Light–fertilizer interactions regulate morphophysiological performance and nutrient dynamics in Plumeria rubra under subtropical nursery conditions

Plumeria rubra L. is an important ornamental species cultivated in subtropical nursery production; however, its response to combined light and fertilization management remains insufficiently understood. This study evaluated the effects of light intensity and fertilization on plant growth, physiological performance, and substrate nutrient dynamics under container conditions. A split-plot completely randomized design (CRD) was used, with two light environments (full sun and shade) as the main factor and five cumulative fertilizer rates (0, 90, 105, 120, and 150 g/plant) as the subplot factor. Plant growth, physiological traits, and leachate properties were assessed at 180 days after treatment (DAT). Both light intensity and fertilization significantly affected plant growth and physiological responses (p < 0.05), whereas a significant light × fertilizer interaction was detected only for SPAD. Plant height increased from 103.58 cm in the unfertilized control to 136.59 cm at 120 g/plant, followed by a decline at the highest fertilizer rate. Leaf number increased progressively with fertilization, reaching 29.9 leaves at 150 g/plant. SPAD values increased with fertilizer application under both light environments, while NDVI increased from 0.69 in the control to approximately 0.80 in fertilized plants. Fertilization significantly increased NO 3 - and Ca 2+ concentrations, whereas light intensity affected Ca 2+, Na +, pH, and total dissolved salts in the leachate. Principal component analysis explained 76.04% of the total variation in the first two dimensions and showed that increasing fertilizer rates were associated with greater plant growth and physiological performance. A fertilizer rate of 120 g/plant was associated with the greatest plant growth and physiological performance. Fertilization had a greater effect on plant growth and substrate nutrient dynamics than light intensity, providing guidance for nutrient management in nursery production of P. rubra.

The floriculture industry constitutes a significant component of global horticulture, driven by sustained demand for ornamental plants with high aesthetic value and adaptability to diverse production systems. This sector has experienced continuous growth, supported by the diversification of plant species and cultivars, advances in propagation and production technologies, and the increasing adoption of practices aimed at improving resource-use efficiency and reducing environmental impacts (Salachna, 2022). In the United States, floriculture crop sales reached approximately USD 6.71 billion in 2024, with container-grown ornamentals representing a substantial share of production (USDA NASS, 2026). Production is regionally concentrated, with Florida leading national output at approximately USD 1.13 billion, surpassing other major producing states such as California (USDA NASS, 2026). Beyond their economic relevance, ornamental plants contribute to urban ecosystem services and human well-being through their roles in landscape design, environmental quality, and socio-cultural value (Francini et al., 2022; Beckerman, 2024).

The genus Plumeria L. (Apocynaceae) comprises laticiferous shrubs and small trees native to the Neotropics, particularly Mexico, Central America, and the Caribbean, and widely cultivated in tropical and subtropical regions (Koeser et al., 2014; Malaspina et al., 2024). Taxonomically, the genus is classified within the tribe Plumerieae and subtribe Plumeriinae and is recognized for its ornamental flowers and ecological adaptation across the Caribbean basin, a key center of diversity (Tiernan et al., 2021). However, species delimitation within Plumeria remains unresolved due to extensive synonymy, inconsistencies in historical classification, and the presence of numerous endemic taxa described based on limited morphological differentiation (Tiernan et al., 2021). Current estimates suggest approximately 11–18 accepted species, although a larger number of published names exist, many corresponding to cultivated forms or taxonomic synonyms (Malaspina et al., 2024; Antesco et al., 2025). Morphologically, the genus is characterized by thick succulent branches, milky latex, and terminal inflorescences bearing large, salverform, and fragrant flowers, with leaves typically clustered at the apical portions of branches (Brown, 2008; Antesco et al., 2025). From a floricultural perspective, Plumeria species are highly valued due to their ornamental diversity, fragrance, vibrant colors, and adaptability to both landscape and container production systems, with applications ranging from landscaping and floral arrangements to perfumery and cultural products such as leis (Koeser et al., 2014; Verma, 2016).

Among the species within the genus, Plumeria rubra L. is one of the most widely cultivated and commercially relevant taxa (Supplementary Material 2). This deciduous laticiferous tree, native to Mexico and Central America, has been extensively distributed across tropical and subtropical regions, where it is commonly grown in both landscape and container production systems (Koeser et al., 2014; Malaspina et al., 2024). Botanically, P. rubra exhibits a spreading growth habit, thick succulent branches with latex exudation, and large lanceolate leaves concentrated toward the apical regions of the stem (Verma, 2016; Antesco et al., 2025). Its terminal inflorescences produce numerous five-petaled, highly fragrant flowers displaying a wide range of colors, including white, yellow, pink, and red, which has supported the development of numerous cultivars (Brown, 2008; Malaspina et al., 2024). From a production standpoint, P. rubra represents a primary genetic resource for ornamental breeding and is considered the most economically important species within the genus due to its ornamental versatility, extended flowering period, and adaptability to different cultivation conditions. In addition, its phytochemical diversity and associated biological activities have attracted scientific interest, further enhancing its value beyond ornamental use (Bihani, 2021; Malaspina et al., 2024).

Light intensity is a central abiotic factor regulating plant growth and physiological performance, acting both as an energy source for photosynthesis and as a signal controlling plant development. It directly influences carbon assimilation, chlorophyll synthesis, and the efficiency of the photosynthetic apparatus within the photosynthetically active radiation (PAR) range (Paradiso and Proietti, 2022). Variations in light intensity and spectral composition affect pigment content, stomatal conductance, and biomass accumulation, ultimately shaping plant productivity and metabolic balance (Sytar et al., 2020; Eghbal et al., 2024). While optimal light conditions enhance photosynthetic efficiency and growth, both low and excessive irradiance can limit plant performance by restricting carbon fixation or inducing photoinhibition and oxidative stress (Wang et al., 2025). High irradiance levels are associated with the generation of reactive oxygen species (ROS) and damage to photosynthetic structures (Roeber et al., 2021), whereas moderate shading can improve chlorophyll content, light-use efficiency, and growth, particularly in tropical ornamental species (Munoz-Salas et al., 2025). In addition to its role in photosynthesis, light regulates plant development through photoreceptor-mediated pathways that control photomorphogenesis, plant architecture, and adaptive responses such as shade avoidance (Kami et al., 2010). Despite its importance, information on the response of P. rubra to varying light intensities under container nursery conditions remains limited.

Nutrient management represents another critical factor controlling plant growth and physiological responses in ornamental production systems. In container-based cultivation, where root volume is restricted and substrate properties determine nutrient availability and retention, fertilization practices influence plant performance. Adequate nutrient supply enhances photosynthetic capacity, chlorophyll accumulation, and biomass production, thereby improving plant quality and commercial value (Furtini Neto et al., 2015; Khoddamzadeh et al., 2025). However, excessive or unbalanced fertilization can increase substrate salinity and electrical conductivity, leading to nutrient leaching and reduced nutrient-use efficiency, particularly under conditions of high irrigation or rainfall (Balliu et al., 2021; Khoddamzadeh et al., 2025). Nutrient availability also interacts with environmental factors such as water quality and salinity, influencing plant metabolism and stress responses (Munoz-Salas et al., 2025). Although general fertilization recommendations are available for ornamental crops, including P. rubra (Koeser et al., 2014), quantitative assessments of fertilizer dose effects on growth, physiological performance, and substrate nutrient dynamics remain scarce. In this context, recent advances in precision agriculture highlight the importance of integrating sensor-based diagnostics, soil characterization, and data-driven decision-making to optimize fertilizer and irrigation management in specialty crops (Khoddamzadeh et al., 2026). emphasize that combining proximal and remote sensing technologies with adaptive management strategies enables site-specific nutrient application, improves resource-use efficiency, and enhances crop performance while reducing environmental impacts.

Despite the horticultural importance of P. rubra, the combined effects of light intensity and fertilization on plant performance and nutrient dynamics under nursery conditions have not yet been investigated. This gap is critical in subtropical production environments such as Florida, where high solar radiation and frequent rainfall can simultaneously influence plant physiology and nutrient leaching. In addition, the integration of morphological, physiological, and substrate chemical variables through multivariate approaches remains largely unexplored for this species.

To address this gap, the objectives of this study are to (i) evaluate the effects of light intensity and fertilization on the morphological and physiological performance of P. rubra during nursery production, (ii) quantify their effects on substrate leachate chemical properties and nutrient dynamics, and (iii) integrate morphological, physiological, and leachate variables using multivariate analyses to identify the primary drivers of plant performance and treatment differentiation. The findings are expected to provide practical insights for optimizing nursery management practices in Plumeria production, improving plant quality, enhancing fertilizer-use efficiency, and reducing environmental impacts associated with nutrient losses in container-based ornamental systems. This study may support more efficient nursery production and contribute to the sustainability and competitiveness of the ornamental horticulture industry. Based on the roles of nutrient availability and irradiance in regulating plant growth and physiology, an optimum fertilization rate and light intensity were expected to enhance the morphological and physiological performance of Plumeria rubra under nursery conditions.

Experimental site and plant material

The experiment was conducted at the United States Department of Agriculture – Agricultural Research Service, Subtropical Horticulture Research Station (USDA-ARS SHRS) in Miami, Florida, USA (25°37′ N, 80°25′ W), at approximately 1.5 m above sea level.

Three-year-old Plumeria rubra plants were obtained from the USDA-ARS germplasm collection. Prior to treatment application, plants were selected based on uniformity in size, vigor, and overall physiological condition to minimize experimental variability. Each plant was transplanted into 5-gallon plastic containers (18.9 L) filled with a standardized USDA nursery substrate consisting of 37.5% pine bark, 30% coir pith, 25% perlite, and 7.5% sand (v/v). Irrigation was applied using an overhead irrigation system twice daily for approximately 3 min per irrigation event to maintain adequate substrate moisture throughout the experimental period.

Experimental design and treatment application

The experiment was conducted under subtropical nursery conditions to evaluate the effects of light intensity and fertilizer rate on the growth, physiological responses, and leachate properties of Plumeria rubra. A split-plot arrangement in a completely randomized design was used, with light conditions (full light and shade) assigned to the main plots and fertilizer rates (0, 90, 105, 120, and 150 g/plant) assigned to the subplots. Measurements were collected at the end of the experiment (180 DAT). This design facilitated the implementation of treatments applied at different management levels while reducing variability associated with treatment application (Altman and Krzywinski, 2015). The experimental layout and treatment randomization were generated using the inti package (Lozano-Isla, 2026).

The main factor consisted of two light environments defined by the average photosynthetically active radiation (PAR) recorded throughout the experiment. Plants grown under full sun were exposed to direct solar radiation and received an average PAR of 1700 μmol m -2 s - ¹, whereas shaded plants received an average PAR of 790 μmol m -2 s - ¹ under a black polyethylene shade cloth with 50% light reduction and 70% UV stabilization (Green-Tek ®, Janesville, WI, USA). Midday PAR was measured periodically with a LightScout Quantum PAR Meter (Spectrum Technologies Inc., Aurora, IL, USA), and monthly averages were used to characterize each light environment.

The subplot factor consisted of five fertilizer treatments, including a non-fertilized control (F0) and cumulative application rates of 90 (F1), 105 (F2), 120 (F3), and 150 g/plant (F4). A controlled-release granular fertilizer (Sunniland ® Azalea, Camellia, and Gardenia Fertilizer, 8–4–8 with micronutrients; Sunniland Corporation, Sanford, FL, USA) was incorporated into the upper 5 cm of the substrate. Fertilizer applications were divided into two events, at 0 days after treatment (0 DAT) and 120 DAT, to improve nutrient availability throughout the experiment and enhance fertilizer use efficiency. The fertilization schedule is summarized in Table 1.

Within each light environment, fertilizer treatments were randomly assigned to the subplots. One container-grown plant was considered an experimental unit, and five replicates were established for each treatment combination, resulting in a total of 50 experimental units. Measurements were collected before treatment application (0 DAT) and at the end of the experiment (180 DAT). The complete dataset and details of the statistical analyses are provided in Supplementary Material 1.

Morphological and physiological measurements

Morphological and physiological measurements were evaluated before treatment application (0 DAT) and at the end of the experiment (180 DAT). Because Plumeria rubra is a woody ornamental species grown over several months in nursery production (Brown, 2008; Lim, 2014; Bihani, 2021), measurements were taken at the start and end of the study to compare the initial condition of the plants with their response after 180 days under the experimental treatments. At each evaluation date, five plants per treatment combination were randomly selected for morphological and physiological assessments.

Plant height (cm) was measured from the substrate surface to the apical meristem of the main shoot using a measuring tape. The number of leaves per plant was determined by counting all fully expanded leaves present at the time of evaluation.

Leaf chlorophyll status was assessed using a non-destructive optical method with a portable SPAD-502 chlorophyll meter (Konica Minolta, Osaka, Japan). Measurements were taken on the third fully expanded leaf of each plant to ensure consistency among samples. Three readings per leaf were recorded and averaged to obtain a representative chlorophyll index value, following the protocol described by Lozano-Isla et al. (2024).

Plant canopy vigor and greenness were further quantified using the Normalized Difference Vegetation Index (NDVI) measured with a GreenSeeker™ handheld optical sensor (Trimble Inc., Sunnyvale, CA, USA). During data acquisition, the sensor was positioned at a constant distance of approximately 45 cm above the plant canopy, and five NDVI readings per plant were collected and averaged to represent the physiological condition of each individual plant, following standardized procedures reported in previous horticultural studies (Khoddamzadeh et al., 2025; Munoz-Salas et al., 2025; Nogueira Souza Costa and Khoddamzadeh, 2025).

To minimize variability associated with diurnal fluctuations in light conditions and plant physiological activity, all measurements were conducted between 09:00 and 14:00 hours under relatively stable environmental conditions, following the procedures described by Neupane et al. (2023).

Leachate collection and nutrient analysis

Leachate samples were collected at each sampling interval to monitor nutrient mobility and salt accumulation within the container substrate. Substrate solution was obtained using the pour-through extraction method described by LeBude and Bilderback (2009). Briefly, containers were first irrigated until the substrate reached field capacity, after which 350 mL of deionized water was applied to displace the substrate solution and collect the resulting leachate.

The collected leachate was analyzed to determine the concentrations of nitrate (NO 3 -, ppm), potassium (K +, ppm), calcium (Ca 2+, ppm), and sodium (Na +, ppm). Ion concentrations were quantified using a LAQUAtwin ion meter (Horiba Scientific, Kyoto, Japan). Prior to each measurement session, the instrument was calibrated according to the manufacturer’s guidelines using two-point calibration with ion-specific standard solutions. To prevent cross-contamination, electrodes were rinsed with distilled water between successive samples.

In addition to ion concentrations, substrate solution pH and total dissolved salts (ppm) were measured using an ExStik EC500 conductivity meter (EXTECH Instruments, FLIR Commercial Systems, Goleta, CA, USA).

Statistical analysis

Experimental data were recorded, curated, and processed using the statistical environment R version 4.5.3 (R Core Team, 2026). Linear mixed-effects models were fitted to evaluate the effects of fertilizer level, light regime, and their interaction on growth, physiological, and leachate variables. Model fitting was performed using the lme4 2.0–1 package (Bates et al., 2015), applying restricted maximum likelihood (REML) to estimate variance components and appropriately partition fixed and random sources of variation.

In the model, light intensity (L) and fertilization rate (F) were treated as fixed effects, with light intensity assigned to the main plots and fertilization rate applied within subplots. Block (B) was included as a random effect to account for the hierarchical structure of the split-plot experimental design. Additionally, the interaction between block and light intensity (L×B) was considered as a random term representing the main plot error, while the residual term accounted for subplot-level variability. The statistical model was defined as follows:

where Y i j k represents the observed value of the response variable for the i -th light intensity level, the j -th fertilization rate, and the k -th block; μ is the overall mean; L i is the fixed effect of light intensity (main plot factor); B k is the random effect of block; (L × B) i k ∼ N (0, σ L B 2) corresponds to the main plot error term; F j is the fixed effect of fertilization rate (subplot factor); (L × F) i j represents the interaction between light and fertilization; and ϵ i j k ∼ N (0, σ 2) is the residual (subplot error) term.

Model assumptions were verified for each response variable, including residual normality, homogeneity of variance, and outlier diagnostics. Associated diagnostic plots are presented in Supplementary Material 1 (ESM_1). The significance of fixed effects was assessed using Type III analysis of variance implemented through the car 3.1–5 package (Fox et al., 2019), which is appropriate for unbalanced designs and models including interactions. Pairwise comparisons of estimated marginal means were conducted using Tukey’s test as implemented in the emmeans 2.0.2 package (Searle et al., 1980; Lenth and Piaskowski, 2017), maintaining a significance threshold of p < 0.05.

Multivariate relationships among response variables were explored using Principal Component Analysis (PCA), performed using the FactoMineR 2.13 R package (Lê et al., 2008). Prior to PCA, variables were standardized to ensure comparability across measurement scales. Graphical representations of both univariate and multivariate analyses were generated using ggplot2 4.0.2 package (Wickham, 2016) and inti 0.6.92 package (Lozano-Isla, 2026).

All data and statistical analyses were reproducible, and the complete code used for data processing and model fitting is provided in Supplementary Material 1.

Morphological and physiological responses to light intensity and fertilization

To evaluate plant performance under nursery conditions, morphological and physiological variables were assessed at the end of the experiment (180 DAT) in Plumeria rubra (Figure 1). No significant light × fertilizer interactions were detected for plant height, number of leaves, and NDVI (p > 0.05). In contrast, SPAD values were significantly influenced by the interaction between both factors (p < 0.05) (Supplementary Material 1 – Section 3).

For plant height, both light intensity (F = 315.65, p < 0.001) and fertilizer rate (F = 7.64, p < 0.001) significantly affected plant growth, whereas the light × fertilizer interaction was not significant (F = 2.40, p = 0.05) (Figure 1A). Although plant height was similar under full sun (118.87 cm) and shade (117.59 cm), fertilization significantly enhanced plant growth. Averaged across light treatments, plant height increased from 103.58 cm in the unfertilized control (F0) to a maximum of 136.59 cm at F3, before declining to 120.06 cm at F4. These results indicate that plant growth was optimized at the intermediate fertilizer rate (F3). Similarly, the number of leaves was significantly affected by both light intensity (F = 45.50, p < 0.001) and fertilizer rate (F = 14.40, p < 0.001), whereas the light × fertilizer interaction was not significant (F = 1.81, p = 0.1271) (Figure 1B). Averaged across light treatments, plants grown under full sun produced more leaves than plants grown under shade (22.5 vs. 21.4 leaves). Averaged across fertilizer treatments, leaf number increased progressively from 14.1 leaves in the unfertilized control (F0) to 19.8, 19.8, and 26.1 leaves in F1, F2, and F3, respectively, reaching a maximum of 29.9 leaves at F4.

SPAD values were significantly affected by light intensity (F = 655.66, p < 0.001), fertilizer rate (F = 10.37, p < 0.001), and the light × fertilizer interaction (F = 3.19, p = 0.0139) (Figure 1C). Under full-sun conditions, SPAD values increased progressively from 35.15 in the unfertilized control (F0) to 44.67 at F4. Under shade conditions, SPAD values increased from 37.07 at F0 to a maximum of 44.36 at F3, followed by a decline to 41.62 at F4. For NDVI both light intensity (F = 1596.73, p < 0.001) and fertilizer rate (F = 5.43, p < 0.001) significantly affected plant vigor, whereas the light × fertilizer interaction was not significant (F = 0.04, p = 0.9973) (Figure 1D). Averaged across light treatments, NDVI values were greater under full sun than under shade (0.78 vs. 0.76). Fertilizer application also increased NDVI, with values rising from 0.69 in the unfertilized control (F0) to 0.78, 0.80, 0.78, and 0.79 in F1, F2, F3, and F4, respectively.

Leachate collection and nutrient analysis

To evaluate nutrient availability in the substrate, leachate chemical properties were monitored at the end of the experiment (180 DAT) (Figure 2). No significant light × fertilizer interactions were detected for nitrate (NO 3 -), potassium (K +), calcium (Ca 2+), sodium (Na +), pH, and total dissolved salts. Significant main effects of fertilizer dose were observed for NO 3 - and Ca 2+ concentrations, whereas K + concentrations were not affected by either light intensity or fertilizer dose. Light intensity significantly affected Ca 2+, Na +, pH, and total dissolved salts, while fertilizer dose had no significant effect on Na +, pH, or total dissolved salts (Supplementary Material 1 – Section 4).

For nitrate (NO 3 -) concentration, fertilizer rate significantly affected leachate NO 3 - levels (F = 2.77, p = 0.0309), whereas light intensity (F = 2.72, p = 0.0741) and the light × fertilizer interaction (F = 1.49, p = 0.2094) were not significant (Figure 2A). Although not statistically significant, NO 3 - concentrations were numerically greater under full sun (717.80 ppm) than under shade (581.06 ppm). Fertilizer application significantly influenced nitrate concentrations. NO 3 - concentrations increased from 360.82 ppm in the unfertilized control (F0) to 520.89 ppm in F1 and reached the highest values at intermediate fertilizer rates, with 861.63 ppm and 884.16 ppm in F2 and F3, respectively. Thereafter, NO 3 - concentrations declined to 619.67 ppm at the highest fertilizer rate (F4).

For potassium (K +) concentration, neither light intensity (F = 1.28, p = 0.2849), fertilizer rate (F = 0.57, p = 0.6825), nor the light × fertilizer interaction (F = 0.37, p = 0.8315) significantly affected leachate K + levels (Figure 2B). Although not statistically significant, K + concentrations were numerically greater under shade (68.82 ppm) than under full sun (54.02 ppm). Across fertilizer treatments, K + concentrations ranged from 26.60 ppm in the unfertilized control (F0) to 61.76 ppm, 66.11 ppm, 83.64 ppm, and 68.99 ppm in F1, F2, F3, and F4, respectively.

For calcium (Ca 2+) concentration, both light intensity (F = 44.74, p < 0.001) and fertilizer rate (F = 3.29, p = 0.0137) significantly affected leachate Ca 2+ levels, whereas the light × fertilizer interaction was not significant (F = 0.89, p = 0.4744) (Figure 2C). Averaged across fertilizer treatments, Ca 2+ concentrations were greater under full sun (112.49 ppm) than under shade (92.01 ppm). Fertilizer application also significantly influenced Ca 2+ concentrations. Averaged across light environments, Ca 2+ concentrations increased from 82.49 ppm in the unfertilized control (F0) to 94.65 ppm, 103.68 ppm, and 110.98 ppm in F1, F2, and F3, respectively, reaching the highest concentration at F4 (119.44 ppm).

For sodium (Na +) concentration, light intensity significantly affected leachate Na + levels (F = 39.63, p < 0.001), whereas fertilizer rate (F = 2.09, p = 0.0873) and the light × fertilizer interaction (F = 0.95, p = 0.4358) were not significant (Figure 2D). Averaged across fertilizer treatments, Na + concentrations were greater under full sun (22.51 ppm) than under shade (19.19 ppm). Although fertilizer rate did not significantly affect Na + concentrations, values tended to increase from 16.74 ppm in the unfertilized control (F0) to 20.64 ppm in F1, 21.12 ppm in F2, and 23.77 ppm in F3, before decreasing to 23.99 ppm in F4.

For leachate pH, light intensity significantly affected substrate pH (F = 3871.20, p < 0.001), whereas fertilizer rate (F = 1.27, p = 0.2876) and the light × fertilizer interaction (F = 0.37, p = 0.8284) were not significant (Figure 2E). Averaged across fertilizer treatments, pH values were greater under shade (7.58) than under full sun (7.47). Although fertilizer rate did not significantly affect pH, values ranged from 7.64 in the unfertilized control (F0) to 7.61, 7.47, 7.36, and 7.55 in F1, F2, F3, and F4, respectively.

For total dissolved salts, light intensity significantly affected leachate salinity levels (F = 45.26, p < 0.001), whereas fertilizer rate (F = 0.52, p = 0.7223) and the light × fertilizer interaction (F = 0.48, p = 0.7526) were not significant (Figure 2F). Averaged across fertilizer treatments, salinity values were greater under shade (572.05 ppm) than under full sun (482.85 ppm). Although fertilizer rate did not significantly affect salinity, values ranged from 558.64 ppm in the unfertilized control (F0) to 505.73 ppm, 555.72 ppm, 551.81 ppm, and 465.33 ppm in F1, F2, F3, and F4, respectively.

Multivariate integration of morphophysiological traits and leachate chemistry

To evaluate the combined responses of plant growth, physiological performance, and substrate nutrient dynamics, principal component analysis (PCA) was performed using all measured variables. The analysis enabled simultaneous evaluation of relationships among variables and visualization of treatment distribution under different light conditions and fertilization levels. Principal component analysis revealed that the first two dimensions explained 76.04% of the total variation, with Dim 1 and Dim 2 accounting for 58.90% and 17.14%, respectively (Figure 3). The variables with the highest contributions to Dim 1 were Na + (13.2%), number of leaves (12.4%), Ca 2+ (12.2%), NDVI (12.2%), plant height (11.4%), and pH (11.1%), whereas Dim 2 was primarily associated with salt concentration (39.2%), K + (21.6%), and SPAD (16.6%). Variable contributions, and cos 2 values are provided in Supplementary Material 1.

The PCA biplot of variables (Figure 3A) showed that Dim 1 was associated with plant growth and physiological variables. Plant height, number of leaves, SPAD, NDVI, and potassium (K +) were oriented on the positive side of this axis. In contrast, pH and salt concentration were oriented on the negative side. Nitrate (NO 3 -), calcium (Ca 2+), and sodium (Na +) were also located on the positive side of Dim 1 but with a different orientation compared to growth-related variables. The variables Na + (0.88), number of leaves (0.86), Ca 2+ (0.85), NDVI (0.85), and plant height (0.82) showed positive correlation with Dim 1, whereas pH showed a negative correlation (-0.81). For Dim 2, salt concentration had the highest correlation (0.82), followed by K + (0.61) and SPAD (0.53). Dim 2 separated variables related to substrate chemistry and environmental conditions. Salt concentration showed a positive contribution along this axis, while calcium and nitrate were oriented in the opposite direction.

The PCA of individuals (Figure 3B) showed a gradient along Dim 1 associated with increasing fertilization and variation in morphophysiological traits. Treatments were distributed from left to right, with F0 located on the negative side, indicating lower growth and physiological performance. As fertilization increased, treatments shifted toward the positive side of Dim 1. Along Dim 2, light conditions separated treatments, with shade treatments located in the upper part of the plot and full sun treatments in the lower part. This pattern indicates that, under increasing fertilization, shade conditions were associated with higher plant height, leaf number, chlorophyll-related indices (SPAD, NDVI), and K +, whereas full sun conditions were more closely related to leaf number and higher concentrations of Ca 2+, Na +, and NO 3 -. The highest cos 2 values on Dim 1 were observed for Na + (0.78), number of leaves (0.73), NDVI (0.72), Ca 2+ (0.72), plant height (0.67), and pH (0.67), whereas salt concentration showed a cos 2 value of 0.67 on Dim 2. Among treatments, shade-F3 was positioned in the upper-right quadrant, indicating a combination of high morphophysiological performance and favorable nutrient status.

Plumeria rubra is an economically important ornamental species widely cultivated in tropical and subtropical regions for landscape use and container production. Despite its commercial value, information on nutrient management and fertilization recommendations are limited and often based on general practices developed for other ornamental crops. In particular, limited information is available regarding the combined effects of fertilization and light environment on plant performance during nursery production. In the present study, both light intensity and fertilization influenced plant growth and physiological performance, although their effects were independent, with only SPAD showing a significant light × fertilizer interaction. Fertilization was associated with greater variation in plant morphology, physiological traits, and substrate nutrient dynamics than light environment. Plant height, leaf number, SPAD, and NDVI increased with fertilizer application, indicating improved plant performance under greater nutrient availability. Light intensity also influenced SPAD, NDVI, Ca 2+, Na +, pH, and salinity, highlighting its role in shaping physiological responses and substrate chemistry. Principal component analysis further showed that increasing fertilizer rates were associated with improved morphophysiological performance, whereas light environment contributed to differences in nutrient distribution. These findings provide new information on the responses of P. rubra to nursery management practices and may help refine fertilization strategies for subtropical ornamental production.

Plant performance in response to fertilization under contrasting light environments

Plant growth and physiological responses in P. rubra were influenced by both light intensity and fertilization, although fertilization generally produced larger changes in plant performance than light environment (Figure 1). Plant height increased with fertilizer application up to F3 and declined at the highest fertilizer rate (F4), indicating a non-linear response to nutrient availability (Figure 1A). Similarly, leaf production increased progressively with fertilization, reaching its highest values at F4 (Figure 1B). Physiological traits also responded positively to fertilizer application. SPAD values increased under both light environments, although the response varied with light intensity, resulting in a significant light × fertilizer interaction (Figure 1C). NDVI increased substantially between the unfertilized control and fertilized treatments and remained relatively stable thereafter (Figure 1D), suggesting that canopy greenness reached a plateau once adequate nutrient availability was achieved.

The increase in plant height up to intermediate fertilization levels followed by a decline at the highest fertilizer rate is consistent with nutrient-response patterns reported in ornamental species, where optimal growth occurs under moderate nutrient supply, while excessive fertilization may reduce growth efficiency due to nutrient imbalances and osmotic effects in the root zone (Furtini Neto et al., 2015). Similar non-linear responses have been reported in container-grown ornamentals, where intermediate fertilization rates maximize vegetative growth, whereas higher fertilizer inputs result in diminishing returns rather than proportional increases in biomass accumulation (Khoddamzadeh et al., 2025). Although light intensity significantly affected plant height and leaf number, the magnitude of these differences was relatively small compared with the response to fertilization. This suggests that P. rubra can maintain growth under both full-sun and shaded nursery conditions, reflecting a degree of adaptability to contrasting light environments.

In contrast to morphological traits, SPAD values were significantly influenced by both fertilization and the interaction between light intensity and fertilizer rate. Under full sun, SPAD values increased continuously with increasing fertilizer application, whereas under shade they increased up to F3 and declined slightly at F4. Increased SPAD values with fertilization are consistent with the role of nutrient availability, particularly nitrogen, in chlorophyll synthesis and leaf greenness (Paradiso and Proietti, 2022). Differences between light environments may reflect adjustments in chlorophyll accumulation and leaf physiology that allow plants to optimize light capture under contrasting irradiance conditions (Sytar et al., 2020).

NDVI values increased with fertilization and stabilized at intermediate to high fertilizer rates, indicating that nutrient availability enhanced canopy vigor and greenness but that additional fertilizer provided limited benefits once plants reached adequate nutritional status. Similar responses have been reported in horticultural crops, where NDVI is positively associated with plant nutritional status and vegetative growth but becomes less responsive at higher canopy densities because of spectral saturation effects (Atik and Akdemir, 2022; Costa et al., 2023; Neupane et al., 2024). Because leaf area and biomass were not measured, NDVI should be interpreted as an indirect indicator of canopy greenness and vigor rather than a direct measure of biomass accumulation.

Nutrient dynamics in the substrate solution under fertilization and light conditions

Leachate chemical properties responded differently to fertilization and light intensity (Figure 2). Fertilizer application significantly affected nitrate (NO 3 -) and calcium (Ca 2+) concentrations, whereas light intensity significantly influenced Ca 2+, sodium (Na +), pH, and total dissolved salts. In contrast, potassium (K +) concentrations were not significantly affected by either factor, and no significant light × fertilizer interactions were detected for any leachate chemical variable. These results indicate that nutrient availability in the substrate solution was regulated by both fertilizer inputs and environmental conditions, although the magnitude of the response varied among nutrients.

The chemical characteristics of the leachate are consistent with previous studies in ornamental nursery systems, where nutrient concentrations do not always respond proportionally to fertilizer inputs because they are influenced by substrate properties, irrigation practices, nutrient retention, and fertilizer release dynamics (Chen et al., 2001; Marfà et al., 2002; LeBude and Bilderback, 2009). In ornamental crops, nitrate is frequently the nutrient most susceptible to leaching losses, whereas the availability of other ions depends on nutrient mobility, substrate interactions, and plant uptake patterns (Chen et al., 2001). Similarly, nutrient availability in container substrates is regulated by substrate decomposition processes, cation exchange capacity, water-holding capacity, and interactions between nutrients and the growing medium, all of which influence nutrient retention and mobility within the root zone (Marfà et al., 2002; Criscione et al., 2022; Huntley, 2023; Rotowa et al., 2025). Water availability further influences nutrient transport because nutrient movement depends on the continuity of the aqueous phase within the substrate, which regulates nutrient diffusion and redistribution throughout the root zone. Under favorable moisture conditions, water facilitates nutrient transport, whereas reduced connectivity of water-filled pores can limit nutrient mobility and availability (Bauke et al., 2022). These mechanisms help explain the results observed in the present study, where fertilizer application significantly increased NO 3 - and Ca 2+ concentrations but had little effect on K +, Na +, pH, or salinity. Moreover, the absence of significant light × fertilizer interactions suggests that nutrient availability in the substrate solution was primarily regulated by the independent effects of fertilization and light environment rather than by a combined response.

The nitrate pattern observed in this study, characterized by increasing concentrations at intermediate fertilizer rates followed by a decline at the highest fertilization level, is consistent with nutrient dynamics reported in ornamental nursery systems receiving controlled-release fertilizers. In container-grown ornamentals, nitrate availability reflects the balance among fertilizer release, plant uptake, nutrient retention within the substrate, and leaching losses rather than fertilizer application alone (Chen et al., 2001; Pitton et al., 2023). Similar responses have been reported in woody nursery crops, where fertilization enhances plant growth and nutrient accumulation, but nutrient concentrations measured in the substrate solution do not necessarily increase proportionally because nutrients may be retained within the growing medium or incorporated into plant biomass (Pitton et al., 2022; Rotowa et al., 2025). Previous studies have also demonstrated that nutrient concentrations in leachate are influenced by substrate water retention characteristics and water movement through the container profile, which regulate nutrient transport and losses from the root zone (Criscione et al., 2022). Water-mediated transport processes play a central role in nutrient redistribution because dissolved ions move through interconnected water films within the substrate matrix, and changes in moisture conditions can alter nutrient mobility and accessibility throughout the root zone (Bauke et al., 2022). As a result, the decline in nitrate concentrations observed at the highest fertilizer rate may indicate greater nutrient utilization or retention within the plant substrate system rather than reduced fertilizer availability.

The behavior of potassium, calcium, and sodium in this study generally agrees with previous reports showing that nutrient responses vary according to ion mobility, environmental conditions, plant demand, and nutrient-specific transport and regulatory processes within the plant (White and Broadley, 2003; Gilliham et al., 2011; Islam et al., 2025). Calcium concentrations were significantly greater under full sun and increased progressively with fertilizer application, whereas sodium was affected primarily by light intensity and potassium remained relatively stable across treatments. One possible explanation for the higher Ca 2+ concentrations under full-sun conditions is the close relationship between calcium transport and transpiration-driven xylem flow (White and Broadley, 2003; Gilliham et al., 2011). Increased irradiance can enhance transpiration and water movement through the plant, thereby promoting Ca 2+ transport and accumulation in aboveground tissues and the root-zone solution (Montanaro et al., 2006). In contrast, potassium is highly mobile and participates in multiple physiological processes, including stomatal regulation, osmoregulation, ion transport, enzyme activation, and maintenance of cellular ion homeostasis, which can result in variable responses to environmental conditions (Ahammed et al., 2022; Islam et al., 2025). This may explain why K + concentrations showed only numerical variation among treatments despite changes in fertilization and light environment. Sodium concentrations were consistently greater under full sun but were not affected by fertilizer rate, suggesting that Na + dynamics were more closely associated with environmental conditions than with nutrient inputs. The tendencies observed for Ca 2+, Na +, and K + under contrasting light environments may reflect differences in ion mobility, transport processes, and physiological regulation within the substrate plant system.

The limited variation in pH and total dissolved salts observed in this study aligns with patterns commonly reported in nursery production systems, where these variables are regulated by interactions among substrate properties, irrigation water, and nutrient. Although pH was significantly affected by light intensity, fertilizer application did not alter substrate pH, indicating that the controlled-release fertilizer did not substantially modify substrate chemical conditions during the production cycle. Similarly, total dissolved salts were influenced by light environment but did not increase significantly with fertilizer rate. This absence of salt accumulation suggests that nutrient release and drainage remained balanced throughout the experiment. Recent studies have shown that ion concentrations and electrical conductivity are strongly influenced by substrate moisture conditions because water availability regulates ion solubility, mobility, and distribution within the pore solution (Kim and Park, 2024). Therefore, the relatively stable salinity values observed across fertilization treatments likely reflect interactions among nutrient release, water movement, and substrate retention processes rather than fertilizer input alone. These results indicate that the substrate system maintained favorable chemical conditions throughout the experiment while supporting enhanced plant growth under fertilized treatments.

Integrated responses of plant performance and nutrient dynamics

The multivariate analysis showed that fertilization was the main factor associated with variation in plant growth, physiological traits, and nutrient dynamics (Figure 3). The distribution of treatments along Dim 1 followed a clear fertilization gradient, with the unfertilized control (F0) located on the negative side of the axis and fertilized treatments progressively shifting toward the positive side. This pattern indicates that increasing nutrient availability was associated with improvements in plant performance and changes in substrate nutrient composition. Plant height, SPAD, NDVI, and potassium (K +) were oriented on the positive side of Dim 1, whereas pH and salt concentration were located on the opposite side. Number of leaves, nitrate (NO 3 -), calcium (Ca 2+), and sodium (Na +) also contributed substantially to Dim 1 but exhibited a different orientation relative to the growth-related variables, indicating differences in their association with plant performance. These results suggest that fertilization was the primary factor structuring treatment distribution, while light environment contributed to secondary variation among treatments.

The multivariate analysis indicated that fertilization structured treatment distribution along a gradient defined by plant height, SPAD, NDVI, and K +, while pH was oriented in the opposite direction. This inverse relationship between fertilization and pH is consistent with acidification processes reported in container substrates, where ammonium transformations and nitrification contribute to the release of H + and gradual changes in substrate pH. Studies using controlled-release fertilizers in nursery systems have shown that leachate pH may vary throughout the production cycle as a result of nutrient release patterns, nitrification processes, and substrate chemical interactions (Merhaut et al., 2006).

The close association among plant height, leaf number, SPAD, and NDVI observed in the PCA indicates that morphological growth and physiological performance responded similarly to increasing fertilizer availability. Similar multivariate patterns have been reported in ornamental production systems, where nutrient supply explains a substantial proportion of the variation in chlorophyll-related indices and vegetative growth parameters (Costa et al., 2023; Neupane et al., 2024). In Chrysobalanus icaco, fertilization increased chlorophyll content and leaf production, while NDVI and plant growth responded positively to improved nutritional conditions (Costa et al., 2023). Likewise, studies in ornamental and horticultural crops have shown that NDVI and SPAD are closely associated with plant nutritional status and canopy development, supporting their use as nondestructive indicators of plant performance (Neupane et al., 2024).

In the present study, the alignment of SPAD, NDVI, plant height, and leaf number along the positive side of Dim 1 suggests that these variables captured a common response to fertilization. Because biomass accumulation was not measured directly, these relationships should be interpreted as associations among growth and physiological indicators rather than evidence of increased biomass production. Nevertheless, the PCA supports the conclusion that moderate fertilization promoted coordinated improvements in plant vigor and canopy development. Similar relationships among optical indices and vegetative growth have been reported in ornamental nursery crops receiving controlled-release fertilizers (Khoddamzadeh et al., 2025).

A similar coordination between physiological traits and plant performance has been observed under favorable environmental conditions in Musa haekkinenii, where increased chlorophyll-related indices were associated with enhanced vegetative growth and photosynthetic performance (Munoz-Salas et al., 2025). In contrast, pH and salt concentration were separated from the growth-related variables in the ordination space, indicating that changes in leachate chemistry were not directly coupled with plant performance. This pattern agrees with studies showing that nutrient leaching and salinity dynamics may vary independently of plant growth because they are regulated by substrate characteristics, nutrient retention, and water movement through the root zone (Pitton et al., 2023; Neupane et al., 2024).

Although NO 3 -, Ca 2+, and Na + were positioned on the positive side of Dim 1, their orientation differed from that of plant height, SPAD, NDVI, and leaf number. This separation suggests that greater nutrient concentrations in the substrate solution did not necessarily translate into proportional increases in plant growth. Similar observations have been reported in container nursery systems, where nutrient availability in the root zone is influenced by nutrient retention, plant uptake efficiency, and leaching processes, resulting in a partial decoupling between nutrient concentrations in leachate and plant performance (Pitton et al., 2022, 2023).

Light intensity contributed to treatment separation along Dim 2, where shade treatments were located in the upper portion of the PCA and full-sun treatments in the lower portion. Shade conditions were associated with plant height, SPAD, NDVI, and K +, whereas full-sun treatments were more closely associated with NO 3 -, Ca 2+, and Na + concentrations. This separation indicates that light environment influenced the relationship between plant performance and nutrient dynamics, even though fertilization explained most of the overall variation among treatments.

The proximity of K + to plant height, SPAD, and NDVI in the PCA suggests that potassium availability in the leachate followed a pattern similar to that of the measured morphophysiological variables. However, because nutrient concentrations were not determined in plant tissues, no conclusions can be drawn regarding potassium uptake or accumulation within plants.

In contrast, the association of Ca 2+ with full sun is consistent with its transport through the transpiration stream, which increases under higher irradiance and evaporative demand (White and Broadley, 2003). The association of NO 3 - and Na + with full-sun treatments observed in the PCA may suggest differences in ion dynamics reported in previous studies; however, processes such as mass flow, transpiration, and water flux were not measured and therefore cannot be confirmed from the present results.

Among all treatment combinations, shade-F3 was positioned in the upper-right quadrant of the PCA, indicating an association with relatively high values of plant height, leaf number, SPAD, NDVI, and favorable nutrient availability. This positioning coincides with the univariate analyses, which identified F3 (120 g/plant) as one of the most effective fertilizer rates for promoting plant growth. Comparable responses have been reported in ornamental nursery systems where moderate fertilizer inputs improve plant performance and nutrient use efficiency, whereas excessive fertilizer applications provide limited additional benefits and may increase nutrient losses (Pitton et al., 2023; Khoddamzadeh et al., 2025).

Limitations and future perspectives

This study evaluated morphological, physiological, and leachate chemical responses of Plumeria rubra under contrasting light environments and fertilization regimes during nursery production. Although these variables provided useful indicators of plant performance and nutritional status, flowering traits were not assessed. Because the ornamental value of P. rubra is closely linked to its floral display, future studies should include reproductive variables such as flowering time, flower number, inflorescence development, and flower quality to provide a more complete evaluation of commercial performance. At the same time, canopy density and foliage quality remain important attributes because consumers often select Plumeria for both its flowers and attractive vegetative appearance.

Environmental characterization was limited to light conditions, and other microclimatic variables such as air temperature, relative humidity, rainfall, and vapor pressure deficit were not monitored during the experiment. These factors may influence plant growth, nutrient availability, and substrate chemistry. Measurements related to transpiration, water relations, and nutrient transport were also not included. As a result, interpretations regarding ion mobility, calcium transport, and nutrient movement within the substrate should be viewed as possible explanations that warrant further investigation.

Leachate analyses described nutrient availability in the substrate solution but did not quantify nutrient uptake, nutrient allocation within plant tissues, or nutrient use efficiency. In addition, destructive measurements such as shoot and root biomass, leaf area, stem diameter, and tissue nutrient concentrations could not be performed because the plants belonged to a USDA germplasm collection. Future experiments using plant material designated for destructive sampling could incorporate these measurements and provide additional information on nutrient acquisition, allocation patterns, and plant nutritional status under different fertilization regimes.

The experiment was conducted at a single location during one production cycle using one controlled release fertilizer source and a single Plumeria species. Although these conditions reflect common nursery production practices, caution should be exercised when extending the results to other environments, fertilizer formulations, substrates, or genotypes. Studies conducted across multiple locations, production cycles, fertilizer sources, and Plumeria genotypes would help determine the consistency of these responses and improve fertilization recommendations for nursery production. Evaluating P. rubra under field conditions may also provide insight into whether similar responses occur across different production systems.

Future research could integrate physiological, nutritional, and biochemical measurements with flowering assessments to provide a broader characterization of plant responses to fertilization and light environment. Expanding these evaluations across a wider range of genotypes and production conditions may contribute to the development of management practices that improve plant quality, increase fertilizer use efficiency, and maintain efficient nutrient management in ornamental nursery systems.

This study shows that both fertilization and light intensity independently influenced the growth, physiological performance, and substrate nutrient dynamics of P. rubra. Fertilization was the main factor associated with variation in plant performance, increasing plant height, leaf number, SPAD, and NDVI. Plant growth was optimized at 120 g/plant (F3), whereas higher fertilizer application provided limited additional benefits. Fertilizer significantly affected NO 3 - and Ca 2+ concentrations, while light intensity influenced Ca 2+, Na +, pH, and total dissolved salts. Principal component analysis indicated that fertilization was associated with improved morphophysiological performance, while light environment contributed to differences in nutrient distribution patterns. The combination of shade and 120 g/plant fertilizer was associated with favorable growth, physiological performance, and nutrient status. These findings provide practical information for fertilization management of P. rubra under nursery conditions.