Introduction

The perennial herb Silene baccifera (L.) Roth (berry catchfly) of the genus Silene in the family Caryophyllaceae is distributed across mountainous regions and fields throughout Korea (NIBR, 2025). Commonly referred to as “Nambangbyeolkkot” or “Dunggulbyeolkkot” in Korean, this species is characterized by dense, curly trichomes over the entire plant (FCEC, 2003; RHS, 2022). It grows to a height of 2–5 cm, bears white flowers from July to August, and produces black mature seeds in September. The young shoots and leaves of S. baccifera are traditionally harvested and cooked as a vegetable, and in some regions, it has been utilized as a nutritional supplement (Hassan et al. 2024;). Phytoecdysteroids, phenolic compounds, flavonoids, and triterpenoids have been reported in the aerial parts of S. baccifera, and its fruit juice contains high total phenolic and total flavonoid contents, exhibiting both antioxidant and antimicrobial activities (Kulakov et al. 2022). Consequently, in traditional medicine, it has been applied as a poultice or decoction for the treatment of bone fractures, muscle injuries, tuberculosis, and scrofula, effects that are closely associated with its pharmacological properties (Bahadori et al. 2021). Related species such as Silene jenisseensis and Silene arenarioides have been found to contain saponin compounds, including jenisseensoside A–D, and phytoecdysteroids such as 20-hydroxyecdysone and ponasterone A, which confer multiple pharmacological activities including anti-inflammatory and analgesic effects, promotion of muscle regeneration and recovery, immune modulation, skin regeneration, and wound healing. These bioactivities indicate their high potential for use as natural antimicrobials, functional food ingredients, topical preparations, and anticancer adjuvants (Chandra, 2015; Mamadalieva, 2014). Silene baccifera also possesses ornamental value due to its slender climbing stems, white flowers, and distinctive black fruits. Some regional floristic surveys and horticultural records have reported its potential for use as a garden ornamental (Donovan, 2022). According to the Korean Red List of Threatened Species, it is currently classified as Least Concern (LC) (NIBR, 2025). However, domestic databases and floristic inventories provide only limited information on the distribution and morphology of S. baccifera, with little attention given to its propagation or physiological traits (NIBR, 2025). Despite its broad occurrence in mountainous and field habitats, the ecological adaptations and reproductive biology of this species remain poorly understood, particularly regarding seed dormancy and germination behavior. Previous studies within the genus Silene (e.g., S. latifolia, S. vulgaris, S. nutans, S. italica, and S. hicesiae) have reported a wide range of dormancy types—from non-dormant to physiological and combinational—depending on environmental conditions and geographic distribution (Baskin and Baskin, 2014;Vandelook et al, 2008). Yet, no systematic investigation has been carried out to determine the dormancy class or germination ecology of S. baccifera. Understanding its germination physiology is essential not only for elucidating species-specific dormancy mechanisms but also for supporting future applications in ornamental horticulture, restoration ecology, and the conservation of native Silene populations in Korea. Therefore, this study aimed to identify the dormancy type of Silene baccifera seeds and systematically evaluate their germination responses to temperature (alternating and constant), light, and GA₃ treatments to clarify the physiological mechanisms underlying seed germination.

Materials and Methods

Plant material and seed collection

Seeds of Silene baccifera (berry catchfly) used in this study were collected on November 1, 2023, from plants cultivated within the Forest Biological Resources Utilization Center of the Korea National Arboretum, Yangpyeong-gun, Gyeonggi-do, Korea. The experiment was initiated after the seeds were collected, after ripened for two weeks by dry storage, and cleaned, followed by storage in a seed chamber at 0 °C for approximately four months.

Seed morphology

To investigate external morphology, seed length and seed width were measured using a Vernier caliper immediately after seed collection. For internal morphological observation, seeds were sectioned with a stainless razor blade, and cross-sections were photographed using a USB microscope (AM4515T Dino-Lite Premier, AnMo Electronics Co., New Taipei City, Taiwan) at a magnification of 78.8×.

Physical dormancy test

Physical dormancy was assessed by imbibing 10 seeds per replicate (three replicates) in distilled water at room temperature (22–25 °C) on two filter papers (ADVENTEC No. 1) in 90 × 15 mm Petri dishes. Seed weight was recorded at 3, 6, 9, 12, 24, and 72 hours to calculate percentage increase. Procedures for assessing physical dormancy and quantifying seed imbibition followed established methods (ISTA, 2023;Baskin and Baskin, 2014). The imbibition percentage was calculated using the following formula:

where:

Germination temperature experiment

Optimal germination temperature was determined by surface sterilizing seeds in 2000 mg·L^-1 benomyl for 24 h, then rinsing twice and incubating them under four temperature regimes: a constant 4 °C treatment and three day/night regimes at 15/6 °C, 20/10 °C, and 25/15 °C. Day/night conditions were implemented by providing light for 12 h followed by 12 h of darkness, synchronized with the day and night temperatures, respectively. Each treatment used 20 seeds per replicate with three replicates. Germination percentage was recorded weekly for 12 weeks.

GA₃ bioassay for physiological dormancy diagnosis

GA₃ effects on dormancy release were evaluated by soaking seeds in GA₃ at concentrations of 0, 10, 100 and 1000 mg·L^-1 for 24 h. Seeds were sterilized in 2000 mg·L^-1 benomyl for 12 h and rinsed twice with distilled water. They were incubated in a growth chamber at 15/6 °C and 25/15 °C and germination percentage was recorded every week for 12 weeks.

Data collection and analysis

Data collected from the experiments were analyzed using analysis of variance (ANOVA) with SAS 9.4 (SAS Institute Inc., Cary, USA). Mean comparisons among treatments were performed using Tukey’s honestly significant difference (HSD) test at p ≤ 0.05. Graphs were prepared using SigmaPlot v10.0 (Systat Software Inc., San Jose, CA, USA).

Results and Discussion

The seeds of Silene baccifera are spherical with an areolate surface pattern, having a thousand-seed weight of 1.751 g and a length and width of 1.32 ± 0.06 mm and 0.99 ± 0.02 mm, respectively (Table 1; Fig. 1A–C). Sectioning showed a peripheral (curved) embryo that hugs the seed perimeter adjacent to the nutritive tissue (Fig. 1C). Both the axis and cotyledons were completely formed at maturity, indicating the absence of morphological dormancy (MD) (Martin, 1946;Baskin and Baskin, 2014). A 72-h imbibition test showed a 39.8% increase in mass, confirming that the seed coat is water-permeable and that physical dormancy (PY) is unlikely (Fig. 2) (ISTA, 2023;Finch-Savage and Leubner-Metzger, 2006).

Germination exhibited pronounced temperature dependence across day/night regimes. At 25/15 °C and 20/10 °C, cumulative germination increased slowly at first and then rose sharply after week 5, reaching 86.6% and 81.6% by week 12, respectively. By contrast, germination at 15/6 °C remained minimal, around 5.0%, and no germination was observed at 4 °C (Fig. 3). This lag-then-surge pattern at warmer thermo-periods is consistent with conditional dormancy that progressively weakens during incubation under favorable temperatures (Batlla and Benech-Arnold, 2015). Consistent with this pattern, Mediterranean congeners show similar thermal windows: for example, Silene colorata reached 90% germination at 15 °C and 69% at 20 °C under a 12/12 h light/dark regime, whereas performance declined at 5 °C (Brullo and Salmeri, 2022). More broadly across the genus, species from warmer climates tend to germinate at higher test temperatures, indicating climate-linked shifts in thermal requirements (Zani and Müller, 2017).

Following GA₃ treatments (0, 10, 100, and 1000 mg·L^-1), no substantial improvement in final germination percentage was observed at 25/15 °C, although the onset of germination was accelerated (Fig. 4). By contrast, at 15/6 °C, GA₃ at 1000 mg·L^-1 elevated final germination to 47.6 ± 6.7% at 12 weeks versus 5% in the untreated control—a more than 9-fold increase— demonstrating that GA₃ can partially overcome the low-temperature constraint. This marked enhancement supports the well-established criterion that a significant increase in germination following GA₃ application is diagnostic of physiological dormancy (PD) (Baskin and Baskin, 1998). Mechanistically, these responses align with PD governed by ABA–GA antagonism: seasonal temperature cues reduce ABA influence and/or enhance GA biosynthesis/signaling, thereby lowering the germination threshold under warm diurnal regimes; when temperatures are suboptimal, exogenous GA₃ can compensate for this threshold and promote radicle protrusion (Gonai et al. 2004;Tuan et al. 2018). Comparable findings in the congeneric alpine species Silene elisabethae, where GA₃ markedly improved germination across various temperature and light regimes, further corroborate the role of GA₃ as an effective dormancy-breaking agent in Silene species (Mondoni et al. 2009). However, even under the warm diurnal regimes and GA₃ treatment, final germination did not exceed ~80%. This persistent ~20% of ungerminated seeds is consistent with a non-deep physiological dormancy (PD) fraction that varies in depth within the seed cohort—i.e., a risk-spreading strategy in which a portion remains dormant to contribute to the soil seed bank and buffer against temporal environmental uncertainty (Baskin and Baskin, 2014). Similar non-deep PD and conditional dormancy behavior has been documented in congeners (e.g., Silene hicesiae), supporting the interpretation that a dormant fraction can be maintained despite otherwise favorable cues (temperature and GA) (Carruggio et al, 2021;Gianella et al, 2021).

Therefore, based on the comprehensive results of the dormancy assays, it was determined that seeds of Silene baccifera do not possess physical dormancy (PY), as the seed coat did not restrict water uptake, and no morphological dormancy (MD) was present, since embryos were fully developed upon anatomical observation. These findings suggest that this species possesses physiological dormancy (PD) as the sole dormancy type.

The results for Silene baccifera in this study clearly delineate a thermal window in which germination accelerates at 20/10–25/15 °C but remains stagnant at the lower regime of 15/6 °C, and they show that GA₃ at 1000 mg·L^-1 under 15/6 °C increased final germination from 5% to 47.6 ± 6.7% by week 12, partially offsetting the low-temperature constraint. This accords with the general pattern that dormancy is aligned with late-spring to early-summer diurnal conditions favorable for seedling establishment (Rosbakh et al. 2023;Fernández-Pascual et al. 2021). Operationally, it suggests sowing and nursing within the 20/10–25/15 °C day/night range while allowing for a latency of several weeks; when production must proceed under cooler conditions, a short high-concentration GA₃ priming can improve final percentages and synchrony (Ma et al. 2018). In addition, because longer incubation—especially at suboptimal temperatures—elevates the risk of damping-off in cool, wet substrates, identifying and maintaining an optimal temperature regime, together with sanitation (seed disinfection, sterile or pasteurized media, adequate drainage, and regular monitoring), is critical to maximize germination and minimize disease losses (Lamichhane et al. 2017; NC State Extension; UW–Madison Extension). In sum, this study provides empirical evidence directly linking the germination ecology/physiology of S. baccifera with propagation practice, thereby establishing a practical baseline for reliable seedling production and scalable mass-propagation protocols.

Moreover, S. baccifera has documented edible and medicinal uses (Cavero and Calvo, 2015), and antimicrobial and antioxidant activities in its fruit extracts further support its value as a functional raw material (Gajić et al. 2020). Established in vitro propagation methods in related species (Silene schimperiana, Silene cretacea) (Ghareb et al. 2020;Kritskaya et al. 2016) and the expanding market for edible flowers (RDA, 2018) further support its industrial potential. Thus, this study provides baseline data for stable seedling production and propagation of S. baccifera.