ABSTRACT
Powdery mildew (Podosphaera pannosa) is a major disease of roses which reduces yield and quality through growth of mycelia on different parts of the rose plant which affects their salability. The disease is wide spread where roses are grown both in greenhouse and field. This study was carried out to evaluate the effects of Bacillus spp and sodium nitrophenolate on powdery mildew of roses in greenhouse conditions. The trial was done in an already established bush of roses in the greenhouses at Kikuyu in Kiambu County and at Naivasha in Nakuru County both are in Kenya. The experiment was conducted on a variety known as A one which is grown in different farms in Kenya. Weekly foliar application of Real subtilis® (Bacillus subtilis) at the rate of 2ml/L, Hatake (Bacillus amyloliquefaciens) 3.0g/L, Atonik® (sodium nitrophenolate) 1ml/L and Meltatox® (Dodemorph acetate) 2.5ml/L. In the second experiment, foliar application of Bacillus amyloliquefaciens was done at different concentrations and at various intervals of application as follows 1.5g/L, 4 days, 3.0g/L, 4 days, 4.5g/L 4 days, 1.5g/L, 7 days, 3.0g/L, 7 days, 4.5g/L, 7 days, 1.5g/L, 10 days, 3.0g/L, 10 days, 4.5g/L, 10 days and no treatment as control. The experiment was laid out in a randomized complete block design with four replications for each treatment in plots with 44 plants. Data was collected on incidence and severity of powdery mildew on weekly basis while data on stem length, bud diameter, bud length and marketable grade were collected daily for twenty weeks and six weeks for the first and the second experiments respectively. In the first experiment, Bacillus subtilis, B. amyloliquefaciens, sodium nitrophenolate and dodemorph acetate significantly reduced incidence of powdery mildew from 83% to 55% with dodemorph acetate posting the best results followed by B. amyloliquefaciens, sodium nitrophenolate and Bacillus subtilis. In the second experiment, at different rates and at various intervals Bacillus amyloliquefaciens reduced incidence of powdery mildew with the rate of 1.5g/L at four and seven day intervals gave better results than ten days interval. Disease severity was reduced significantly from 15.8% to 2.1% by application of dodemorph acetate and B. amyloliquefaciens while application of B. subtilis and sodium nitrophenolate did not significant ly at P ≤0.05 reduce severity of powdery mildew. Applications at the rate of 1.5g/L and 3.0g/L at four days and weekly applications had the highest reduction in severity in the second experiment. Dodemorph acetate had the highest area under disease progress curve (932) followed by Bacillus amyloliquefaciens (988) in the first experiment while application of Bacillus amyloliquefaciens at the rate of 1.5g/L at the interval of seven days showed the highest AUDPC (799.8). Application of the test products did not significantly affect the yield and quality paramenters in the first experiment but in the second experiment, foliar application of Bacillus amyloloquefaciens at different rates and at various intervals improved the quality. Marketable stems significant ly increased in the second experiment after the application of Bacillus amyloliquefaciens at different rates and at various intervals. Foliar application of Bacillus Spp and sodium nitrophenolate controlled powdery mildew of roses. Application of the same products did not have significant effects on quality and yield parameters of flowers such as flower bud length, stem length and bud diameter and number of stems produced in the first experiment, however, there was improvement in yield and quality in the second experiment. Growers of roses should be encouraged to apply Bacillus spp and sodium nitrophenolate in managing powdery mildew. The optimal concentration of Bacillus amyloliquefaciens should be established and how sodium nitrophenolate induce resistance to plants against phytopathogens should be investigated.
Key words: Rosa hybrida, Podosphaera pannosa, Bacillus subtilis, Bacillus amyloliquefaciens, sodium nitrophenolate.
TABLE OF CONTENTS
DECLARATION ii
DECLARATION OF ORIGINALITY iii
DEDICATION iv
ACKNOWLEDGEMENT v
ABBREVIATIONS vi
TABLE OF CONTENTS vii
LIST OF TABLES x
LIST OF FIGURES xi
GENERAL ABSTRACT xii
CHAPTER ONE: INTRODUCTION
1.1 Background information 1
1.2 Problem statement 2
1.3 Justification 4
1.4 Objectives of the study 4
1.5 Hypothesis 5
CHAPTER TWO: LITERATURE REVIEW
2.1 Taxonomy and morphology of rose plant 6
2.2 Growing requirements and establishment 7
2.3 Constraints to production of roses 8
2.4 Powdery mildew of roses 10
2.4.1 Host range and distribution 10
2.4.2 Causal agent of rose powdery mildew 11
2.4.3 Symptoms of powdery mildew 12
2.4.4 Epidemiology of powdery mildew of roses 14
2.5 Management of powdery mildew 15
2.5.1 Management of plant diseases with antagonistic organisms 19
2.5.2 Management of plant diseases using Bacillus spp 20
2.6 Management of plant diseases using nitrophenolates 22
CHAPTER THREE
MANAGEMENT OF POWDERY MILDEW OF ROSES USING BACILLUS SUBTILIS, BACILLUS AMYLOLIQUEFACIENS AND SODIUM NITROPHENOLATE
3.1 Abstract 23
3.2 Introduction 25
3.3 Materials and methods 26
3.3.1 Description of the experimental site and the crop 26
3.3.2 Experimental design and layout 27
3.3.3 Application of treatments 28
3.3.4 Assessment of incidence and severity of powdery mildew 28
3.4 Data analysis 29
3.5 Results 30
3.5.1 Efficacy of Bacillus subtilis, B. amyloliquefaciens and sodium nitrophenolate on incidence and severity of powdery mildew 30
3.5.2 Frequency and rate of application of Bacillus amyloliquefaciens on incidence and severity of powdery mildew 32
3.4 Discussion 37
3.5 Conclusion 40
CHAPTER FOUR: 41
EFFECT OF BACILLUS SUBTILIS, BACILLUS AMYLOLIQUEFACIENS AND SODIUM NITROPHENOLATE ON QUALITY OF ROSE FLOWERS
4.1 Abstract 41
4.2 Introduction 42
4.3 Material and methods 43
4.3.1 Description of the experimental site and the crop 43
4.3.2 Experimental design and layout 43
4.3 Data collection 43
4.3 Data analysis 44
4.5 Results 44
4.5.1 Efficacy of Bacillus spp. and sodium nitrophenolate on quality and marketable grade.44
4.5.2 Effect of rate and interval of application of Bacillus amyloliquefaciens on quality and marketable grade of harvested stems 46
4.6 Discussion 53
4.7 Conclusion 57
CHAPTER FIVE
GENERAL DISCUSSION, CONCLUSIONS AND RECOMMENDATIONS
5.1 General discussion 58
5.2 Conclusion 60
5.3 Recommendations 61
REFERENCES 62
LIST OF TABLES
Table 3. 1: Incidence of powdery mildew of roses after treatments application (season I) 31
Table 3. 2: Incidence of powdery mildew of roses at different rates and intervals of application of Bacillus amyloliquefaciens 32
Table 3. 3: Incidence of powdery mildew of roses after application of different concentrations of Bacillus amyloliquefaciens 33
Table 3. 4: Severity of powdery mildew expressed as percentage after treatments application (season I) 31
Table 3. 5: Severity of powdery mildew of roses at different rates and intervals of application of Bacillus amyloliquefaciens 34
Table 3. 6: Severity of powdery mildew of roses after application of different concentrations of Bacillus amyloliquefaciens 35
Table 3. 7: Effect of various treatments on mean incidence and area under disease progress curve (AUDPC) (season I) 32
Table 3. 8: Area under disease progress curve of powdery mildew of roses at different rates and intervals of application of Bacillus amyloliquefaciens (Season II) 36
Table 4. 1: Stem length of roses after application of Bacillus subtilis, B. amyloliquefaciens and sodium nitrophenolate 45
Table 4. 2: Marketable grade of harvested rose stems at different rates and interval of application of Bacillus amyloliquefaciens 52
Table 4. 3: Marketable grade of roses after application of different concentrations of Bacillus amyloliquefaciens 53
LIST OF FIGURES
Figure 2. 1: Life cycle of Podosphaera pannosa causing powdery mildews on roses 12
Figure 3. 1: P. M after Bacillus amyloliquefaciens (3.0g/L, 4days application 36
Figure 3. 2: P. M before Bacillus amyloliquefaciens application 36
Figure 3. 4: P.M 7 weeks after Bacillus amyloliquefaciens (1.5g/L) application 37
Figure 3. 3: P. M before Bacillus amyloliquefaciens application 37
ABBREVIATIONS
AFA Agricultural Food Authority
AIPH International Association of Horticultural Producers
ANOVA Analysis of Variance
AUDPC Area Under Disease Progress Curve
ASL Above Sea Level
CO2 Carbon IV oxide
CV Coeficient of Variation
EC Electrical Conductivity
EU European Union
GA Gibberellic Acid
IAA Indole Acetic Acid
ICM Integrated Crop Management
ISR Induced Systemic Resistance
JETRO Japan External Trade Organization
KEPHIS Kenya Plant Health Inspectorate Service
KFC Kenya Flower Council
LSD Least Significant Difference
PDS Plant Disease Severity
PGPR Plant Growth Promoting Rhizobacteria
pH Potential of Hydrogen ions
UV-B Ultra Violet – B
USDA United States Department of Agriculture
VOC Volatile Organic Compounds
CHAPTER ONE
INTRODUCTION
1.1 Background information
Roses (Rosa Spp.) are one of the most popular garden plants and the most economically important ornamental crop traded as cut flowers worldwide (Wen et al., 2006; Debener and Linde, 2009; Leus et al., 2018; United States Department of Agriculture-USDA, 2020). About 70% of the trade is done in the European Union markets. Large scale production is reported in countries such as Ecuador, Kenya and Colombia (Blom and Tsujita, 2003; International Association of Horticultural Producers-AIPH, 2019). Roses are grown for cut flower markets which are largely for its aesthetic value (Farooq and Kama, 2020; Tiwari et al., 2020). It is estimated that annual production for cut flowers ranges between 18 trillion stems, potted rose plants to be 80 million and garden roses to be 220 million (Pemberton et al., 2003; Agricultural and Food Authority-AFA, 2019).
In Kenya, roses is one of the major cut flowers grown and exported. Kenya is the leader in export of rose cut flower to the European Union (EU) with a market share of about 38% (Kenya trade.org, 2022). About half of the total export goes through the Dutch auction, although direct sales are also available. In some countries like United Kingdom, supermarkets are the main outlets for rose flowers (Kenya Flower Council, 2020). More markets are also coming up and they include Russia, Germany and Asian continent. About a quarter of the produced flowers are delivered directly to these markets which give opportunity for value addition through grading, sleeving, labelling and bouquet making (Adeola et al., 2018).
The main growing areas are Mt. Kenya, Nairobi, around Lake Naivasha, Kericho, Nakuru, Kitale, Athi River, Kiambu, Thika, Eastern Kenya, Uasin Gishu and Trans Nzoia. According to Kenya Flower Council, an estimate of 500,000 people including over 90,000 flower farms employees depend on floriculture industry which in turn have got impact on livelihoods of over two million people. Production level comprises of small, medium and large scale farming (KFC 2020).
Roses grown in the greenhouse are affected by many fungal diseases including powdery mildew caused by Podosphaera pannosa (Sphaerotheca pannosa) which can easily be seen on the leaves, stems and upcoming shoots, floral parts and buds. The disease is associated with grayish or whitish patches on the affected parts of the plant (Eken, 2005; Matysiak, 2021). Powdery mildew causes leaf chlorosis, leaf curling, premature leaf drops and in severe cases death of affected plants (Shetty et al., 2012; Scott, 2021), which leads to serious economic losses on productivity, quality and marketability of the produce (Suthaparan et al., 2010; Lima et al., 2019).
McGrath et al. (1996) pointed out that the most common method of disease control in roses is use of fungicides, however, their continued use may result in environmental pollution and development of resistant strains of the pathogen. Fungicides have also shown negative impacts on the beneficial microorganisms and insects, and this calls for softer alternatives to fungicides. Eken, (2005) and Khakimov et al., (2020) contends that in the recent past, bio-fungicides have been used in the management of powdery mildew disease.
1.2 Problem statement
Powdery mildew is one of the major diseases of roses grown in greenhouses. The greenhouse environment is ideal for the growth and development of the disease all year round. The high level of repetitive fungicide application needed to lower the powdery mildew pressure normally result to faster fungicides resistance (Daughtrey and Benson, 2005; Kumar and Chandel, 2018; Wanasiri et al., 2020). It is estimated that 40% of pesticide usage is directed in controlling powdery mildew in greenhouses which depicts high increased cost of production (Debener and Byrne, 2014;
Sambucci et al., 2019). Continuous application of manmade fungicides has given rise to the growth of resistance of some of the most crucial pesticide molecules such as dimethyl inhibitors and sterner restrictions on usage of others (McGrath, 2001; Gao et al., 2009; Ishii et al., 2021). Growers continue to incur losses on production and quality of roses which in turn lowers their income due to powdery mildew infections. It is estimated that 30%-42% production losses may occur when infection levels are high (Sudheendra, 2014; Linde and Shishkof, 2014). Annual monetary losses amounting to approximately Kenya shillings 74.5 billion have been reported (AFA, 2019). Other monitary losses results from labour charges, acquisition and application of fungicides to treat the disease. Millions of stems are lost due to powdery mildew infections and loss to biodiversi ty (Debener and Byrne, 2014; Ribes et al., 2018). There are also reports of losses due to rejection of flowers in the European Union markets as a result of non compliance to sanitary and phytosanitary standards (Pizano, 2019). Losses resulting from rejection due to excess residue on the products have been reported (Toumi et al., 2016). Several strategies which includes biological control agents, such as Tilletiopsis pallescens (Ken and Leslie, 1997; Amin et al., 2018; Tahir et al., 2018; Verma et al., 2020), anhydrous milk fat and soybean oil emulsion Chee et al., (2011, 2018); Kamel and Afifi, 2020; Wurms et al., 2021), synthetic fungicides (Scarito et al., 2007), sodium bicarbonate (Salamone et al., 2009; Shetty et al., 2021) have been used in an effort to manage powdery mildew in greenhouse rose crop in Kenya and globally. However, the pathogen is still causing losses due to resistance to current molecules and inability of rose growers to fully explore other methods of disease management. Therefore, there is need for use of safer alternatives for disease control with no negative effects to both the environment, human beings and non target organisms.
1.3 Justification
Because of the losses incurred by growers and the negative effects of fungicides on workers, environment and other animals, it is important to explore other means of management of greenhouse powdery mildew on roses such as biological control agents like Bacillus subtilis, Bacillus amyloliquafasciens and sodium nitrophenolate. These microbial antagonists do not harm animals, human beings and they also help in the conservation of the environment and increases consumer acceptability (Tjosvoldo and Koike, 2001; Kumar and Chandel, 2018; Abhiram et al., 2018). They are also easy to apply by the farmer as well as enhancing botanical progression and improve yield of the crop, have no residual effects, hydrolyses faster, have long term effects on the target organism and exhibit numerous modes of action (Whipps, 2001; Almoneafy et al., 2012; Abhiram et al., 2018; Campos et al., 2019). Several reports have been fronted by various researchers indicating that Bacillus spp and sodium nitrophenolate have been used in managing various diseases of plants (Abbas et al., 2019; Drobek et al., 2019; Saxena et al., 2019). An effective control will save rose growers from huge losses incurred in terms of money and labour and in return increase production level and income to workers. It is against this background that the evaluation of Bacillus subtilis, Bacillus amyloliquefaciens and sodium nitrophenolate is to be conducted to determine their effects on the control of the disease.
1.4 Objectives of the study
The broad objective was to improve the quality of roses through management of powdery mildew using Bacillus Spp and sodium nitrophenolate.
The specific objectives were
i. To evaluate potential of Bacillus subtilis, Bacillus amyloliquefaciens and sodium nitrophenolate in the management of powdery mildew on greenhouse roses.
ii. To evaluate the effect of Bacillus subtilis, Bacillus amyloliquefaciens and sodium nitrophenolate on quality of rose flowers.
1.5 Hypothesis
i. The antagonistic activities of Bacillus subtilis, Bacillus amyloliquefaciens and sodium nitrophenolate on powdery mildew of roses enhances management of the disease.
ii. The quality of rose flowers is improved through application of Bacillus subtilis, Bacillus amyloliquefaciens and sodium nitrophenolate due to enhanced production of growth hormones and improved tolerance to biotic and abiotic stress.
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