Effects of Foliar and Soil Applications of Cold-Pressed Neem

Oil Solutions on Vigor and Phytotoxicity in Cucurbits

Katie Cyr

aUniversity of Wisconsin – Marathon County Wausau, WI; bStoney Acres Organic Farms Athens, WI

Introduction:

With increasing concern from both farmers and the public about the use of inorganic pesticides on crops, many farmers are turning to organic pesticides as a safer, more eco-friendly alternative. Neem oil, known for its insecticidal and antifungal properties, is one of these pesticides. Neem oil is derived from the neem tree (), native to and other parts of southern Asia. The most active and well known insecticidal component of neem oil is the chemical azadirachtin. It has been shown to be an growth regulator, and has anti- feedant and oviposition deterrent properties (Caldwell, et al. 2013; Musabyimana, et al. 2001).

Neem oil is most effective when applied to young seedlings to protect them from pests during early and more vulnerable stages of growth.

Neem oil and other oil-based pesticides tend to have a lower phytotoxicity than most commercial insecticides, and are relatively inexpensive. Oils also have little risk to human health and tend to break down naturally over time, leaving little or no trace in the fruit and vegetable products of plants (Finger, et al. 2002). Because they aren’t water soluble, neem oil and other oils are most often applied in an oil/water emulsion mix. Another solution is extracting the active pesticide ingredients (such as azadirachtin) from the oil and applying them separately (Caldwell, et al. 2013). Azadirachtin is water-soluble, so the chemical is often extracted from the neem oil and applied separately as part of a neem-based insecticide. Many oils possess some phytotoxic

1 properties that can cause leaf necrosis, chlorosis, leaf drop, and scorching when applied to the surface of (Baudoin, et al. 2006). The application of oils to the leaves of plants can cause a reduction in their net photosynthesis, including reduction in net CO2 assimilation and transpiration. This is thought to be caused by the oil creating a barrier that blocks the stomates from absorbing CO2 and releasing oxygen and H2O. In experiments where oils caused a phytotoxic effect in plants, an oily residue was often observed on the leaves (Baudoin, et al.

2006; Finger, et al. 2002). Reduction in net CO2 assimilation was shown to correlate with an increase in concentration of oil applied in an oil/water emulsion mixture (Moran, et al. 2003).

Eventually, the decrease in net CO2 assimilation can lead to a decrease in sugar production and storage and can reduce crop yields (Finger, et al. 2002). Many experiments have been conducted to determine an optimal concentration of oil to be applied to plants in order to reduce leaf phytotoxicity while still maintaining beneficial insecticide or anti-fungal properties (Baudoin, et al. 2006; Finger, et al. 2002; Moran, et al. 2003; Musabyimana, et al. 2001).

While azadirachtin is the main insecticidal ingredient in neem oil, other components of neem oil have been shown to have insecticidal properties as well. When extracted from the neem oil solution, azadirachtin has shown to be less effective against and fungi than unmodified neem oil (Chowdhury, et al. 2002). It has also been shown that an aqueous neem seed kernel extract tends to have a lesser degree of phytotoxicity than an independent application of azadirachtin, indicating that that other properties of neem oil may decrease its phytotoxicity towards plants (Chowdhury, et al. 2002). The application of neem oil products to plants appears to be more effective as an insecticide than azadirachtin-based products.

Neem oil can be applied as either a foliar spray (applied to the surface of the plant’s leaves) or a soil treatment in which the plant absorbs the active ingredients into the vascular

2 system of its roots. In the case of foliar application, the oil properties of a neem solution have the potential to have phytotoxic effects on the plant’s leaves. A soil application of neem oil could reduce the pesticide’s chances of being phytotoxic to the surface of the plant’s leaves, but the hydrophobic properties of neem oil may reduce or prevent its absorption by the plant’s roots. The xylem tissue of the roots of plants, which intake water and hydrophilic substances, do not allow for the absorption of hydrophobic substances. The application of water-soluble neem pesticides has been shown to reduce feeding damage in certain plants, but these pesticides may not contain all of the insecticidal properties of cold-pressed neem oil (Shivashankar, et al. 2000). The purpose of the experiment performed by my research group was to compare the phytotoxic effects of foliar and soil applications of neem oil on cucurbits, as well as the affects that they have on the plants’ vigor.

Materials and Methods:

Plants: For this experiment, three cucurbit were used: Amish Heirloom muskmelon

(melon), Chicago Warted Hubbard squash (squash), and Black Beauty zucchini (zucchini). Seeds were planted under greenhouse conditions at the University of Wisconsin Marathon County in

5.08x5.08 cm cells filled with Jiffy Mix Organic Seed Starting Mix (JiffyGroup, Inc.) on Day 1

(May 3rd). Sixty cells of each species were planted, with three seeds per cell. After 9 days of growth, the plants were thinned out to one plant per cell (Day 9).

Neem oil: Cold-pressed neem oil (Ahimsa organic neem oil – OMRI listed) was prepared in a 1% v/v neem oil/water emulsion solution, with 0.1% Seventh Generation Liquid Dish Soap (Seventh

Generation, Inc.) as an emulsifier. The neem solution was mixed at a ratio of 37 ml neem oil and

3.7 ml emulsifier per 1 gallon of water (3785.41 ml).

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Treatment: Each species of plant was separated into three groups, each receiving one of three treatment types. The control groups (control) received regular watering using a hand-controlled watering can. The foliar treated groups (foliar) received the 1% v/v neem oil/water treatment applied to the leaves using a hand-controlled watering can. The soil of the foliar treated plants received regular watering with water, which was applied using a hose with a slow flow while avoiding the leaves. The cell trays of the soil treated groups (soil) were placed in a tray filled with the neem oil solution, and the soil was allowed to absorb the solution until fully saturated.

The soil was considered saturated when the upper surface of the soil had an oily sheen and was moist. The number of plants in the treatment groups were as follows: melon – 19 control, 20 foliar, and 20 soil; squash – 18 control, 18 foliar, and 17 soil; zucchini – 24 control, 20 foliar, and 20 soil. The treatments were applied on Days 20, 22, and 25. On Day 26, the plants were transplanted to outdoor fields at Stony Acres Organic Farm in Athens, WI. The plants were organized using a randomized complete block system design at 60 cm within row spacing, with 9 plants per block and 15 replications.

Data Collection: Data were collected on Days 33, 41, and 47. For each plant, the number of leaves and number of damaged leaves were recorded. Each leaf on the plant was rated for damage on a 0-5 scale as follows: 0 – no visible damage; 1 – scorched leaf margins only; 2 –

<25% of the leaf area damaged; 3 – 25-75% of the leaf damaged or leaf cupped but mostly green/healthy; 4 – 75-95% of the leaf damaged or leaf cupped and sick looking (yellowing); 5 – the leaf mostly or entirely dead. These scores were then added up to calculate the total damage score per plant, which was recorded. The data collected from the plants was then used to calculate the average percent of leaves damaged per plant and the average leaf damage score per plant. On the second and third days of data collection (Days 41 and 47), the number of flowers

4 per plant was also recorded; flowers weren’t yet present during the first week of measurements.

The number of leaves and flowers per plant was used as a measure of the plant’s vigor, whereas the percent of damaged leaves and average damage score per leaf were more direct measurments of phytotoxicity.

Results:

Our primary goal was to compare the degrees of vigor and phytotoxicity of foliar and soil treated plants to the water control treated plants, and comparisons between the different plant types were not of interest. Data for each dependent variable (number of leaves, number of flowers, % of leaves damaged, and overall damage score) were analyzed for each crop and sampling date separately using one-way ANOVA followed by pairwise comparisons. We used an alpha level of

0.05 for the ANOVA. Tests of a priori hypotheses of pairwise comparisons of means were conducted using Bonferroni adjusted alpha levels of 0.0167 per test (0.05/3). All analyses were performed using the Data Analysis Add-In in Microsoft Excel 2010.

Average Number of Leaves/ Plant: On the first day of data collection (Day 33), the foliar and soil treatments averaged fewer numbers of leaves per plant compared to the control treatment (Table

1). This can be seen most prevalently in the zucchini and melon plants, and was most significant in the foliar treatment. Compared to the control treatment, the foliar treatment resulted on average in 0.6 leaves less per zucchini plant and 1.6 leaves less per melon plant. Among the melon plants, the soil treatment resulted in 0.86 leaves less on average than the control treatment on Day 33.

This slower production of leaves became more apparent over the next two weeks, as can be observed in Figures 1 and 2. The melon plants were affected most significantly by both neem treatments. By Day 47, the foliar treatment melon plants had 4.74 fewer average leaves than the control melon plants, while the soil treatment plants had 7.38 fewer average leaves (Table 1, Figure

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2). While all three treatment groups grew additional leaves over time, the soil and foliar groups added leaves at a slower rate than the control group.

Average Number of Flowers/ Plant: Flowers were not yet present on any of the plants on Day 33

(Table 2). By Day 41, it became apparent that the foliar treatment resulted in fewer average flowers per plant than control or soil treatment plants, which were significantly less than the control treatment among the zucchini and melon plants. By Day 47, the average number of flowers per plant was significantly less in the foliar group for all three plant types (Table 2). The foliar treatments averaged 2.51 flowers less than the control treatment among the squash plants

(Figure 3), 2.5 flowers less among the zucchini plants, and 2.06 flowers less among the melon plants on Day 47 (Table 2). There were almost no significant differences between the control and soil treatments for any of the plants on either Day 41 or 47, the only exception being the melon plants on Day 41.

Average Percent of Leaves Damaged per Plant: When compared to the control and soil treatment plants, the foliar treatment plants sustained a significantly higher average percentage of leaves damaged per plant among all three plant types on Day 33 (Table 3). Among the melon plants, the foliar treatment resulted in over three times more leaves damaged on average compared to the control plants on Day 33 (Figure 5). The foliar treatments for the squash and zucchini had slightly less than twice the percentage of damaged leaves compared to their control groups (Table 3, Figure 4). Over time, the percentage of damaged leaves decreased in all three treatment groups. This decrease was greatest for the foliar-treated group, which had percentages very similar to the control and soil-treated groups during the second and third weeks of data collection (Table 3). The zucchini plants were the only group that maintained statistically significant percentages of damaged leaves between the foliar and control treatments by Day 47

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(Figure 4) This steep decrease in percent of damaged leaves between Day 33 and Day 47 can be observed in Figures 4 and 5. The soil treatment plants showed little significant difference from the control plants over the testing period, the exceptions being the squash on Days 33 and 47 and the zucchini on Day 47.

Average Leaf Damage Score per Plant: The patterns found in the average damage score per leaf, both over time and between the treatment groups, were very similar to the trends found in the average percent of leaves damaged per plant. On the first day of observations (Day 33), the average damage score per leaf was significantly higher among the foliar treatments for both the zucchini and melon plants, compared to their respective control groups (Figures 6 and 7). The foliar treatment zucchini plants averaged 1.21 damage points higher than the control zucchini plants, while the foliar treatment melon plants averaged 3.85 damage points higher than the control melon plants (Table 4). Over time, the average damage scores of all three treatment groups decreased. The greatest decrease over the three-week period was among the foliar treatment plants, which is apparent in Figures 6 and 7. By Day 47, only the zucchini plants maintained a significant difference between the foliar treatment and control treatment damage scores (Table 4, Figure 7). There was no significant difference in damage score among any of the treatment groups for the squash plants, and there was very little significant difference in damage scores between the soil and control treatments for all three plants on all three of the observation days. The only exception was Day 47 for the melon plants, in which the average leaf damage score for the soil treatment plants was significantly less than the control plants.

Discussion:

Overall, the foliar treatment plants showed the greatest degree of phytotoxicity, especially earlier in the life cycle of the plants. Foliar-treated plants showed the greatest degree

7 of scorching, yellowing, and cupping, averaging a much higher percentages of leaves damaged and leaf damage scores than the control and soil treatment groups. This is likely due to the phytotoxic effects that oils tend to have on the surface of leaves. The neem oil could have covered the stomates of the leaves, causing a decrease in transpiration and a loss in moisture that would explain the scorching and cupping of the leaves. As the plants grew, the new leaves that emerged would not have received the neem solution treatment. Since they wouldn’t suffer the phytotoxic effects of the neem oil, these healthier leaves would reduce the average number of damaged leaves and the average damage score per leaf (Figures 4, 5, 6, and 7).

On average, the soil treatment groups did not differ significantly to the control groups in terms of average number of leaves damaged and average damage score per leaf. Both the control and soil treatment groups averaged relatively low numbers of damaged leaves and leaf damage scores, and these measurements were almost always significantly lower than the foliar-treated groups during the first day of measurements. This can indicate one of two possibilities: that the absorption of the neem oil solution through the plant’s roots does not cause any phytotoxic effects to the plant, or, that the plants did not absorb the neem oil solution. While the emulsifier in the neem oil solution reduced the hydrophobic nature of the neem oil, there is still a chance that the plants were unable to absorb the neem oil portion of the solution. This would affect the efficacy of the neem oil as a systemic insecticide if insects were to feed on the plants.

In terms of plant vigor, both the foliar and soil treatments reduced the average number of leaves per plant compared to the control treatment, indicating that both had negative effects on the plants’ vigor (Table 1, Figures 1 and 2). Because the foliar treatment resulted in greater phytotoxicity towards plants, it would have reduced the absorption rate of CO2, and the dispersal of nutrients throughout the plant. The foliar treatment plants’ leaves also had a greater degree of

8 leaf death, which would reduce the photosynthetic rate of the plant and lessen its ability to produce new leaves. The soil treatment, however, did not appear to have any phytotoxic effects on the leaves of the plants. This would indicate an alternative cause for reduced vigor. Previous studies have shown that azadirachtin can have phytotoxic effects on plants (Chowdhury, et al.

2002). This reduction in leaf vigor among the soil treatment plants supports the theory that some of the neem oil solution was taken up by the plants’ roots. While this contradicts the results of the average percent of damaged leaves and leaf damage score for the soil treatment plants, it could provide evidence that, while not phytotoxic to the leaves of a plant directly, the soil treatment of neem oil can still reduce the vigor of a plant through other effects.

When looking at the average number of flowers per plant (Table 2, Figure 3), the foliar treatment reduced the number of flowers per plant much more than the soil treatment did. This pattern contrasts the pattern shown by the number of leaves per plant. The reduced transfer of nutrients in the foliar treatment plants due to their damaged leaves would result in fewer nutrients that could be devoted to plant reproduction. Increased leaf phytotoxicity has been shown to reduce crop yields among grape crops (Finger, et al. 2002). Additional measurements of the final crop yield among the treatment groups, such as number of fruits per plant and size of fruits, would have provided more accurate information on the effects of foliar-applied neem solution on the fruit maturity and crop yield of cucurbits. The soil treatment groups averaged about the same number of flowers as the control groups. This would indicate that soil treatments of neem oil do not have an effect on the number of flowers grown by the plants or the amount of nutrients that could be devoted to reproduction by the plant.

Throughout the observation period, the melon plants showed the greatest sensitivity to the foliar application of neem oil. On average, they had the greatest percent of leaves damaged

9 and damage score per leaf in the foliar treated groups throughout the experiment (Tables 3 and 4,

Figures 5 and 7). The squash, on the other hand, had the lowest sensitivity to the foliar treatment, with the lowest average percent of leaves damaged and damage score per leaf in the foliar treated groups throughout the observation period (Tables 3 and 4). These measurements support the hypothesis that the Hubbard winter squash would be a more ideal plant on which to use a neem- based foliar spray than the melons and zucchini, while other, less phytotoxic methods might be ideal for the Amish muskmelon.

In many experiments involving the foliar application of an oil-based pesticide to a plant, the amount of phytotoxic damage to a plant is positively related to the concentration of oil in the solution applied to the plant and the amount applied to the plant (Baudoin, et al. 2006; Finger, et al. 2002; Moran, et al. 2003). A balance between efficacy and phytotoxicity needs to be found when determining the concentration of oil to use in an oil-based pesticide solution. In an experiment involving the use of soybean oil as a pesticide against European red mites, soybean oil solutions of 4% and above had phytotoxic effects on apple trees, and left a residue on the leaves (Moran, et al. 2003). Sprays with a concentration below 4% still had significant effects in reducing mite population without having phytotoxic effects. The oil’s residue that was observed to remain on the leaves, which has also been observed in other experiments involving oil-based pesticides (Baudoin et al. 2006; Finger et al. 2002), is believed to be the cause of reduced phytotoxicity among plants treated with oil-based pesticides. During the foliar application of the neem oil solution in our experiment, an oily sheen was visible on the leaves. This layer of oil was less visible two days after the solution was applied, but still could have been present on the leaves and responsible for the high degree of phytotoxicity among the foliar-treated plants. A

10 reduction in the concentration of neem oil used in our solution would have likely reduced the phytotoxic effects of applying neem to the leaves.

The application of neem oil to plants in order to prevent against pests and infections is an increasingly popular practice among farmers. Both foliar and soil applications of different neem products have proven to be effective in many experiments. The foliar application of neem oil solutions causes a greater amount of phytotoxicity than soil applications, although the hydrophobic nature of neem oil may affect its root uptake in soil applications. Further studies are needed in order to determine which of these methods is more effective in and infection prevention.

Tables and Figures:

Table 1: Average number of leaves per plant (mean +/- SEM). Within a row, means followed by a different letter differ significantly from the control plants for that date, using Bonferroni adjusted alpha levels of 0.0167 per test

(0.05/3)

Control Foliar Soil F df P

Squash Day 33 5.13 ± 0.12 a 5.13 ± 0.19 a 4.40 ± 0.24 a 4.51 2,42 0.017

Day 41 8.60 ± 0.33 a 8.21 ± 0.52 a 6.20 ± 0.30 b 9.20 2,41 0.0005

Day 47 13.07 ± 0.85 a 13.57 ± 1.22 a 10.27 ± 0.63 a 2.94 2,41 0.064

Zucchini Day 33 6.53 ± 0.45 a 5.93 ± 0.28 a 5.00 ± 0.20 b 3.40 2,42 0.043

Day 41 10.47 ± 0.80 a 9.27 ± 0.27 a 9.60 ± 0.58 a 0.69 2,42 0.51

Day 47 17.21 ± 0.88 a 13.85 ±0.60 a 14.67 ± 0.94 a 3.23 2,40 0.05

Melon Day 33 3.73 ± 0.15 a 2.13 ± 0.22 c 2.87 ± 0.17 b 16.56 2,42 <0.001

Day 41 6.20 ± 0.19 a 4.67 ± 0.29 b 4.60 ± 0.34 b 9.32 2,41 <0.001

Day 47 11.07 ± 0.47 a 6.33 ± 0.49 b 3.69 ± 0.50 b 20.03 2,42 <0.001

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Table 2: Average number of flowers per plant (mean +/- SEM). Within a row, means followed by a different letter differ significantly from the control plants for that date, using Bonferroni adjusted alpha levels of 0.0167 per test

(0.05/3)

Control Foliar Soil F df P

Squash Day 33 0 0 0 ------

Day 41 0.53 ± 0.17 a 0.21 ± 0.15 a 0.067 ± 0.067 a 2.05 2,41 0.14

Day 47 3.80 ± 0.36 a 1.29 ± 0.27 b 3.33 ± 0.37 a 11.12 2,41 <0.001

Zucchini Day 33 0 0 0 ------

Day 41 0.87 ± 0.18 a 0.067 ± 0.067 b 1.07 ± 0.28 a 5.58 2,42 0.007

Day 47 3.14 ± 0.32 a 0.64 ±0.36 b 4.07 ± 0.41 a 18.75 2,40 <0.001

Melon Day 33 0 0 0 ------

Day 41 0.13 ± 0.12 a 0 b 0.27 ± 0.35 b 9.32 2,41 <0.001

Day 47 3.13 ± 0.32 a 1.07 ± 0.34 b 2.40 ± 0.39 a 6.90 2,42 0.003

Table 3: Average percent of leaves damaged per plant (mean +/- SEM). Within a row, means followed by a different letter differ significantly from the control plants for that date, using Bonferroni adjusted alpha levels of 0.0167 per test (0.05/3)

Control (%) Foliar (%) Soil (%) F df P

Squash Day 33 32.22 ± 3.60 a 52.89 ± 5.12 b 92.22 ± 4.26 c 39.64 2,42 <0.001

Day 41 26.61 ± 3.71 a 26.97 ± 2.32 a 19.05 ± 2.74 a 1.49 2,41 0.24

Day 47 15.32 ± 1.81 a 14.06 ± 1.67 a 8.34 ± 1.22 b 3.95 2,41 0.027

Zucchini Day 33 38.63 ± 3.04 a 68.94 ± 3.74 b 39.89 ± 3.95 a 18.34 2,42 <0.001

Day 41 16.78 ± 1.13 a 30.18 ± 2.71 b 16.32 ± 2.29 a 12.23 2,42 <0.001

Day 47 11.50 ± 0.90 a 17.87 ± 1.62 b 19.67 ± 3.47 b 3.18 2,40 0.052

Melon Day 33 25.0 ± 4.11 a 92.2 ± 4.26 b 31.0 ± 5.30 a 51.95 2,42 <0.001

Day 41 23.89 ± 1.99 a 33.44 ± 4.34 a 21.69 ± 2.30 a 3.68 2,42 0.034

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Day 47 13.46 ± 2.15 a 31.92 ± 7.41 a 16.57 ± 4.04 a 3.63 2,42 0.035

Table 4: Average leaf damage score per plant (mean +/- SEM). Within a row, means followed by a different letter differ significantly from the control plants for that date, using Bonferroni adjusted alpha levels of 0.0167 per test (0.05/3)

Control Foliar Soil F df P

Squash Day 33 0.79 ± 0.12 a 1.31 ± 0.19 a 0.88 ± 0.12 a 2.69 2,42 0.080

Day 41 0.54 ± 0.087 a 0.69 ± 0.10 a 0.40 ± 0.10 a 1.66 2,41 0.20

Day 47 0.51 ± 0.038 a 0.47 ± 0.063 a 0.48 ± 0.050 a 1.18 2,41 0.32

Zucchini Day 33 1.17 ± 0.11 a 2.38 ± 0.21 b 0.73 ± 0.21 a 26.47 2,42 <0.001

Day 41 0.54 ± 0.13 a 1.10 ± 0.10 b 0.37 ± 0.072 a 8.79 2,42 <0.001

Day 47 0.26 ± 0.031 a 0.68 ± 0.060 b 0.17 ± 0.038 b 12.44 2,40 <0.001

Melon Day 33 0.39 ± 0.072 a 3.46 ± 0.25 b 0.60 ± 0.13 a 95.51 2,42 <0.001

Day 41 0.53 ± 0.059 a 1.02 ± 0.19 b 0.42 ± 0.072 a 6.59 2,42 0.003

Day 47 0.30 ± 0.065 a 0.57 ± 0.11 a 0.23 ± 0.052 b 3.63 2,42 0.035

Figure 1: Average number of leaves per plant for zucchini plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

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18 SEM)

- 16 14 12 10 Control 8 Foliar 6 * Soil 4

2 # of Leaves/ Plant (Mean +/ Plant Leaves/ (Mean of # 0 33 41 47 Day #

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Figure 2: Average number of leaves per plant for melon plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

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12

SEM) - 10

8 * Control 6 * * Foliar * 4 Soil * *

2 # of Leaves/ Plant (Mean +/ Plant Leaves/ (Mean of # 0 33 41 47 Day #

Figure 3: Average number of flowers per plant for squash plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

4

4 SEM) - 3 3 2 Control 2 Foliar 1 * Soil 1

0 # of Flowers/ Plant (Mean +/ Plant (MeanFlowers/ of # 0 33 41 47 Day #

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Figure 4: Average percent of leaves damaged per plant for zucchini plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

80 - * 70 60 50 40 Control

SEM) * 30 Foliar * 20 * Soil 10

0 % of Leaves Damaged/ Plant (Mean +/ of % Plant LeavesDamaged/ 33 41 47 Day #

Figure 5: Average percent of leaves damaged per plant for melon plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

120 -

100 *

80

60 Control

SEM) Foliar 40 Soil 20

0 % of Leaves Damaged/ Plant (Mean +/ of % Plant LeavesDamaged/ 33 41 47 Day #

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Figure 6: Average leaf damage score per plant for zucchini plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

3 - * 2

2

1 Control SEM) * Foliar 1 * Soil 0 * Leaf Damage Score/ +/ Plant(MeanDamage Leaf Score/ 0 33 41 47 Day #

Figure 7: Average leaf damage score per plant for melon plants. Bars marked with an asterisk differ significantly from the control plants for that date, using Bonferroni adjusted levels of 0.0167 per test (0.05/3)

4 - * 3 3 2

2 Control SEM) 1 Foliar * 1 Soil

0 * Leaf Damage Score/ +/ Plant(MeanDamage Leaf Score/ 0 33 41 47 Day #

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