A novel technique for treating seeds with biocontrol agents for the sustainable management of bacterial fruit blotch of watermelon

View the project final report

• 2014 annual report

Commodities

• Fruits: melons
• Vegetables: cucurbits

Practices

• Crop Production: biological inoculants
• Pest Management: biological control

Watermelon seed treatments with biocontrol agents that are antagonistic against Acidovorax citrulli significantly reduced bacterial fruit blotch (BFB) seedling transmission. 10⁸ CFU/flower was the optimal inoculation concentration for seed treatment. When watermelon seeds treated with biocontrol agents and PBS (negative control) were challenged with 10⁷ CFU/ml of A. citrulli the mean BFB seedling transmission was 80%. By comparison, seedlots from flowers treated with Bacillus mojavensis RRC101 and Bacillus spp. #24 and #35, had significantly lower BFB seedling transmission percentages of 19.4%, 53.2%, and 41.7%, respectively. These results demonstrate the potential for seed treatments with biocontrol agents to improve BFB management.

Introduction

The main goal of this project was to optimize a technique for applying biocontrol agents to watermelon seeds to reduce the seed-to-seedling transmission of bacterial fruit blotch of cucurbits (BFB). Acidovorax citrulli is the causal agent of BFB, a sporadic but devastating threat to watermelon fruit production worldwide. Outbreaks of this disease can result in severe fruit rot and significant economic losses (Burdman and Walcott, 2012). Infested cucurbit seeds are the primary source of inoculum for BFB outbreaks that develop in seedling transplant facilities and fruit production fields. Currently there are no commercially available sources of BFB resistance (Bahar and Burdman, 2010). Therefore disease management relies heavily on foliar-applied chemicals (e.g. Kocide, Mankocide, Nu-Cop) and pathogen exclusion. However, the efficacy of foliar applied chemicals can be limited under conditions of high rainfall and relative humidity.

A. citrulli exclusion relies on production of pathogen-free seeds in regions of countries with cool and dry climates that are unfavorable for BFB development. Despite these efforts, infested watermelon seed remain the most important source of inoculum for BFB outbreaks. One reason for the lack of effectiveness of seed treatments is that A. citrulli cells can be located in the embryos of watermelon seeds where externally applied chemicals cannot penetrate (Dutta et al., 2012). Additionally, seeds containing as little as 10³ colony forming units (CFU) can result in BFB seed-to-seedling transmission under transplant house conditions (Dutta et al., 2012). Hence, an effective treatment must penetrate deep into the seed to eliminate resident bacterial cells.

One solution to this problem is to deposit biocontrol agents into internal tissues of watermelon seeds. On seed germination, the antagonists would rapidly colonize seed tissues and prevent the establishment of A. citrulli populations. Ideally, this would reduce BFB seed-to-seedling transmission and limit BFB outbreaks. Using recombinant DNA techniques, we generated a non-pathogenic strain of A. citrulli (AAC00-1ΔhrcC) that colonized germinating watermelon seedlings at wild-type levels (Johnson et al., 2011). When applied externally to naturally infested watermelon seeds, AAC00-1ΔhrcC significantly reduced BFB seed-to-seedling transmission (Johnson et al., 2011). To improve the efficacy of biocontrol seed treatment, we propse to deposit biocontrol bacteria within seeds by applying cell suspensions to the stigmas of female watermelon flowers. A. citrulli can penetrate the pistil of the female flowers and become deposited in the seed endosperm without inducing BFB symptoms on fruits (Walcott et al., 2003). By exploiting the “pistil invasion pathway”, we hypothesize that biocontrol agents will be more effective in limiting BFB seedling transmission.

The main objective of this study was to optimize a flower inoculation technique for efficiently delivering biocontrol bacteria into watermelon seeds. The specific objectives were:

1. To optimize the concentration of bacteria applied to watermelon flowers to maximize seed infestation.

2. To determine the efficacy of the flower treatments with biocontrol agents in reducing seed-to-seedling transmission of BFB.