Biopesticides Potential to Protect Tomato (Solanum lycopersicum L.) Production from Early Blight Disease (Alternaria solani) and Leaf Miners (Tuta absoluta)

1. Introduction

Tomatoes (Solanum lycopersicum L.) are a major vegetable crop in sub-Saharan Africa (Fufa et al., 2009) providing important nutritional benefits to consumers (León-García et al., 2017). In Tanzania’s 64% of the tomato crop is produced by smallholder farmers whom this crop has the potential to reduce poverty by increasing incomes for produce, as it contributes to the total harvested vegetables (URT, 2012, Mutayoba and Ngaruko, 2018). While tomato is one of the most important vegetables in Tanzania, the current fruit yield is very low (7.5 to 8.4 t/ha) compared to the developed countries (40 to 100 t/ha) depending on the location, growing season, the cultivar used and crop management practices (FAO, 2009, Heuvelink and Dorais, 2005). The impact of diseases is reported to be a major limiting factor for tomato production (Lynch, 1999) and these include early blight (Alternaria solani), late blight (Phytophthora infestans), leaf spot (Septoria lycopersici), fusarium wilt (Fusarium oxysporum, F. lycopersici), bacterial wilt (Pseudomonas solanacearum) (Dimitrios et al., 2018). Insects are also a major constraint with leaf miners (Tuta absoluta) being the most damaging pest causing 80 to 100% yield loss (Abada et al., 2008; Brévault et al., 2014).

Currently, small holder farmers use chemical fungicides such as ridomil gold (4% mefenoxam and 64% mancozeb), fungozeb 80 WP (mancozeb 80%), ivory M72 (64% mancozeb and 8%metalaxy), equation pro as prophylactic measures against early blight disease; and insecticides like radiant (Spinetoram) and snow thunder (30 g/l thiamethoxam, 10 g/l emactin benzoate) for controlling leaf miners (Tescari et al., 2014; Mushobozi and Gautam, 2017; Nuwamanya et al., 2023). Most of these are harmful to humans and other living organisms in the ecosystem (Sithanantham et al., 2002; Ngowi et al., 2007; Mushobozi and Gautam, 2017). It is reported that overuse of pesticides has increased in recent years due to a lack of information regarding the chemicals, lack of alternatives and resistance of pests to some pesticides (Abhilash and Singh, 2009; Nuwamanya et al., 2023).

Some smallholder farmers use plant extracts such as lantana (Lantana camara), myrrh (Commiphora swynnertonii) and pyrethrin (Chrysanthemum cinerariifolium) and animal waste to manage diseases in common beans (Mkindi et al., 2020). For example, early blight disease reduction in tomato has been reported using Allium sativum extract (Nashwa and Abo-Elyousr, 2012). Similarly, rabbit urine (LD50) significantly reduced the survival of insect pests for the first, second and third instars (Kemunto et al., 2022). Biopesticides, including plant extracts, offer numerous benefits in disease and pest management in agriculture, including decreased toxicity, increased safety, higher selectivity, and resistance prevention when used in combination with chemical pesticides (Patel et al., 2019; Kemunto et al., 2022). However, the absence of standardized formulations appropriate for smallholder farmers hinders their adoption and laboratory, screen house and field research trials are required to identify formulations that effectively control early blight disease-causing pathogens and leaf miners (Tuta absoluta). Here we test seven (7) plants extracts and two (2) bio products against the early leaf blight (Alternaria solani) and the leaf miner (Tuta absoluta) in laboratory, screen house and field trials and undertake a cost benefit analysis of using plant extracts for controlling pest and disease in tomato compared with using synthetics pesticides.

2. Results and Discussion

2.1. Effectiveness of Biopesticides on Invitro Alternaria solani Mycelial Growth

Analysis of variance and mean separation test on in vitro A. solani mycelia growth indicated significant (P<.001) differences among all tested biopesticides ranging from 20 to 100% reduction in mycelia growth compared with the control. The inhibition rate was highest for the positive control ridomil gold (100%), followed by 10% and 5% hot peppers (98.26% and 97.44%, respectively), which were not significantly (p>.05) different from the positive control (Table 1). The reduction in mycelia growth due to plant extracts and rabbit urine on A. solani is likely due to the bioactive metabolites contained in them (Abd-El-Khair and Haggag, 2007; Gotora et al., 2014;Mwelasi, 2015). For example, Azadirachta indica contains azadirachtin, nimbidin, salannin, azadiradione and beta-sitosterol which exhibit potent antifungal properties (Iqba et al., 2003; Anwar et al., 2007). These compounds inhibit fungal growth through disruption of cell wall membranes, inhibition of cell wall synthesis, inhibit spore germination and interfere with metabolic pathway (Wedge et al., 2002;Kumar et al., 2006).Ginger contains gingerol, paradol, shogaols and zingerone that possess significant antifungal activity against A. solani (Khatun et al.,2015;Alam et al.,2016). These compounds act through various mechanism such as disrupting fungal cell walls, affecting cell membrane integrity and blocking fungal growth (Yoshinda et al., 2011; Satyal et al., 2013; Raut and Karuppayil, 2014). The most active compounds in hot pepper for controlling A. Solani include capsanthin and flavonoids like quercetin and rutin that disrupting the integrity of fungal cell membrane and inhibiting the growth (Matsuoka et al., 2003; Giriraju et al., 2013; Koleva-Gudeva et al., 2013). Lantana contains sesquiterpenes and phenolic like quercetin and caffeic acid which penetrate the microbial membrane and enter the fungal cell, resulting in a notable reduction in the synthesis of essential components, including ergosterol (the primary component of fungal membranes), glucosamine (an indicator of growth) and proteins (Brul and Klis, 1999; Gopieshkhanna and Kannabiran, 2007 and Abd-El-Khair and Haggag, 2007). However, rabbit urine contains phenolic acids (garlic, caffeic, ferulic, o-coumaric, cinnamic, and salicylic acids) that disrupt fungal cell membranes, inhibit enzyme activity, and interfere fungal metabolism (Martin, 1982; Singh et al., 2012; Gotora et al., 2014). The higher concentration of compounds led to a greater inhibitory effect on fungal (Mohana and Raveesha, 2007; Yanar et al., 2011; Kalidindi et al., 2015; Zhao et al., 2022).

2.2. The Influence of Biopesticides on Tomato Leaf Miner Population in the Screen House Experiment

2.2.1. Effect of Biopesticides on Tomato Leaf Miner Population

Data analysis on the effect of different biopesticides on leaf miner population generally showed a significant decrease in leaf miner numbers. The result was outstanding on water formulated biopesticides, which included 10% lantana (80%), 10% hot pepper (76.67%) and ginger (71.67%) had good performance (Table 2). The effect with all the biopesticides indicated to be concentration dependent with 10% being most effective than 5% concentrations. Comparing the biopesticides effect with the negative control shows that all the biopesticides had pesticidal effect.

Several reports indicated that lantana, ginger, hot pepper has pesticidal effects aided by active compound contained in them (Liambila et al., 2021: Liambila, 2023). For example, lantana has chemical compounds like phenolic and terpenes, tetra-terpenoids with different mode of actions that can interfere the Tuta absoluta larvae’s regular metabolic processes, disrupt feeding and inhibits the synthesis of ecdysteroid hormones (Liambila et al., 2021: Liambila, 2023). Hot pepper contains flavonoids and phenolic acid that have repelling effect, which lessens the attraction of tomato plants to larvae and have poisonous effects, which result in death or stunted growth of leaf miner larvae (Kashiwagi, 2005; Mendoza, 2023). Neem contains azadirachtin, a tetranortriterpenoid compound that interferes the growth and development of larvae, it is antifeedant and it directly poisons larvae, causing their demise up to 100% (Nisbet, 2000; Kihampa, 2010; dos Santos et al., 2011; Roychoudhury, 2016; Mulugeta et al., 2020). Moreover ginger, a plant well-known for its volatile components, might have impacted negatively on tomato leaf miner (Tuta absoluta) due to gingerol, paradol, shogaols, and zingerone that contained it (Gopieshkhanna and Kannabiran, 2007; Abd-El-Khair and Haggag, 2007). These compounds in ginger are reported to reduce the desire for leaf miners’ larvae to feed on plant tissues (Gopieshkhanna and Kannabiran, 2007; Abd-El-Khair and Haggag, 2007).

2.2.2. Evaluation of the Influence of Biopesticides on Tomato Leaf Damage Caused by Leaf Miners in the Screen House Experiment

The result on evaluation of tomato plants larvae damage were analysed and indicated that, there was significantly more damage on the negative control due to leaf miner tunnelling, blotching and discoloration as compared to all other treatments. Biopesticides that resulted in the least tomato leaf damage were 10% lantana (7.83%), 10% ginger (11%), 10% hot pepper (13.53%) and 10% neem (16%) and these did not differ significantly from the positive control radiant (1.67%) (Table 3). According to the scale by Lopez et al. (2020), these treatments represented the very low damage which are acceptable by tomato growers. Damage was recorded on the remaining treatments ranged 20% to 80.70% which is considered moderate to severe damages (Lopez et al., 2020).

The effects of leaf miner tunnelling, blotching and discoloration affects photosynthesis ultimately affecting plant growth, development and performance (Liambali 2023). Tomato fruit yield and quality are both significantly impacted by direct feeding of the leaf miner as well as secondary pathogens entering host plants through wounds made by the pest (Chhetri, 2018). The use of biopesticides can disrupt the leaf miner life cycle and discourage them from feeding (Kashiwagi, 2005; Liambila et al., 2021: Liambila, 2023; Mendoza, 2023,). Azadirachtin, a deterrent and active compound in neem, has been shown to have a high mortality rate against T. absoluta and could explain the activity reported (Kubo et al., 2012; Frank et al., 2014). L. camara also had good insecticidal efficacy and repellence, with higher mortality rates of larvae (Liambila et al., 2021), suggesting it as an environmentally friendly alternative to synthetic pesticides.

2.3. Field Evaluation of Four Biopesticides on Early Blight Disease Incidence, Severity and Tomato Growth Parameters

2.3.1. Early Blight Disease Incidence

Early blight disease incidence in Kilala showed that all the treatments were significantly (p < 0.05) different and lower than the negative control which suffered 84.4% damage. The most effective treatments, 10% ginger (37.8 %), 5% hot pepper (33.3%) and 10% hot peppers (26.7%) were not significantly different (p > 0.05) from the positive control (24.4%) (ridomil gold). The result in Mailisita indicated that, 5% hot pepper (31.1%), 10% hot pepper (28.9%), 10 % lantana (31.1%) were more effective in reducing disease incidence but differed significantly (p < 0.05) with the positive control (ridomil gold) (Table 4). For both sites, disease incidence was slightly higher in Kilala (84.4%) compared to Mailisita (77.8%) on negative control. The effect of sites (blocks) did not affect disease incidence (Table 4).

Here we have shown that biopesticides and especially plant extracts have the potential to reduce early blight disease incidence by over 50% in the field and across two field sites. Metabolites reported from hot pepper and lantana, include phenolics, flavonoids and terpenoids which could explain the efficacy while gingerol, shogaol, and zingerone in ginger are known to inhibit fungal growth (Brul and Klis, 1999; Gopieshkhanna and Kannabiran, 2007; Abd-El-Khair and Haggag, 2007; Giriraju et al., 2013; Raza et al., 2016; Ahmad et al.,2017;). Moreover, hot pepper, ginger, and lantana extracts trigger a plant’s innate defence responses and enhance its ability to resist pathogens (Abd-El-Khair and Haggag, 2007; Nashwa and Abo-ElyouSr, 2012; Sallam et al., 2022) and are also deterrent to some pests that may vector early blight pathogens (Spochacz et al., 2018). Biopesticides can target specific pathogens while having limited effects on beneficial organisms and contribute to healthier ecosystems and more sustainable agricultural practice (Shuping & Eloff, 2017; Lengai and Muthomi, 2018 and Tembo et al., 2018).

2.3.2. Early Blight Disease Severity

Analysis of variance on early blight disease severity data from Kilala showed that there were no significant differences among 10% ginger (39.8%), 5% hot pepper (40.9%) ,10% hot pepper (40.0%) and positive control (ridomil gold) (32.3%). Disease severity in Mailisita showed that, the positive control (27.2%) differed slightly with 10% hot pepper (31.1%) but differed significantly from the negative control (79.3%) and rest of the treatments (10% lantana (42.2%), 10% ginger (39.6%), 5% hot pepper (39.5%). Comparing the efficacy of the treatments from two sites, biopesticides were more effective in Kilala than Mailisita with three treatments (10% ginger, 5% and 10% hot pepper) performing as well as the positive control whereas in Mailisita only one biopesticides (10% hot pepper) performed as well as the positive control (Table 5). This might be due to the high rainfall at Mailisita during the experiment providing conditions that encourage fungal growth (Singh et al., 2020).

The bioactive components of ginger (gingerol, shogaol, and zingerone), lantana (lantadene A, lantadene B, terpenoids, and flavonoids), and hot pepper (flavonoids and phenolic acids) extracts inhibit fungal growth and initiate plant defence responses that increase resistance to pathogens (Abd-El-Khair and Haggag, 2007; Nashwa and Abo-ElyouSr, 2012; Langai et al. 2017; Fuentefria et al., 2018; Vaou et al., 2021; Abdule et al., 2022). Plant extracts offer additional benefits such as their natural abundance, low cost, non-persistence, and low adverse environmental consequences (da Cruz Cabral et al., 2013). These results showed that biopesticides are promising means for disease management for smallholder farmers in developing countries.

2.3.3. To Evaluate the Effect of Biopesticides to Control Early Blight, Leaf Miners and Their Impact on Tomato Growth and Fruits Yield

The effect of different treatments on plant height, number and weight of tomato fruits per plant on both sites were significantly greater than the negative control (p < 0.05). The plant height was relatively higher in Mailisita (123.3 cm) compared to Kilala village (114.3cm) (Table 6). This might be due to high rainfall in Mailisita that boosted plant growth by enhancing stronger root systems and nutrient absorption (Zafar et al., .2024). Similarly, higher number and weight of tomato per plant was observed in Mailisita. These data suggests up to 50% increase in plant growth through the use of plant extracts either as growth regulator or bio-control agent for disease (Abdel-Kader & El-Mougy, 2016). Plant extracts sprayed to control pests and diseases could act additionally as a nutrient supplement boosting plant growth and yield (Mkindi et al., 2020). Moreover, biochemical ingredients from hot peppers, lantanas, and ginger might affect plant hormone levels including auxins and gibberellins, which are essential for cell elongation and other unique modes of action (Badr et al., 2021; Chtioui et al., 2022; Sohrabi et al., 2024; Manish et al., 2024). Alterations in these hormones could lead changes in plant growth (Ashraf et al. 2018).

2.3.4. Effect of Biopesticides on Tomato Leaf Damage Caused by Leaf Miners

Field assessment of biopesticides for reducing tomato leaf miner (Tuta absoluta) damage on tomato leaves revealed that all biopesticides tested at Kilala were significantly more effective than the negative control. However, their efficacy was still lower than that of positive control , which exhibited only 4.4% damage compared to 10% ginger (32.8%), 10% hot pepper (29.8%), 5% hot pepper (29.1%) and 10% lantana (32.8%) (Table 7). The greater efficacy of the positive control might be due to the fact that radiant is a systemic insecticides (Tescari et al., 2014) whereas, the plant extract have a multitude of effects including toxic, sub lethal, antifeedant or neurotoxic activity, ultra structural malformation, and effects on prooxidant/antioxidant balance (Spochacz et al., 2018).

For the Mailisita site, 10% ginger (24.4%) and 5% hot pepper (28.3%) were also effective at controlling the insect but significant less so than the positive control (radiant with 10.0%). The rest of the treatments were all significantly better at controlling the insect than the negative control which had a higher leaf damage of 72.2%. The pest pressure on both sites was similar in Kilala (76.8%) and Mailisita (72.2%) on the highly susceptible control (Tanya). This suggests that all the biopesticides used in this experiment had negative effect on leaf miner (Tuta absoluta) larvae. Biopesticides (neem, hot peeper, ginger, lantana) disturb larvae life cycle and discourage them from feeding, resulting in death and reducing damage (Kashiwagi, 2005; Langai et al. 2018; Ibrahim et al., 2019; Rahardjo et al.,2019; Liambila et al., 2021: Liambila, 2023; Mendoza, 2023).

2.4. To examine the Cost-Benefit Analysis of Using Biopesticides as Means of Controlling Early Blight and Leaf Miners

2.4.1. Treatment Advantage

Analysis of variance on treatment advantage showed that, there were significant differences on revenue accrued between negative control and all treatments applied including the conventional tomato production method with synthetic pesticide. Among the test treatments 5% hot pepper (2818.48 USD) had the highest treatment advantage revenue followed by the positive control (2611.79 USD), 10% hot pepper (2585.31 USD), 10% lantana (2459.66 USD), conventional tomato production method (2458.53 USD), 10% ginger (2018.81 USD), 10% rabbit urine (1519.37 USD) and the negative control was the least (Table 8). This low treatment advantage revenue exhibited on the negative control indicated the importance of pest and disease management in tomato production. Analysis on total cost of production showed no significant difference on all the treatments and, positive and negative control. This indicates that biopesticides for this case are expensive as the conventional pesticides though, all were profitable (Malinga and Laing, 2023). However, biopesticides are cheaper than chemical pesticides when locally produced especially for small scale agricultural use or for domestic pest management (Agboola et al., 2022; Ayilara et al., 2023). The treatment advantage accrued in this study is the result of price fetched by organically produced tomato at the marketplace. This results give an opportunity for smallholder farmers to invest on medicinal plants production, which will help in lowering the cost of these important plants for pest and disease management.

2.4.2. Cost-Benefit Ratio

Result on cost-benefit analysis indicated that, 5% hot pepper (1:3.5), positive control (1:3.3), 10% hot pepper (1:3.1), 10% lantana (1:3.0), conventional tillage practices (1:2.7), 10% ginger (1: 2.4), rabbit urine (1:2.0) had their cost benefit ratio greater than 1.0 whereas the negative control (1:-0.4) had its cost-benefit ratio below 1.0 (Table 8). According to the profitability index rule, a result of more than 1.0 typically considered financially viable and likely to be successfully; a reading of 1.0 indicates that the costs and benefits are equal; and a reading of less than 1.0 indicates that the costs outweigh the benefits (Gharib et al., 2017). The 5% hot pepper (1:3.5) treatment can be considered the most beneficial biopesticide used in this study even compared to the positive control-radiant (pest control) and ridomil gold (Alternaria solani). Generally, all the biopesticides were an effective means of pest and disease management in the study.

3. Materials and Methods

3.1. Study Location

A laboratory study to assess the efficacy of botanical extracts on A. solani was conducted at the Nelson Mandela African Institution of Science and Technology (NM-AIST), Arusha, in 2023. The field and screen-house experiments were conducted at Mailisita located at 3.3717014 S, 37.28944444 E and at altitude 970 m .a .l, Hai district, Kilimanjaro. Another field experiment was conducted at Kilala- Arumeru district, Arusha located at 3.366667 S, 36.85 E. Mailisita and Kilala sites’ temperature and rainfall averages ranged 17–29 ⁰C and 15–25 ⁰C while rainfall was 500–1800 mm and 500 mm–1200 mm, respectively. Both sites were selected due to their suitability for tomato growing and, leaf miner pests and early blight disease were the common biotic constraints in these areas.

3.2. Collection of Materials and Preparation of Biopesticides

Plant materials were collected locally (Table 9) and thoroughly washed with running tap water. They were dried in shade in the screen house for four (4) days, followed by grinding into a powder using pestle and mortar. The plant pounded materials were dissolved in water as extracting solvent using a ratio of 1:10 (w/v), placed on the shaker for 12 hours then filtrated by two layers of cheesecloth before passing into Whitman’s No.2 filter paper to obtain infranatant as described by Tadele & Emana, (2017), and Abubakar & Haque, (2020). The infranatant of each mixture was diluted to make 5% and 10% concentrations using autoclaved distilled water.

Jeevamrutham was prepared according to Kaur, (2020). Cow dung (1kg), cow urine (1l), jaggery (200 g), common bean flour (200 g), and fertile soil (100 g) were all combined in the bucket (20 L). For nine (9) days in the shade, the bucket was stirred vigorously three times daily for 10 to 15 minutes with a wooden stick, and it was covered with a junk sack to ensure aeration. Water was added to the mixture to bring the volume up to 20 l after 9 days and the solution was sterilized with 0.2 µ disposable syringe filters that was used in the laboratory experiment with 5% and 10% concentrations. Additionally, One (1) litre of rabbit urine was collected and diluted to formulate 5% and 10% concentrations followed by sterilization with 0.2 µ disposable syringe filters.

3.3. Laboratory Evaluation of Biopesticides on the Growth Rate of Mycelium in Alternaria solani

The laboratory experiment was laid out in a factorial complete randomised design with 20 treatment combinations (7 plant extracts, 2 bio products and 2 concentrations), including negative (media inoculated pathogen) and positive (ridomil gold) controls, replicated thrice with three observations each. Prepared biopesticides in 3.2 were used in the laboratory experiment. Alternaria solani isolates used in this experiment were cultured from NM-AIST maintained cultures on highly susceptible Tanya tomato variety. Isolation and multiplication of inoculum (A. solani) was done to obtain enough inoculum for 180 petri dishes of 90 mm diameter. Preparation of full-strength potato dextrose agar (PDA) was done by autoclaving the media at 121⁰C for 15 minutes in 1l bottle and allowed to cool to 40 °C. Each biopesticides added in the media petri dishes by ratio of 1:4 v/v (5 ml biopesticides: 20 ml media), shook for 10 minutes before solidification. Inoculation was done using mycelium agar plug (MAPs, 5 mm in diameter) from a full-grown petri dish (Wonglom et al., 2019). All the petri dishes were incubated at room temperature of 24 to 25 °C. Data were collected by measuring the colony growth size (mm) starting from one day after inoculation for seven consecutive days, as described by (Wonglom et al., 2019). The inhibition rate for each treatment was calculated using the formula by Wonglom et al. (2019) as in equation 1.

$$\mathrm{M}\mathrm{y}\mathrm{c}\mathrm{e}\mathrm{l}\mathrm{i}\mathrm{a}\mathrm{l} \mathrm{g}\mathrm{r}\mathrm{o}\mathrm{w}\mathrm{t}\mathrm{h} \mathrm{i}\mathrm{n}\mathrm{h}\mathrm{i}\mathrm{b}\mathrm{i}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\left(\%\right)={\displaystyle \frac{\mathrm{G}\mathrm{r}\mathrm{o}\mathrm{w}\mathrm{t}\mathrm{h} \mathrm{i}\mathrm{n} \mathrm{c}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l}-\mathrm{G}\mathrm{r}\mathrm{o}\mathrm{w}\mathrm{t}\mathrm{h} \mathrm{i}\mathrm{n} \mathrm{t}\mathrm{r}\mathrm{e}\mathrm{a}\mathrm{t}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t}}{\mathrm{G}\mathrm{r}\mathrm{o}\mathrm{w}\mathrm{t}\mathrm{h} \mathrm{i}\mathrm{n} \mathrm{c}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l}}} \mathrm{X} 100$$

(1)

The data from mycelial growth inhibition rate (%) were subjected to analysis of variance, a mean separation test was done by using Bonferroni multiple comparison test to identify effective treatments using GenStat 21st Version (64 bits), and Statistical software from Visual Statistics and Information (VSNi) located at Heslington, United Kingdom.

3.4. To Assess the Efficacy of Biopesticides on Tomato Leaf Miner Populations and Leaf Damage

The screen-house experiment was conducted to investigate the efficacy of biopesticides in controlling leaf miner tomato crop damage using a factorial complete randomized design. The experiment used 20 treatment combinations together with negative (Unsprayed plants) and positive (Radiant 120 SC) controls, replicated six times, totalling 120 plastic pots. The preparations of plant extract and animal waste were prepared as in section 3.3 above. Tanya tomato variety was raised in the nursery for three weeks then transplanted into the plastic pot using 2 kg sterilized forest compost soil. The soil was sterilized to remove other organisms that could interrupt the experimentation. The pots were covered by fine-meshed netting of 0.4 mm size cage to prevent leaf miners from moving out from the pots and other insects from getting into the pots. Irrigation using tap water was done regularly. The introduction of 10 larvae of a leaf miner (Tuta absoluta) was done one week after transplanting. The larval stage was considered to be the most damaging stage of the pest. Then the leaf miners were left to adopt to the environment for 1 day before start spraying and evaluation. The spraying of 30 ml of each biopesticide was done using the same concentrations as above in section 3.3 was done. The same sprays were repeated three times at an interval of 5 days.

Data collection was done on leaf miner one day after each spray making three total measures. Tomato leaf damage was assessed and ranked as “mines” or “punctures” using the damage index established by Lopez et al. (2020), on the percentage of the leaf area damaged as follows: very low (0-20%), low (20-40%), moderate (40-60%), high (60-80%) and severe (80-100 %). Both data for leaf miner and tomato leaf damage % were recorded on excel sheet. Data analysis, data on leaf miner and tomato leaf damage % were visualized for normality and subjected to analysis of variance, then a mean separation test was done by using the Bonferroni multiple comparison test to identify effective treatments using GenStat 21st Version (64 bits), Statistical software from Visual Statistics and Information (VSNi) located at Heslington , United Kingdom.

3.5. Evaluation of Biopesticides in Controlling Early Blight in the Field

Field experiment was conducted at Mailisita and Kilala. Each site of area 84 m² was cleared and ploughed and the experiment set up using a randomized complete block design with three replications. Nursery was prepared and Tanya tomato variety was used. Tomato seedlings were transplanted to the experimental plots after 1 month. The size of each block was 12 m² with each having six plots of 2 m² where each plot was transplanted with 13 tomato seedlings at a spacing of 50 cm x 30 cm. Temperature and precipitation were recorded throughout the experiment (Table 10).

The four best treatment combinations from the laboratory experiment (5% hot pepper, 10% hot pepper, 10% Lantana, and 10% ginger) were used, with negative and positive controls. Each treatment was applied 8 times in each block, on a weekly basis starting from one week after transplanting. Field management was done according to smallholder farmers’ practices to reflect field conditions.

Data collection was done at the fruiting stage on early blight disease severity by randomly selecting five plants in each plot for every treatment as described by (Weber and Halterman, 2012). Infected leaves were classified into five categories (0, 1, 2, 3 and 4) according to blighted area of leaves, where by 0 = no infected leaves, 2 = ≥25% or less, 3 = 26-50%, 3 = 51-75% and 4= 76-100% (Weber and Halterman, 2012). The data was used to calculate percentage disease severity using the equation 2 of Ibrahim et al., (2004).

$$\mathrm{Disease} \mathrm{severity} \left(\%\right)=\sum (T/N)X100$$

(2)

where T = number of infected leaves per plant; N = total number of leaves per plant.

Disease incidence in tomato plants involved counting of tomato plants to quantify the proportion of plants affected by early blight disease within a population (Madden and Hughes, 1995). Tomato leaf damage was assessed and ranked as “mines” or “punctures” using the damage index established by Lopez et al. (2020), on the percentage of the leaf area damaged as follows: very low (0-20%), low (20-40%), moderate (40-60%), high (60-80%) and severe (80-100 %). Data on plant height, number of tomato fruits per plant and total tomato fruit weight per plant were collected as described by (Balemi, 2008).This was done by randomly selecting five plants in each plot for every treatment once at the harvesting stage. Data were subjected to analysis of variance, and then the mean separation test was done using the Bonferroni multiple comparison to identify effective treatments by GenStat 21st Version (64 bits), Statistical software from Visual Statistics and Information (VSNi) located at Heslington, United Kingdom.

3.5. To Perform Cost-Benefit Analysis of Using Biopesticides as Means of Controlling Early Blight in Tomato

3.5.1. Cost-Benefit Analysis

The examination of the cost-benefit analysis involved in the tomato production was observed. The evaluation survey was conducted in the two sites of Mailisita and Kilala, using a structured questionnaire administered (attached annex 1) to 20 smallholder farmers in each site as described by (Mkindi, 2021). The selection of 20 smallholder farmers was based on the total average number 200 and 250 smallholder farmers from Mailisita and Kilala villages, respectively. The respondents in both sites were selected at random from pre-selected tomato farmers with a history of tomato growing in these areas. Second part was the cost benefit analysis involved the two experimental sites of Mailisita and Kilala, where the use of biopesticide was done. The activities and cost attached to each site and on each biopesticide were evaluated as described by (Sheshma et al., 2022).

The costs of obtaining plant materials and animal excretes, processing them and cost of tomato production for both conventional and experimental sites production were recorded as average for each item as described by (Sheshma et al., 2022; Malinga and Laing, 2023). It involved the evaluating cost based on conventional tomato production that included costs of purchasing fungicides and insecticides, transport, spraying, and protective gears and other common costs. Furthermore, the cost production based on using biopesticides (10% lantana, 5% hot pepper, 10% hot pepper and 10% ginger) also evaluated. It included cost of collecting / purchasing botanical plants /animal excretes, transportation, plant extracts preparation, cost of protective gears, plant extracts spray, times of application and common costs. The sum of these expenses indicated the entire cost of plant protection and revenue accrued from the tomato production.

The net benefit was calculated by deducting the total cost of tomato production from the total income, which was calculated by multiplying the total yield per hectare by the current market price (Sheshma et al., 2022; Malinga and Laing, 2023).

$$\mathrm{N}\mathrm{e}\mathrm{t} \mathrm{b}\mathrm{e}\mathrm{n}\mathrm{e}\mathrm{f}\mathrm{i}\mathrm{t}=\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l} \mathrm{i}\mathrm{n}\mathrm{c}\mathrm{o}\mathrm{m}\mathrm{e}-\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l} \mathrm{c}\mathrm{o}\mathrm{s}\mathrm{t} \mathrm{o}\mathrm{f} \mathrm{p}\mathrm{r}\mathrm{o}\mathrm{d}\mathrm{u}\mathrm{c}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}$$

(3)

Treatment advantage over the control was calculated by deducting the control treatment’s income from each sprayed treatment income as described by (Sheshma et al., 2022).

$$\mathrm{T}\mathrm{r}\mathrm{e}\mathrm{a}\mathrm{t}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t} \mathrm{a}\mathrm{d}\mathrm{v}\mathrm{a}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{g}\mathrm{e}=\mathrm{E}\mathrm{a}\mathrm{c}\mathrm{h} \mathrm{s}\mathrm{p}\mathrm{r}\mathrm{a}\mathrm{y}\mathrm{e}\mathrm{d} \mathrm{t}\mathrm{r}\mathrm{e}\mathrm{a}\mathrm{t}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{s}\mathrm{’} \mathrm{i}\mathrm{n}\mathrm{c}\mathrm{o}\mathrm{m}\mathrm{e}-\mathrm{C}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l} \mathrm{t}\mathrm{r}\mathrm{e}\mathrm{a}\mathrm{t}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t} \mathrm{i}\mathrm{n}\mathrm{c}\mathrm{o}\mathrm{m}\mathrm{e}$$

(4)

Cost-benefit ratio was calculated according to (Sheshma et al., 2022). Each treatment was derived by subtracting the additional income of production from the net income of production, then were divided by total cost of tomato production for each treatment.

$$\mathrm{C}: \mathrm{B} \mathrm{ratio}={\displaystyle \frac{\mathrm{A}\mathrm{d}\mathrm{d}\mathrm{i}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\mathrm{a}\mathrm{l} \mathrm{i}\mathrm{n}\mathrm{c}\mathrm{o}\mathrm{m}\mathrm{e} \mathrm{f}\mathrm{r}\mathrm{o}\mathrm{m} \mathrm{p}\mathrm{r}\mathrm{o}\mathrm{d}\mathrm{u}\mathrm{c}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}}{\mathrm{C}\mathrm{o}\mathrm{s}\mathrm{t} \mathrm{o}\mathrm{f} \mathrm{p}\mathrm{r}\mathrm{o}\mathrm{d}\mathrm{u}\mathrm{c}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}}}$$

(5)

Data were subjected to analysis of variance, and then mean separation test was done by using Bonferroni multiple comparison test to identify effective treatments by GenStat 21st Version (64 bits), Statistical software from Visual Statistics and Information (VSNi) located at Heslington, United Kingdom.

4. Conclusions

The current study indicated that extracts of 5% hot pepper, 10% hot pepper, 10% ginger, and 10% lantana proved successful in controlling early blight (A. solani) and leaf miners in both laboratory, screen house and field experiments. These plant extracts may be used instead of fungicides, which will lower the cost of fungicides and their residual effects that pollute the environment and climate. Moreover, the study reveals significant differences in treatment advantage revenue between conventional tomato production methods and biopesticides, with the 5% hot pepper treatment having the highest revenue. The cost-benefit ratio analysis shows all biopesticides treatments are financially successful. We therefore recommend the research on identifying the best time of application, assess the adjuvants and additives that can be used in combination with plant extracts to enhance their efficacy. Nevertheless, this research recommends further experiments to evaluate the best method of applying plant extracts that can influence their distribution, coverage, and penetration into plant tissues.