Introducing Beneficial Entomopathogenic Nematodes for Biological Control and Enhanced Plant Resistance to Improve Pest Management in Cucurbit Crops

Materials and Methods

This project was conducted at Ronin Farm, which is a 15-acre diversified vegetable farm located in Bryan, TX. This project was developed in close collaboration with the owner, Mr. Brian Light, who expressed strong interest in improving pest management on his farm through sustainable strategies and incorporating biological control with beneficial nematodes as part of his management plan. Mr. Light selected the plant varieties for this project based on previous experience with produce quality and plant performance in Brazos county, TX. The entomopathogenic nematodes selected for this project are Heterorhabditis bacteriophora and Steinernema riobrave, which are suitable for southern climates and cucumber beetle control (Arbico Organics, USA). 

This project was conducted over two field seasons in 2021 and 2022, using a randomized complete block design, based on a previous EPN study (Ellers-Kirk et al., 2000). Each block was divided into 2 rows and each row received one treatment: 1) plants treated with EPNs and 2) untreated control plants. Blocks were replicated 4 times each for cucumbers, watermelon or bitter melon, and summer squash. Our plots were planted in April of each year (4/20/21 and 4/27/22) and plants were grown and maintained following typical cultivation practices on Ronin Farm. Seeds were planted directly into raised rows and thinned as needed. For cucumber and bitter gourd, we planted ~4 m long rows with 10 plants each, spaced 30 cm apart. For watermelons, we planted ~9 m long rows with 5 plants each, spaced 1.5 m apart. For Summer squash, we planted ~7 m long rows with 10 plants each, spaced 60 cm apart. Crops were adjacent, and all rows were 1 m apart. Cucumber and bitter gourd vines were trained onto trellises to promote airflow to leaves and facilitate easier surveys and harvest. Rows were covered with straw mulch to suppress weeds and retain moisture and plants were irrigated as needed.

EPN treatments were applied twice per year to trigger plant resistance and target soil-dwelling pests like cucumber beetles. Plants were treated with EPNs at 1 true leaf (~2 weeks after planting) and when they began to flower and set fruit (~6 weeks). Each treatment plant received ~200,000 free-living EPNs in 100 mL water and control plants each received 100 mL water added to soil at the plant base.

Insect surveys and damage assessments

To quantify numbers of pest insects, we conducted weekly visual surveys of all experiment plants from germination until final harvest (Skidmore et al., 2019). Each plant was observed for 60 seconds to search for insects. If detected, insects were sight identified (to species, family, or order as possible) and counted. Examples of insects in surveys included striped (Acalymma vittatum), spotted (Diabrotica undecimpunctata howardi), and banded (Diabrotica balteata) cucumber beetles, melon aphids (Aphis gossypii), squash bugs (Anasa tristis), squash vine borers (Melittia cucurbitae), leaf footed bugs (Leptoglossus zonatus), and silverleaf whiteflies (Bemisia tabaci). We also assessed cucumber beetle control using traps to monitor adult beetle emergence from soil. Traps (25 cm², 12 cm tall) were placed on the soil surface between plants in each row (4 per row for each plant species) 5 weeks after planting, allowing time for any adult beetles to lay eggs, and were monitored twice weekly for 4 weeks (Ellers-Kirk et al., 2000). 

During surveys, we also quantified numbers of beneficial insects present on plants to determine whether EPN introductions influenced the beneficial insect community. Examples of insects surveyed (sight identification) included pollinators like honey bees (Apis mellifera), squash bees (Peponapis pruinosa), and other native bees (Osmia, Agapostemon, Bombus, Megachile sp., etc.), as well as natural enemies like wasps (e.g. Ichneumonidae, Braconidae, Vespidae, etc.), flies (Syrphidae), and beetles (e.g. Coccinellidae, Carabidae, etc.).

In separate weekly surveys, we estimated % damage (insect or pathogen) area on leaves, flowers, and fruits.

Plant growth and yield data

To determine the effects of EPN introduction on plant biomass, we quantified root length density following previously described methods (Ellers-Kirk et al., 2000). Briefly, soil cores were taken near plant stems during the final week of harvest and cm root length per mm soil was calculated. This was replicated three times per row for each plant species. We also recorded plant heights, numbers of leaves, and numbers of male and female flowers.

Yield data were collected throughout the season as fruits were harvested and total weights were recorded.

Data analysis

Data were analyzed using the software program R. Data on insect numbers, damage estimates, plant growth, and yield were analyzed using generalized liner mixed models.