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© 2006 Plant Management Network.
Accepted for publication 9 May 2006. Published 18 July 2006.


Managing Botrytis Gray Mold in Greenhouse Tomatoes Using Traditional and Bio-Fungicides


David M. Ingram, Central Mississippi Research & Extension Center, Mississippi State University, 1320 Seven Springs Road, Raymond 39154; and Charles W. Meister, IR4 Regional Field Coordinator, FSHN, IFAS, University of Florida, P.O. Box 110720, Gainsville 32611


Corresponding author: David M. Ingram. davidi@ext.msstate.edu


Ingram, D. M., and Meister, C. W. 2006. Managing Botrytis gray mold in greenhouse tomatoes using traditional and bio-fungicides. Online. Plant Health Progress doi:10.1094/PHP-2006-0718-01-RS.


Abstract

Eight fungicide products were evaluated over a three-year period to compare their efficacy in managing Botrytis gray mold in greenhouse tomatoes. Disease severity ratings indicated that STBX-016, Decree 50 WG, and Serenade ASO significantly reduced gray mold in two of three years as compared to the inoculated control. Milsana significantly reduced gray mold severity in only one of three years. Pristine and Switch reduced disease severity in the one year tested but Scala and Prestop did not. Switch, Milsana, and STBX-016 significantly increased marketable yield per plant in one of three years, as did Decree 50 WG in two of three years.


Introduction

Gray mold, caused by Botrytis cinerea, is probably the most ubiquitous disease of greenhouse tomato (5) and also affects many ornamental crops grown in greenhouses (3). There are approximately 850 acres of greenhouse-grown tomatoes in the United States (1). Greenhouse tomatoes account for approximately 37% of all fresh market production in the United States (Mike Bledsoe, personal communication). In Mississippi seedlings are started in a propagation house for subsequent transplanting in the production greenhouse after six weeks of growth (7). Plants are grown in a variety of media including ground pine bark, perlite, rice hulls, and sand. These media are usually contained within plastic bags that lay in the drainage canals or upright plastic bags in the row. Typical plant arrangement consists of 2 to 4 plants per bag or one plant per 4 ft2 of floor space. Most growers begin transplanting tomato seedlings into greenhouses near mid-October. During November through March, temperatures in Mississippi are cool enough to require supplemental heating. During this period, greenhouses are closed to conserve heat and venting outside air with lower humidity into the greenhouse is seldom practiced. Humidity levels are commonly 90% or greater for most of the day and night. Freestanding water is also common in many operations. These environmental conditions are ideal for the initiation and proliferation of gray mold (4). In addition, many days during this period are cloudy and rainy, resulting in an inability to maintain dry canopy conditions. Greenhouse tomatoes tend to grow rapidly, producing great amounts of lush, succulent tissue, which shades the lower plant canopy and results in highly susceptible plant tissues.

Gray mold affects all aboveground plant parts (5). The most characteristic sign is the fuzzy, gray-brown mold growth on plant parts (Fig. 1). Fruiting structures of the fungus are easily observed with a hand lens. Gray mold often affects lower stems of greenhouse tomatoes resulting in rapid wilting and plant death (Fig. 2). Another aspect of gray mold is the occurrence of "ghost spots" on fruit (Fig. 3). The spores germinate and penetrate the fruit surface, but infection aborts and leaves a necrotic fleck surrounded by a whitish halo.


 

Fig. 1. Botrytis gray mold on greenhouse tomato leaf.

 

 

Fig. 2. Botrytis gray mold lesion on greenhouse tomato stem.

 

Fig. 3. Botrytis ghost spots on greenhouse tomato fruit.


Currently, no greenhouse tomato cultivars exhibit resistance to gray mold. Cultivars commonly grown in Mississippi include ‘Trust’ and ‘Match’; these routinely become infected with gray mold, and losses can be significant if control measures are not implemented (D. Ingram, unpublished data). Chemical control for greenhouse tomatoes is limited to use of two fungicides: dichloran (Botran), for which restrictions limit use in greenhouses; and fenhexamid (Decree), which has a supplemental label for the control of gray mold in the United States and Canada. Chlorothalonil (Exotherm Termil) was formerly labeled for use on greenhouse tomatoes, but the new formulation can only be used on non-food crops.

Reduced efficacy of dichloran and chlorothalonil against gray mold has been observed in Mississippi in recent years and development of strains of Botrytis resistant to commonly used fungicides in greenhouse ornamental crops has been reported (6,9). A number of bio-fungicides (synonymous with biological control products) have recently been developed, but their efficacy against gray mold has not been evaluated. Several biofungicide products approved for organically grown tomatoes are effective against early blight (8) and may be effective tools for managing Botrytis gray mold in greenhouse tomatoes.

The objective of this research was to determine the efficacy of traditional fungicides and bio-fungicides used alone or in combination for the control of Botrytis gray mold in greenhouse tomatoes.


Fungicide Efficacy Trials

All experiments were conducted at the Mississippi State University Truck Crops Experiment Station located in Crystal Springs, MS. The greenhouse (24 × 96 ft) is set up to carry 24 rows (12 on each side oriented east-west), each with six, 7.5-gallon plastic bags filled with perlite. Seeds of the greenhouse tomato cultivar Trust (De Ruiter Seeds, Inc., Lakewood, CO) were germinated in a commercial potting mix (Redi-Earth, Marysville, OH) in 72-cell pack trays placed on a bench in a propagation house. After six weeks of growth two seedlings were transplanted into each perlite bag in the greenhouse in early November of 2002, 2003, and 2004. Treatments were arranged in a randomized complete block design with four (2004 and 2005) or six (2003) replications. Plants were fertilized by an injection system (H. E. Anderson Company, Muskogee, OK) that employed two plastic 55-gal drums each with specific fertilizer components (Total Gro, Winnsboro, LA) and calcium nitrate. Initially, plants were fertigated three times per day. Watering was increased to six times per day by the end of the growing season to accommodate plant needs. Greenhouse controls were set to maintain a minimum temperature of 65°F and a maximum temperature of 87°F.

The Botrytis cinerea isolate used in this study was collected from a greenhouse tomato sample exhibiting symptoms of gray mold. Inoculum was produced on half strength potato dextrose agar (Difco, Becton Dickinson Microbiology Systems, Sparks, MD). Petri plates (3.5-inch diameter) with Botrytis were incubated on the laboratory bench at approximately 72°F with a 12-h photoperiod for 14 days. To obtain inoculum, Botrytis cultures were flooded with sterile water containing several drops of Tween 20 per quart (Fisher Scientific Company, L.L.C., Houston, TX), and spores were dislodged using a bent glass rod. The resulting suspension was strained through two layers of cheesecloth and the spore concentration was adjusted to 3 × 106 spores/oz.

After transplanting in the greenhouse, plants were grown for about four weeks to the stage of 4 to 5 flower clusters per plant. Plants then were tied to trellising. Some lower leaves had been pruned, which resulted in wounds in the stems, and provided infection courts for germinating spores of Botrytis. Plants were inoculated by initially spraying with a spore suspension (2003 and 2004) and approximately one week later, placing one open petri dish containing spores of Botrytis under the canopy of plants in the center of each row (2003-2005). Natural airflow and splashing water from an overhead sprinkler system disseminated these spores (3).

All fungicides were mixed and applied in distilled water using a 1-quart hand-held commercial sprayer (Selig Commercial, Cartersville, GA). Application rates were calculated on basis of per acre rates suggested by each company (Table 1). There was no non-inoculated control treatment because gray mold could not be prevented in control plants placed randomly among inoculated plants in the greenhouse. Fungicides were first applied just prior to inoculum application, and additional applications were made as noted in Table 1. Fungicides were applied to the point of runoff. Inoculated controls received distilled water only.


Table 1. Application rates and times for products evaluated for the control of Botrytis gray mold in greenhouse tomatoes, Crystal Springs, MS, 2003-2005.

Treatment
(and source)
Rate
(per stated unit)
Application dates
2003 2004 2005
Copper sulfate pentahydrate (STBX-016, Revised formulation of Phyton 27, Phyton Corporation, Bloomington, MN) 20 fl oz in 100 gal/acre January 23
February 7
March 18
January 30
February 13
March 1
February 15
March 1
March 8
March 15
Fenhexamide
Decree50 WG, Arysta LifeScience Corporation, Cary, NC)
4 lb/acre January 23
February 7
March 18
January 30
February 13
March 1
Bacillus subtilis strain QST 713
(Serenade ASO, AgraQuest Inc., Davis, CA)
6 qt/acre January 23
February 7
March 18
January 30
February 13
March 1
February 15
March 1
March 8
March 15
Extract of Reynoutria sachalinensis
(Milsana, KHH Biosci Inc., Raleigh, NC)
1% solution in 50 gal/acre January 23
February 7
March 18
January 30
February 13
March 1
February 15
March 1
March 8
March 15
Pyrimethanil
(Scala, Bayer CropScience, Research Triangle Park, Raleigh, NC)
27 fl oz/acre January 23
February 7
March 18
Cyprodinil and fludioxonil
(Switch 62.5 WG, Syngenta Crop Protection Inc., Greensboro, NC)
4 lb/acre January 23
February 7
March 18
Boscalid and pyraclostrobin
[BAS 516 (Pristine), BASF Corporation, Research Triangle Park, Raleigh, NC]
0.64 lb/acre January 23
February 7
March 18
Gliocladium catenulatum
(Prestop, AgBio Inc., Westminster, CO)
1% solution (w/v) January 23
February 7
March 18
January 30
February 13
March 1
-
Inoculated control Spray entire plant to runoff with 3 × 106 spores/oz of Botrytis cinerea (2003 and 2004) or open petri plates with actively sporulating Botrytis cultures (2003-2005) January 24
February 2
March 25
(spore suspension applied on first date with petri plates used on next two dates)
February 3
February 9
March 16 (spore suspension applied on first date with petri plates used on next two dates)
February 15 (only petri plates of Botrytis cultures used

Disease severity was determined by visual estimates of percentages of plant tissue with gray mold symptoms and ranged from 0 (no symptoms) to 100 (entire plant symptomatic). Disease severity was rated multiple times and the area under the disease progress curve (AUDPC) was calculated using midpoints of the visual rating scale (2). Cull fruit weight and total fruit weight were determined in all three years, and incidence of ghost spots was determined in 2004, but data are not presented due to lack of statistical significance.

All data were subjected to analysis of variance using SAS version 9.1 for PC (SAS Institute Inc., Cary, NC). Fisher’s Protected Least Significant Difference Test (P = 0.05) was used to separate means.


Effect of Fungicides on Gray Mold in 2003

Disease severity ratings were made on 14 April, 29 April, and 29 May, 2003. STBX-016, Milsana, Decree 50 WG, Serenade, Switch 62.5 WG, and BAS 516 significantly reduced the severity of gray mold as compared to the untreated, inoculated control (Table 2). Scala did not reduce severity of gray mold. AUDPC data indicated that STBX-016 and Decree 50 WG were equally effective in slowing the progression of gray mold. Marketable fruit yield was significantly improved when plants were treated with STBX-016, Milsana, Decree 50 WG, and Switch 62.5 WG.


Table 2. Disease severity ratings, area under the disease progress curve and marketable yield of ‘Trust’ greenhouse tomatoes treated with fungicides, 2003.

Treatmentw Gray mold
(%)x
Gray mold
AUDPC
y
Marketable yield
(lb/plant)
STBX-016         15.0 az          204 e          11.8 a
Milsana         26.7 a          349 cd          11.4 a
Decree 50 WG         16.7 a          187 e          11.2 ab
Switch 62.5 WG         26.7 a          347 cd          11.4 a
Serenade         20.0 a          232 de          10.7 abc
Scala         53.3 c          496 ab          10.7 abc
Bas 516 (Pristine)         40.0 b          390 bc          10.0 bc
Inoculated control         53.3 c          535 a            9.6 c

 w See Table 1 for application rates and schedules.

 x Plants within a plot were visually rated for percentages of tissues that exhibited gray mold symptoms. 0 = no symptoms and 100 = entire plant symptomatic. Data presented are from the final severity rating in the trial.

 y AUDPC (area under the disease progress curve) calculated using midpoints of the visual rating scale.

 z Means in a column followed by the same letter are not significantly different according to Fisher’s Protected Least Significant Difference Test (P = 0.05).


Effect of Fungicides on Gray Mold in 2004

Disease severity ratings were made on 30 March, 6 April, 20 April, 29 April, and 5 May 2004. STBX-016, Decree 50 WG, and Serenade each significantly reduced the severity of gray mold as compared to the untreated, inoculated control (Table 3). Only STBX-016 and Decree 50 WG significantly slowed the development of gray mold based on AUDPC. Decree 50 WG was the only product that significantly increased marketable fruit yield in 2004. Ghost spot (Fig. 3) was prevalent in 2004. Number of symptomatic fruit and ratings of severity of ghost spot were the same across all treatments.


Table 3. Disease severity ratings, area under the disease progress curve and marketable yield of ‘Trust’ greenhouse tomatoes treated with fungicides, 2004.

Treatmentw Gray mold
(%)x
Gray mold
AUDPC
y
Marketable yield
(lb/plant)
STBX-016          17.5 b          389 b          9.5 ab
Milsana          26.3 ab          405 ab          8.7 b
Prestop          23.8 ab          439 a          9.8 ab
Decree 50 WG          18.8 b          257 b        10.6 a
Serenade          18.8 b          410 ab          9.6 ab
Inoculated control          30.0 a          533 a          8.4 b

 w See Table 1 for application rates and schedules.

 x Plants within a plot were visually rated for percentages of tissues that exhibited gray mold symptoms. 0 = no symptoms and 100 = entire plant symptomatic. Data presented are from the final severity rating in the trial.

 y AUDPC (area under the disease progress curve) calculated using midpoints of the visual rating scale.

 z Means in a column followed by the same letter are not significantly different according to Fisher’s Protected Least Significant Difference Test (P = 0.05).


Effect of Fungicides on Gray Mold in 2005

In 2005, STBX-016 was used as a standard and was alternated with either Serenade or Milsana. Disease severity ratings were made on 8 March, 15 March, 23 March, 28 March, 4 April, 13 April, 19 April, and 29 April 2005. No significant differences were observed for any of the factors measured in this study (Table 4). White mold (Sclerotinia sclerotiorum) occurred sporadically throughout the greenhouse resulting in random plant death, which complicated the ratings on gray mold. Ratings of the percentage of leaf area affected by Botrytis ranged from about 8 to 20%. None of the control products significantly reduced the percentage of leaf area with gray mold symptoms.


Table 4. Disease severity ratings, area under the disease progress curve and marketable yield of ‘Trust’ greenhouse tomatoes treated with fungicides, 2005.

Treatmentw Gray mold
(%)x
Gray mold AUDPCy Marketable yield (lb/plant)
STBX-016 20.0 a          630 a          5.3 a         
Milsana 13.8 a          566 a          6.2 a         
Serenade 15.0 a          569 a          6.5 a         
STBX-016 alternated
with Serenade
7.5 a          550 a          6.0 a         
STBX-016 alternated
with Milsana
16.3 a          488 a          6.5 a         
Inoculated control 15.0 a          663 a          5.3 a         

 w See Table 1 for application rates and schedules.

 x Plants within a plot were visually rated for percentages of tissues that exhibited gray mold symptoms. 0 = no symptoms and 100 = entire plant symptomatic. Data presented are from the final severity rating in the trial.

 y AUDPC (area under the disease progress curve) calculated using midpoints of the visual rating scale.

 z Means in a column followed by the same letter are not significantly different according to Fisher’s Protected Least Significant Difference Test (P = 0.05).


There appeared to be a trend where the untreated, inoculated control plants produced lower marketable fruit yields than plots treated with fungicides. The greatest increases in marketable fruit yield were observed with Serenade applied alone or STBX-016 alternated with Milsana.


Summary

Few fungicide products are available for management of Botrytis gray mold on greenhouse tomatoes. STBX-016 is a copper-based fungicide which is a revised formulation of Phyton 27. Serenade uses a strain of Bacillus subtilis as a bio-control agent and Milsana is an extract from the giant knotweed plant, which induces plant resistance. These bio-pesticide products were shown to be effective against gray mold on greenhouse tomatoes. In addition, STBX-016 and Milsana were shown to increase marketable fruit yield in 2003. STBX-016, Milsana, and Serenade were tested in all three years. These products would be beneficial for use in an integrated disease management program for greenhouse tomatoes.


Acknowledgments

This work was funded by IR4 grant appropriations to Mississippi State University, Central Mississippi Research and Extension Center. Phyton Corporation, Arysta LifeScience Corporation, AgraQuest Inc., Khh Biosci Inc., Bayer Crop Science, Syngenta Crop Protection Inc., BASF Corporation, and AgBio Inc. each donated product. We gratefully acknowledge the pre-submission review provided by Maria Tomaso-Peterson and Robert G. Pratt.


Literature Cited

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