Maize (Zea mays L.) is ranked as the third most important food crop by production globally, after rice and wheat. Several biotic (diseases, pests) and abiotic (unfavorable climatic conditions) factors affects its production. Maize lethal necrosis (MLN) disease outbreak within East Africa threatens production of maize. Information on interactions of viruses causing MLN with plant parasitic nematodes is lacking. This study was carried out to determine i) the effect interaction of of plant parasitic nematodes with viruses causing MLN on disease development in maize fields and ii) the effect of lesion nematodes (Pratylenchus spp.) on MLN disease development in the greenhouse.
For the field study, four counties in Kenya were visited, farms selected at random, MLN scored and both maize leaf and soil samples collected and analyzed for presence of viruses causing MLN disease and for parasitic nematodes. Snowball sampling or chain-referral sampling technique was used to sample MLN infected farms across the selected regions. Variance analysis was used to measure significant differences (P< 0.05) in MLN disease incidence and severity due to interaction between viruses and nematode populations. In the greenhouse study, two maize varieties, were used H614D and Emph 1101. Variety H614D is known to be susceptible to both MLN and Pratylenchus spp. whereas maize variety Emph 1101 is susceptible to MLN but resistant to Pratylenchus nematodes. The two maize varieties were subjected to three distinct treatments: single inoculation with MCMV and SCMV; combined MCMV + SCMV inoculation; the third treatment was the addition of Pratylenchus nematodes to the previous two treatments. Disease severity and incidence were recorded weekly over a period of two months.
Survey results indicated no significant effect of combined infestation of parasitic nematodes (Pratylenchus spp., Tylenchus spp., Meloidogyne spp. and Helicotylenchus spp.,) on MLN disease severity in the field. However, there was significant effect of Pratylenchus to MLN severity in the greenhouse experiment. The development of MLN disease in maize varieties Emph 1101 and H614D infected with Pratylenchus spp. nematodes was studied under a greenhouse experiment. MLN disease severity was higher in H614D than in Emph 1101. Plants inoculated with MLN+Pratylenchus recorded a significant difference across the two varieties on area under disease progress curve (AUDPC). There is need for nematodes management even though the field experiment indicated no significant effect of parasitic nematodes on MLN disease developement. There is also need for an open field study to evaluate the effect of Pratylenchus spp. on the development of MLN disease.
TABLE OF CONTENTS
DECLARATION OF ORIGINALITY ii
TABLE OF CONTENTS v
LIST OF TABLES ix
ABREVIATIONS AND ACRONYMS xi
1.1 Global maize production 1
1.2 Maize production in Kenya 2
1.3. Maize production constraints 3
1.4 Statement of the problem 5
1.5 Justification 5
1.6 Objectives 6
1.6.1 Broad objective 6
1.6.2 Specific objectives 6
1.7 Hypotheses 7
2.1 History of maize cultivation 8
2.1.1 Maize production in Kenya 8
2.2 Maize lethal necrosis disease 9
2.2.1 Etiology of Maize lethal necrotic disease 9
2.2.2 Host-range for MLN viruses 10
2.3 Maize chlorotic mottle virus 10
2.3.1. Transmission of MCMV 11
2.3.2 Maize chlorotic mottle disease symptoms 12
2.3.3. Impact of MCMVon crop yield 12
2.4. Distribution and transmission of SCMV 13
2.5. Management of Maize lethal necrotic disease 13
2.6 Impact of Maize lethal necrosis on maize yields 14
2.7 Plant parasitic nematodes infecting maize 14
2.8 Management of Pratylenchus 17
2.9 Plant parasitic nematodes and plant viruses interactions 18
INTERACTION OF PLANT PARASITIC NEMATODES AND VIRUSES CAUSING MAIZE LETHAL NECROSIS DISEASE IN SELECTED MAIZE FIELDS IN KENYA
3.1 Abstract 20
3.2 Introduction 21
3.3 Materials and methods 23
3.3.1 Survey regions 23
3.3.2 Plant parasitic nematodes extraction 24
3.3.3 Virus assays 25
3.3.4 Data analysis 26
3.4 .2 Association of MLN, MCMV, SCMV and plant parasitic nematodes 28
3.5 Discussion 31
DEVELOPMENT OF MAIZE LETHAL NECROSIS DISEASE IN PLANTS INFECTED WITH LESION NEMATODES (PRATYLENCHUS SPP.)
4.1 Abstract 33
4.3 Materials and methods 35
4.3.1 Experimental design 35
4.3.2 Multiplication of Pratylenchus nematodes inoculum 36
4.3.3 Preparation and multiplication of Pratylenchus nematode cultures 36
4.3.4 Assessing the development of Maize lethal necrosis disease in lesion nematode infected plants 38
4.3.5 Inoculation of experimental plants with MLN viruses and lesion nematodes 39
4.3.6 MLN disease severity assessment 40
4.3.7 Detection of MLN disease-causing viruses 40
4.3.8 Estimation of lesion nematode populations 41
4.3.9 Data analysis 42
4.4 Results 42
GENERAL DISCUSSION, CONCLUSION AND RECOMMENDATIONS
5.1 Discussion 52
5.2 Conclusion 53
5.3 Recommendation 53
LIST OF TABLES
Table 3. 1 Sampled counties GPS coordinates, farms and level of disease severity across sampled farms and MLN average scores for the counties 28
Table 3. 2 Description of correlations (n=12) with significance at 0.05 and 0.01 levels (2-tailed) for MLN and Pratylenchus spp., Helicotylenchus spp., Tylenchus spp., and Meloidogyne spp. 29
Table 3. 3 Description of correlations matrix (n=12) with significance at 0.01 and 0.05 levels (2- tailed) for MCMV and Pratylenchus spp., Helicotylenchus spp., Tylenchus spp.,and Meloidogyne spp. 29
Table 3. 4 Description of correlation matrix (n=12) with significance at 0.01 and 0.05 levels (2- tailed) for SCMV and Pratylenchus spp., Helicotylenchus spp., Tylenchus spp., and Meloidogyne spp. 30
Table 3. 5 Description of correlation matrix (n=12) with significance at 0.01 and 0.05 levels (2- tailed) for plants infected with Pratylenchus spp., Helicotylenchus spp., Tylenchus spp., and Meloidogyne spp. 30
Table 4. 1 Description of treatments (viruses and their combinations) to evaluate the effect of viruses causing maize lethal necrosis (MLN) disease and Pratylenchus nematodes on maize varieties H614D and Emph 1101 38
Table 4. 3 Effect of presence of nematodes (Pratylenchus spp.) on development of MLN disease among the virus treatments in susceptible maize hybrid ‘H614D’ in season one. 44
Table 4. 4 Effect of presence of Pratylenchus on disease development among the virus treatments in susceptible ‘H614D’ in season two. 44
Table 4. 5 Effect of presence of Pratylenchus on disease development among the virus treatments in Pratylenchus-resistant maize variety Emph 1101. 46
Table 4. 6 Effects of MLN, MCMV, SCMV and Pratylenchus on root and above ground plant growth and the Area Under Disease Progression Curve (AUDPC) among treatments for H614D and Emph 1101 maize varieties 48
LIST OF FIGURES
Figure 3. 1 Maize fields showing symptomatic farm infected with maize lethal necrosis disease; Maize crop in the first image shows mid-leaf chlorosis while second image shows severe leaf chlorosis and necrosis being symptoms of MLN. 27
Figure 4. 1 Stunted growth symptom expression in plants inoculated with; (A) Maize lethal necrosis disease-causing viruses MCMV+SCMV, (B) Maize lethal necrosis disease- causing viruses+Pratylenchus, (C) SCMV, (D) SCMV+Pratylenchus and (E) Healthy plants (F) MLN positive plants 43
ABREVIATIONS AND ACRONYMS
BRI Biotechnology Research Institute
DAS Double Antibody Sandwich
ELISA Enzyme-Linked Immunossorbent Assay
KALRO Kenya Agricultural and Livestock Research Organization
MCMV Maize chlorotic mottle virus
MLN Maize Lethal Necrosis
SCMV Sugarcane mosaic virus
UoN University of Nairobi
ICIPE International Centre of Insect Physiology and Ecology
1.1 Global maize production
Maize (Zea mays L.) is an important cereal crop grown throughout the world in different agroecological environments, ranking third after wheat and rice in terms of production (Wheeler and Reynolds, 2013). Maize was among the first cultivated plants between 7,000-10,000 years ago, as documented by Mexico archaeological sites of corn cobs and fossil pollen (Piperno and Flannery, 2001; Smith, 2015). Maize distribution from Mexico to other regions of Latin America, Caribbean, United States, Canada, Asia and Africa by European explorers was rapid, leading to its evolution and cultivation for human food and animal feeds (Brown and Darrah, 1985; Gibson and Benson, 2002; Vollbrecht and Sigmon, 2005).
Cereal grains are the main targets from a family of cultivated grasses which provide needed nourishment to humankind more than other foods and accounts for almost half of the total caloric requirement (Ranum et al., 2014). Several cereal crops have been utilized for food; however, maize, rice, and wheat are the ones mainly utilized as human food sources and accounts for the highest consumption (Olugbire et al., 2021). However, maize global human consumption is lower than the stated consumption percentage as a result of wastage, other non-food products usage, and processing as animal feeds (Ranum et al., 2014). Its cultivation cuts across entire Africa making maize the dominant cereal, accounting for about 56% of the total food crops harvest area yearly. Maize is one of the prefered source of calories, and is used as a primary weaning food for children in more than 20 developing countries globally. The highest percentage of maize is milled and packaged as flour, a process that removes the most nutritious outer layer of maize grains resulting in the loss of minerals and vitamins (Uchendu et al., 2016). Different maize types, based majorly on colour ranging from yellow to red to black, have been developed and adopted across the world. Yellow maize is of high preferrance in United States while white is mostly utilized in the southern parts of USA, Central America and Africa (Ranum et al., 2014). Regions where white maize is more prefered for food have social status misperception of yellow maize due to it having been associated with food-aid programs for the poor communities while yellow maize is prefered for animal feeds (Louw et al., 2010).
Maize production is dominated by North America and Asia, mainly by 4 countries which account for two thirds of global production; United States, China, Brazil and Argentina (FAOstat, 2014). Sub-Saharan Africa utilize maize as a major staple for income and food to more than 300 million smallholder farmers (Kadjo et al., 2016). In 2021, more than 650 million people consumed an average of 43 kg yr-1 of maize, representing a 35% increase since 1960 (Shiferaw et al., 2011).
1.2 Maize production in Kenya
Maize is the main food crop to over 90% of Kenya’s population. It accounts for 65% of total staple food intake with an average person consumption of 77 kgs of maize and its products per year (Ariga et al., 2010; FAOSTAT, 2014). Maize is mainly produced in the Rift Valley, including Uasin Gishu and Trans Nzoia counties.
Maize production in Kenya fluctuate over years which result in supply shortage, and ensuring sufficent supply of the crop is vital to national food security (Yearbook, 2013). However, there was an increased production in 1994, 2001 and 2003 of which average annual production accounted for 2.3 million tonnes which could not meet annual consumption demand of 2.6 million tonnes in the same period (Kariuki et al., 2018). The country relies on food import due to 40 percent of its population being food insecure (Faostat and Production, 2016; Mutimba et al., 2010). Maize is widely consumed in Kenya’s parts of central, Rift Valley, western and eastern regions. Morever, maize has been used in these regions as a source of income, improving the local’s living standards. Consequently, factors that threaten maize production inherently impact food security. Average annual consumption rates of maize in Kenya are amongst the greatest in East Africa which accounts yearly per-capita consumption of approximately 77 kg (Koskei et al., 2020). A survey by world bank in 2015 indicated that 38% of the population in Kenya cultivates maize of which 70% is produced by smallholders for the domestic market, with the remainder produced by large scale, commercial organizations for the export market. The majority of smallholder maize production is for subsistence rather than for income generation, indicating that most families are dependent on maize as their main source of food (Simiyu, 2014). Of the 1.6 million ha of land under annual maize cultivation, 80% is owned by smallholder farmers.
1.3. Maize production constraints
Maize in Kenya is mainly produced under rainfed conditions and, therefore, erratic and lack of rainfall are the principal abiotic causes of low yields (Nyoro et al., 2004). Small scale farm holders dominate most of Kenya’s maize production. These farmers are strained by lack of enough resources and therefore cannot afford agricultural inputs like fertilizers and quality seeds. The high cost and inaccessibility of certified seed has affected the adoption of improved varieties leading to poor yields (Shiferaw et al., 2011). An estimate of about 30-1005 bags of maize is lost every year due to weeds infestation, such as striga, and decline in soil fertility (Jamil et al., 2012; Manyong et al., 2007).
Pests and disease infestation during cultivation and storage are the main biotic factors that limit crop production in Kenya (Pingali et al.,2001). Main pests affecting maize production include fall armyworm, stem borers and locusts. Fall armyworm causes 21-53%, stem borers resulting in about 15% of losses every year, while the larger grain borers may cause upto 100% losses of stored maize (Day et al., 2017). Diseases affecting maize production include fungal diseases such as Fusarium and Gibberella stalk rots and ear rots both affecting the roots, stalk and ears: others include anthracnose stalk rot, leaf blight and southern rust. Maize production is also affected by diseases caused by viruses including maize streak and the current maize lethal necrosis (MLN) disease (Savary et al., 2019). In the recent years, MLN has emerged as an important viral disease of maize. It is caused by a combination of two viruses, that is, Maize chlorotic mottle virus (MCMV) and Sugarcane mosaic virus (SCMV), resulting in significant yield losses in Kenya (Wangai et al., 2012). Nematodes have also been recorded across the globe as major pests of maize. Plant parasitic nematodes infecting maize have been studied across species and level of pathogenicity, correlation of population densities and impact to yields, determination of environmental influence to severity and management strategies (Norton, 1983, Tylka, 2007, Kimenju, 2008 and Bekker et al., 2016). Limited research on biotic and abiotic management is amongst constrains affecting maize production. There is need to carry out more research on crop diseases, pests and their interaction as well as developing resistant crops in order to address food security.
1.4 Statement of the problem
Although almost the entire Kenyan population is dependent on maize as main food crop, animal feed, and income generation, the country produced 42.1 million bags in the year 2020 which is less than the national demand of 52 million bags. The deficit is complimented by imports of maize from other countries, such as Uganda and Tanzania (Wamalwa, 2020). Among other factors, MLN poses a high threat to maize production across the country and East African region with over 80% crop loss (Wangai et al., 2012). A combination of MCMV and SCMV led to the outbreak of MLN in Kenya (Wangai et al., 2012). The loss due to this outbreak was very high, thus calling for more research on its epidemiology and management. Generally, different pathogens are kown to interact and affect disease severity in any given crop (Belval et al., 2019), and this is also suspected to be the case for MLN disease. Currently, there is minimal research on synergies between MCMV, SCMV and other pathogens (fungal, bacterial, nematodes and other viruses). There is need to study the role of plant parasitic nematodes associated with maize in the development of MLN disease.
Maize lethal necrosis diseases has had a devastating impact locally and globally, thus threatening food security (Wangai et al., 2012). Interaction between different pathgens infecting a crop have been shown to lead to increased disease severity or reduced level of disease resistance. Despite the adverse impact of MLN disease, information on the effect of other maize pathogens on MLN development is limited. For instance, decrease in maize yields due to damage caused by parasitic nematode have been documented at a range of 0 to 10% in the United States of America and up to 50% in Kenya (Tylka, 2007; Kimenju, 2008). However, there are no studies to show how the nematodes may interact with MLN-causing viruses and their effect on infected maize. In addition, MCMV is known to be quite stable in soil (Jiang et al., 1992), and there is likelihood that root- infecting nematodes may aid in transfer of the virus into plant roots leading to MLN disease development, but there is no documented evidence. There is therefore need to conduct more research on effects of parasitic nematodes on MLN disease development.
This study was carried out to determine the effects of common plant parasitic nematodes on MLN disease development on infected maize plants in the field and how different plant parasitic nematode species affect individual viruses causing MLN disease. The study results will add on to the understanding of MLN disease epidemiology, leading to more effective MLN disease management, thus improving maize yields, and resulting in increased income, animal feeds and food for current and future population.
1.6.1 Broad objective
To enhance management of Maize lethal necrosis (MLN) disease by determining the role of plant parasitic nematodes in disease development.
1.6.2 Specific objectives
i. To determine the effect of interaction of plant parasitic nematodes with viruses causing Maize lethal necrosis on disease development in maize fields in major maize growing regions of Kenya
ii. To determine the effect of lession nematode infestation on severity of Maize lethal necrosis disease.
i. High infestation of maize by plant parasitic nematodes in the field results in increased level of MLN disease severity.
ii. Lesion nematode (Pratylenchus spp.) infestation significantly increase maize lethal necrosis disease severity in maize.
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