Description

This plant communication apparatus was first used by Gagliano et al (2013) for chilli plants (Capsicum annuum, Solanaceae) for discrimination of adult conspecific and a fennel plants despite blocking of common signaling through the apparatus.

This experimental set-up prevents above and below ground contact as well as chemical and light-mediated signals normally exchange by plants. The apparatus can be used to search for alternative signaling modalities or control specific signaling.

Chilli seeds are classically arranged in a circle around the adult plant sealed the central cylindrical box. Seeds and adult plants in each unit are housed within 2 different sized square boxes, one inside the other, with the air in between the two boxes removed using a vacuum pump (not included).

Two different sized square boxes are placed inside the other, with the air in between the two boxes removed using a pump to create a vacuum and thus avoid interference between adjacent experimental units at any time. The MazeEngineers  unit is made with colorless cast acrylic material (Moden Glas), which transmits 92% of visible light, but is opaque to ultraviolet and infrared wavelengths

Communication research has mainly focused on animals, as their signal-mediated interactions frequently involve auditory or visual displays of body parts which readily attract attention. Plant communication, on the other hand, has been a controversial topic, but in recent times it has gained more attention. Plant communication mechanisms take various forms, some of them are readily accepted by many researchers like communication through chemicals, contact, and light while others are still questioned (Karban 2008). Many studies have proven that plants are highly sensitive organisms capable of interacting and facilitating each other by actively gaining information from their surrounding environment. Plants modify their behavior according to the processing and evaluation of information about their neighbors above as well as below ground, and also about the resources available in their surroundings. Similar to animal social systems, plants facilitate kin selection process by favoring their relatives over strangers by using the information to recognize and prevent competition (Gagliano et al., 2012).

Plants use volatile organic compounds (VOCs) as a warning signal for other plants against herbivores. Such air-borne communication allows the undamaged neighboring plants to respond by increasing their defense mechanisms. Plants can detect changes in the levels of different wavelengths in the sunlight to become aware of future competition in their proximity. This is a beneficial response as plants are unable to invest in growth and defense simultaneously due to the presence of limited resources. This allows the plant to change their morphology and behavior accordingly for future growth before the resources become limited (Gagliano et al., 2012).

Plant communities are shaped by competition as well as facilitative or positive interactions. Plant facilitate each other by improving growth, nitrogen fixation, nurse cropping, controlling pests or attracting beneficial organisms such as insects or mycorrhizae. Facilitative interactions mainly use mechanisms that focus on the effects of light and shade, chemical signals, physical proximity, and contact. For example, shade from perennial canopies protect young seedlings and smaller species from extreme temperatures, excessive water loss and also provide desirable soil texture and chemistry. Recent research provides many examples of plant association with benefactor species for protection either by visual or auditory concealment, which protects them from predation by herbivores (Gagliano et al., 2013).

Research Studies

In a recent study by Gagliano et al., it was shown that plants could detect and affect their neighbors by pathways other than light, chemicals or physical contact. The study used seeds of chili plant Capsicum annuum and measured their germination rates in the presence of an adult chili plant and an adult fennel plant. Florence fennel plant (Foeniculum vulgare) inhibits the germination of chili seeds by emitting volatile chemicals. It was observed that the germination rates of chili seeds were higher in the presence of fennel plant with its signals partially or entirely blocked.

In another study by Gagliano et al., germination rates of chili seeds in the presence of a good neighbor, Basil plant (Ocimum basilicum) were measured. The basil plant positively facilitates the chili seeds by increasing its germination rates as it produces a large number of volatile compounds that act as natural insecticides and kill competitive weeds.  Gardeners are commonly seen growing basil plants with chili plants as it keeps the soil moist and acts as an organic living mulch. It was observed that basil plant increases the rate of chili seeds germination when allowed open contact between the two but also when the basil plant was placed inside a sealed box. This shows that there are other signal modalities yet to be identified apart from chemical signals.

The plant communication chamber consists of two square boxes made up of transparent acrylic which allows transmission of visible light (92%) but blocks transmission of ultraviolet and infrared wavelengths. The boxes are of different sizes, one large with dimensions of 44×44×65 cm and one small with the dimensions of 32×32×45 cm, placed one inside the other. The space between the two boxes is occupied by air which is removed with the use of a vacuum pump. In the center of the experimental chamber, a sealed cylindrical box with dimensions of 18×30 cm is placed which is used for chemical isolation.

Florence fennel plant (Foeniculum vulgare) will be used as one of the heterospecific neighbors (bad neighbor) because it produces chemicals from its roots and aerial parts that inhibit growth and kill its neighbors. Hence, it is grown in isolation.  Basil plant (Ocimum basilicum) will be used as the other heterospecific neighbor (good neighbor) because it produces chemicals that act as insecticides that inhibit the growth of weeds.  Seeds of chili plant (Capsicum annuum) will be used for observing effects of the neighbors on their germination rates.

Set up a Controlled Environment Room (CER) of 5.30 m². Fit high-density discharge lamps in the room. Maintain the temperature at 18°C during the day and 13°C during the night. All the seeds and plants are exposed to identical nutrients, temperature and 12 h light: 12 h dark cycle. Place chili seeds between layers of 2 mm thick felt to trap moisture and retain darkness in a petri dish. Inspect and water the seeds every 24 hours.

Heterospecific Neighbor Experiment

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.

Follow-up Germination Experiment

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.

Neighbor Identity Experiment

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 4^th 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.

Follow-up Growth Experiment

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 7^th day, use a fine paintbrush and lightly brush away the top coco fibers to expose the seed. Monitor and record the germination rates for each treatment for the first 20 days until the rate becomes constant for at least one of the treatments. Monitor and record emergence rates, maximum stem height and number of leaves over the course of the entire experiment. At the end of day 38, record the number of branches and also carefully wash the roots of all the seedlings and photograph the roots against a scale bar to calculate maximum root length.

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 described in the literature for a long time, but the idea is still rejected by many scientists. Plant researchers suggest, based on the evidence so far that communication in plants occurs through mechanisms using light, chemicals, and physical contact. But there has been evidence of other mechanisms involved in communication through different studies. More studies are required to further discover the mechanisms involved and gather more evidence in favor of plant communication.