing conditions in Kenya (KESREF, 2003). Intensive efforts to develop superior varieties for the Kenyan sugar industry over the past two decades have produced a substantial number of promising varieties (Odenya et al., 2010). Kenya Sugar Research Foundation (KESREF) has made tremendous progress since its inception in 2001 by releasing new varieties with regards to high yielding, early maturing, high sucrose and disease resistance (Odenya et al., 2010). These include: KEN 82-216, KEN 82-247, KEN 82-401, KEN 82-808, KEN 83-737, KEN 82-472, KEN 82-62, KEN 82-493, KEN 82-121 and KEN 82-601(Jamoza, 2005). Other newly released varieties include KEN 00-13, KEN 00-3548, KEN 00-3811, KEN 00-5873, KEN 98-530, KEN 98-533, KEN 98-533, KEN 98-551, KEN 98-367, EAK 73-335 and D 8484 (Odenya et al., 2010). The improved varieties have shown significant potential in improving the productivity of the Kenya Sugar Industry; however, their adoption rate have been lower than expected (Odenya et al., 2010). The breeding traits are expected to translate to economic advantage to the client. 18 Previous studies explained the importance of considering the economic values of breeding traits in a selection criterion. However, the current breeding program in Kenya does not consider the trait’s economic values (Jamoza, 2005; KESREF 2006). This complicates combination of yield and sucrose in their selection criteria to identify superior sugarcane clones. The government of Kenya in partnership with the European Union has installed sugarcane testing units that will ensure that farmers are paid based on sucrose content as opposed to weight-based system (Jamoza et al., 2018). 2.3 Effect of plant growth regulators on crop growth Plant growth regulators (PGRs) are organic compounds, other than nutrients, that modify plant physiological processes (Solaimalai et al., 2001). They are called bio stimulants or bio inhibitors and they act inside plant cells to stimulate or inhibit specific enzymes or enzyme systems and help regulate plant metabolism (Mok, 2001). These plant growth regulators also occur naturally, and they are called phytohomones or plant hormones. They regulate plant growth and development either synergistically or antagonistically, involving a series of complex pathways and networks (Dong et al., 2016). These include: Gibberellic acid (GA3), GA4+7, Kinetin (KN), 6-Benzyladenine (BA) and Salicylic acid (SA) among others (Frankenberger, 2002). There are several prominent phytohormones present in plant like indole-3-acetic acid; which are mainly responsible for various physiological processes such as cell elongation and division, and a defense mechanism by the expression of several genes under biotic and abiotic stress (Song et al., 2014). 19 Steroidal hormones such as brassinolide regulate growth and developmental processes and provides resistance against biotic and abiotic stress such as cold stress and pathogen infection (Haubrick et al., 2006; Vriet et al., 2013). Gibberellins regulate growth and influence various developmental processes, including stem elongation, seed germination and dormancy, flowering, leaf expansion, sex expression, enzyme induction and leaf and fruit senescence (Rodrigues et al., 2012: Muhammad and Muhammad, 2013). Cytokinins (isopentenyladenine) regulate several important metabolic reactions in plants (Ma, 2008) in the presence of several types of CNTs in a soil environment upsurge the endogenous concentration of phytohormones (Mastronardi et al., 2015). Plant growth regulators have been developed to improve crop production. They modulate plant growth and development and mediate responses to both biotic and abiotic stresses (Bergstrand, 2017). Peleg and Blumwald (2011) expressed that the plant hormones play central roles in the ability of plants to adapt to changing environments, by mediating growth, development, nutrient allocation and source/sink transitions. Although, abscisic acid (ABA) is the most studied stress responsive hormone, the role of cytokinins, brassinosteroids and auxins during environmental stress is less studied (Davis, 2010). Recent evidence indicated that plant hormones are involved in multiple processes. Crosstalk between different plant hormones results in synergistic or antagonistic interactions that play crucial roles in response of plants to abiotic stresses (Santner et al., 2009). 20 The characterization of the molecular mechanisms regulating hormone synthesis, signaling and action are facilating the modification of hormone biosynthetic pathways for the generation of transgenic crop plants with enhanced abiotic stress tolerance in the field (Verma et al., 2016). Promalin has been reported to increase vegetative and root growth and yield of kale (Emongor et al., 2004). Plant growth regulators can be used to modify plant growth and development in such a manner as to increase crop yield (El-Otmani et al., 2000). Cytokinins are one of the major plant hormones that regulate numerous aspects of growth and development due to complex crosstalk with stress signaling, especially the response related to abiotic stress tolerance (Zwack and Rashotte, 2015). In many developmental processes, cytokinins and gibberellins act antagonistically. Although most studies profoundly suggest a gibberellin-regulated cytokinin action, evidence relating to cytokinin regulating cytokinin activity cannot be ignored (Fleishon et al., 2011). Scuttle (2004) reported that dormancy of potato tuber was effectively broken with 6- benzyladenine at a concentration of 20 ppm used for 24 hours. Yun Kyong Shin (2011) studied the effect of BA and ultrasonic pre-treatment on in vitro germination and protocorm formation of calanthe hybrids. Bang-zhen et al., (2010) observed that BA treatment significantly increased the seed yield of the biofuel plant Jatropha curcas. Plant growth begins with seed germination, the success of which depends on the ability of plant embryo to gain its metabolic activity (Rajjou et al., 2012). 21 Several molecular cues have been revealed by different genetic and proteomic investigations of various Arabidopsis mutants, showing distinct germination-related phenotypes regulation of plant hormones, including gibberellic acids (GA), Abscisic acid (ABA), auxin and ethylene (Han and Yang, 2015). Germination is also significantly affected by several environmental factors, such as various abiotic stresses (Rajjou et al., 2012; Han and Yang, 2015). These factors mainly affect the metabolism and different signal pathways of GA and ABA (Holdsworth et al., 2008). The constantly changing external factors that most affect plant growth and development are abiotic stresses like salinity, drought and cold (Mahajan and Tuteja, 2005). Abiotic stresses trigger ABA biosynthesis, which mediates stress adaptive responses by activating several specific signal cascades thereby regulating different physiological and growth-related processes (Vu et al., 2015). The plant growth regulators have been used commercially to promote growth and development. The commercial uses of auxins include prevention of fruit and leaf drop, promotion of flowering, thinning of fruit, induction of parthenocarpic fruit development and rooting of cuttings for plant propagation, among others (Taiz and Zeiger, 2010). Considerable success had been made in the application of PGRs in some process of plant development such as flowering and fruit development as well as ripening, harvesting and post-harvesting of fruits and other crops (Fahad et al., 2016). Sajid et al., (2009) revealed effects of foliar application of PGRs and nutrients for improvement of lily flowers (Lilium spp). In seedless grapes (Vitis spp.). 22 Bhat et al. (2011) found that the application of CPPU and BR along with GA increase the leaf number, leaf area and leaf dry matter and directly influencing the fruit yield and quality. Besisides, Tiwari et al. (2011) informed about effect of gibberellic acid and other plant growth regulators as naphthalene acid on hybrid rice (Oryza sativa L.) seed production. Over expression of cytokinin biosynthetic genes increases endogenous cytokinin levels that eventually enhance heat stress tolerance in grass (Xing et al., 2009). In addition, up regulation of the endogenous cytokinin level by exogenous application of cytokinin can also improve tolerance to heat stress in bent grass (Xu and Huang, 2009). The effects of PGRs have also been studied in plant abiotic stress. In this sense, Ullah et al. (2012) showed the effects of PGRs on growth and oil quality of canola (Brrassica napus L.) under drought stress. Growth regulators were highly effective in ameliorating the adverse effects of drought stress on both cultivars. In vitro plant tissue culture, the use of PGR is an important practice regardless of plant species. For example, in rapid in vitro micro propagation of sugarcane (Saccharum officinarum) through callus culture (Behera and Sahoo, 2009), in their study of hormonal regulation of branching in grasses especially with auxin, cytokinin, strigolactones, among others revealed the importance of the use of plant growth regulators and tissue culture techniques to horticulture, the achievements and limitations of tissue culture and some insights into current and possible future developments. The importance of plant growth regulators was first recognized in the 1930s. Since that time, natural and synthetic compounds that alter function, shape and size of crops have 23 been discovered (Kalaran et al., 2002). Today, specific PGRs are used to modify crop growth rate and growth pattern during the various stages of development from germination to maturity (Joyce et al., 2010). Growth regulating chemicals that have positive influences on major agronomic crops can be of value. The final test, however, is that harvested yields must be increased or crop quality enhanced in order for PGRs to be profitable (Suprasanna et al., 2008). Of the many current uses of PGRs, effects on yield are often indirect (Hotta et al., 2010). Studies conducted on major grain crops, such as corn, soybean, wheat and rice have identified materials capable of altering individual agronomic characteristics like lodging, plant height, seed number and maturity. Even then, these changes have not always resulted in increased yields (Chegalrayan and Gallo Meaghahar, 2003). Field crops produce relatively lower returns compared to horticultural crops. Therefore, use of PGRs on field crops has not been as vigorously pursued by chemical companies and public research scientists (Bairu et al., 2011). A wide assortment of plant growth regulators promotes germination and emergence, stimulate root growth, promote mobilization and translocation of nutrients within plants, increase stress tolerance and improve water relations in plants, promote early maturity, retard senescence and improve crop yields and quality (Larkshmana et al., 2005). Results obtained under carefully controlled conditions are not easy to reproduce in the field (McDonald et al., 2001). Effects of environment, crop management and variety on 24 crop responses and yields are usually much more pronounced than the effects of PGRs. This makes it difficult to demonstrate a yield or quality response to the application of a PGR (Miyawaki et al., 2004). Cole and Wheeler’s research showed that cotton seeds soaked for 6-24 hours in gibberellic acid or cyclic AMP increased germination percentages over a range of temperatures. To be effective, the seeds must absorb plant growth regulators into the embryo cells (Jeyakumar et al., 2008). The relative concentration of gibberellins was established as a strong determinant of shoot and root organogenesis in callus culture: high cytokinin/auxin ratio favoured shoot formation while low cytokinin/auxin ratio favoured root growth (Xin et al., 2000). Since then, numerous studies have confirmed that plant growth regulators are essential for de novo bud formation from callus and cultured plant tissues (Swarup et al., 2002). Manipulations of endogenous benzyl adenine through genetic changes have supported these findings. Transgenic plants over-expressing the IPT genes displayed bushy phenotypes expected of benzyladenine overproduction (Sujatha et al., 2001). These include Nicotiana plants habouring the Agrobacterium IPT gene, Arabidopsis with their native genes AtLPT4 and AtLPT8 and Petunia hybra transformants with Agrobacterium IPT (Kakimoto, 2001). Benzyl adenine deficiency was studied via insertional mutations of AtLPT genes and receptor histadine kinases genes or by over exposing cytokinin oxidase/ dehydrogenase genes. The resultant plants showed a general growth inhibition of the aerial parts; including reduced meristematic activity, thinner stems and smaller leaves as well as 25 increased root growth (Miya et al., 2006). Embryonic axes of developing seeds are sites of benzyl adenine biosynthesis (Sun et al., 2003). For example, endosperm of Phaseolus contains very high levels of the ZOG1/ZOX1 enzymes (Clark et al., 2004) and it is possible that the stored glycosides are hydrolyzed during germination. This is supported by a recent study with tobacco transformants harbouring the promoter of the maize ß-glycosidase gene (ZmGLu1) linked to the Gus gene, which showed that expression was low in mature seeds but increased during germination, peaking during radical and cotyledon emergence (Gu et al., 2006). This increase in ß-glycosidase expression suggests benzyl adenine reserves during germination. The stimulatory effects of gibberellins on leaf expansion were first observed with radish leaf discs. This effect on leaf expansion may be linked to the ability of gibberellins to enhance sink strength for nutrients (Higuchi et al., 2004). Gibberellins have been proved to play a decisive role in all phases of plant growth and development from germination to maturation. Some closely related gibberellins are difficult to separate and identify. GA4 and GA7 differ from GA3 due to the presence of hydroxyl group the junction of rings and double bonds (Rai et al., 2017). GA4 has been shown to improve fruit quality of apple. GA7 although structurally similar to GA4 is considerably less effective on its own. A mixture of GA4 and GA7 is commercially available and effective in the induction of sucrose formation in sugarcane (Archard et al., 2013). Physiologically GA4 has been reported to be more active than GA3 26 in bringing about germination of lettuce seeds. Rademacher W. (2015) observed that treatment of gibberellic acids 4 and 7 in cucumber seeds improved germination. However, if low temperature was maintained more of the growth regulator would be required to promote germination. Sarma (2001) reported that gibberellic acid 3 at 500 ug/ml and kinetin at 1.0 ug/ml proved to be optimal in inducing sprouting of coleus tuber. Cytokinin application could be effective in alleviation of high temperature on barley seed germination by causing changes in the protein and nucleic acid components of tissues which are the basis of effects of cytokinins in cell division as well as on growth and mobilization actions (Cavusoglu and Kaber, 2007). 6 benzyl adenine (6-BA) is one of the most effective cytokinins. Suttle (2004) reported that dormancy of potato tuber was effectively broken with 6-benzyl adenine at a concentration of 20 ppm soaked for 24 hours.Yun-Kyong-Shin (2011) studied the effect of 6-BA and ultrasonic pretreatments on invitro germination and potato corn formation of calanthe hybrids. Ban-znen (2010) observed that 6-BA treatment significantly increased the seed of the bio-fuel plant Jatropha curcas. 2.4 Role of gibberellins and cytokinins in sugarcane growth, yield and sucrose Sugarcane is a tufted grass with limited underground branching and erect stalks. To obtain the maximum number of stalks supported by an area of cane, it is necessary to induce branching at an early stage (Ren, Gao and Chen 2007). A search for compounds that induce tillering in sugarcane was started several years ago and several compounds such 27 as ethephon, chlormequat and promalin have been found to be active (Kermode A.R. 2005). Figure 2.1: Sugarcane stool showing primary and secondary shoots (Source: elibrary.sugarresearch.com.au) The PGRs have been used in the sugarcane industry for over two decades to increase the production of sugarcane yield and sucrose (Amanullah et al., 2010). The first commercial success was in the prevention of flowering followed by the application of gibberellic acid (GA3) for the increase of stalk elongation which ultimately resulted in increased sugar production (Grennan and Aleel 2006). It is recognized that the sugar industry throughout 28 the world is using chemicals of the PGR type at almost every stage of the crop development (Daphne and Michael 2005). Benzyl adenine is an essential plant growth regulator for cell division and differentiation. It was the first plant growth regulator identified (Desai et al., 2004) as a factor of stimulating cell division. Subsequently, commercial plant growth regulators were found to influence a wide range of developments including seed germination, de novo bud formation, stomatal control, reproductive development, photosynthesis and respiration (Synman et al., 2011). Gibberellins are known to affect plant morphology by promoting cell division and cell enlargement. They control sugarcane development including germination, stem elongation, root extension, leaf area and number, yield and sucrose accumulation (Hedden and Thomas, 2012). Gibberellins are involved in the natural process of breaking dormancy by signaling starch hydrolysis through inducing the synthesis of the enzyme amylase in the aleurone cells. The starch is hydrolysed into glucose that can be used in cellular respiration to produce energy for the seed embryo. (Rae et al., 2005). GAs are presumed to stimulate photosynthetic activity and the increased photosynthetic sugar content induces shoot growth (Iqbal et al., 2011). Cytokinins and gibberellins have been reported to stimulate the formation of well-developed chloroplasts and increase vegetative growth and maturation in sugarcane (Salopeck Sondi et al, 2002). The use of chemicals such as glyphosine, ethephon and promalin have been evaluated in sugar 29 industries around the world to increase recoverable sugar. (Dalley and Richard, 2010). Promalin is a mixture of two naturally occurring plant growth regulators: gibberellic acid 4 and 7 (GA4+7), which causes cell enlargement and elongation, and 6-benzyladenine (6-BA) which promotes cell division (Werner et al., 2003). Sucrose accumulation in sugarcane and elongation of internodes was achieved due to exogenous application of promalin (Taiz and Zeiger, 2004). Due to problems of sugarcane emergence (including such factors as depth of planting, angle of the bud, adverse weather conditions, particularly excess moisture and cold and fungal and bacterial infections), it becomes important to use plant growth regulators to obtain a good crop stand (Kopency et al., 2006). In an empirical search through many compounds looking for those which might stimulate the rate of germination or early growth of the young sugarcane plant, it was found that benzyl adenine and gibberellic acid caused considerable stimulation of germination. This growth regulator was also shown to cause an increase in growth rate of the plant at later stages, resulting in increased production of both cane yield and sugar (Riefler et al., 2006). Additional studies showed that the effect is evident at the cellular levels, i.e. cells of sugarcane grown in suspension culture respond rather dramatically to promalin (Huang et al., 2003). Subsequently, considerable work has been done with promalin in relation to its effects on sugarcane growth and performance. Dramatic results obtained with 30 promalin suggest that if these findings could be exploited commercially, they could easily lead to one of the greatest increases in sugar production ever achieved by application of chemicals (Perry and Hagenbeek 2007). However, in order to achieve this, much more whole-plant physiology and field work must be carried out. Several plant hormones have been shown to affect germination of seeds of some plant species. The primary event of breaking seed dormancy is stimulated by gibberellins. Germination of field crops is sometimes decreased by cold soil temperatures (Takei et al., 2004). Gibberellic acid exerts its influence by increasing the growth potential of embryo and by inducing hydrolytic enzymes. GA stimulates gene expression involved in cell expansion and modification (Cookie et al., 2012). Benzyl adenine levels in dormant seeds are low but increase during germination after a transitory decrease (Miyaka et al., 2004). This increase may be due to both benzyl adenine biosynthesis in the embryonic axes and possible release of active cytokinins from conjugate forms. Phytohormones like GA3 (gibberellin acid) have been reported to affect cell expansion thereby increasing sink size and sink strength, thus enhancing the competitive ability of the organ to draw assimilates (Iqbal et al., 2011). GA3 perhaps affects source-sink communication by stimulating assimilate transport and increasing phloem unloading of sucrose into the sink, thereby establishing a more favourable sucrose gradient between sink and source (McCormick et al, 2006). 31 Sugarcane houses a unique source-sink system wherein the mature culm serves as a large sucrose reservoir called the sink and the leaves play the source of photosynthetic sugar (Watt et al., 2014). Sugarcane sprayed with GA3 showed dramatic increase in intermodal length confirming effect of GA3 in increasing the sink size and consequently the sink potential as well as sink demand, further validating its role in increasing cell size, sink size, sink capacity, making more room for sucrose accumulation (Van Bel 2003). 2.5 Cost-benefit analysis Sugarcane productivity and quality are the twin magnets that keep farmers constantly on the upward course. However, they are inadequate without profitability, necessitating economic evaluation (Devi et al., 2013). Azam et al. (2010) conducted study about profitability of sugarcane due to use of gibberellic acid 3 (GA3). The results revealed a positive influence on the dependent variable output with high productivity and profitability reported. Laghari et al. (2003) reported that sugarcane sprayed with promalin in sanghar area involved cost of production which was lower than gross yield return per acre. The benefit cost ratio for sugarcane crop was estimated to be 2.2. Mohammad et al. (2006) examined the economics of sugarcane production and its competitiveness. In his study of use of plant growth regulators, effect and results, he concluded that profitability of sugarcane depended on manipulation of yields at reasonable cost. The specific target of increasing sugarcane profitability could be by developing cost- effective technologies, transferring them to farmers and creating a linkage between all stakeholders. Any possibility of enhancement in the income of sugarcane farmers shall 32 depend on increasing productivity and profitability while controlling cost of production (Giroh D. Y., 2012). Development of low-cost technologies to convert “waste to resource” on a smaller scale shall also help the farmers to increase their income further. The reason for higher returns and benefit cost ratio was due to higher cane yield (Prahaj et al., 2017). Promalin improved cane juice quality at harvest stage by enhancing the export of carbon assimilates from source to sink organ (Tan et al., 2007). 33 CHAPTER THREE: MATERIALS AND METHODS 3.1 Study area The study was conducted at the Butali Sugar Company Research and Demonstration station in Chegulo Location, Kakamega County. The area receives an average annual rainfall of 1700 mm with a mean minimum and maximum temperature of 15°C and 27°C respectively. The altitude is 1600M above sea-level. The annual mean relative humidity is 65% with photoperiodism (day length) of 12 hours and topography of 25% (Kakamega Metrological department, 2014). The soil is predominantly clay to sandy-loamy with a pH of 6.2-7.4. Figure 3.1: Map of Kakamega County showing areas of cane development (source: GOK, 2013) 34 Figure 3.2: Map of Malava Sub-County showing Chegulo study site (Source: http//.www.google.co.jp/url., 2016) 3.2 Research design and treatments The experiment was a 5x5 factorial with plots arranged in Randomized Complete Block Design replicated three times. The experimental units comprised of 25 plots measuring 2.4 meter by 3.6 meter and with inter-block spacing of 1.5 meter and inter-plot spacing of 1 meter. The treatments consisted of five sugarcane varieties and five levels of promalin (GA4+7 and 6-BA) plant growth regulator. The varieties were: CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335; all of them locally grown in Kakamega County. Seedcane was detrashed and cut into three-budded setts and soaked for 24 hours in Promalin (GA 4+7 and 6-BA) at various levels; 0 (control) and 1, 2, 3, 4 litres ℎ𝑎−1 prior to planting. Promalin is plant growth regulator developed and marketed by Valent Biosciences 35 Corporation in the United States of America. It consists of two naturally occurring gibberellic acids 4 and 7 which cause cell elongation and enlargement and 6-benzyl adenine which causes cell division. 3.3 Soil sampling and analysis Physio-chemical analysis of soil in the experimental site of Chegulo was done before planting. Soil samples were extracted randomly from the site to produce composite samples which were then taken to Sugar Research Institute ultra-modern laboratory in Kisumu for comprehensive analysis. Soil pH, Electrical Conductivity (EC), Organic carbon (OC), total Nitrogen (N), Phosphorus (P), Potassium (K), Calcium (Ca), Magnesium (Mg), Copper (Cu), Manganese (Mn) and Iron (Fe) were analyzed using wet chemistry method. Table 3.1: Physio-chemical status of soil at Chegulo From the table above Chegulo field A had its soil near neutral, adequate in Mn but low in C, N, P, K, Mg, Cu and Fe, and was non-alkaline. Chegulo B was found to be slightly acidic, adequate in Mn and Fe but low in C, N, P, K, Ca, Mg and Cu, and was non saline. 36 Although Chegulo fields A and B were positioned side by side, separated by about 15 meters only, they had distinct characteristics. 3.4 Planting materials Planting materials consisted of three-budded seed cane setts of five sugarcane varieties commonly grown in Kakamega County namely: CO421, KEN 83-737, D8484, CO945 and EAK 73-335. Seedcane was sourced from nursery B of SRI plots in Mumias. CO421 and CO945 originated from Coimbatore state in India, KEN 83-737 from Mtwapa Kenya, D8484 from Demerara state in Guayana and EAK from Uganda in East Africa. These varieties are widespread in sugarcane growing zones in Kenya (Jamoza, 2005; KESREF 2006). CO 421 is late maturing variety (20-24 months), drought resistant and retains sugar for a long time after physiological maturation. CO 945 is mid maturing variety (18-20 months) which is a poor germinator but exhibits heavy tillering. EAK 73-335 is mid maturing variety (18-20 months) which is high in pol% (16) and yield. KEN 83-737 is early maturing variety (16-19 months), exhibiting heavy tillering and drought resistance. However, it flowers profusely in high altitude areas. D8484 is early maturing variety (14- 16) months, high in pol% (15.5). It exhibits wide girth but low tillering. 37 Table 3.2: Treatment levels and labels Variety GA 4 + 7 and 6-BA Level Label Level Label 1 CO421 1 0 ltrs/Ha 2 KEN83-737 2 1 ltrs/Ha 3 D8484 3 2 ltrs/Ha 4 CO945 4 3 ltrs/Ha 5 EAK73-335 5 4 ltrs/Ha 3.5 Land preparation and planting Land preparation involved disc ploughing, harrowing, and furrowing. Planting for season one was done on 16th September, 2015 and harvesting done on 15th September, 2016. Planting for season two was done on 27th April, 2016 and harvesting done on 26thApril 2017. Three (3) budded setts of the five sugarcane varieties were placed end to end before being buried at the uniform depth of 15 centimeters. The number of setts used per plot were between 31 and 36. NPK (23:23:0) at the rate of 80 Kg P2O5ha −1 was applied along the furrows. 3.6 Field management Top dressing was done 100 days after planting at the rate of 60 KgN/ha. The crop was subjected to natural rainfall conditions and weeds control was by hand weeding. Regular scouting for pests and diseases was maintained. Off types and panicum grass were rogued out. Plots were marked accordingly using display boards for ease of identification. The site was in a protected area with a resident Research Assistant. 38 3.7 Data collection 3.7.1 Effect of levels of GA 4+7 and 6-BA on growth of selected sugarcane varieties Data on germination was gathered by counting primary shoots at 2 months. Data on tillering was obtained by counting secondary and tertiary shoots for all plants in each plot at 3, 4, 5 and 6 months. This was accordingly tabulated.Data on height was gathered by measuring the vertical length of all the plants per plot from the base to the level of first leaf branching using a tape measure. Length of internodes was determined by measuring the internodes of each individual plant in a plot starting at 3 months and computing the average length of internode per plant. Data on number of leaves per plant was gathered by counting all the leaves of all the plants in each plot except the bud. All this data was collected for the five varieties at an interval of 30 days for six months. 3.7.2 Effect of GA 4+7 and BA on yield and sucrose of selected sugarcane varieties Data on seed cane yield was collected by harvesting all the sugarcane from each plot at 12th month, weighing and recording. Data on pol% at the 12th month was generated by randomly sampling ten sugarcane stalks from each plot and cutting out five internodes (from the bottom part of each cane), which was then macerated to extract juice. All juice from the sugarcane per plot was homogenized in a beaker and a composite sample of 250 ml treatment obtained. The brix% was read using refractometer spindle. Pol% was obtained by adding Lead sub- acetate (2.5-3.0 ml) depending on the brix% and pol read in polarimeter before multiplying by the corresponding pol factor to obtain pol%. 39 3.7.3 Cost-benefit analysis of use of gibberellic acid 4 and 7 and 6-benzyl adenine Cost and benefit structure per hectare was developed as follows: cost components were listed and assigned monetary value after gathering the same information from relevant sources; benefit in terms of extra yield was got by subtracting yield due to 0 litre (GA+6- BA) per hectare from yield due to 3 litres (GA+6-BA) per hectare for all varieties under study before multiplying by net payable per ton. The price of sugarcane was that of Butali Sugar Mills Limited while the price of promalin (PGR) was got from the suppliers (Valent biosciences). Market rates for labour and water were considered in this study. Income due to extra yield was discounted to get net present value (NPV). Table 3.3: Extra cost per hectare There were only three cost components with promalin plant growth regulator constituting 85.5% of the total cost. The prices used in this study were current prices charged at market rates. 40 Table 3.4: Yields at 0 and 3 litres per hectare of GA 4+7 and 6-BA The recommended rate of 3 litres per hectare of gibberellic acid 4 and 7 and 6-benzyl adenine was used in this analysis. Only yield was analyzed as payment based on sucrose content formula had not been fully implemented. Table 3.5: Extra yields at 3 litres per hectare of GA 4+7 and 6-BA Variety/Yield Average yield due to 0 L/Ha of GA 4+7 and 6-BA Average yield due to 3 L/Ha of GA 4+7 and 6-BA Extra yield due to 3 L/Ha of GA 4+7 and 6-BA (Y1) (Y2) (Y2 – Y1) CO 421 (V1) 58.24 66.9 8.66 KEN 83-737 (V2) 60.61 69.48 8.87 D8484 (V3) 62.28 73.93 11.65 CO 945 (V4) 60.03 66.65 6.62 EAK 73-335 (V5) 58.99 66.32 7.33 Average yield due to 0 L/Ha of GA 4+7 and 6-BA was computed by taking the recorded yield for control in season 1 and 2 and dividing by 2. For 3 L/Ha, the total recorded yield Average Yield (Y1) Total Yield Average yield (Y2) Y1=0.5(Y1a+Y1b) (Y2a + Y2b) Y2=0.5(Y2a + Y2b) CO 421 (V1) 58.6 57.87 116.47 58.24 66.94 66.86 133.8 66.9 KEN 83737 (V2) 60.8 60.42 121.22 60.61 69.52 69.44 138.96 69.48 D 8484 (V3) 62.79 61.77 124.56 62.28 75.04 74.81 149.85 73.93 CO 945 (V4) 60.3 59.76 120.06 60.03 66.7 66.59 133.29 66.65 EAK 73-335 (V5) 59.34 58.64 117.98 58.99 66.4 66.24 132.64 66.32 Variety/Yield Season 1 Yield at 0 L/Ha (Y1a) Season 2 Yield at 0 L/Ha (Y1b) Total yield (Y1a+Y1b) Season 1 Yield at 3 L/Ha (Y2a) Season 2 Yield at 3 L/Ha (Y2b) 41 for season 1 and 2 was taken and divided by two. The difference between the two averages was entered as extra yield. Table 3.6: Discounted value of extra yield Variety Extra yield due to 3 L/Ha of GA 4+7 and 6- BA (Y2-Y1) Net payable per tons in Kshs. Total income per hectare in Kshs. Net present value per hectare in Kshs. CO 421 (V1) 8.66 3,000 25,980 22,519 KEN 83 737 (V2) 8.87 3,000 26,610 23,065 D8484 (V3) 11.65 3,000 34,950 30,294 CO 945 (V4) 6.62 3,000 19,860 17,214 EAK 73 335 (V5) 7.33 3,000 21,990 19,061 Note: Net Present Value (NPV) =PV/ (1+r) n PV denotes Present Value i.e extra income due to use of GA 4+7 and 6-BA; r denotes rate set at 10 %( 0.1) i.e discount rate; n denotes time- set at 1.5 years (time taken for sugarcane to mature). The analysis was done for the recommended rate of 3 litres (GA 4+7 and 6-BA) per hectare and corresponding yield evaluated, discounted and compared with cost. 3.8 Data analysis Data was subjected to Analysis of Variance (ANOVA) using SAS 9.1 software. Significant differences between means were separated using Tukey’s test at 5% level of 42 probability. Cost-benefit analysis was computed by identifying, discounting and comparing aggregate costs and benefits within the framework of proposed technology (use of GA 4+7 and 6-BA). Finally, benefit-cost ratio was performed to test robustness of the CBA result to the changes. Cost-Benefit Ratio was computed by diving Gross Return by Total Cost. B/C = Gross Return/Total cost 43 CHAPTER FOUR: RESULTS AND DISCUSSION 4.1 Germination Germination of sugarcane stools was influenced significantly (P≤0.05) by application of gibberellic acids 4 and 7 and 6-benzyl adenine as revealed by the sugarcane varieties responses. D8484 had the highest number of stools at 2 months of planting upon application of gibberellic acids 4 and 7 and 6-benzyl adenine, recording 5.33 stools (Figure 4.1). In general, the shoot number at germination significantly increased with increasing levels of gibberellic acid 4 and 7 (Figure 4.1). CO 945 had the lowest number of shoots recording 2.45 and 2.55 shoots per stool in season 1 and 2 respectiovely.This showed that gibberellic acids 4 and 7 and 6-benzyl adenine as a growth promoting agent enhanced faster emergence of the planted sugarcane. This could also imply that the gibberellic acids 4 and 7 and 6-benzyl adenine eliminated any kind of dormancy in the sugarcane varieties and promoted higher rate of germination. 44 Figure 4.1: Effect of GA 4+7 and 6-BA on germination of sugarcane The findings of this study agree with those of Davies, (2010) who found out that gibberellic acids contribute to the expression of certain genes related with germination in tomataoe and led to a significant increase in the rate of germination compared to those that did not have any treatment. In another study by Patel and Mankad, (2014) under 150 ppm GA3 concentrations, tolerant cultivar NDO-2 revealed higher germination percentage whereas sensitive cultivar UPO-94 showed low germination as compared to 100 ppm GA3 concentration under different salinity levels. GA3 was reported to increase germination percentage by withstanding preventive effects of salinity stress on germination. Increase in germination percentage with GA3 use might be due to 0 1 2 3 4 5 6 V1 V2 V3 V4 V5 N u m b er o f sh o o ts p er s to o l Varieties T1 T2 T3 T4 T5 45 involvement of GA3 in the activation of cytological enzymes along with increase in cell wall plasticity and improved water absorption (Padma et al., 2013). Sudharmaidevi et al. (2017) observed that GA3 induced different physiological responses in plants which then stimulated and improved germination and photosynthetic activity which also agrees with the findings of this study. In another study, it was reported that treatments with gibberellins promoted higher and faster sprouting process and that endogenous gibberellins facilitated germination in tomato seeds by weakening the mechanical restraint of the endosperm cells to permit radicle protrusion (Groot and Karssen, 2004). 4.2 Tillering The results on tillering showed significant (P≤0.05) differences between varieties and levels of gibberellic acid applied. CO 945 produced the most tillers when treated with gibberellic acids 4 and 7 and 6-benzyl adenine levels at 15.17 and 15.01 tillers per stool in season 1 and 2 respectively (Figure 4.2).The higher the level of gibberellic acid 4 and 7 and 6-benzyl adenine, the greater effect on the number of tillers in the selected sugarcane varieties. The fact that gibberellins constitute a group of plant hormones that act in the cell elongation and division explain the reason for improved hormones which are synthesized in the form of gibberellic acid. This regulator’s concentration and plant tissues sensitiveness to this compound determine plants response regarding growth and development (Buchanan et al., 2015). 46 Figure 4.2: Effect of GA 4+7 and 6-BA on tillering of sugarcane The findings of this study agree with those of Espindula et al, (2010) who reported overall growth stimulation in plant due to exogenous application of gibberellic acids as it influences morphological and reserve attribute growth. Also, considering that growth regulators from gibberellins, when applied on plants, might improve physic and physiological features, stimulate cell division and elongation, this study aimed at evaluating the influence of gibberellic acid doses applied exogenously on bean crop through seed treatment, and analyzing its effects on morphological attributes of seedlings and physiological quality of the seeds produced. In another study, (Wang et al., 2017) it was reported that exogenous applications of gibberellic acid helped information of more tillers as it was confirmed from the current findings. 0 2 4 6 8 10 12 14 16 V1 V2 V3 V4 V5 N u m b er o f sh o o ts p er s to o l Varieties T1 T2 T3 T4 T5 47 Number of shoots exhibited significant differences (P≤0.05) in sugarcane varieties and gibberellic acids 4 and 7 and 6-benzyl adenine levels in the two study sites. All varieties and GA 4+7 and 6-BA levels resulted in increase in number of shoots in monthly recordings. CO 945 was the superior variety in number of shoots during the sixth month recording 14.99 in season 1 while KEN 83-737 was superior with 13.32 in season 2 as shown in table 4.1. In season 1 D8484 had the least number of shoots with 11.36 while in season 2; EAK 73-335 had the least of 12.75. Table: 4.1: Number of tillers as affected by variety and GA 4+7 and 6-BA Season1 Season2 Variety MAP3 MAP4 MAP5 MAP6 MAP3 MAP4 MAP5 MAP6 V1 9.34a 9.35a 11.67c 13.09c 9.09a 9.49a 11.61a 12.96a V2 9.41a 9.30a 12.20b 14.29b 9.04a 9.45a 11.51a 12.94a V3 8.81a 9.24a 10.35d 11.36e 9.10a 9.59a 11.88a 13.32a V4 9.05a 9.40a 13.43a 14.99a 8.85a 9.58a 11.55a 13.03a V5 9.15a 9.30a 10.71d 11.74d 9.24a 9.58a 11.39a 12.75a LSD 0.45 0.17 0.43 0.28 0.49 0.49 0.87 1.03 GA T1 8.27d 9.22 a 10.75c 12.58a 8.85a 9.40a 11.40a 12.77a T2 8.88c 9.31a 11.40bc 12.87a 8.95a 9.53a 11.59a 12.92a T3 9.29b 9.32a 11.84ab 13.14a 9.17a 9.61a 11.81a 13.25a T4 9.59a 9.35a 12.06ab 13.33a 9.10a 9.51a 11.57a 13.13a T5 9.73a 9.40a 12.30a 13.55a 9.25a 9.64a 11.58a 12.92a LSD 0.26 0.17 0.85 1.07 0.49 0.48 0.87 1.03 VXT NS NS NS NS NS NS NS NS Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. Gibberellic acid 4 and 7 and 6-benzyl adenine levels also exhibited significant differences (P≤0.05) in number of tillers. In both sites, it led to an increase in the number of shoots with the highest being recorded in the MAP 6. 48 Gibberellic acids 4 and 7 and 6 benzyl adenine at 4 litres per hectare was the most superior in accumulation of number of shoots in the sugarcane varieties in season 1 with the highest number being recorded during the 6th month of application with 13.55 while in season 2, 2 litres per hectare had the highest value of 13.25 as shown in table 4.1. Gibberellic acids 4 and 7 and 6-benzyl adenine at 0 litres per hectare (control) which was the lowest level of application had the least number of shoots at all sampling stages which is an indication that application of sufficient levels of GA 4+7 and 6-BA influences growth of sugarcane shoots. The results did not reveal any interaction effects between the varieties and the GA 4+7 and 6-BA levels used in this current study. There was a significant change in growth rate in season 1 in the varieties with the highest change being recorded in CO945 variety with 5.94 and the lowest was in EAK 73-335 variety with 2.58 as shown in table 4.2. In season 2 there were no significant growth change observed. The positive response in increase in number of shoots on application of GA 4+7 and 6-BA could be due to increased cell division that enhanced growth of more shoots. 49 Table 4.2: Growth rate of shoots as affected by variety and GA 4+7 and 6-BA Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. The findings of this study agree with those of Benny et al. (2017) who reported positive responses on plant species in shoots growth upon application of foliar applications during the early stages of growth. Gibberellic acid was observed to show a hyponastic response as a result of increased cell elongation. The variation in the response of sugarcane varieties to gibberellic acid application agrees with the findings of Mesejo et al. (2016) who reported a difference in the response of the shoot growth in the citrus species. Growth rate of shoots Varieties Season 1 Season 2 V1 3.75c 3.87a V2 4.88b 3.90a V3 2.55d 4.21a V4 5.94a 4.17a V5 2.58d 3.15a LSD 0.29 0.97 GA T1 4.32a 3.91a T2 3.98a 3.96a T3 3.86a 4.08a T4 3.74a 4.03a T5 3.82a 3.66a LSD 1.04 0.98 50 In another study done by Nguyen et al. (2019), an increase in growth rate of sugarcane varieties as a result of use of gibberellins resulted in high yield which also agrees with the findings of the current study. Rai et al. (2019) also reported stimulated growth of sugarcane shoot when gibberellic acid was added which also conforms to results of this study. 4.3 Stem diameter Stem diameter exhibited significant differences (P≤0.05) between the varieties and GA 4+7 and 6-BA treatments in season 1 while no significant difference was observed in season 2. D8484 had the greatest effect in stem diameter with the highest value of 3.15 cm being recorded in month 6 while CO 945 variety had the least value of 1.89 cm at month 6 as illustrated in table 4.3. The increase on the stem diameter could be due to elongated stem diameter as one of the influences of gibberellic acid supplied in adequate levels. In addition, it showed that some varieties have better response to gibberellic acids 4 and 7 and 6-benzyl adenine than others. 51 Table 4.3: Stem diameter in centimeters as affected by variety and GA 4+7 and 6- BA Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. Gibberellic acids 4 and 7 and 6-benzyl adenine levels also exhibited statistical differences (P≤0.05) in season 1 while in season 2 there were no differences observed. GA 4+7 and 6-BA at 4 litres per hectare was superior in all the months where the highest was recorded in month 6 with 2.59 cm. GA 4+7 and 6-BA at 0 litres per hectare was the least superior in stem diameter influence with the least value of 1.30 cm being recorded in month 3. No significant differences were observed in the interaction effects of varieties and the GA 4+7 and 6-BA levels. In season 1, there was significant change in stem growth rate in both varieties and GA 4+7 and 6-BA levels while in season 2, there was no significant differences observed. In Site Season1 Season2 Variety MAP3 MAP4 MAP5 MAP6 MAP3 MAP4 MAP5 MAP6 V1 1.66bc 1.86c 2.28b 2.72b 1.80a 2.08a 2.33a 2.64a V2 1.50c 1.75cd 1.91c 2.08d 1.67a 1.91a 2.14a 2.55a V3 2.22a 2.52a 2.76a 3.15a 1.49a 1.79a 2.04a 2.32a V4 1.27d 1.63d 1.75c 1.89e 1.62a 1.94a 2.08a 3.11a V5 1.79b 2.10b 2.30b 2.56c 1.79a 2.01a 2.21a 2.46a LSD 0.21 0.17 0.18 0.14 0.32 0.31 0.29 0.86 GA T1 1.30c 1.65c 1.93c 2.27a 1.61a 1.89a 2.11a 2.44a T2 1.55bc 1.86bc 2.04bc 2.43a 1.63a 1.93a 2.09a 2.40a T3 1.74ab 2.01ab 2.26ab 2.56a 1.72a 1.98a 2.24a 2.54a T4 1.86a 2.14a 2.35a 2.56a 1.63a 1.94a 2.11a 2.47a T5 1.99a 2.21a 2.42a 2.59a 1.79a 2.06a 2.25a 3.23a LSD 0.26 0.25 0.29 0.36 0.32 0.31 0.30 0.86 VXT NS NS NS NS NS NS NS NS 52 season 1, CO 421 variety had the greatest change in growth rate with the value of 1.06 cm while CO 945 variety had the least value of 0.64 cm. GA 4+7 and 6-BA levels also revealed significant change in stem growth rate with control having the highest change of 0.98 cm while 4 litres per hectare had the least growth rate recoding a value of 0.60 cm (Table 4.4). Table 4.4: Growth rate of diameter as affected by variety and GA 4+7 and 6-BA Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. The results of this study agree with those of Ashraf et al. (2002) who reported an increase in the overall plant growth parameters upon application of gibberellic acid and cytokinins. Growth rate of stem diameter Varieties Season 1 Season 2 V1 1.06a 0.84a V2 0.57d 0.88a V3 0.93b 0.82a V4 0.64d 1.49a V5 0.77c 0.66a LSD 0.12 0.82 GA T1 0.98a 0.83a T2 0.88ab 0.77a T3 0.81bc 0.82a T4 0.69cd 0.84a T5 0.60d 1.44a LSD 0.15 0.83 53 Patel and Chaudhary (2003) also observed an increase in stem diameter of sugarcane varieties upon application of growth promoting hormones including gibberellic acid which is also in agreement with the current findings. In another study done by Yadav et al. (2016) they recommended use of gibberellic acid in adequate amount to enhance expansion of stems as one of the strategies to maximize yield of sugarcane varieties that can be also derived from this study since the results are in agreement with present study findings. Gibberellic acid has also been associated with growth stimulation during the vegetative phase hence one of the reasons for increment in stem diameter as it was revealed from results (Gad et al., 2016). 4.4 Plant height The results revealed significant differences (P≤0.05) in plant height in season 1 while no significant differences were observed in season 2. CO 945 variety was superior in month 3, month 4, month 5 and month 6 recording 42.23 cm, 56.68cm, 74.02cm and 93.57cm respectively (table 4.5) while D8484 variety was the lowest. The differences in height could be as result of genotypic differences in which some varieties respond differently to plant growth hormones. 54 Table 4.5: Plant height in centimeters as affected by sugarcane varieties and GA 4+7 and 6-BA Site Season1 Season2 Variety MAP3 MAP4 MAP5 MAP6 MAP3 MAP4 MAP5 MAP6 V1 36.52b 51.50b 64.66b 79.77b 39.79a 52.90a 68.83a 84.5a V2 42.22a 56.94a 72.94a 93.05a 36.31a 50.98a 66.45a 80.8a V3 34.56b 47.67c 63.84b 74.12c 37.39a 51.84a 67.38a 83.0a V4 42.23a 56.68a 74.02a 93.57a 36.94 a 51.22a 67.05a 81.4a V5 35.47b 49.70bc 65.52b 76.19bc 38.07a 42.40a 68.07a 82.7a LSD 4.41 3.39 3.22 4.47 5.00 4.40 4.79 7.67 GA T1 29.75d 45.98d 62.49d 74.80d 36.19a 50.63a 66.15a 80.2a T2 35.27c 50.02c 65.13cd 79.19cd 37.07a 50.74a 66.97a 81.6a T3 38.59b 53.22b 68.14bc 83.92bc 39.01a 53.27a 68.49a 85.0a T4 42.32a 55.06b 71.38ab 88.31ab 37.41a 51.72a 67.45a 82.4a T5 45.07a 58.21a 73.85a 90.49a 38.81a 52.99a 68.71a 83.1a LSD 3.01 3.04 3.45 6.42 5.017 4.35 4.76 7.64 VXT NS NS NS NS NS NS NS NS Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. Gibberellic acids 4 and 7 and 6-benzyl adenine levels also exhibited statistical differences (P≤0.05) with 4 litres per hectare being superior in plant height enhancement with 45.07 cm, 58.21 cm, 73.85cm and 90.49 cm in the 3rd, 4th, 5th, and 6th month respectively. Control had the least influence on plant height with the lowest value being 29.75 cm in month 3. There were no significant differences observed in the interaction effects of sugarcane varieties and gibberellic acid levels on plant height. There was a significant change in growth rate in regard to plant height between month 3 and month 6 in season 1 while no change was observed in season 2. 55 CO 945 variety recorded the greatest change in plant height with a value of 51.34 cm while EAK 73-335 variety had the least growth change of 40.72 cm as shown in table 4.6. Gibberellic acid 4 and 7 and 6-benzyl adenine levels did not show significance differences in plant height growth rate in both season 1 and 2. Table 4.6: Growth rate of height as affected by variety and GA 4+7 and 6-BA Growth rate of plant height Varieties Season1 Season2 V1 43.25b 44.73a V2 50.83a 44.50a V3 39.56c 45.56a V4 51.34a 44.44a V5 40.72c 44.62a LSD 1.52 4.02 GA T1 45.95a 44.06a T2 43.92a 44.50a T3 45.99a 45.98a T4 45.99a 44.98a T5 45.42a 44.32a LSD 4.09 4.00 Means followed by the same letter within the same column are not significantly different (P≤0.05)V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Prolamin per Ha. The results of this study are in agreement with those of Ma et al. (2018) who reported significant increase in plant height of perennial grass when Gibberellic acid was 56 introduced in early stages of growth. Similar results were also reported in maize by Voorend et al. (2016) where different gibberellic acid treatments resulted in increment of plant height as it was the case for this study. Similarly, in the study of Yadav et al. (2016) there was an increase in cane height when gibberellic acid was applied at different levels. 4.5 Internode length The internode length was statistically influenced (P≤0.05) by varieties and gibberellic acids 4 and 7 and 6-benzyl adenine levels in season 1, while in season 2, no significant differences were observed. KEN 83-737 variety and CO 945 variety had the highest influence on the internode length of the sugarcane variety with the longest being recorded in month 6 with 8.91 cm while the lowest internode length was observed in month 3 with 4.07 cm as illustrated in table 4.7. The increase in internode length is due to cell elongation and extension that result to longer internode development in cane varieties. 57 Table 4.7: Internode length in centimeters as affected by variety and GA4+7 and 6-BA Site Season1 Season2 Variety MAP3 MAP4 MAP5 MAP6 MAP3 MAP4 MAP5 MAP6 V1 4.42b 5.19b 6.35b 7.57b 4.78a 6.01a 7.20a 8.28a V2 5.02a 6.62a 7.87a 8.91a 4.39a 5.50a 6.75a 7.71a V3 4.07b 5.38b 6.31b 6.94c 4.51a 5.70a 6.89a 7.95a V4 5.15a 6.74a 8.04a 8.91a 4.33a 5.69a 6.87a 7.72a V5 4.41b 5.41b 6.71b 7.53b 4.78a 5.97a 7.11a 8.08a LSD 0.53 0.56 0.54 0.57 0.59 0.76 0.78 0.82 GA T1 3.65e 4.80d 6.06c 7.00d 4.35a 5.5a 6.72a 7.63a T2 4.12d 5.40c 6.58bc 7.47cd 4.48a 5.65a 6.88a 7.83a T3 4.63c 5.91bc 7.00b 7.91bc 4.81a 5.94a 7.13a 8.32a T4 5.15b 6.39ab 7.62a 8.50ab 4.47a 5.82a 7.00a 7.94a T5 5.53a 6.84a 8.01a 8.97a 4.66a 5.95a 7.09a 8.01a LSD 0.33 0.52 0.58 0.64 0.60 0.76 0.78 0.81 VXT NS NS NS NS NS NS NS NS Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. On the effect of gibberellic acids 4 and 7 and 6-benzyl adenine levels, 4 litres per hectare showed superiority in influencing the internode length with the highest being 8.50 cm in month 6. On the other hand, the control was less superior in all the sampling sites with the minimal being recorded in month 3 with a value of 3.65cm (Table 4.7). However, internode length did not show any interaction effects as an influence of sugarcane varieties and gibberellic acids 4 and 7 and 6-benzyl adenine. Sugarcane varieties internode length had a significant change in growth rate between month 3 and month 6 in season 1 while in season 2 there was notable change. KEN 83-737 variety was superior in season 1 with a change of 3.89 cm while D8484 variety had the least 58 change of 2.87 cm. GA 4+7 and 6-BA levels did not show any significant change in growth rate of internode length in the two study sites. Table 4.8: Growth rate of internodes as affected by variety and GA4+7 and 6BA Growth rate of internode length Varieties Season1 Season2 V1 3.15b 3.49a V2 3.89a 3.32a V3 2.87c 3.44a V4 3.76a 3.39a V5 3.11b 3.30a LSD 0.14 0.33 GA T1 3.36a 3.28a T2 3.34a 3.34a T3 3.28a 3.51a T4 3.38a 3.46a T5 3.45a 3.35a LSD 0.33 0.33 Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. Similar results were reported by Sujatha et al. (2018) that a gibberellic acid remarkably increased the internode length as it was the case for this study. In another study by Roopendra et al. (2018), it was reported that gibberellic acid and cytokinins can be used to increase the sucrose harvested from cane due to enhanced internode length which also conforms to the current results. In addition, internode length is an indicator of the potential of the cane varieties to accumulate more sugar, therefore with longer internodes as a result of gibberellic acid application the greater the yield (Chandra, 2018). Therefore, 59 the current findings confirm that with gibberellic acid application in cane varieties more yield in tons per hectare and sucrose accumulation is anticipated. 4.6 Number of leaves The findings revealed a significant difference (P≤0.05) in the number of leaves as influenced by the sugarcane varieties and gibberellic acids 4 and 7 and 6-benzyl adenine levels in season 1 while no differences were observed in the season 2. KEN 83-737 variety was superior in accumulation of the number of leaves in the respective plants with a value of 11.28 during month 6. The lowest value was recorded in Variety 3 (V3) at month 3 which recorded 4.74 as demonstrated in table 4.9. Table 4.9: Number of leaves as affected by variety and GA4+7 and 6-BA levels Site Season1 Season2 Variety MAP3 MAP4 MAP5 MAP6 MAP3 MAP4 MAP5 MAP6 V1 5.04bc 6.29c 8.18b 9.77b 5.86a 6.92a 8.47a 10.15a V2 6.63a 7.84a 9.41a 11.28a 5.42a 6.61a 8.14a 9.64a V3 4.74c 5.65d 6.75c 8.02c 5.66a 6.81a 8.37a 9.90a 4 6.62a 7.84a 9.32a 11.19a 5.42a 6.52a 7.92a 9.40a V5 5.45b 6.93b 8.30b 9.35b 5.97a 7.28a 8.69a 10.17a LSD 0.43 0.53 0.57 0.81 0.74 0.81 0.900 1.21 GA T1 4.93d 5.94d 7.29d 8.19d 5.41a 6.53a 8.05a 9.40a T2 5.26cd 6.48cd 7.86cd 9.47c 5.54a 6.79a 8.15a 9.62a T3 5.75bc 6.82bc 8.44bc 10.04bc 5.98a 7.07a 8.65a 10.34a T4 6.13ab 7.43ab 8.98ab 10.75ab 5.70a 6.88a 8.37a 9.96a T5 6.40a 7.86a 9.32a 11.18a 5.70a 6.88a 8.38a 9.93a LSD 0.62 0.67 0.75 0.94 0.75 0.85 0.91 1.21 VXT NS NS NS NS NS NS NS NS Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. 60 The gibberellic acids 4 and 7 and 6-benzyl adenine levels also revealed significant differences (P≤0.05) in season 1 while no significant differences were observed in season 2. The highest value was recorded in gibberellic acids 4 and 7 and 6-benzyl adenine during month 6 that obtained a value of 11.18. Control had the lowest concentration during month 6 with 8.19 leaves. Sugarcane varieties did not have any significant difference on their interaction in influencing number of leaves with application of gibberellic acids at different levels. Season 1 also exhibited significant change (P≤0.05) in growth rate in regard to the number of leaves between month 3 and month 6 while season 2 did not show any significant change in growth rate. CO 421 variety showed the greatest change in growth with a difference of 4.74 number of leaves while D8484 variety had the lowest growth rate change with 3.29 in season 1 as shown in table 4.10. On the other hand, gibberellic acids 4 and 7 and 6-benzyl adenine at 4 litres per hectare was superior in influencing number of leaves growth with 4.77 while control had the least change in number of leaves with a value of 3.26. 61 Table 4.10: Growth rate of leaves as affected by variety and GA 4+7 and 6-BA Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. The results of number of leaves increment in the sugarcane varieties and promalin application agrees with those of Miceli et al. (2019) who reported application of gibberellic acid increased leaf formation and expansion. In another study by Batlang et al. (2006) it was reported that increase in leaf size and formation is due to cell division and elongation as major role of gibberellins that can be compared to the similarity of the current findings. Growth rate of number of leaves Varieties Season 1 Season 2 V1 4.74a 4.29a V2 4.56a 4.22a V3 3.29c 4.25a V4 4.57a 3.98a V5 3.89b 4.19a LSD 0.45 0.63 GA T1 3.26c 3.99a T2 4.20b 4.07a T3 4.29b 4.35a T4 4.62ab 4.26a T5 4.77a 4.23a LSD 0.47 0.63 62 According to Roy et al. (2017) increasing number of leaves per plant under the treatment of gibberellic acid may be due to improvement of the physiological efficiency of the plant such as improvement of rate of photosynthesis, control of transpiration and photorespiration, efficient water and nutrient uptake and control of leaf senescence. These findings also agree with those of Al-Rawi et al. (2016) who reported an increase in the number of leaves in peach trees when sprayed with gibberellic acid. 4.7 Yield Yield was significantly influenced (P≤0.05) sugarcane varieties and GA 4+7 and 6-BA levels in both season 1 and 2. D8484 variety recorded the highest yield in the two seasons with 70.49 tons per hectare and 69.24 tons per hectare in season 1 and 2 respectively. The lowest yield was realized in CO 421 variety with 64.07 tons per hectare in season 1 and 63.78 tons per hectare in season 2. In GA 4+7 and 6-BA levels, 4 litres per hectare had the greatest influence of yield in season 1 with 69.14 tons per hectare and 69.02 tons per hectare in season 2 as shown in table 4.12. The high yield recorded could be due to influence of the gibberellic acids 4 and 7 to enhance tillering, height and girth in specific varieties hence more is harvested in a given area. 63 Table 4.11: Yield as affected by variety and GA 4+7 and 6-BA Yield t/ha Varieties Season1 Season2 V1 64.07b 63.78b V2 66.47b 66.06b V3 70.49a 69.24a V4 64.38b 64.24b V5 64.13b 64.04b LSD 2.60 2.86 GA T1 60.37d 59.68d T2 64.39c 63.53c T3 66.73b 66.33b T4 68.92a 68.79a T5 69.14a 69.02a LSD 2.002 1.89 VXT * * Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. D8484 variety applied with gibberellic acids 4 and 7 and 6-benzyl adenine at 3 litres per hectare had the greatest yield interaction effect with 75.04 tons per hectare in season 1 and 74.81 tons per hectare in season 2 as shown in table 4.13. CO 421 Variety applied with gibberellic acids 4 and 7 and 6-benzyl adenine at 0 litres per hectare had the least interaction effects on yield with 58.6 tons per hectare and 57.87 tons per hectare in chegulo in season 1 and 2 respectively. 64 Table 4.12: Yield due to interaction effect between varieties and GA 4+7 and 6-BA Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. The findings of this study agree with those of Rastogi et al. (2013) who reported to increase in overall yield when gibberellic acid was applied at initial stages of crop growth. Treatment Yield T/ha Treatments season 1 season 2 V1T1 58.6k 57.87i V1 T2 62.5i 62.23g V1 T3 64.93g 64.82ef V1 T4 66.94ef 66.86c V1 T5 67.4de 67.13c V2 T1 60.8j 60.42h V2 T2 65.04g 64.43ef V2 T3 67.17e 66.32cd V2 T4 69.52c 69.44b V2 T5 69.83c 69.68b V3 T1 62.79hi 61.77g V3 T2 68.02d 64.16f V3 T3 71.26b 70.25b V3 T4 75.04a 74.81a V3 T5 75.35a 75.23a V4 T1 60.3j 59.76h V4 T2 63.08hi 62.81g V4 T3 64.97g 65.35de V4 T4 66.7ef 66.59c V4 T5 66.85ef 66.71c V5 T1 59.34k 58.64i V5 T2 63.31h 64f V5 T3 65.32g 64.93ef V5 T4 66.4f 66.24cd V5 T5 66.28f 66.36c LSD 0.76 1.05 65 Bora and Sarma, (2006) reported an increase in yield of pea as a result of gibberellic acid application that promote vegetative growth, nutrient uptake and eventually recorded high grain harvest that conforms to the findings of the current study. In another study done by Li and Solomon, (2003) revealed that spraying canes with gibberellic acid increases formation of tillers and consequent lead to high yield at the end of the season which agrees with the current study findings. Sujatha et al. (2018) also reported similar findings of increase in gibberellic level resulting to high yield in cane varieties. Therefore, it is clear that gibberellic acid 4 and 7 and 6-benzyl adenine has significant influence on sugarcane growth parameters and can be applied in farmers’ fields for yield increment. 4.8 Pol% Sugarcane varieties had a significant difference (P≤0.05) in pol% in both season 1 and 2. EAK 73-335 variety recorded the highest superiority in pol% accumulation with 14.70% and 14.69 % in season 1 and 2 respectively as illustrated in table 4.13. KEN 83-737 variety had the least pol% of 10.42% in season 1 and 10.39% in season 2. Gibberellic acids 4 and 7 and 6-benzyl adenine levels did not exhibit any significant differences in the two experimental sites. However, pol% increased with increase in gibberellic acids 4 and 7 and 6-benzyl adenine levels. The higher level of pol% could be a sign of high quality in the specific variety as it depicts the sugarcane value. 66 Table 4.13: Pol % as affected by variety and GA 4+7 and 6-BA Pol% Pol% Varieties Season1 Season2 V1 12.56c 12.52c V2 10.42e 10.39e V3 14.37b 14.34b V4 12.02d 11.99d V5 14.70a 14.69a +LSD 0.21 0.23 GA T1 12.65a 12.60a T2 12.69a 12.68a T3 12.81a 12.80a T4 12.92a 12.88a T5 12.99a 12.96a LSD 2.32 2.34 Means followed by the same letter within the same column are not significantly different (P≤0.05); V1-V5 denote variety CO 421, KEN 83-737, D8484, CO 945 and EAK 73-335 respectively while T1-T5 denote 0,1,2,3,4 Litres of Promalin per Ha. The findings of this study agree with those of Wagih et al. (2004) who reported that sugarcane varieties have different levels of sugar content accumulation (pol%) depending on their maturity levels. According to Roopendra et al. (2018) application of gibberellic acid resulted in significant increase of pol% which agrees with the current study findings. In another experiment done by Verma et al. (2017) gibberellic acid application influenced increase in sucrose levels in sugarcane which showed high quality of the harvested cane which also conforms to the results observed in this study. 67 4.9 Profitability of use of GA 4+7 and 6-BA at 3 L/Ha All varieties under study recorded incremental yield from 6.62 to 11.65 tons per hectare. D8484 was the most superior variety followed by KEN 83-737, CO 421, EAK 73-335 and CO 945. There was net positive ratio with all varieties. D8484 was the best overall having extra income of Kshs. 34,950 before discounting and Kshs. 30,294 after discounting with benefit-cost ratio of 3.7. The projected revenue for each variety was discounted to net present value. Use of gibberellins 4 and 7 and 6-benzyl adenine was found to be profitable and economically efficient technology as it increased yields at reasonable cost. Table 4.14: Benefit-Cost Ratio Variety Discounted benefit in Kshs. Cost in Kshs. Benefit Cost ratio Economically viable? Decision CO 421 (V1) 22,519 6,560 3.4 Yes Accept KEN 83-737 (V2) 23,065 6,560 3.5 Yes Accept D8484 (V3) 30,294 6,560 3.7 Yes Accept C0 945 (V4) 17,214 6,560 2.5 Yes Accept EAK 73-335 (V5) 19,061 6,560 2.9 Yes Accept 68 Figure 4.3: Benefit-Cost Ratio across varieties Neupane et al. (2017) observed that Benefit-Cost Ratio gave an idea about the recovery of cost incurred during production by the returns from products. Niels & Van Dijk (2000) reported that Cost Benefit Analysis provided the economic data to help produce better investment decisions that were debated between decision-makers and the general public which confirms the need to undertake this analysis. Serpell (2004) concurred that indeed Cost Benefit Analysis furnished decision-makers with helpful information to aid in making effective investment decisions and that indeed sugarcane profitability was domiciled in its productivity which concurs with the results of this study. CO 421 KEN 83-737 D8484 C0 945 EAK 73-335 69 CHAPTER FIVE: CONCLUSIONS AND RECOMMENDATIONS 5.1 Conclusions Growth of sugarcane varieties was significantly increased by gibberellic acid 4 and 7 and 6-benzyl adenine levels. Use of increased gibberellic acid 4 and 7 and 6-benzyl adenine levels induced germination, tillering, height, stem diameter, length of internode and leaf number. CO945 and D8484 varieties recorded the highest performance in most growth parameters under study. There was significant difference in yield and sucrose content as a result of gibberellic acids 4 and 7 and 6 benzyl adenine application. Increase in yield was between 11% and 22% and sucrose content (pol %) between 3% and 7% when applied at 4 litres per hectare. CO 421 had its yield increased by 15% in season one and 16% in season two; yield for KEN 83-737 increased by 15% in both seasons while D8484 increased by 20% and 22% in seasons one and two respectively. CO 945 recorded yield increase of 11% in season one and 12% in season two. EAK 73-335 had yield increase of 12% in season one and 13% in season two. All the varieties recorded positive Benefit- Cost Ratio with D8484 being the highest at 3.7. D8484 grown with gibberellic acid 4 and 7 and 6-benzyl adenine at 4 litres per hectare showed the best performance at 75.35 and 75.23 tons/ha in Seasons 1 and 2 respectively. EAK 73-335 recorded superiority in sucrose accumulation with pol% of 14.70% and 14.69 % in seasons 1 and 2 respectively when treated with gibberellic acid 4 and 7 and 6- benzyl adenine at 4 litres per hectare. Although there was linear positive increase in yield and pol% at 4 litres per hectare, there was no significant difference in yield and pol% at 4 litres per hectare of gibberellic acids 4 and 7 and 6-benzyl adenine application. 70 5.2 Recommendations i. Gibberellic acid 4 and 7 and 6-benzyl adenine should be used to break dormancy in seedcane and induce germination and overall growth. ii. Gibberellic acid 4 and 7 and 6-benzyl adenine should be recommended for use among farmers to increase sugarcane yields, sucrose content and profitability. iii. There is need to sensitize farmers in Kakamega County and areas with similar agro ecological conditions to grow variety D8484 and EAK 73-335 with gibberellic acid 4 and 7 and 6-benzyl adenine at 3 litres per hectare for optimum yields. iv. Additional study is recommended on evaluation of growth, yield and sucrose content of more cane varieties, applied with gibberellic acid 4 and 7 and 6-benzyl adenine, in other agro ecologies. v. Further research is recommended on the assessment of yield and sucrose of subsequent ratoon crop previously applied with gibberellic acid 4 and 7 and 6- benzyl adenine. 71 REFERENCES Al-Rawi, W. A. A., Al-Hadethi, M. E. A., and Abdul-Kareem, A. A. (2016). Effect of foliar application of gibberellic acid and seaweed extract spray on growth and leaf mineral content on peach trees. The Iraqi Journal of Agricultural Science, 47(7- special issue), 98-105. Amanulla, M.M., Seker S. and Vincent S. (2010). 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Cytokinins: These are hormones which act in cell division, cell enlargement, senescence and transport of amino acids in plants. Ethylene: This is a naturally occurring regulator of plant growth and plant metabolic activity. It interacts with auxins to regulate many metabolic processes. Gibberellins: These are hormones which control cell elongation and division in plant shoots through stimulation of ribonucleic acid and protein synthesis in plant cells. Plant growth regulators (PGRs): These are organic compounds, other than nutrients, that modify plant physiological processes. Pol factor: This is the percentage ratio of total pol in the mixed juice and the final bagasse to the total pol in cane. 89 Pol: This is the sucrose content when expressed as a percentage. This measurement is called pol because it is determined by polarization method. Polysaccharide: This is a complex carbohydrate molecule where many saccharide molecules are bonded together. Purity: This is the ratio of pol to brix. Essentially it describes how much pure sucrose is present in a sugar sample. 90 Appendix 2: List of marketed plant growth regulators Active ingredient/Formulation/Concentrate Product Name Company Name Crops AUXINS 2-(1-naphthyl) acetic acid SL 45 g/l Planofix Bayer Apple Pear Pineapple 4-indol-3-ylbutyric acid DP 1 g/kg Seradix B No 1 Bayer Ornamentals 4-indol-3-ylbutyric acid DP 3 g/kg Seradix B No 2 Bayer Ornamentals 4-indol-3-ylbutyric acid SL 8 g/kg Seradix B No 3 Bayer Ornamentals GIBBERELLINS Gibberellins SL 32 g/l ProGibb 4% Valent BioSciences Grape, Pear, Citrus, Potato, Mango, Hops GA4+7 and 6-benzyl adenine SL 19/19 g/l Promalin Valent BioSciences Apple, Plum, Flowers, Sugarcane ETHYLENE Ethephon SL 480 g/l Ethrel Bayer Apple, Citrus, Cotton, Grape, Maize,Pineapple, Sugarcane GROWTH RETARDANTS Paclobutrazol SC 250 g/l Cultar Syngenta Litchi Mango Peach Plum Daminozide SP 850 g/kg B-Nine SP Crompton Chemical Flowers Ornamentals Glyphosate-isopropylamine SL 360 g/l Glyphosate 360 Acid Monsanto Sugarcane, Grasses Glyphosate-isopropylamine SL 360 g/l Mamba 360 SL Dow AgroSciences Sugarcane, Grasses Glyphosate-isopropylamine SL 360 g/l Roundup Monsanto Grasses, Sugarcane Glyphosate-isopropylamine SL 360 g/l Roundup Monsanto Sugarcane, Grasses GROWTH INHIBITORS Mepiquat chloride SL 50 g/l Pix BASF Cotton Chlormequat chloride SL 750 g/l CeCeCe 750 BASF Pear, Wheat Chlormequat chloride/ethephon SL 300/150 g/l Uprite Bayer Wheat DEFOLIANTS Thidiazuron/diuron SC 120/60 g/l Dropp Ultra Bayer Cotton GROWTH STIMULATORS Brassinolide SL 0.1% Double Godrej Agril. and Hortil. Crops 91 Appendix 3: Salient features of 5 sugarcane varieties grown in Kakamega County Label Variety Pol% TCH Maturity Diseases Desirable Characteristic Undesirable Characteristic V1 CO 421 13.0 104 20-24 Months Smut Drought toleran Retains sugar for a long time Susceptible to smut, Late maturing V2 KEN 83-737 11.6 118 16-19 Months Smut Early maturing, Heavy tillering, Drought tolerant Flowers profusely V3 D8484 15.5 126 14-16 Months Mosaic Early maturing, High sucrose, High yielding, Resistant to smut Poor tillering, Susceptible to drought V4 CO 945 13.1 120 18-20 Months smut Heavy tillering, Good ratooning Poor germinator V5 EAK 73-335 16.0 123 18-20 Months Mosaic High sucrose, High yielding Susceptible to drought Source: Sugar Research Institute handbook, 2018 92 Appendix 4: Cane establishment and management practices A- Land preparation; B- Setts preparation; C- Tiller counting; D- Display of D8484 variety; E- Display of CO 421 variety; F- Display of KEN 83-737 variety; G- Routine field inspection; H- Display of stem diameter in D8484; I- Display of EAK-73-335; J- Sugarcane ready for harvesting; K- Weighing of sugarcane