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This plant communication apparatus was first employed by Gagliano et al. (2013) to study how chili plants (Capsicum annuum, Solanaceae) distinguish between adult conspecifics and fennel plants, even when common signaling pathways were blocked.
The experimental setup prevents both above- and below-ground contact as well as chemical and light-mediated signals typically exchanged by plants. This allows researchers to explore alternative signaling methods or control specific signaling pathways.
In this setup, chili seeds are arranged in a circle around an adult plant within a sealed central cylindrical box. Seeds and adult plants in each unit are housed within two nested square boxes. The air between these two boxes is removed using a vacuum pump (not included) to prevent interference from adjacent experimental units.
The apparatus is made of colorless cast acrylic material (Moden Glas), which transmits 92% of visible light while being opaque to ultraviolet and infrared wavelengths. This ensures the exclusion of specific wavelengths and prevents unwanted signals between experimental units.
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Communication research has primarily concentrated on animals, whose interactions often involve auditory or visual displays that attract attention. Plant communication, although a more controversial topic, has garnered increased interest recently. Accepted mechanisms of plant communication include chemical signals, physical contact, and light-based signals, while others remain debated (Karban, 2008). Numerous studies have shown that plants are highly sensitive organisms, capable of interacting and facilitating each other by actively gathering information from their environment. They adjust their behavior based on this information about their neighbors, both above and below ground, and the resources available in their surroundings. Similar to animal social systems, plants engage in kin selection by favoring relatives over strangers, thus minimizing competition (Gagliano et al., 2012).
Plants use volatile organic compounds (VOCs) as warning signals to alert neighboring plants of herbivore threats, prompting them to bolster their defenses. This air-borne communication allows undamaged plants to prepare for potential attacks. Additionally, plants can detect variations in sunlight wavelengths to anticipate future competition, enabling them to adjust their growth and defense strategies in response to the availability of resources (Gagliano et al., 2012).
Plant communities are shaped by both competition and facilitative interactions. Facilitation includes processes like improving growth, nitrogen fixation, nurse cropping, pest control, and attracting beneficial organisms such as insects or mycorrhizae. These interactions often rely on mechanisms involving light and shade effects, chemical signals, and physical proximity. For instance, shade from perennial canopies can protect seedlings and smaller plants from extreme temperatures and water loss while enhancing soil conditions. Recent research highlights various examples of plant associations with benefactor species that offer protection through visual or auditory concealment, shielding them from herbivore predation (Gagliano et al., 2013).
Research Studies
In a recent study by Gagliano et al., it was demonstrated that plants can detect and influence their neighbors through mechanisms beyond light, chemical signals, or physical contact. The research involved chili plant (Capsicum annuum) seeds and examined their germination rates in the presence of an adult chili plant and an adult fennel plant (Foeniculum vulgare). Fennel plants are known to inhibit chili seed germination by emitting volatile chemicals. However, the study found that chili seed germination rates were higher when the signals from the fennel plant were partially or completely blocked.
Another study by Gagliano et al. investigated the effect of a beneficial neighbor, the basil plant (Ocimum basilicum), on chili seed germination rates. Basil plants positively influence chili seeds by increasing germination rates, likely due to their production of volatile compounds that act as natural insecticides and suppress competitive weeds. This is a common practice among gardeners who grow basil alongside chili plants, as basil helps maintain soil moisture and serves as a living mulch. The study found that basil plants enhanced chili seed germination rates even when the basil was enclosed in a sealed box, indicating the presence of unidentified signaling modalities beyond chemical signals.
Florence fennel plant (Foeniculum vulgare) will serve as one of the heterospecific neighbors (negative influence) due to its production of growth-inhibiting chemicals from both its roots and aerial parts, necessitating its isolation. Basil plant (Ocimum basilicum) will be used as the other heterospecific neighbor (positive influence) for its production of insecticidal chemicals that suppress weed growth. The experiment will monitor the germination rates of chili plant seeds (Capsicum annuum) in response to these neighbors.
Establish a Controlled Environment Room (CER) spanning 5.30 m², equipped with high-density discharge lamps. Maintain daytime temperatures at 18°C and nighttime temperatures at 13°C. Ensure consistent nutrient exposure, temperature conditions, and a 12-hour light:12-hour dark cycle for all seeds and plants involved. Place chili seeds within layers of 2 mm thick felt in petri dishes to preserve moisture and maintain darkness. Monitor and water the seeds every 24 hours as needed.
Set up 15 experimental units individually in the plant communication chamber. Each experimental unit consists of 8 petri dishes with 20 chili seeds in each petri dish. Arrange the petri dishes at a distance of c.10 cm from each other in a circle around the sealed central cylindrical box. Expose the experimental units to the following treatments.
Control – Place the cylindrical box in the center of the experimental unit with no plant inside it.
F open – Place an adult fennel plant in the center of the experimental unit without the central box.
B open – Place an adult basil plant in the center of the experimental unit without the central box.
F closed – Place an adult fennel plant in the center of the experimental unit inside the cylindrical box.
B closed – Place an adult basil plant in the center of the experimental unit inside the cylindrical box.
F masked – Place an adult fennel plant in the center of the experimental chamber inside the cylindrical box covered in black plastic.
B masked – Place an adult basil plant in the center of the experimental chamber inside the cylindrical box covered in black plastic.
Control masked – Place the cylindrical box covered in black plastic in the center of the experimental unit with no plant inside it.
Repeat the whole experiment 3 times, each time using 15 experimental units for each treatment as described above. Every day, move each petri dish randomly around the central box and rearrange them. Also move the experimental chamber randomly to different positions in the CER. Do this by transferring each experimental unit one at a time to a separate room. Open the experimental chamber made up of two different sized boxes. Remove all the petri dishes and inspect the seeds. Take the base along with the sealed central cylindrical box outdoors and open it to aerate the plant. Do this for all the control treatments as well. Monitor the temperature inside the boxes every day.
Set up 15 experimental units individually in the plant communication chamber. Each experimental unit consists of 12 petri dishes with 25 chili seeds in each petri dish. Arrange the petri dishes at a distance of c.8 cm from each other in a circle around the sealed central cylindrical box. Expose the experimental units to the following treatments.
F open – Place an adult fennel plant in the center of the experimental unit without the central box.
B open – Place an adult basil plant in the center of the experimental unit without the central box.
F masked – Place an adult fennel plant in the center of the experimental chamber inside the cylindrical box covered in black plastic.
B masked – Place an adult basil plant in the center of the experimental chamber inside the cylindrical box covered in black plastic.
Control masked – Place the cylindrical box covered in black plastic in the center of the experimental unit with no plant inside it.
Repeat the whole experiment 4 times, each time using 15 experimental units for each treatment as described above. Randomly move the experimental units and chamber around the CER every day as described above.
Set up experimental units with 8 chili seeds by individually sowing them into small pots of size 3×3×7 cm filled with coco fiber substrate in the plant communication chamber. Arrange the pots at a distance of c.10 cm from each other in a circle around the sealed central cylindrical box. Expose the experimental units to the following treatments.
Control – Place the cylindrical box in the center of the experimental unit with no plant inside it.
Chili – Place the cylindrical box in the center of the experimental unit with adult chili plant inside it.
Fennel – Place the cylindrical box in the center of the experimental unit with adult fennel plant inside it.
Basil – Place the cylindrical box in the center of the experimental unit with an adult basil plant inside it.
Water and fertilize coco fiber substrate every 4th day. Repeat the whole experiment 4 times as described above. Randomly move the experimental units and chamber around the CER every day as described above.
Set up 15 experimental units individually in the plant communication chamber. Each experimental unit consists of 12 petri dishes with 25 chili seeds in each petri dish. Arrange the petri dishes at a distance of c.8 cm from each other in a circle around the sealed central cylindrical box. Expose the experimental units to the following treatments.
F open – Place an adult fennel plant in the center of the experimental unit without the central box.
B open – Place an adult basil plant in the center of the experimental unit without the central box.
F masked – Place an adult fennel plant in the center of the experimental chamber inside the cylindrical box covered in black plastic.
B masked – Place an adult basil plant in the center of the experimental chamber inside the cylindrical box covered in black plastic.
Control masked – Place the cylindrical box covered in black plastic in the center of the experimental unit with no plant inside it.
At day 14, transfer 240 seedlings from all three treatments individually into small pots filled with an identical mixture of sterilized soil and sand in the ratio of 3:1 respectively. Place the pots in a glasshouse with no fennel plants. Repeat the whole experiment 3 times for all treatments as described above. Randomly move the experimental units and chamber around the CER every day as described above.
Heterospecific Neighbor Experiment & Follow-up Germination Experiment
Monitor and record the germination rates every other day for 12 days until the rate reaches 90% or reaches a constant for at least one of the treatments.
Neighbor Identity Experiment
On the 7th day, gently remove the top layer of coco fibers using a fine paintbrush to expose the seeds. Monitor and document the germination rates of each treatment continuously for the initial 20 days until a stable rate is observed in at least one treatment. Throughout the experiment, track the emergence rates, measure the maximum stem height, and count the number of leaves. On day 38, assess the number of branches and meticulously cleanse the seedlings’ roots. Capture photographs of the roots alongside a scale bar to measure the maximum root length accurately.
Follow-up Growth Experiment
Record stem height over the course of the experiment. At day 38, carefully wash the roots of all the seedlings and photograph the roots against a scale bar to calculate maximum root length.
Plant communication has been a topic of discussion in scientific literature for decades, yet it remains controversial among researchers. Current understanding posits that plants communicate through mechanisms such as light, chemicals, and physical contact. However, emerging evidence from various studies suggests the existence of additional communication pathways. Further research is needed to explore and substantiate these mechanisms, advancing our understanding of plant communication in the scientific community.
Karban R (2008). Plant behavior and communication. Ecology Letters 11(7), 727-39. DOI: 10.1111/j.1461-0248.2008.01183.x
Gagliano M, Renton M, Duvdevani N, Timmins M, Mancuso S (2012). Out of sight but not out of mind: alternative means of communication in plants. PLoS One 7(5), e37382. DOI: 10.1371/journal.pone.0037382
Gagliano M, Renton M (2013). Love thy neighbor: facilitation through an alternative signaling modality in plants. BMC Ecology 13, 19. DOI: 10.1186/1472-6785-13-19
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