Mechanisms Underlying Intercropping Success in the Field 

In many gardening circles, some of the more seasoned practitioners will recommend to novice gardeners the technique of intercropping their plants to assist their primary plant in staying alive and healthy. Intercropping entails combining the desired plant and planting them alongside an herbaceous plant, which act as the ‘sacrificial’ plant (They are sometimes referred to as companion plants), which either deter or attract insects towards these plants instead. This technique is relatively easy to implement too, allowing any novice gardening practitioner a relatively simple and natural way to deter pests from their gardens. 

For the experiment, we acquisitioned 3 species of seeds from the “Back to Roots”, (BTR), brand for consistency. Solanum lycopersicum var. Cerasiforme or Red Cherry tomatoes, Ocimum basilicum or Sweet Basil and Anetheum graveolens or Mammoth Dill seeds. The S. lycopersicum seeds came from three BTR envelopes, both consisted of 100mg of seeds from Lot A and packaged in 2025. One envelope of BTR O. basilicum seeds weighing 300mg of seeds were from lot B and were packaged in 2025. One envelope of BTR A. graveolen seeds weighed in at 1g and were from lot A and packaged in 2025. The seeds were germinated by surface sterilizing for 30 minutes in a 75% water and 25% bleach solution. Seeds were then placed individually into cell trays (10.6 cm wide by 9 cm tall by 14.3 cm long) filled with General Pro-Mix from (BX, Canada). (N =110 S. lycopersicum seeds, N =120 A. graveolen seeds and N =130 O. basilicum seeds for a goal of 100 total seedlings to transplant to the field). 

Planted seeds from all three species were placed into an incubator fridge for approx. 3 weeks. Temperature was maintained at 26 degrees celsius and lights were set to be in between the hours for 6am -9pm. Seeds were placed in cell trays and bottom watered minimally on a daily basis. After the incubation period, sprouts were moved out to begin hardening them for the transplantation into the field plot. To harden the seedlings, we took them into a shaded area first, moving them as needed before placing them outside with more direct sunlight over the course of 10 days. Sprouts were then placed into the exterior growing area with a protective netting placed over them. Once they were sufficiently hardened, we planted the seedlings into the field plot. A split plot layout was utilized to section off the field; 4 Rows were utilized with 18 trial specimens per row Each row was separated by 2.5 meters. Rows were labeled with associated specimen information such as species being tested and associated specimen ID groups.  All plants were watered initially to help with transplanting success. 

Field Plot 

The field plot measured 20 meters by 11 meters and was tilled prior to planting. Once tiled, garden fencing (71.12cm x 15.24m ,Everbilt, Vietnam) was added to exclude larger vertebrate herbivory. Rows were separated representing trial type, (ie. whether they are in a control block or a companion block). This was done in a randomized order. Control group rows(rows 2 and 4), where only the S. lycopersicum was present, had specimens separated by 1 meter between each specimen. For sections with intercropped specimens (Rows 1 and 3), each specimen was separated by 1 meter with companion plants being grouped within 10-14cm of the focal S. lycopersicum. Rows 1 and 3 consisted of test specimen groups, with specimens 1-9 and 47-54 which had S. lycopersicum intercropped with both O. basilicum and A. graveolen. Specimens 10-18 in row 1 and 37- 45 in row 3 consisted of S. lycopersicum paired with O. basilicum. Rows 2 and 4 were exclusively solo S. lycopersicum specimens, 19-36 in row 2 and 55-72 in row 4. If any specimens were damaged or destroyed by outside factors (ie, Birds) they were replaced and provided a new ID. this was done during the first 3 weeks of the experiment. Specimens that died after this period were deemed as such on the report.

Field plants measurements

For the purpose of identifying differences in S. lycopersicum growth in solo and companion groups, we took health measurements every three days, measuring each plant’s growth, and damage measurements on the focal S. lycopersicums. Growth included height which was measured to the nearest 0.1 cm, stem diameter which was measured to the nearest 0.01 mm, leaf number (and leaflet number during early trial period), flowering number and fruiting number. 

Health evaluation was determined by the number of damaged leaves, number yellowing leaves, and visually monitoring for insect damage. We also measured the pH levels and general dryness of the soil once a week utilizing a soil meter (IRTOV, China). For Leaf health, we utilized a chlorophyll meter from the (GOYOJO, China) to measure chlorophyll In SPAD, Nitrogen levels in mg/g, leaf humidity in RH% and the leaf’s temperature in celsius. 

To determine if there was a correlation between the solo planted S. lycopersicum and the companion planted ones, we observed the number of insects that interacted with individual plants, taking account of how many insect species interacted with them along with the number of individuals within a specific species (ie. number of Manduca sexta larvae or Hornworms). Leaves damaged by insect activity were identified and counted thrice weekly, Mondays, Wednesdays and Fridays, until the end of the trial period.

Oviposition Assay

To test if the presence of companion plants affected the oviposition of M. sexta, we conducted a test wherein we placed a test cage consisting of 2 tomato plants, one alone and one grouped with Basil and Dill. These groups were separated by ~30.5cm n apart, splitting the cage into 2 parts, the lone tomato plant side and the companion plants side. The Basil, tomato, and Dill  on the companion plant side were separated from each other by ~5-10cm, depending on sizes of plants. A food cup and paper towel to assist the M. sexta in returning onto the wall was placed at the center of the cage and that of other trial cages. This cage tested how the visual presence of the companion plants affected how the M. sexta in their choice of egg laying environment.

To determine how neighboring plants affect how M. sexta determine what is a viable location for laying their eggs, we tested how plant volatiles from other plants affect oviposition on host plants. This was done by having a cage set up with 2 tomato plants on the interior of the cage on opposite sides, ~30.5cm apart. One side was a lone tomato plant while the opposite side had the companion plants, Basil and Dill, sitting on the outside of the cage against the mesh wall. We added an additional paper towel layer (may be adjusted to use cheese cloth or other permeable material) to further reduce visibility of these plants. In both tests, the female was removed from the cage after a 24h period and the number of eggs deposited on each plant were counted and allowed to hatch in lidded 4oz cups to verify the female had mated. Females that did not survive the assay period or did not mate were discarded, with the total limit of N = 1 moth per trial had laid fertilized eggs during the oviposition choice test. To determine if eggs were fertilized, we waited for them to hatch for a period of 9 Days. If they had not hatched by the 9th day, the eggs were discarded.

In a controlled environment with isolated cues, basil and dill deterred M. sexta oviposition. This is likely due to dilution of the tomato host plant visual and volatile cues in the presence of the other companion plants. However, in the field plot, basil and dill did not protect from leaf herbivory. This indicates in more complex settings, presence of basil and dill not enough to prevent leaf damage.  Future studies may look at how larger scale assays may be able to utilize companion plants in a more effective manner, potentially in individual pots rather in than in a field to reduce inconsitansies with soil quality and water retention.