Abstract
In indoor crop production, light intensity is typically maintained constant throughout the photoperiod. Dynamic lighting strategies, such as dimming or turning off lights during periods of high electricity prices, can help reduce energy costs. However, fluctuating light conditions may negatively affect plant growth. In this study, we quantified the effects of frequent light-dark cycles on lettuce growth and physiological responses. Two lettuce cultivars, ‘Rex’ and ‘Rouxai’, were grown under seven treatments that differed in the number of light-dark cycles per 24-h period (1–96 cycles): 16 h light/8 h dark (1 cycle; control), 8 h/4 h (2 cycles), 4 h/2 h (4 cycles), 1 h/30 min (16 cycles), 30 min/15 min (32 cycles), 20 min/10 min (48 cycles), and 10 min/5 min (96 cycles). Across all treatments, light intensity during the light periods was maintained at 300 µmol m-2 s-1. The ‘16 h/8 h’ and ‘8 h/4 h’ treatments resulted in the greatest plant growth in both cultivars. In contrast, the ‘1 h/30 min’ and ‘20 min/10 min’ treatments resulted in the lowest leaf area and shoot biomass in ‘Rouxai’. Plants under these treatments exhibited a slow increase in photosynthetic rate (Pnet) following transitions from dark to light. This slow photosynthetic induction, combined with the frequent light-dark cycles, likely attributed to the reduced growth. Similarly, the ‘20 min/10 min’ treatment resulted in the lowest steady-state Pnet and shoot mass in ‘Rex’. However, both cultivars grown under the most frequent light-dark cycle (‘10 min/5 min’) showed rapid photosynthetic induction and comparable growth to those grown under the control treatment. Pigment levels, including total chlorophylls, carotenoids, and anthocyanins, decreased under frequent light-dark cycles (32–96 cycles per 24-h). Overall, frequent light-dark cycles reduced lettuce growth and pigmentation. However, when the dark period was brief (5 min), the negative effects on plant growth were mitigated, likely due to rapid photosynthetic induction. The duration of dark periods and cycle frequency should be carefully considered to minimize growth reductions when implementing non-static lighting strategies in indoor production.
1 Introduction
In natural environments, plants are continually exposed to fluctuations in light conditions caused by diurnal changes in solar angle, cloud movement, wind, and canopy shading. During transitions from low light or darkness to high light, photosynthesis increases gradually before reaching a steady-state rate, a process termed photosynthetic induction (). Due to biochemical and stomatal limitations, adjustments in photosynthetic activity typically lag behind changes in light intensity, resulting in reductions in light use efficiency and carbon assimilation rate of approximately 10–40% during the induction period compared with steady-state conditions (; ; ).
To cope with fluctuating light, plants have evolved various mechanisms that provide photoprotection while modulating photosynthetic efficiency. During sudden increases in light intensity, absorbed energy can temporarily exceed the capacity of photosynthetic carbon assimilation, leading to excess excitation pressure and over-reduction of the photosynthetic electron transport chain (). To protect photosynthetic apparatus from photodamage, excess energy is safely dissipated as heat through non-photochemical quenching (NPQ) (). When light intensity subsequently decreases, thermal dissipation is slowly relaxed, allowing photosynthetic efficiency to increase (). Under fluctuating light, a higher fraction of the absorbed light energy is dissipated via photoprotective mechanisms, often leading to reduced photosynthetic efficiency and plant growth. The balance between maximizing photosynthetic efficiency and maintaining adequate photoprotection is important for overall plant productivity (; ). Consistent with this, previous studies have shown that rapid relaxation of photoprotective mechanisms increases photosynthesis and crop productivity ().
Beyond short-term regulation, long-term exposure to fluctuating light induces extensive acclimation responses at both molecular and structural levels. Transcriptomic studies have revealed reprogramming of genes related to photosynthesis, photoprotection, primary and secondary metabolisms, and circadian regulation, with responses differing by time of day and leaf development stage (). At the same time, plants often develop structural and morphological adjustments under fluctuating light, such as reduced leaf thickness and chloroplast anatomy (; ; ).
The rate of photosynthetic induction is a key determinant of time-integrated carbon assimilation under fluctuating light (). Photosynthetic induction is strongly influenced by the duration, frequency, and amplitude of light fluctuations. During transitions from darkness or low light to high light, photosynthetic induction rate is typically dependent on the relaxation of three limiting processes: (1) a rapid phase in which limitations on ribulose 1, 5-bisphosphate (RuBP) regeneration are relaxed as the photosynthetic electron transport rate increases within seconds to 1–2 minutes; (2) activation of Calvin cycle enzymes, particularly Rubisco, which is generally completed within ~10 minutes; and (3) the slow opening of stomata that typically require 10–60 minutes to reach full stomatal conductance (). Importantly, the relaxation kinetics of these processes depend on the prior light history of the leaf, for example, how long leaves have remained in darkness. Short dark periods allow partial retention of Rubisco activation and stomatal opening, enabling faster photosynthetic induction upon dark-to-light transition (). However, longer dark periods lead to greater Rubisco deactivation and stomatal closure, thereby prolonging induction time ().
Plant responses to fluctuating light can vary widely among species and across different light regimes. In Arabidopsis thaliana, acclimation to fluctuating light mimicking natural conditions reduced leaf thickness, growth rate, and photosynthetic capacity (). In tomato, exposure to brief lightflecks (peak light intensity of 1000 μmol m-2 s-1 for 20 s) every 5 min over 16 h per day for three weeks accelerated photosynthetic induction after darkness compared with plants grown under constant light (). More recently, examined tomato responses to sinusoidal fluctuating light and observed thinner leaves and lower NPQ, although stomatal conductance, photosynthetic capacity, and overall plant growth were not significantly affected. These studies have primarily focused on sunflecks or stepwise changes in light intensity occurring over seconds to tens of minutes. In contrast, few studies have examined long-term exposure to repeated square-wave light regimes with frequent light-dark cycles, which are relevant for developing cost-effective lighting strategies in indoor crop production.
In indoor farming, constant light levels are usually implemented throughout the photoperiod using electric lighting, such as light emitting diodes (LEDs). Despite the high crop productivity and various environmental benefits, the energy required for crop lighting and environmental control remains a major cost of indoor farming, limiting the economic viability and wider adoption (). Implementing dynamic lighting, such as dimming or turning off lights during periods of high electricity prices, can help reduce energy costs (). However, studies examining the effects of frequent light-dark cycles on photosynthetic physiology and plant growth under controlled environments are limited. Therefore, the objectives of this study were to: (1) quantify the effects of fluctuating light with different frequencies of light-dark cycles on lettuce growth and morphology, and (2) examine the associated photosynthetic responses and the mechanisms linking photosynthetic responses to plant growth.
2 Materials and methods
2.1 Plant materials and growing conditions
Two lettuce (Lactuca sativa) cultivars, green butterhead ‘Rex’ and red oakleaf ‘Rouxai’, were used in this study. Seeds (Johnny’s Selected Seeds, Fairfield, ME, USA) were sown in 0.9 L plastic pots (10 cm × 10 cm × 9 cm) filled with a soilless substrate containing sphagnum peat moss (80-90%) and perlite (PRO-MIX LP15, Premier Tech Horticulture, Rivière-du-Loup, Canada). The containers were moved into an air-conditioned walk-in growth room after sowing and placed under seven light treatments (see details below). Six containers per cultivar were included in each treatment. Seedlings were thinned to one plant per container and fertigated daily with a nutrient solution made with deionized water, a water-soluble fertilizer (12N-1.75P-13.3K; Jack’s Nutrients FeED 12-4–16 RO; JP Peters, Inc., Allentown, PA, USA), and Epsom salt (MgSO4). The nutrient solution contained (in mg L-1) 150 N, 21.9 P, 166.2 K, 87.6 Ca, 36.9 Mg, 15 S, 0.255 B, 0.255 Cu, 1.875 Fe, 0.6 Mn, 0.015 Mo, and 0.45 Zn. The pH of the nutrient solution was adjusted to 5.8 using a 0.5 M potassium hydroxide solution.
The air temperature and relative humidity in the growth room were measured with a temperature and humidity probe (EE08-SS, Apogee Instruments, Inc., Logan, UT), and the CO2 level was measured with a CO2 probe (GMP252, Vaisala Inc., Louisville, CO) throughout the experiment. Additionally, type-J thermocouples were installed to monitor air temperature in each treatment area. All sensors were connected to a data logger (CR1000X, Campbell Scientific, Inc., Logan, UT). Data was measured every 30 seconds and recorded as hourly and daily averages. A small air mixing fan (model CFM-9238B-140-473; Mouser Electronics, Mansfield, TX) was installed in each experimental unit to improve air circulation throughout the experiment. Air temperature was uniform across treatments with minimal variation. The average air temperature was 26.1 ± 0.59 °C (mean ± SD), relative humidity was 51.7 ± 4.5%, and CO2 level was 482.4 ± 72.7 μmol mol-1.
2.2 Light treatments
Lettuce plants were exposed to seven fluctuating light treatments with varying frequencies of light-dark cycles (ranging from 1 to 96 cycles) per 24 hours (Figure 1). The light treatments included: 16 h light and 8 h dark (one cycle per 24-h, ‘16 h/8 h’; control), 8 h light and 4 h dark (two cycles per 24-h, ‘8 h/4 h’), 4 h light and 2 h dark (four cycles per 24-h, ‘4 h/2 h’), 1 h light and 30 min dark (16 cycles per 24-h, ‘1 h/30 min’), 30 min light and 15 min dark (32 cycles per 24-h, ‘30 min/15 min’), 20 min light and 10 min dark (48 cycles per 24-h, ‘20 min/10 min’), and 10 min light and 5 min dark (96 cycles per 24-h, ‘10 min/5 min’). In all treatments, the ratio of light to dark duration within each cycle was kept at 2:1, resulting in a cumulative light duration of 16 h and cumulative dark period of 8 h per day in all treatments. The light treatments were randomly assigned to seven experimental compartments (83 cm × 61 cm × 61 cm), which were created using two double-layer metal shelves. The shelves were lined with opaque plastic sheeting, and the compartments were separated using reflective foils to minimize light contamination among the treatments. Each light treatment was provided by two light emitting diode (LED) fixtures (Alina Luminaires, RAYN Growing Systems, Middleton, WI) installed at the top of each treatment compartment. The light fixtures were programmed to emit broad-spectrum white light (peak wavelengths at 453 nm and 591 nm; Figure 2), following the light on/off cycles described above. The blue (400–500 nm):green (501–600 nm):red (601–700 nm):far-red (701–800 nm) ratio of the white light was 24.1:44.7:28.7:2.5 (each component expressed as % of total photons within the 400–800 nm range). Photosynthetic photon flux density (PPFD; 400–700 nm) was measured at canopy level (40 cm below the light fixtures) using a spectroradiometer (PS300; Apogee Instruments, Logan, UT) at 12 locations in each light treatment area. The average PPFD was maintained at 300 µmol m-2 s-1 in all treatments during the light periods. The daily light integral (DLI) in each treatment was 17.28 mol m-2 d-1. Plants were stacked using layers of foam boards, which were periodically removed to maintain a consistent distance between the top of the plant canopies and the light fixtures.
Figure 1
Figure 2
Plants within each treatment were rotated every 1–2 days to minimize potential effects of spatial variation in light intensity (the standard deviation of the light intensity in each treatment ranged from 48.8 to 60.6 µmol m-2 s-1).
2.3 Leaf photosynthetic responses to light-dark cycles
Leaf photosynthetic responses to light-dark cycles were measured at 27–32 days after planting (DAP). The measurements were made on the most recently fully expanded leaves using a portable gas exchange system with a 2 × 3 cm clear-top chamber (LI-6800; LI-COR, Lincoln, NE). Plants were measured under white light provided by the same type of LEDs described above (Figure 2). Light intensity at the leaf surface inside the clear-top chamber was measured using a spectroradiometer (PS300; Apogee Instruments) and adjusted to 300 µmol m-2 s-1.
During measurements, the LEDs were programmed to match the same light-dark cycle as each corresponding light treatment. Photosynthetic responses were continuously monitored over a light-dark-light cycle for five light treatments with relatively short cycles: ‘4 h/2 h’, ‘1 h/30 min’, ‘30 min/15 min’, ‘20 min/10 min’ and ‘10 min/5 min’. For example, in the ‘1 h/30 min’ treatment, photosynthetic responses were measured during 1 hour of light, followed by 30 min of dark, and then one additional hour of light, to determine the steady-state net photosynthetic rate (Pnet) and how quickly photosynthesis reached steady state during the induction period (i.e., the dark to light transition). In the ‘4 h/2 h’ treatment, photosynthetic responses were measured only during the last hour of the 4-h light period, followed by 2 hours of dark, and then one additional hour of light, due to the longer cycle duration. For the ‘16 h/8 h’ and ‘8 h/4 h’ treatments, leaf Pnet was measured only during the light phase due to the long duration of each light-dark cycle. Throughout the measurement period, plants being measured followed the same light and dark schedule as those in the corresponding light treatments, ensuring that the photosynthetic responses remained representative and unaffected by the measurement process.
Gas exchange data were recorded every 30 s at the steady-state and every 5 s during the light-to-dark and dark-to-light transition phases to capture the response kinetics. The air temperature within the leaf cuvette was maintained at 25 °C, CO2 concentration was 450 µmol mol-1, leaf vapor pressure deficit was 1.3 kPa, and flow rate was 700 µmol s-1. In each replicate study, measurements were repeated three times on three different plants per lettuce cultivar per treatment. The average leaf Pnet was calculated from the response curves for three intervals: the first 10 minutes of the light period, the entire light period of a single light-dark cycle, and the steady-state phase under each light treatment.
2.4 Leaf light response curves and chlorophyll fluorescence measurements
Leaf light response curves were constructed at 33–35 DAP on three plants per cultivar for only three light treatments (‘16 h/8 h’, ‘1 h/30 min’ and ‘10 min/5 min’) due to the short interval between when the leaves of those lettuce cultivars became sufficiently large and sturdy for clamping in the gas exchange chamber and when plants reached mature, harvestable size. These treatments were selected to represent a range of light-dark cycles frequencies, from least to most frequent. Measurements were conducted on the most recently fully expanded leaves using a portable gas exchange system equipped with a fluorometer (LI-6800; LI-COR). The fluorometer provided a light source containing 80% red light and 20% blue light with intensity ranging from 0 to 1500 µmol m-2 s-1. Plants were first given 30 min to acclimate to a light intensity of 1500 µmol m-2 s-1. Net photosynthetic rate was then measured in descending order of light levels: 1500, 1200, 800, 500, 250, 100, 50, 0 µmol m-2 s-1. At each light level, plants were given 5 to 6 min for photosynthetic rate to reach a steady state. The CO2 concentration within the cuvette was maintained at 450 µmol mol-1, and leaf vapor pressure deficit was 1.4 kPa.
Chlorophyll fluorescence was measured on most recently fully expanded leaves to detect any stress or photodamage induced by the fluctuating light cycles. Plants were dark-adapted for 2 hours before measurements with a fluorometer (OS1p; Opti-Science, Hudson, NH). The maximum quantum yield of photosystem II (PSII) was estimated as the ratio of variable fluorescence to maximum fluorescence (Fv/Fm). Plants were harvested twice in each replicate study (see details below). Chlorophyll fluorescence measurements were made one day before each harvest, at 4 weeks and 5 weeks after planting, on both lettuce cultivars.
2.5 Chlorophyll and carotenoid concentrations
Chlorophyll and total carotenoid concentrations were measured for each plant by sampling three leaf disks from light exposed regions of the most recently mature leaves using a cork borer (8.43 mm in diameter). The leaf disks were immediately put into 5 mL dimethyl sulfoxide solution and bathed at 65 °C until the disks became transparent. The extracted solution was then measured at 480 nm, 649.1 nm, and 655.1 nm for light absorbance using a spectrophotometer (GENESYS™ 180 UV–Visible Light; Thermo Fisher Scientific, Waltham, MA). The chlorophyll and total carotenoid concentrations (µg mL-1) were calculated following the equations by .
Chlorophyll a (Chla) concentration = 12.47 × A655.1 − 3.62 × A649.1
Chlorophyll b (Chlb) concentration = 25.06 × A649.1 − 6.5 × A655.1
Total carotenoid concentration = (1000 × A480 − 1.29 × Chla − 53.78 × Chlb)/220
The concentration of chlorophylls in µg mL-1 was converted to micromoles per square meter of leaf area (µmol m-2) using the molar mass of chlorophyll a or chlorophyll b, and the leaf disk area per sample. The concentration of total carotenoids was converted to milligrams per square meter of leaf area (mg m-2). The chlorophyll and total carotenoid concentrations were measured at both harvests.
2.6 Anthocyanin concentration
Anthocyanin concentration was quantified using both extraction and image analysis methods at each harvest. For the extraction method, five representative leaf disks were collected per plant from leaves that were fully exposed to light using a cork borer with 9.41 mm diameter. The leaf disks were immediately transferred into vials containing 5 mL extraction solution (5% 3M HCl + 15% H2O + 80% C2H6O) () and then placed in a refrigerator at 6 °C for 16 h. Subsequently, light absorbance of the extracted solution was measured at 530 nm and 653 nm using the spectrophotometer (GENESYS™ 180 UV-Visible Light; Thermo Fisher Scientific). The anthocyanin index was determined using the method described by :
Anthocyanin index = A530 nm − A653 nm × 0.24.
For image analysis method, top-down photos were taken before each harvest using a digital camera positioned 152 cm above the plants. The RGB images were analyzed using an open-source Python program to determine the normalized difference anthocyanins index (NDAI), an index developed by for estimating anthocyanins concentration. NDAI is calculated based on the optical properties of anthocyanins, specifically their high absorptance in the green and low absorptance in the red region of the spectrum. The Python program distinguishes the plant objects from the background and extracts the pixel intensity of the green and red color channels from the RGB images (Supplementary Figure 1). NDAI is then calculated as (Ired − Igreen)/(Ired + Igreen), where I represents the pixel intensity (). Image-based NDAI analysis was conducted for the red oakleaf cultivar ‘Rouxai’ only.
2.7 Growth parameters
To quantify the effects of fluctuating light cycles on plant growth at different stages, lettuce plants were harvested twice, at 4 weeks (young plant stage) and 5 weeks (mature stage) after planting. Three plants per cultivar from each light treatment were harvested each time. At harvest, shoot fresh mass (g plant-1) of each plant was measured. Total leaf area (cm2 plant-1) of each plant was determined using a leaf area meter (LI-3100C; LI-COR, Lincoln, NE). Then, plants were oven-dried at 60 °C for five days to obtain shoot dry mass (g plant-1). Leaf mass per area (LMA, g cm-2) was calculated as the ratio of shoot dry mass (g plant-1) to total leaf area (cm2 plant-1). The occurrence of tip burn was assessed through visual observations, with severity graded on a scale from 0 to 5, where 0 indicated the absence of tip burn and 5 represented the most severe form of tip burn.
2.8 Statistical analysis
The experiment was conducted three times following a randomized complete block design. Each replicate was treated as a block and the light treatments were randomized in each replication. Light response curves were fitted using nonlinear regression (exponential rise to maximum) in SigmaPlot (SigmaPlot 15.0; Systat Software Inc., Palo Alto, CA). Data were analyzed using Analysis of Variance (ANOVA) in Statistical Analysis Systems (SAS Institute, Cary, NC). Mean separation was performed using Fisher’s least significant difference test (LSD, P < 0.05). Data from the two lettuce cultivars were analyzed separately.
3 Results
3.1 Total leaf area and plant biomass
At the young-plant stage (harvested 4 weeks after sowing), light treatments with fewer light-dark cycles per 24-h period generally resulted in greater total leaf area and shoot biomass (Figures 3, 4; Supplementary Figure 2). The ‘16 h/8 h’ (control) and ‘8 h/4 h’ treatments, which had only one and two light-dark cycles per 24-h, respectively, resulted in the highest total leaf area and shoot fresh and dry mass in both cultivars (Figure 4; Supplementary Figure 2). The ‘4 h/2 h’ treatment (four cycles per 24-h) resulted in similar total leaf area and shoot fresh and dry mass in ‘Rex’ young plants as those in the ‘16 h/8 h’ and ‘8 h/4 h’ treatments but significantly reduced plant growth in ‘Rouxai’. Treatments with16 to 48 light-dark cycles per 24-h (‘1 h/30 min’, ‘30 min/15 min’, and ‘20 min/10 min’) tended to reduce leaf area and shoot biomass in both cultivars, with greater reductions under more frequent cycles in ‘Rex’. However, total leaf area of ‘Rouxai’ young plants under the ‘30 min/15min’ treatment did not differ significantly from the control, and plant growth was greater than in the ‘1 h/30 min’ treatment and the ‘20 min/10min’ treatments (Figures 4b, d). Interestingly, lettuce grown under the most frequent light-dark cycles (‘10 min/5 min’; 96 cycles per 24-h) showed total leaf area and shoot biomass comparable to those under the ‘16 h/8 h’ and ‘8 h/4 h’ treatments in both cultivars (Figure 4; Supplementary Figure 2).
Figure 3
Figure 4
At the mature plant stage (harvested at 5 weeks after sowing), there were no significant differences in total leaf area among treatments in ‘Rex’ (Figure 4a). However, shoot fresh and dry mass exhibited similar treatment responses to young plants, with highest values under treatments with fewer dark-light cycles per 24-h period (‘16 h/8 h’, ‘8 h/4 h’, and ‘4 h/2 h’) and increasingly greater reductions under more frequent cycles (‘1 h/30 min’ ‘30min/15 min’, and ‘20 min/10 min’) (Figure 4c; Supplementary Figure 2a). The ‘10 min/5 min’ treatments had similar shoot mass as the control. Both total leaf area and shoot biomass in mature ‘Rouxai’ plants showed similar responses as young plants. The highest total leaf area and shoot fresh and dry mass were observed in the ‘16 h/8 h’ and ‘8 h/4 h’ treatments, while the lowest values were found in the ‘1 h/30 min’ and ‘20 min/10 min’ treatments. Compared to the control, the ‘10 min/5 min’ treatment resulted in significantly reduced leaf area (by 16.6%) but similar shoot fresh and dry mass (Figures 4b, d; Supplementary Figure 2b).
The effects of different light-dark cycles on LMA differed between ‘Rex’ and ‘Rouxai’. In ‘Rex’, the highest LMA was observed under the ‘16 h/8 h’ (control) treatment in both young and mature plants (Figure 4e). Treatments with 16 to 96 cycles per 24-h (‘1 h/30 min’, ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’) resulted in the lowest LMA, although there were no significant differences among these treatments (Figure 4e). In contrast, in mature ‘Rouxai’ plants, more frequent cycles (16–96 cycles per 24-h) resulted in the highest LMA, whereas the control treatment resulted in the lowest LMA (Figure 4f). LMA did not differ significantly among treatments in young ‘Rouxai’ plants (Figure 4f).
3.2 Pigment concentrations
Pigment concentrations, including chlorophylls, total carotenoids, and anthocyanins, decreased under more frequent light-dark cycles in both cultivars (Figures 3, 5; Supplementary Figure 3). Specifically, in ‘Rex’ young plants, chlorophyll a, chlorophyll b, total chlorophylls, and total carotenoids concentrations were highest under the less frequent light-dark cycles (‘16 h/8 h’, ‘8 h/4 h’, and ‘4 h/2 h’) and reduced progressively under more frequent cycles (Figures 5a, c). The most frequent light-dark cycles treatments (‘20 min/10 min’ and ‘10 min/5 min’) resulted in the lowest chlorophylls and total carotenoids concentrations (Figures 5a, c). Similarly, in ‘Rouxai’ young plants, chlorophylls and total carotenoids concentrations were significantly reduced in the ‘30 min/15 min’, ‘20 min/10 min’ and ‘10 min/5 min’ treatments compared to treatments with less frequent light/dark cycles (Figures 5b, d).
Figure 5
Anthocyanins concentration was the highest in the ‘8 h/4 h’ treatment in ‘Rouxai’ young plants, followed by the ‘16 h/8 h’ control treatment, and lowest in the ‘30 min/15 min’, ‘20 min/10 min’ and ‘10 min/5 min’ treatments, based on both the anthocyanins index obtained from anthocyanins extraction and the NDAI values from image analysis (Figures 5e, f). A close correlation was observed between anthocyanin concentrations estimated from extraction and image-based NDAI values (analysis not shown), indicating that the extraction method reliably reflected anthocyanin responses at the whole-canopy level.
Similar trends in chlorophyll a, chlorophyll b, total chlorophylls, total carotenoids, and anthocyanins concentrations were observed in mature plants of both cultivars (Supplementary Figure 3).
3.3 Leaf photosynthetic responses
3.3.1 Leaf photosynthetic responses to light-dark cycles
Induction kinetics. Different light/dark cycles markedly affected leaf photosynthetic responses in both lettuce cultivars. Both ‘Rex’ and ‘Rouxai’ plants grown in the ‘4 h/2 h’ and ‘1 h/30 min’ treatments exhibited slow induction kinetics during the dark-to-light transition (Figures 6A–D); Pnet gradually increased for more than 20 minutes before reaching a relatively steady state (Figures 6a–d). In contrast, photosynthetic rate of ‘Rex’ grown in the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments showed rapid increases during the light induction phase (Figures 6E, G). Time to reach a near-steady-state maximum Pnet was approximately 1.0 min, 0.75 min, and 0.5 min in the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments, respectively (Figures 6e, g). Similarly, Pnet in ‘Rouxai’ plants grown in the ‘30 min/15 min’ and ‘20 min/10 min’ treatments increased rapidly over ~0.5 min upon transition from dark to light (Figures 6f, h). However, Pnet continued to increase slowly over an additional ~20 minutes to before reaching a maximum (Figures 6F, H). In the ‘10 min/5 min’ treatment, ‘Rouxai’ plants showed a rapid induction similar to ‘Rex’, with Pnet quickly reaching its maximum within 0.5 min (Figures 6I, J, 6i, j).
Figure 6
Average Pnet over different time intervals. In ‘Rex’, the average Pnet during the first 10 min of the light period was highest in the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments, but was significantly reduced in the ‘4 h/2 h’ and ‘1 h/30 min’ treatments, with the lowest values observed in the ‘4 h/2 h’ treatment. (Table 1). This was consistent with the slower photosynthetic induction under treatments with less frequent light-dark cycles (and therefore longer dark period within each cycle) (Figure 6). As described in the ‘Materials and Methods’, this parameter was not measured in the treatments with long light-dark cycle durations (16 h/8 h’, ‘8 h/4 h’).
Table 1
| Cultivar | Pnet (µmol m-2 s-1) | 16h/8h | 8h/4h | 4h/2h | 1h/30min | 30min/15min | 20min/10min | 10min/5min |
|---|---|---|---|---|---|---|---|---|
| ‘Rex’ | First 10-min average of the light period | — | — | 5.5 ± 0.38 c | 6.2 ± 0.28 bc | 7.6 ± 0.19 a | 7.5 ± 0.09 a | 7.4 ± 0.32 ab |
| Entire light period average for one light-dark cycle | — | — | — | 8.6 ± 0.21 a | 8.3 ± 0.24 a | 8.0 ± 0.08 ab | 7.4 ± 0.32 b | |
| Steady-state Pnet | 10.0 ± 0.27 ab | 10.1 ± 0.21 a | 10.4 ± 0.16 a | 9.5 ± 0.25 bc | 9.3 ± 0.19 cd | 8.6 ± 0.17 e | 8.7 ± 0.32 de | |
| ‘Rouxai’ | First 10-min average of the light period | — | — | 2.8 ± 0.49 b | 2.4 ± 0.43 b | 4.7 ± 0.26 a | 5.5 ± 0.29 a | 5.7 ± 0.24 a |
| Entire light period average for one light-dark cycle | — | — | — | 5.7 ± 0.36 ns | 6.1 ± 0.25 ns | 6.2 ± 0.26 ns | 5.7 ± 0.24 ns | |
| Steady-state Pnet | 6.6 ± 0.28ns | 6.8 ± 0.26ns | 6.8 ± 0.17 ns | 6.9 ± 0.3 ns | 6.8 ± 0.27 ns | 6.9 ± 0.19 ns | 6.9 ± 0.17 ns |
Average leaf net photosynthetic rate (Pnet) during the first 10 min of the light period, the entire light period of one light-dark cycle, and the steady state rate for ‘Rex’ and ‘Rouxai’ under different light treatments.
Data represent mean ± SE (n=9; 3 plants per replicate x 3 replicate studies). Within each photosynthetic parameter, different letters indicate significant differences among treatments at P < 0.05. ns indicates non-significance at P < 0.05. For the16 h/8 h and 8 h/4 h treatments, only steady state Pnet values were obtained due to the long light/dark cycle durations. See Figure 1 legend for treatment details.
Similarly, the average Pnet of ‘Rex’ over the entire light period was calculated only for the treatments with more frequent cycles (‘1 h/30 min’, ‘30min/15 min’, ‘20min/10 min’, and ‘10 min/5 min’). The ‘1 h/30 min’ treatment had the highest average Pnet over the entire light period, although it did not differ statistically from the ‘30 min/15 min’ and ‘20 min/10 min’ treatment. The ‘10 min/5 min’ treatment showed the lowest average Pnet over the entire light period, despite exhibiting rapid photosynthetic induction (Table 1; Figure 6).
Steady-state maximum Pnet of ‘Rex’ was measured for all treatments and was highest in the ‘16 h/8 h’, ‘8 h/4 h’, and ‘4 h/2 h’ treatments. The ‘1 h/30 min’ treatment showed comparable steady-state values to the control treatment but was significantly lower than those in the ‘8 h/4 h’ and ‘4 h/2h’ treatments (Table 1). Steady-state Pnet was further reduced in the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments, by 7%, 14%, and 13% relative to the control, respectively (Table 1).
In ‘Rouxai’, the average Pnet during the first 10 min of the light period followed a similar response pattern to that of ‘Rex’, with the highest values in the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments and significantly lower values in the ‘4 h/2 h’ and ‘1 h/30 min’ treatments (Table 1). However, the average Pnet over the entire light period and the steady-state maximum Pnet showed no significant differences among treatments in ‘Rouxai’ (Table 1).
3.3.2 Light response curves
Leaf light response curves were constructed for three treatments, ‘16 h/8 h’, ‘1 h/30 min’ and ‘10 min/5 min’, representing the least frequent, intermediate, and most frequent light-dark cycles, respectively (see Materials and Methods). In ‘Rex’, Pnet was significantly reduced in the ‘1 h/30 min’ and ‘10 min/5 min’ treatments at all measured PPFD levels above 50 µmol m-2 s-1 compared to ‘16 h/8 h’ control, with the lowest Pnet in the ‘10 min/5 min’ treatment at light levels above the light compensation point (Figure 7a). Similarly, Pnet in the ‘10 min/5 min’ treatment was significantly reduced at all PPFD levels above 50 µmol m-2 s-1 compared to ‘16 h/8 h’ in ‘Rouxai’. However, there were no significant differences between the 16 h/8 h’ and ‘1 h/30 min’ treatments across all PPFD levels (Figure 7b).
Figure 7
3.3.3 Maximum quantum yield of PSII
Fv/Fm ranged from 0.807 to 0.819 in young plants and from 0.81 to 0.83 in mature plants across both cultivars, indicating an absence of photoinhibition in all treatments (Supplementary Figure 4).
4 Discussions
4.1 Growth reductions were mitigated under the most frequent light-dark cycles with short dark periods
Our results showed that total leaf area and shoot biomass were generally reduced when the frequency of light-dark cycles increased from 1 cycle (‘16 h/8 h’) to 48 cycles (‘20 min/10 min’) per 24-h in both ‘Rex’ and ‘Rouxai’. Notably, the most frequent light-dark cycles (96 cycles, ‘10 min/5 min’) resulted in growth comparable to the control in both cultivars (Figures 4a-d; Supplementary Figure 2). In general, frequent light-dark cycles tend to negatively affect plant growth, as plants are naturally adapted to a single light-dark cycle per 24 hours. Manipulation of light-dark cycles might alter various metabolic, morphological, and physiological processes and impair plant normal growth and development (; ; ; ). For instance, found that frequently alternating light between 400 µmol m-2 s-1 and darkness every 15 minutes resulted in significantly lower shoot biomass, total leaf area, and chlorophyll content in lettuce than constant light at 200 µmol m-2 s-1 over a 16-h photoperiod. Similarly, showed that fluctuating light intensities, applied by alternating light intensities between 900 µmol m-2 s-1 for 1 minute and 90 µmol m-2 s-1 for 4 minutes, significantly reduced the projected leaf area of Arabidopsis compared with plants grown under constant light providing the same daily light integral.
However, previous studies have found that plant growth is not always reduced under fluctuating light intensities or frequent light-dark cycles. grew plants under various light-dark cycles and found that 3 cycles per 24-h (4 h light/4 h dark) and 12 cycles per 24-h (1 h light/1 h dark) resulted in the greatest dry biomass in tomato seedlings and cocklebur plants, respectively. In both species, however, treatments with less frequent or more frequent cycles than these optima generally reduced growth compared with a single daily light-dark cycle (12h light/12h dark) (). Similarly, reported that 4, 6 and 8 light-dark cycles significantly increased lettuce shoot and root fresh mass compared with one light-dark cycle per 24-h. Nevertheless, the underlying physiological mechanisms, such as photosynthetic responses, were not examined in these studies.
A recent study found that plant dry biomass and total leaf area of young tomato plants decreased linearly as the number of light-dark cycles increased from 1 to 6 cycles per 24-h (). This is largely consistent with the reduced lettuce growth observed in our study under treatments with up to 48 light-dark cycles. Slow photosynthetic induction (e.g., in the 4 h/2 h and 1 h/30 min treatments; Figures 6A–D) and/or reduced steady-state Pnet (Table 1) most likely contributed to the observed growth reductions in these treatments. However, this reduction trend was mitigated in the treatment with the most frequent light-dark cycles (96 per 24-h; 10 min light/5 min dark) (Figures 4a-d; Supplementary Figure 2). This mitigation was likely due, at least in part, to rapid photosynthetic induction under this treatment (Figure 6), as discussed in detail in Section 4.2 below.
4.2 Duration of dark periods strongly affects photosynthetic induction kinetics and morphological acclimation in a cultivar-specific manner
Fluctuating light is generally thought to reduce overall carbon gain because of the lagged responses of stomata and the photosynthetic machinery to changes in light intensity (; ). However, the effects of fluctuating light on carbon assimilation depend strongly on the duration of the dark period within light-dark cycles, likely reflecting the combined effects of short-term photosynthetic induction kinetics and long-term acclimation adjustments. In our study, Pnet increased rapidly following short dark periods in the 10 min light/5 min dark treatment and quickly reached steady state in both lettuce cultivars (Figures 6I, J). Similar rapid initial increases in Pnet were observed in the ‘30 min/15 min’ and ‘20 min/10 min’ treatments during transitions from dark to light (Figures 6E–H).
demonstrated that the exponential phase of photosynthetic induction, i.e., the initial period following a transition from darkness or low light to high light during which Pnet increases rapidly and approximately exponentially, reflects the kinetics of Rubisco activation. After a step increase in light intensity following varying durations of darkness, the relaxation time of this exponential phase closely matched that of Rubisco activation. In addition, Rubisco deactivation in darkness was found to have a relaxation time of approximately 28 minutes (). Thus, longer dark periods lead to greater Rubisco deactivation and consequently require more time for reactivation during dark to light transition. Consistent with this, slow photosynthetic induction over extended periods was observed in the ‘4 h/2 h’ and ‘1 h/30 min’ treatments (Figures 6A–D) in our study. In contrast, short durations (5 min or less) of dark period or low light intensity allow partial retention of Rubisco activation and have minimal impact on stomatal conductance (). Rubisco activation has been reported to be the major limitation to photosynthetic induction during the initial ~10 min of light in various species, including tobacco, rice and wheat (; ; ; ). The short dark duration in the ‘10 min light/5 min dark’ treatment likely allowed partial activation of Rubisco and contributed to the rapid photosynthetic induction (Figure 6) and mitigated growth reductions (Figure 4; Supplementary Figure 2) observed in both lettuce cultivars. Additionally, stomatal conductance (gs) during the induction period, measured as the average gs over the first 10 minutes of the light period, was significantly higher in the ‘10 min/5 min’ treatment than in the ‘4 h/2 h’ treatment (Supplementary Table 1), which may also have contributed to the fast photosynthetic induction after short dark periods.
In the ‘30 min/15 min’ and ‘20 min/10 min’ treatments, rapid initial increases in Pnet were observed in both cultivars during light induction (Figures 6E–H). Similar to responses in the ‘10 min/5 min’ treatment, this rapid response likely results from partial retention of Rubisco activation, as complete Rubisco deactivation requires a longer dark period (~28 min; ). Notably, despite these rapid or comparable photosynthetic induction rates, both steady-state Pnet and the average Pnet of ‘Rex’ over the entire light period of a light-dark cycle generally decreased with increasing cycle frequency, reaching the lowest values in the ‘10 min/5min’ treatment (Table 1). Similarly, leaf photosynthetic capacity (expressed as maximum Pnet at light-saturated intensity) in the ‘10 min/5min’ treatment was also significantly reduced (Figure 7a). Photoinhibition and low light use efficiency have been observed under fluctuating light in previous studies (; ; ). Although photoinhibition was not observed in our study (Supplementary Figure 4), it was possible that a higher fraction of absorbed light was dissipated via photoprotective mechanisms under frequent light-dark cycles, reducing leaf-level Pnet.
The reduced leaf Pnet in ‘Rex’ under frequent light-dark cycles might also be associated with plant morphological acclimation (Figures 4a, c). Biochemical and morphological adjustments, including reduced leaf thickness and increased leaf area, have been observed in plants acclimated to light flecks or fluctuating light, which may lead to increased whole-plant light interception and partially compensate for reductions in leaf-level photosynthesis (; ). Similarly, we observed that ‘Rex’ showed reduced LMA but maintained a similar total leaf area in the ‘10 min/5 min’ treatment compared with the control (Figures 4a, e). In addition, biomass accumulation is often more strongly correlated with radiation capture at the whole-plant level and cumulative carbon gain rather than instantaneous leaf photosynthetic rate (). Rapid photosynthetic induction can improve cumulative CO2 assimilation during light fluctuations (). Collectively, the comparable growth of ‘Rex’ in the ‘10 min/5 min’ treatment relative to the control could likely be attributed to the maintenance of total leaf area and rapid photosynthetic induction.
Although rapid initial photosynthetic inductions were observed in both cultivars in the ‘30 min/15 min’ and ‘20 min/10 min’ treatments, Pnet in ‘Rex’ reached steady-state relatively quickly (within ~1 min), whereas in ‘Rouxai’ it continued to increase gradually over an additional ~20 minutes before reaching steady state (Figures 6E–H). This longer lagging time in ‘Rouxai’, a red-leafed cultivar, may be explained by a slower increase in effective light irradiation upon light induction due to light screening of anthocyanins (), which could in turn delay biochemical processes such as Rubisco activation. However, stomatal limitations likely did not play a role, as stomatal conductance during the induction period in the ‘30 min/15 min’ and ‘20 min/10 min’ treatments was comparable to that in the ‘10 min/5 min’ treatment (Supplementary Table 1). showed that as dark duration increased, the exponential phase of photosynthetic induction accounted for a greater proportion of the overall photosynthetic response in spinach, with a relaxation time of approximately 24 minutes. This is consistent with our observation that photosynthesis in ‘Rouxai’ continued to increase for ~20 minutes before reaching steady state in the ‘30 min/15 min’ and ‘20 min/10 min’ treatments (Figures 6F, H).
Despite the slower photosynthetic induction kinetics in ‘Rouxai’ under treatments with longer dark durations, steady-state Pnet and average leaf Pnet over the entire light period of a light/dark cycle were not negatively affected (Table 1). In contrast to ‘Rex’, which showed reduced LMA while maintaining large total leaf areas particularly at the mature plant stage (Figures 4A, E), ‘Rouxai’ exhibited increased leaf thickness, as indicated by higher LMA, while generally having significantly reduced total leaf area under more frequent light-dark cycles compared to the control (Figures 4B, F). Photosynthetic capacity is generally positively linked to LMA (). The increased LMA in ‘Rouxai’ under more frequent light-dark cycles may have contributed to the minimal reduction in photosynthetic capacity in the ‘1 h/30 min’ treatment and the relatively small reduction observed in the ‘10 min/5 min’ treatment (Figure 7b). In contrast, ‘Rex’ showed larger reductions in photosynthetic capacity under these treatments relative to its control (Figure 7a). However, despite the higher LMA, shoot biomass of ‘Rouxai’ was generally reduced under treatments – compared with the control (Figure 4D). This was probably because the reduced leaf area offset the benefits of increased LMA at the whole-plant scale (Figure 4b). The comparable growth of ‘Rouxai’ in the 10 min/5 min treatment was most likely due to comparable integrated carbon assimilation driven by rapid photosynthetic induction combined with a similar total leaf area as the control.
4.3 Long-term acclimation effects on photosynthetic induction
Beyond the short-term effects of dark period duration on Rubisco activation and stomatal opening, plant long-term acclimation to fluctuating light might have also contributed to the varying photosynthetic induction rates in our treatments. Under long-term exposure to fluctuating light, plants develop systematic acclimation adjustments across different scales, from gene expression to leaf and whole-plant traits (; ; ; ). In our study, increasing the frequency of light-dark cycles was associated with reduced concentrations of chlorophyll, carotenoids, and anthocyanins in both lettuce cultivars (Figure 5). Reduced chlorophyll content may lower light-harvesting capacity and reduce the initial excitation pressure on photosystems following dark-to-light transitions, potentially increasing the efficiency of electron transport and carbon assimilation during light induction (). In addition, decreases in carotenoid and anthocyanin levels may indicate reduced photoprotective capacity, which could allow greater carbon assimilation during light induction under non-stressful conditions (). Frequent light-dark cycles also caused changes in LMA, representing a major modification in leaf structure in both ‘Rex’ and ‘Rouxai’ (Figures 4e, f). Variations in LMA suggest structural acclimation that may alter leaf thickness, mesophyll organization, and internal CO2 diffusion pathways, thereby influencing diffusional limitations during light transitions (; ). Although beyond the scope of the study, the decreases in LMA in ‘Rex’ under frequent light-dark cycles could be partially associated with improved mesophyll conductance, photosynthetic electron transport and Rubisco carboxylation activity (). In contrast, the increases in LMA in ‘Rouxai’ under frequent light-dark cycles may have contributed to its relatively slower photosynthetic induction compared with ‘Rex’. Collectively, these biochemical and structural differences may help explain the slower induction rates under treatments with less frequent light-dark cycles, as well as cultivar-specific differences in induction kinetics.
4.4 Frequent light-dark cycles reduced pigment concentrations in lettuce
Frequent light-dark cycles resulted in significant reductions in pigment concentrations in both lettuce cultivars. Concentrations of chlorophylls (Chlorophyll a, b, and total chlorophylls) and total carotenoids were significantly lower under the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments in both cultivars compared with the control (Figures 5a-d; Supplementary Figures 3a–d). Chlorophylls are the primary photosynthetic pigments, whereas carotenoids act as accessory pigments for light harvesting and photoprotection. Therefore, carotenoids biosynthesis is typically tightly coordinated with chlorophyll accumulation (; ; ). Pigment accumulation in plants is determined by the rates of synthesis and degradation, which are regulated by both internal circadian rhythms and external environmental cues, including light-dark cycles (; ). Many key genes involved in chlorophylls and carotenoids biosynthesis are strongly controlled by circadian rhythms (; ; ; ), and changes in light-dark cycles reset the internal rhythms (). The frequent, short duration light-dark cycles applied in our study might have disrupted the coordination between internal circadian regulation and external cues, therefore disturbing the balance between synthesis and degradation of chlorophylls and carotenoids. In addition, the biosynthesis of chlorophylls requires sustained illumination (). Previous studies have found that frequent light-dark cycles can damage the Chl a/b protein complex in maize and tomato plants () and impair the photosynthetic apparatus in beans (). Consistently, reductions in chlorophyll and carotenoid concentration under frequent light-dark cycles or fluctuating light intensities have been reported in other species (; ; ).
Importantly, the observed decline in chlorophyll and carotenoid concentration was unlikely to result from photobleaching or photodamage. Although photodamage has been observed under extreme fluctuating light conditions where light intensity changes tremendously within seconds or minutes (; ), the light-dark cycle treatments used in our study (‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’) contained relatively low light intensity and did not impose extreme fluctuations. Moreover, high Fv/Fm values (above 0.807 in all treatments) indicated an absence of photosystem II photoinhibition in our study (Supplementary Figure 4), further supporting that the reductions in chlorophylls and carotenoids were not due to damages to the photosynthetic apparatus.
Similarly, anthocyanins concentrations were significantly reduced under the ‘30 min/15 min’, ‘20 min/10 min’, and ‘10 min/5 min’ treatments in ‘Rouxai’ (Figures 5e, f; Supplementary Figure 3e, f). Anthocyanins function as ‘sunscreen’ pigments that protect plants against photoinhibition and photooxidative damage under high light stress (). Therefore, increases in anthocyanin concentration are usually associated with high-light exposure or other stress conditions (such as cold temperature) that reduce light use efficiency (; ). However, neither photoinhibition nor stress-induced reductions in photosynthetic activity were observed in ‘Rouxai’ as indicated by high Fv/Fm values (Supplementary Figure 4) and steady-state Pnet (Table 1). Instead, the decreases in anthocyanin concentrations under frequent light-dark cycles were most likely a result of repeated interruptions of the light and dark periods, as previous studies have found that sustained light exposure is required for anthocyanins production (; ; ). Reduced anthocyanin accumulation may also have contributed to the maintenance of high leaf-level photosynthesis under frequent light-dark cycles (Table 1). Because red coloration is often perceived as a quality attribute in red lettuce cultivars, the effects of light-dark cycles on anthocyanin concentration should be considered when implementing dynamic lighting strategies for indoor lettuce production.
5 Conclusions
Our study showed that frequent light-dark cycles generally reduced growth and pigment concentrations in lettuce, even when the same daily light integral was provided across treatments. However, growth reductions were mitigated under the most frequent light-dark cycles with short dark durations (5 min) in both cultivars, mainly due to the rapid photosynthetic induction following short dark periods and the maintenance of a high total leaf area. Cultivar-specific photosynthetic and morphological acclimation strategies were observed in response to frequent light-dark cycles. Because the duration of darkness or low light can strongly affect photosynthetic induction kinetics, steady-state photosynthetic rates, key morphological traits such as total leaf area, leaf thickness and pigment concentration, and ultimately biomass accumulation, it should be carefully considered when implementing fluctuating light regimes in controlled environment crop production. Further research is needed to identify dark-duration threshold that minimally impact photosynthesis and growth for different crops in controlled environments.
Statements
Data availability statement
The original contributions presented in the study are included in the article/Supplementary Material. Further inquiries can be directed to the corresponding author.
Author contributions
YK: Methodology, Formal analysis, Validation, Writing – review & editing, Data curation, Conceptualization, Writing – original draft, Investigation, Visualization. SZ: Resources, Funding acquisition, Writing – review & editing, Validation, Methodology, Supervision, Project administration, Conceptualization.
Funding
The author(s) declared that financial support was received for this work and/or its publication. This work was supported by USDA-NIFA Hatch Project TEX09862, Accession No. 1026236.
Acknowledgments
We thank Seonghwan Kang, Yilin Zhu, and Hikari Skabelund for their assistance with data collection.
Conflict of interest
The author(s) declared that this work was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
Generative AI statement
The author(s) declared that generative AI was not used in the creation of this manuscript.
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Supplementary material
The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fpls.2026.1791561/full#supplementary-material
Supplementary Table 1Average stomatal conductance (gs) under different light-dark cycles.
Supplementary Figure 1Image analysis of normalized difference anthocyanin index (NDAI).
Supplementary Figure 2Shoot fresh mass of ‘Rex’ and ‘Rouxai’ under different light-dark cycles.
Supplementary Figure 3Pigment concentrations of ‘Rex’ and ‘Rouxai’ at the mature plant stage.
Supplementary Figure 4Maximum quantum efficiency (Fv/Fm) of ‘Rex’ and ‘Rouxai’ at young and mature plant stages.
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Summary
Keywords
fluctuating light, indoor farming, induction kinetics, leafy greens, LED lighting, photosynthetic efficiency
Citation
Kong Y and Zhen S (2026) Rapid photosynthetic induction after short dark periods mitigates growth reduction in lettuce exposed to frequent light-dark cycles. Front. Plant Sci. 17:1791561. doi: 10.3389/fpls.2026.1791561
Received
19 January 2026
Revised
26 February 2026
Accepted
16 March 2026
Published
30 March 2026
Volume
17 - 2026
Edited by
Elias Kaiser, Seoul National University, Republic of Korea
Reviewed by
Marta Joanna Monder, Warsaw University of Life Sciences, Poland
Bingjie Shao, Wageningen University and Research, Netherlands
Updates
Copyright
© 2026 Kong and Zhen.
This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Shuyang Zhen, shuyang.zhen@tamu.edu
Disclaimer
All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.