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Article

The Co-Occurrence of Physiological and Epicotyl Physiological Dormancy in Three Desiccation-Sensitive Castanopsis (Fagaceae) Acorns from China with Specific Reference to the Embryonic Axis Position

by
Jiajin Li
,
Ganesh K. Jaganathan
*,
Han Kang
and
Baolin Liu
Germplasm Conservation Laboratory, School of Health Science and Engineering, University of Shanghai for Science and Technology, Shanghai 200093, China
*
Author to whom correspondence should be addressed.
Forests 2023, 14(12), 2330; https://doi.org/10.3390/f14122330
Submission received: 2 November 2023 / Revised: 22 November 2023 / Accepted: 22 November 2023 / Published: 28 November 2023
(This article belongs to the Section Forest Ecophysiology and Biology)
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Abstract

:
Ecological significance of dormancy in desiccation-sensitive seeds is poorly understood. Quercus exhibits mutually exclusive occurrence of physiological (PD) and epicotyl dormancy (ePD), with no reported co-occurrence or dormancy class in other genera. We aimed to understand the dormancy in three Castanopsis species and document desiccation sensitivity and germination patterns concerning the embryonic axis position. We hypothesized that Castanopsis acorns are recalcitrant and potentially dormant. Fresh and cold-stratified acorns of Castanopsis chinensis, Castanopsis purpurella, and Castanopsis sclerophylla were subjected to desiccation and germination. Seedling emergence and internal morphology was monitored following cold (CS) and warm (WS) stratification. Fresh acorns had radicles emerge only after CS but require WS for shoot emergence. Drying to 20% moisture content led to complete death. In C. purpurella and C. sclerophylla, the embryonic axis was near the scar, and germination occurred by cracking the pericarp near the scar, which contrasts with C. chinensis. Moderate drying relieved dormancy due to the mechanical resistance of the pericarp. All three acorns were desiccation-sensitive and dormant. This is the first explicit report on PD and ePD co-occurrence in desiccation-sensitive seeds, but literature surveys allow for inference of such coexistence. CS alleviated PD and WS relieved ePD. Winter temperatures break PD, and acorns germinate during spring, but shoot emergence is delayed until summer. Our results are instructive for research on the dormancy of desiccation-sensitive species and the reproduction of Fagaceae species in subtropical forests.

1. Introduction

Desiccation-tolerant (DT) or orthodox seeds, dispersed at low moisture content (MC), can tolerate drying to 3%–7% MC and are amenable to ex situ conservation [1]. DT seeds can withstand desiccation, enabling post-dispersal survival in diverse ecosystems, establishing soil seed banks and synchronizing germination with the growing season through dormancy mechanisms [2]. However, such benefits are limited for desiccation-sensitive (DS) or recalcitrant seeds [3] which are dispersed at a high MC of c. 20%–40%. Desiccation negatively impacts DS seeds; thus, immediate germination to establish a ‘seedling bank’ is a prevalent survival strategy [4]. This is achieved by the restricted distribution of DS species in wet ecosystems, e.g., tropical rainforests, where sufficient water for germination occurs year-round [5]. Consequently, dormancy, arresting seed growth until the growing season, is redundant for DS, despite being present in a few species [6,7,8].
The family Fagaceae is globally distributed, adapted to temperate, tropical, and subtropical climates [9]. With over 1000 species encompassing nine genera, Fagaceae is a dominant family in evergreen broad-leaved or mixed coniferous broad-leaved forests [10]. Fagaceae species rely on the successful germination of dispersal units, botanically fruits or nuts (hereafter acorns). However, the germination ecology is complex because most—if not all—species of the dominant genus Quercus and others investigated hitherto show distinct DS characteristics [11,12,13]. The only exception to this generalization is a handful of species from Fagus having DT acorns [14]. Studies on the storage behavior of Fagaceae have received limited attention beyond Quercus. For example, the second largest genus, Castanopsis, consists of 143 species but has distribution restricted to Asian countries [15]; China has 58 Castanopsis species, 30 of which are endemic [16], and limited access to materials hinders research on Castanopsis storage behavior [17]. Several attempts to elucidate the Castanopsis drying response have yielded confusing results. Tian and Tang [18] found that, after drying at 19 ± 2 °C for 72 h, MC of C. fissa acorns declined from 42.7% to 17.3%, leading to complete vitality loss. This contradicts the slow drying rate reported in several Quercus [19] and C. sclerophylla [20], where the MC of acorns reaches c. 20% after 7–20 days of drying in silica gel, resulting in completed acorn death. Thus, knowledge of the drying response of Castanopsis is essential for restoration programs and ex situ conservation.
While many Fagaceae acorns have been known to germinate immediately after dispersal and thus are non-dormant, previous investigations have unveiled the existence of DS, physiological dormancy (PD), and epicotyl physiological dormancy (ePD). In Q. variabilis, Q. wutaishanica, and Q. robur [21], PD occurs due to mechanical restriction of a hard pericarp or hormonal imbalances. Cold stratification (CS) during winter alleviates dormancy, while mechanical scarification promotes germination under empirical conditions [22,23,24]. Some Quercus exhibit ePD, with radicle emergence immediately and shoot emergence delayed to spring or summer, such as Q. alba and Q. prinus [25]. Furthermore, according to the compilation presented by Sun et al. [26], dormancy types in Quercus are mutually exclusive, meaning a species can have either PD or ePD. Unfortunately, substantial efforts have been made to understand dormancy in Quercus, but the second-largest genus Castanopsis has received far less attention. Moreover, the co-occurrence of PD and ePD in one species has not been documented.
Our previous study on C. sclerophylla (data not published) suggests that acorns required over 3 months to germinate (radicle emergence) fully, consistent with results on four Castanopsis species from Malaysia having a mean minimum time for acorns to seedling of 102 days [27]. Acorns with PD require CS for rapid and uniform germination due to long germination time. C. fargesii acorns dispersed before winter generally germinate in the next April, with seedlings emerging around July when the climate warms up [28]. This apparent delay in seedling emergence indicates that Castanopsis acorns may have ePD and require higher ambient temperatures for seedlings growing. Nevertheless, some Castonopsis acorns were reported to have no dormancy, as germination occurred within 30 days [29,30]. The germination ecology of Castanopsis acorns remains unexplored, and recently Baskin and Baskin [31] suggested that it would not be unusual if Castanopsis exhibited both root and shoot dormancy.
It has been shown that the embryonic axis of Quercus is located just below the apex, and germination happens with the elongation of the radicle through the apex [19]. However, previously we found that the embryonic axis of C. sclerophylla acorns is on the opposite side of the apex and located just below the scar [20]. Sun et al. [26] showed the same embryonic axis and germination position of Q. chungii as our result. These observations presumably suggest that the anatomy and morphology of such unusual positioning of the embryonic axis bear no relationship to the genus and are likely species-specific. Nonetheless, Q. chungii has intermediate ePD [26], and whether embryo position would influence dormancy mechanisms in Fagaceae is unclear and formed an important objective of this study.
Because Quercus species have been primarily in the research limelight and, to a large extent, represent generalizations made from Fagaceae regeneration ecology, our study aims to shift this attention to the second dominant genera, Castanopsis. To this end, we investigated Castanopsis chinensis (Spreng.) Hance, Castanopsis purpurella subsp. purpurella, and Castanopsis sclerophylla (Lindl. & Paxton) Schottky, distributed predominantly in south China and other South Asian countries. They often grow in hills or mountain slopes and are considered the dominant species in subtropical evergreen broad-leaved forests of China [32]. We hypothesized that Castanopsis acorns are recalcitrant and potentially dormant and that the embryonic axis position will influence germination characteristics. The aims of this contribution using three Castanopsis acorns were as follows: (1) identify seed storage behavior; (2) determine dormancy type; (3) recognize the embryonic axis position and establish its relationship with germination pattern and seedling establishment; (4) synthesize literature on Castanopisis dormancy and compare with studied species.

2. Materials and Methods

2.1. Sample Site and Acorn Collection

Fresh fruits (acorns) of C. chinensis, C. purpurella, and C. sclerophylla were collected from Wuzhifeng Forestry Farm (25°48′ N–26°02′ E 114°06′–114°26′, Ganzhou, Jiangxi Province, China) at the time of natural dispersal in mid-November to early December 2021. The study site is located at the southern edge of the mid-subtropics and has a humid subtropical monsoon climate, with an average annual temperature of 17.4 °C and an annual rainfall of about 1810 mm. A typical mid-subtropical evergreen, deciduous broad-leaved forest covers the study site [33]. In 2019, the highest and lowest temperatures recorded at the study site were 39.3 °C in August and −3.0 °C in January, respectively. The climate data (temperature and precipitation) were obtained from the website of the Ganzhou Meteorological Bureau [34].
Acorns of all three Castanopsis species were hand-picked after gently shaking the trees. The collection included at least 20 trees distributed in a 6 km radius. Then acorns were placed in woven bags and transported to the University of Shanghai for Science and Technology within three days. Upon arrival, acorns were subjected to a floating test to remove empty and insect-infested acorns. The remaining acorns were disinfected with 1% sodium hypochlorite (NaOCl) solution for 30 min, followed by rinsing with distilled water thrice. After cleaning, all acorns were air dried for half day at room temperature (20 ± 1 °C) and then stored at 5 °C until use. Because we hypothesized DS is a possibility, all experiments began within two days following the model presented in Figure 1.

2.2. Basic Acorn Characteristics

Acorn mass was determined by weighing three replicates of 100 acorns, randomly picked, using a standard balance (0.001 g, BSA224S-CM, Sartorius, Göttingen, Germany). The moisture content of acorns was determined using the standard oven method [35]. For moisture content determination, four replicates of 5 acorns were dried in an oven at 103 °C for 17 h, and the change in acorn mass before and after drying produced the moisture content on a fresh weight basis (%, FWB). The average dimension of acorns was calculated by measuring the height, width, scar diameter, and ratio of scar diameter to width using a Vernier caliper (0.01 mm, MNT-150, Meinaite, Shanghai, China). The thickness of the scar and main pericarp was measured using a thickness gauge (0.001 mm, HT-790, Mitutoyo, Kanagawa, Japan). The hardness of the pericarp was measured by texture analyzer (TA.XT plus, Stable Micro System, Surrey, UK) on the entire scar region, and a quarter of the main pericarp was cut longitudinally. An individual pericarp was placed on the center of the analyzer platform, and a P/100 probe was used, which moved at a speed of 0.5 mm/s with a compression percentage of 50%. The peak force was used to measure the hardness of the pericarp. With no exception, the pericarps of 30 randomly selected acorns for each species were tested, and the values are expressed as the mean of individual measurements in N.

2.3. Acorn Desiccation and Germination Tests

To understand the response of acorns to drying, desiccation experiments were conducted after randomly assigning acorns to various groups. For each species, 6 groups, each consisting of 95 fresh acorns, were dried independently using silica gel with a weight ratio of 5:1 (silica gel:acorns) in completely sealed plastic boxes (l × b × w: 170 × 120 × 25 mm) at room temperature (22 ± 1 °C). One group of acorns from each species was retrieved at 0 (control—undried), 3, 7, 14, 21, and 28 days. After drying, a subset of 20 acorns (4 replicates of 5 acorns) was used for measuring moisture content using the standard oven drying method, and the remaining 75 acorns (3 replicates of 25 acorns) were used for the germination test. Germination tests were carried out by incubating acorns on 1% agar water in closed plastic boxes (l × b × w: 155 × 100 × 42 mm) that contained holes for gas exchange at 15/20 °C (12 h dark/12 h with a light intensity of 60 mol m−2 s−1). Acorns with radicles that emerged to at least 2 mm were concluded as germinated. Acorn germination percentage and pattern were observed for three months (CS acorns) to six months (fresh acorns) until no further germination occurred for three weeks. After ending the germination test, acorns were cut to examine the inside of the acorns. Germination percentages are expressed as mean ± standard deviation (SD). Desiccation sensitivity was quantified using the following quantal response models [36]:
V = V i 1 + e x p b W C − W C 50
where the viability, V, was the response, the moisture content, WC, was the stimulus, Vi was the initial viability (fresh acorns) or biggest viability (CS acorns), WC50 was the moisture content at which half of the initial viability was lost, and b was acorn lot-specific parameters. The desiccation sensitivity model was performed using least-squares regression as computed by the quasi-Newton method.

2.4. Dormancy Breaking Treatments

2.4.1. Effects of Cold Stratification (CS) on Fresh Acorns’ Physiological Dormancy (PD) Release and Desiccation Sensitivity

A total of 1600 fresh acorns were stratified in the dark at 4 °C for six months in airtight plastic boxes [37]. After CS, acorns were dried for 0, 1, 3, 5, 7, and 14 days, then tested for moisture content (four replicates of 5 acorns) and germinated (three replicates of 25 acorns) at 15/20 °C for each drying time. Because the control acorns dried for 21 and 28 days were completely dead, we did not dry the acorns for an extended period following CS. A total of 570 cold-stratified acorns of each species were used in the desiccation experiments.

2.4.2. Effect of Warmer Temperature (Warm Stratification) on Shoot Dormancy Breaking in Root-Emerged Acorns

One thousand undried, cold-stratified acorns were germinated at 15/20 °C until the cotyledonary petiole and radicle growth reached more than 2 mm. After incubation at 15/20 °C for 45 days, 120 acorns with radicles that emerged to 2 mm were selected randomly from all of the germinated acorns, 60 acorns (three replicates of 20 acorns) of which were transferred to a temperature of 25/30 °C for subsequent incubation while another randomly selected 60 root-emerged acorns continued to be incubated at a temperature of 15/20 °C. All root-emerged acorns were incubated on the 1% agar water substrate at different temperatures. The stage of germination and shoot emergence time were monitored every week for two months. These conditions mimic the spring and early summer temperatures at the study site. Our goal of this experiment was to observe if spring temperatures (15/20 °C) were sufficient for leaf emergence.

2.5. Morphological and Anatomical Analysis

For analysis of acorn morphology, photographs of acorns were taken with a camera (EOS 60D, Canon, Tokyo, Japan) at various germination stages. We photographed the germinating acorns weekly until the seedlings were fully developed, i.e., with both root and leaf emerged. Acorns were cut longitudinally with a scalpel along the long axis, and the embryonic axis was observed and photographed. The length of the cotyledonary petiole and root growth was determined using a Vernier caliper. To understand the restriction of pericarp on acorn germination morphology, pericarps of 10 acorns for each species were carefully detached with a scalpel, and acorns without pericarp were incubated at 15/20 °C on 1% agar water.

2.6. Literature Review of Dormancy in Castanopsis

We searched three major databases—Web of Science, China National Knowledge Infrastructure, and China Science and Technology Journal Database—between January and March 2023 for peer-reviewed publications using combinations of search terms relating to Castanopsis, seed (or seedling), acorns, and dormancy. The literature search was conducted for English and Chinese articles, as Castanopsis has highly restricted distribution in China, and a wealth of information is available in journals published in Chinese. We focused on the germination and pre-sowing treatments of Castanopsis in these articles and determined the type and level of seed dormancy based on the data collected in the literature. We followed the dormancy classification system developed by Baskin and Baskin [38].

2.7. Statistical Analysis

Basic acorn traits of three species were statistically compared using analysis of variance (one-way ANOVA) with Tukey’s post hoc test. A correlation analysis was performed for acorn size, scar size, and pericarp thickness to identify the relationship among these parameters among species. Data on acorn germination of different drying periods were analyzed statistically by one-way ANOVA. Subsequently, the mean values for each drying period were compared using Tukey’s post hoc test. Non-linear regressions of moisture content against drying time were performed by an exponential model. The quantification of the desiccation sensitivity of three species was carried out using least-squares regression computed by the quasi-Newton method [36], and the proportion of variance explained, R2, was used for evaluating the fit of the model. Based on the desiccation response fitted curve, critical moisture content (i.e., WC50) was estimated as a measure of the relative level of desiccation sensitivity for the acorn. The effect of drying periods and incubation time and their interaction on mean cumulative germination percentages were analyzed using univariate analysis in a general linear model (GLM) with Tukey’s post hoc test. Data on shoot emergence with different incubation temperatures were analyzed using one-way ANOVA. A significance threshold of p = 0.05 was retained for both ANOVA and univariate analysis. All the analyses were performed on SPSS version 21 and OriginPro 2021 V9.8.0.200.

3. Results

3.1. Acorn Basic Characteristics

With a 100-acorn mass of 56.10 g, C. chinensis acorns were smaller than C. purpurella and C. sclerophylla acorns; both had a 100-acorn mass close to 100 g (Table 1). The moisture content (MC) of freshly collected acorns ranged from 32.7% to 38.0% across species (Table 1). MC was directly proportional to the acorn mass (Pearson correlation coefficient = 0.801, p < 0.01); thus, smaller-sized C. chinensis had the lowest MC, and the heaviest acorns of C. sclerophylla had the highest MC (Table 1). The scar size was also directly proportional to the acorn mass (Pearson correlation coefficient = 0.996, p < 0.01). In particular, the scar region of C. purpurella and C. sclerophylla acorns occupied the larger area of the pericarp compared to C. chinensis. The ratio of the diameter of the scar to the width of both C. purpurella and C. sclerophylla acorns was 0.89, while the ratio for C. chinensis was 0.82 (Table 1). The pericarp characteristics of these three species also differed significantly, and there was a strong correlation between acorn mass and pericarp hardness and thickness. Acorn mass was negatively correlated with scar thickness and main pericarp thickness with Pearson correlation coefficients of −0.899 (p < 0.01) and −0.896 (p < 0.01), respectively. The Pearson correlation coefficients for acorn mass with scar hardness and main pericarp hardness were −0.538 and −0.789, respectively (p < 0.05). The heavier acorns of C. sclerophylla had lesser pericarp hardness and thickness than the other two species with less acorn mass (Table 1).

3.2. Germination of Fresh and Cold-Stratified Acorns

Fresh control acorns of all three species showed no germination within the first month of incubation at 15/20 °C. However, acorns began to germinate progressively during the second month and continued through the third month. There was some significant difference between species during early germination, but germination percentage reached a maximum between five and six months, with no further germination for three weeks. By the end of 6 months, the mean cumulative germination percentage of C. chinensis, C. purpurella, and C. sclerophylla was 94.67% ± 0.94%, 85.33% ± 0.94%, and 82.67% ± 0.94%, respectively (Figure 2a). At the end of the germination test, ungerminated seeds were cut and found to be internally deteriorated or blackened, with fungi on the surface of the acorns.
The MC of all three species dropped marginally during CS compared with the freshly collected acorns but remained above 30% (Figure 2b). CS did not affect the viability of the C. chinensis and C. sclerophylla acorns, and the final germination percentage was 85.33% ± 1.25% and 85.33% ± 0.94% after three months incubation, respectively (Figure 2b). While CS treatments had some negative effects on the viability of C. purpurella acorns, the control acorns had only 37% germination (Figure 2b).

4. Desiccation Response

Fresh (Figure 2a) and cold-stratified (Figure 2b) acorns displayed high sensitivity to desiccation. With an increase in drying time, the MC and the germination percentages of all three species dropped significantly. The MC declined to around 22%–29% (WC50), with only approximately 50% germination across species (Table 2 and Figure 2). Upon continuous drying for one week, the MC reached c. 20%, and none of the acorns of the three species germinated (Figure 2a). The drying rate showed remarkable similarity between species regardless of pericarp thickness, hardness, scar size, and acorn size using an exponential model to fit the observed data of MC with different drying periods, and showed that drying rates were not constant (Figure 2a,b). In the three species, acorn germination followed the typical S-shaped pattern when the moisture content of acorns declined during dehydration (Figure 2c,d). WC50, the moisture content at which half of the initial viability was reached, can be analyzed through the model to quantify the acorns desiccation sensitivity, and the higher the WC50 value, the more sensitive to desiccation for acorns.
The desiccation response of three Castanopsis acorns after CS is similar to that of fresh acorns; as drying progresses, the MC of the acorns drops to around 20% within 5–7 days, and the acorns lost their viability completely (Figure 2b). Drying had contrasting effects on acorn germination. In particular, C. purpurella acorns germinated about 25% more than the control acorns after 1 day of drying (Figure 2b). This also occurred with C. sclerophylla acorns, with 85.33% and 94.67% germination for control and dry 1-day acorns, respectively, but there were no statistically significant differences (Figure 2b).

4.1. Effects of Cold Stratification (CS) in Dormancy Break

The germination of fresh Castanopsis acorns was high (Figure 2a), but the acorns took 5–6 months to germinate. Following CS treatment, the germination time of the acorns was shortened (Table 3). All cold-stratified Castanopsis acorns essentially germinated after 3 months of incubation; germination for C. chinensis and C. sclerophylla acorns was above 80% (Figure 3a,c; Table 3). The final maximum germination of cold-stratified C. purpurella acorns was 60%, which was lower than that of fresh acorns (Figure 3b). In the first 30 days of incubation, the germination percentage of 1-day-dried C. chinensis acorns dried was more than that of the control. However, after 60 days, the cumulative germination percentage of the control C. chinensis acorns, i.e., CS but not dried, gradually exceeded that of 1-day-dried acorns (Figure 3a; Table 3). Yet, the mean cumulative germination percentages of the C. purpurella and C. sclerophylla acorns that had been dried for 1 day were consistently higher than those of the control (Figure 3b,c; Table 3). The C. sclerophylla acorns germinated nearly fully (80%) within 60 days of incubation (Figure 3c; Table 3). The 1-day drying treatment resulted in faster germination and did not affect the overall viability of the C. purpurella and C. sclerophylla acorns.

4.2. Effect of Warm Stratification (WS) on Shoot Emergence

At 15/20 °C, all control and desiccated acorns (fresh, without CS acorns) of all three species showed only the elongation of the radicle and no shoot emergence at the end of the 6-month germination experiment (Figure 4a(a6),b(b6),c(c7)). When CS acorns of all three species were incubated at 15/20 °C, only 1–2 of 20 acorns had elongated shoots after one week. There were only 46.67% ± 2.16% of C. chinensis acorns with elongated shoots within 4 weeks of incubation at 15/20 °C and 80.00% ± 0.82% of acorns had emerged shoots after 2 months (Figure 5). Transferring root-emerged acorns to a higher temperature environment (25/30 °C) significantly reduced the time required for shoot emergence. Acorns of all three species had about 30% of shoot emergence in the first week of incubation at 25/30 °C. The percentage of shoots that emerged acorns of C. chinensis was 91.11% ± 1.25% in 4 weeks at 25/30 °C, but only 4.44% ± 0.94% of acorns had shoot emerged between 4 and 8 weeks (Figure 5).

4.3. Acorn Morphology and Germination Type

In all Castonopsis species studied, three distinctive areas can be observed: (1) the main pericarp, (2) the apex, and (3) the scar (Figure 4). However, the position of the embryonic axis showed a remarkable difference between species. Acorns of C. chinensis are generally conical in shape and consist of a woody dark-brown pericarp and a brown fibrous endocarp. The entire interior of the C. chinensis acorn is filled with the embryo, and the embryonic axis is on the apex side (Figure 4a(a1–a4)). The protrusion of the radicle from the apex region manifests acorn germination. The radicle continues to grow and lengthen and, under appropriate conditions, the shoot emerges near the apex end and grows into stems and leaves (Figure 4a(a5–a9)).
On the other hand, C. purpurella (Figure 4b(b1–b4)) and C. sclerophylla (Figure 4c(c1–c4)) acorns are similar in morphology, with acorns being wide-conical. The brown pericarp, closely adherent to the endocarp, is difficult to separate. Inside the acorn, there is one (occasionally two) embryo(s), and the two cotyledons are irregular and tightly fitted. Unlike C. chinensis, the embryonic axis is located on the opposite side of the apex and below the junction between the main pericarp and the scar. However, in some cases, the embryonic axis is small and uncertain in its position, making it challenging to observe. A small round structure can generally be found as the embryonic axis from the slit gap between two cotyledons below the scar region. Germination is validated as the thickened and elongated embryonic axis breaking through the junction of the main pericarp and the scar, which needs to overcome the resistance of the surrounding tissues to protrude. Simultaneously, the pericarp of some acorns will split or partially fall off. Under suitable conditions, shoots will sprout from the node close to the scar region and grow into stems and leaves.
However, due to the specific location of the embryonic axis, when they germinate, some acorns of C. purpurella and C. sclerophylla were affected by the hard pericarp; hence, the radicle did not extend properly (Figure 6a). When these acorns are visually inspected, no radicle protrudes, and it appears only as a splitting of the pericarp, which would indicate that the acorns have not germinated (Figure 6a(a1)). Upon manually removing the pericarp, the radicle protruded inside the pericarp and showed a curved growth, with the part near the cotyledons thickening to a green color (Figure 6a(a2,a3)). Germination without the pericarp (Figure 6b(b1–b3)) revealed that the radicle could extend directly, and there is no thickening of the embryonic axis near the cotyledons compared with germination with the whole pericarp.

4.4. Literature Review of Dormancy in Castanopsis

A total of 22 articles with 10 species contained information about dormancy in Castanopsis. Acorns of these 10 species were inferred to have different levels of PD. More specifically, five species were deemed to have PD + ePD, and five had PD (Table 4). However, this review shows that a species could receive more than one dormancy class depending on the maturation site. For example, C. chinensis has been reported to be likely PD + ePD, ND, and merely having ePD. We included such species under PD + ePD because acorns from at least one population tend to have PD + ePD, although the levels cannot be determined accurately. However, other forms of dormancy including those caused by a water-impermeable coat (physical dormancy) and underdeveloped embryos (morphological dormancy) are absent. By generalizing the Castanopsis acorn maturation season, germination, and/or emergence times in literature and taking into account the climatic characteristics of Ganzhou, Jiangxi Province, where most of the acorns originated, we developed a theoretical model to describe the entire germination process after acorn maturation and dispersal (Figure 7). This model shows that the sequence of temperatures occurring during winter and spring might act as suitable dormancy-breaking cues under ecological conditions (Figure 7).

5. Discussion

Until now, little attention has been paid to the dormancy and germination ecology of Fagaceae species beyond Quercus. Indeed, to the best of our knowledge, this is the first detailed study addressing the acorn biology of Castanopsis. Freshly collected acorns of all three Castanopsis species were smaller in size compared to the average acorn mass of Fagaceae. According to the data available in the Seed Information Database [17], the average thousand-acorn mass of Castanopsis is 650 g, which is significantly lower than the average Fagaceae acorn mass of 2500 g per thousand acorns (Jaganathan et al. in review). Despite producing smaller acorns, the anatomy of three Castanopsis species displayed a distinguishable scar (hilum), main pericarp, and apex, implying that this characteristic is common among Fagaceae species [19,20,26,58,59]. Furthermore, acorn size influences the pericarp thickness and scar size (Table 1), a feature that appears to be prevalent in Quercus [19,60,61,62].
A complete loss in viability after drying to c. 20% moisture content confirms the desiccation-sensitive behavior of all three species (Figure 2a,c). For most of the Quercus, the drying rate of the acorns varies between species, but it takes at least a week, even when dried above silica gel, to reach the minimum moisture content at which the acorns can survive, i.e., c. 15%–20% [63], although the LT50 (50% germination) occurs after drying the acorns to 25%–35% [64]. However, all three Castanopsis acorns tested here took only three days to reach critical levels, and after seven days of drying, the acorns lost complete viability (Figure 2a,b). Scar size does not control the rate of water loss, as previously observed in some Quercus [19,65]. Taken together, Castanopsis acorns are more sensitive to desiccation, which is possibly related to the lower initial moisture content at dispersal and the small acorn size. The climatic conditions of the region where the acorns matured may also affect the desiccation sensitivity. In C. sclerophylla from Thousand Island Lake, China, 27% of the acorns germinated when the moisture content reached 22.5% after drying for 7 days [20], whereas the moisture content of C. sclerophylla acorns from Ganzhou, Jiangxi Province, China in this study reached 22.9% after 7 days of drying and all the acorns were dead (Figure 2a). Plausibly, the higher summer temperature and annual precipitation in Ganzhou, Jiangxi Province might account for the high desiccation sensitivity of C. sclerophylla acorns in this study (Figure 7). This suggests that acorns maturing in different environments may show different responses to drying, regardless of the reason. This view is shared by Xia et al. [66], who showed that acorns of Quercus from cold regions are more desiccation-sensitive than those from warmer areas. As a common feature across Fagaceae, the maternal environment in which the acorns mature influences their survival during drying.
The absence of germination in all three species for 30 days indicate the presence of PD. This result is further supported by the fact that CS significantly shortened the time needed for completing germination (Figure 3, Table 3). Across species, CS reduced the time required for full germination to less than 2 months. This would be an adaptive strategy for the Castanopsis acorns in the study area because the dispersal time coincides with the long winter season between November and February (Figure 7). Our preliminary studies showed no germination when incubated on 1% agar water at 5 °C for up to 6 months. The depth of PD varies between species, as one-month CS relieved PD in approximately 20% of the acorns across species, and the number of acorns germinating increases with an increase in CS time. This parallels results observed in several Quercus species with PD [25,31,67,68].
It is worth noting that most previous studies have measured successful germination of Fagaceae acorns as a radicle protrusion and establishing root [69,70,71,72], thus limiting our ability to understand the full germination process until leaf emergence. On the other hand, many Quercus species are known to have ePD, with the root emerging immediately after dispersal or after PD is broken, but with delayed shoot emergence [26,51,68,73,74]. It has been described recently in Q. robur that the extended-duration CS and warmer incubation temperature shorten the root emergence time. However, the lag time between radicle and epicotyl emergences was only related to incubation temperature, not CS duration [75]. Existing literature shows that most Castanopsis acorns have delayed germination, with some taking 3–5 months from sowing to seedling emergence (Table 4). Ng [27] reported that C. foxworthyi and Lithocarpus cyclophorus acorns require 227–432 and 151–255 d to germinate (seedling emergence), respectively. Many studies have opted for pre-sowing treatments to reduce germination time, e.g., prolonged CS, GA3 treatment, manual scratching of pericarp treatment, etc. (Table 4). In our study, more acorns had their leaves emerge after experiencing winter → spring → summer temperatures. Thus, the acorns establish roots during spring, and the leaves begin to sprout with the arrival of a warmer climate (Figure 7). Because the acorns are large with rich nutrients, such a slow germination process ensures seedling survival [76]. Thus, we infer that most Castanopsis have PD and ePD. Alternatively, the climatic conditions in which these acorns were maturing could potentially affect their dormancy levels (Table 4).
We found that the WC50 decreased in both C. purpurella and C. sclerophylla after CS (Table 2), indicating that the CS acorns could tolerate lower moisture content than fresh acorns, an effect of the reduced desiccation sensitivity after acorns release PD. This phenomenon was present in C. purpurella and C. sclerophylla but not in C. chinensis and is likely to be related to dormancy due to the unique position of the embryonic axis and the mechanical resistance of the pericarp (Figure 4 and Figure 6). It is interesting to highlight that the moisture content of CS plus one-day-desiccated C. purpurella and C. sclerophylla acorns decreased, but the germination percentage was higher than that of CS acorns (Figure 2b and Figure 3b,c). For C. purpurella and C. sclerophylla acorns, CS alone may not be sufficient to break dormancy, as we found that cold-stratified acorns of C. purpurella and C. sclerophylla with one-day drying treatment had increased germination and shortened germination time compared to cold-stratified acorns without desiccation (Figure 3b,c; Table 3). Slightly dried seeds (one-day-dried CS acorns) showed better germination, further demonstrating reduced desiccation sensitivity (can tolerate lower moisture content) when the dormancy of pericarp restriction was released (Figure 2b and Figure 3b,c). Similar behavior was shown by Yu et al. [30] on C. purpurella acorns from a tropical seasonal rainforest in southern China, where germination percentage peaked at over 90% after 48 h of dehydration and subsequently began to decline as drying progressed. This germination trait could be intimately linked to modifications in the structural characteristics of acorns after desiccation. Thus, moderate drying is beneficial for Castanopsis acorns, but the effect appears to be species-specific.
The three Castanopsis species studied here show distinctive germination characteristics. In this regard, the thickness and hardness of the pericarp of C. chinensis were greater than those of C. purpurella and C. sclerophylla (Table 1). As the embryonic axis of C. chinensis was located at the apex side, the radicle directly protruded through the apex when acorns started to germinate, so the structural characteristics of C. chinensis pericarp tissue had no bearing on its germination (Figure 4a). However, the embryonic axis of C. purpurella and C. sclerophylla was situated near the scar region (Figure 4b,c); the thickness and hardness of the scar region are higher than the main pericarp (Table 1), so the radicle must extend out through the irregular scar region when germinating. Following the Jaganathan and Pharyal (in review) germination model of Fagaceae, C. chinensis shows Type-IA adjacent ligular germination because germination occurs through the apex, and C. purpurella and C. sclerophylla show Type-IS adjacent ligular with germination occurring just under the scar. Due to the restriction of the resistance of the pericarp tissue around the scar, it was difficult for some acorns to extend, and for some acorns, the radicle even grew directly inside the pericarp (Figure 6a), which delayed germination and even affected the determination of whether the acorns germinate or not. In the case of slightly dried acorns, the pericarp structure may have changed during desiccation, such as minor cracks or reduced hardness, making the radicle more easily protruded and, to some extent, releasing the dormancy. Hasnat et al. [51] demonstrated the existence of dormancy in C. Indica acorns, and the germination percentage and seedling quality can be improved by rubbing the pericarp with sandpaper as a pre-sowing treatment method.
Although germination in most Fagaceae species occurs due to radicles extending through the apex, this is not the only germination pattern. Indeed, our previous work on C. sclerophylla showed that the embryonic axis near the scar region breaks open the pericarp and extends [20]. Likewise, working with Q. chungii, Sun et al. [26] reached the same conclusion, demonstrating that this variation is not restricted to Castanopsis. Here, we found that the embryonic axis in C. chinensis acorns is located below the apex, and germination initiates with the radicle extending through the apex and leaves sprout after breaking ePD (Figure 4a). In contrast, the embryonic axis of C. purpurella and C. sclerophylla acorns is located below the scar region (Figure 4b,c). When acorns germinated, the radicle broke through the pericarp tissue near the scar with the shoot, and leaves grew after ePD was released. In Fagaceae, the apex is the weak part of the acorn, and extending the radicle is easily achievable even without external water. The two species with an embryonic axis below the scar have deep PD and ePD. The ecological benefits of these varied germination patterns are unknown, but it nonetheless suggests an adaptive mechanism which remains to be elucidated in future studies.

6. Conclusions

Castanopsis, the second dominant genera in Fagaceae, has not received sufficient attention due to its limited distribution range. Our research has revealed that the mechanical resistance of the pericarp and embryonic axis position are contributing factors to a deeper PD in acorns. Prolonged CS and moderate drying can release PD, while WS alleviates ePD. The ecological changes in Castanopsis acorns during winter, spring, and summer correspond to the process of breaking PD, root emergence after PD break, and shoot emergence after ePD break. All three Castanopsis acorns were desiccation-sensitive and had dormancy; releasing PD can reduce their sensitivity to drying in two species. Thus, our results on the ecological significance of dormancy in desiccation-sensitive seeds of Castanopsis species will be valuable for all the researchers who are interested in seed physiology and ecology as well as forest ecological restoration.

Author Contributions

Conceptualization, J.L., G.K.J. and B.L.; methodology, J.L. and H.K.; software, J.L. and H.K.; validation, J.L.; formal analysis, G.K.J.; investigation, J.L. and G.K.J.; resources, J.L.; data curation, J.L. and H.K.; writing—original draft, J.L.; writing—review and editing, G.K.J.; visualization, J.L.; supervision, G.K.J. and B.L.; project administration, G.K.J. and B.L.; funding acquisition, G.K.J. and B.L. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the National Natural Science Foundation of China (NSFC), grant number 32001119.

Data Availability Statement

The data underlying this article is available in the article.

Conflicts of Interest

The authors declare no conflict of interest.

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Forests 14 02330 g001
Figure 1. Flow chart showing experimental plan and analysis. Dotted arrows indicate morphological and anatomical analysis of seedlings.
Figure 1. Flow chart showing experimental plan and analysis. Dotted arrows indicate morphological and anatomical analysis of seedlings.
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Figure 2. The relationship between moisture content, drying period, and germination in Castanopsis chinensis, Castanopsis purpurella, and Castanopsis sclerophylla for fresh control acorns (a,c) and cold-stratified acorns (b,d). The effect of desiccation on germination percentage (bar chart corresponds to the right y-axis) and moisture content (% fresh weight, scatter, and exponential fitted model correspond to the left y-axis). Germination percentage after desiccation to various moisture content and fitted pattern of the desiccation sensitivity model as calculated by the quasi-Newton method (c,d). Different lower-case letters indicate significant differences in germination percentages at p < 0.05 for each species.
Figure 2. The relationship between moisture content, drying period, and germination in Castanopsis chinensis, Castanopsis purpurella, and Castanopsis sclerophylla for fresh control acorns (a,c) and cold-stratified acorns (b,d). The effect of desiccation on germination percentage (bar chart corresponds to the right y-axis) and moisture content (% fresh weight, scatter, and exponential fitted model correspond to the left y-axis). Germination percentage after desiccation to various moisture content and fitted pattern of the desiccation sensitivity model as calculated by the quasi-Newton method (c,d). Different lower-case letters indicate significant differences in germination percentages at p < 0.05 for each species.
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Figure 3. Mean (±SD) cumulative germination percentages for Castanopsis chinensis (a), Castanopsis purpurella (b), and Castanopsis sclerophylla (c) acorns dried for different times after cold stratification. The label 0–30 d denotes from the start of germination (day 0) to the 30th day after germination; 30–60 d, the 30th day after the start of germination to the 60th day after germination; 60–90 d, the 60th day after the start of germination to the 90th day after germination.
Figure 3. Mean (±SD) cumulative germination percentages for Castanopsis chinensis (a), Castanopsis purpurella (b), and Castanopsis sclerophylla (c) acorns dried for different times after cold stratification. The label 0–30 d denotes from the start of germination (day 0) to the 30th day after germination; 30–60 d, the 30th day after the start of germination to the 60th day after germination; 60–90 d, the 60th day after the start of germination to the 90th day after germination.
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Forests 14 02330 g004
Figure 4. Photographs of Castanopsis chinensis (a), Castanopsis purpurella (b), and Castanopsis sclerophylla (c) acorn structure and different stages of germination. SR, scar; AP, apex; P, pericarp; COT, cotyledons; E, embryonic axis; R, radicle; S, shoot; L, leaves; RT, root. Whole acorns (a1,a2); acorns with removed pericarp (a3); acorns separated along the cotyledon edge (a4); germinated acorns of different stages (a5–a9). In (b), (b1–b9) and in (c), (c1–c9) corresponds to the descriptions provided for (a1–a9).
Figure 4. Photographs of Castanopsis chinensis (a), Castanopsis purpurella (b), and Castanopsis sclerophylla (c) acorn structure and different stages of germination. SR, scar; AP, apex; P, pericarp; COT, cotyledons; E, embryonic axis; R, radicle; S, shoot; L, leaves; RT, root. Whole acorns (a1,a2); acorns with removed pericarp (a3); acorns separated along the cotyledon edge (a4); germinated acorns of different stages (a5–a9). In (b), (b1–b9) and in (c), (c1–c9) corresponds to the descriptions provided for (a1–a9).
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Forests 14 02330 g005
Figure 5. Mean (±SD) cumulative shoot emergence percentages for CS acorns of Castanopsis chinensis, Castanopsis purpurella, and Castanopsis sclerophylla with different incubation temperatures. * shows significant differences in one-way ANOVA in shoot emergence with different incubation temperatures at p < 0.05 for the same incubator time. The label 0–4 w denotes from the start of germination (0 w) to 4th week after germination; 4–8 w, 4th week after the start of germination to 8th week after germination.
Figure 5. Mean (±SD) cumulative shoot emergence percentages for CS acorns of Castanopsis chinensis, Castanopsis purpurella, and Castanopsis sclerophylla with different incubation temperatures. * shows significant differences in one-way ANOVA in shoot emergence with different incubation temperatures at p < 0.05 for the same incubator time. The label 0–4 w denotes from the start of germination (0 w) to 4th week after germination; 4–8 w, 4th week after the start of germination to 8th week after germination.
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Forests 14 02330 g006
Figure 6. Photographs of acorn germination. Castanopsis sclerophylla acorn germinated with the whole pericarp (a), germinated acorn with pericarp, but no root seen protruding (a1), acorns of a1 after peeling the pericarp with the root emerged (a2,a3); acorns germinated without pericarp (b), Castanopsis purpurella (b1), Castanopsis sclerophylla (b2), and Castanopsis chinensis (b3). COT, cotyledons; P, pericarp; R, radicle.
Figure 6. Photographs of acorn germination. Castanopsis sclerophylla acorn germinated with the whole pericarp (a), germinated acorn with pericarp, but no root seen protruding (a1), acorns of a1 after peeling the pericarp with the root emerged (a2,a3); acorns germinated without pericarp (b), Castanopsis purpurella (b1), Castanopsis sclerophylla (b2), and Castanopsis chinensis (b3). COT, cotyledons; P, pericarp; R, radicle.
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Forests 14 02330 g007
Figure 7. Average monthly temperature (blue line with square plots) and rainfall (bars) at the acorn collection site (Ganzhou, Jiangxi) and theoretical model depicting acorn germination time and seedling establishment time. Different colored plots and bars correspond to different seasons.
Figure 7. Average monthly temperature (blue line with square plots) and rainfall (bars) at the acorn collection site (Ganzhou, Jiangxi) and theoretical model depicting acorn germination time and seedling establishment time. Different colored plots and bars correspond to different seasons.
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Table 1. Basic acorn characteristics of 3 Castanopsis species.
Table 1. Basic acorn characteristics of 3 Castanopsis species.
SpeciesC. chinensisC. purpurellaC. sclerophylla
100 acorn mass (g)56.10 ± 0.64 a96.93 ± 1.17 b99.23 ± 1.26 b
Initial moisture content (% FWB)32.69 ± 0.73 a35.49 ± 2.21 ab38.01 ± 1.33 b
Height (mm)10.38 ± 0.83 a13.20 ± 1.06 b13.12 ± 1.05 b
Width (mm)9.90 ± 0.56 a11.84 ± 1.00 b11.96 ± 0.95 b
Scar diameter (mm)8.09 ± 0.77 a10.59 ± 1.25 b10.70 ± 1.14 b
Ratio of scar diameter to width0.817 ± 0.056 a0.893 ± 0.057 b0.888 ± 0.052 b
Scar thickness (mm)0.837 ± 0.151 a0.717 ± 0.138 b0.651 ± 0.143 b
Main pericarp thickness (mm)0.620 ± 0.073 a0.537 ± 0.104 b0.465 ± 0.080 c
Scar hardness (N)20.564 ± 5.46 a11.011 ± 3.63 b7.266 ± 2.36 c
Main pericarp hardness (N)20.769 ± 5.10 a10.154 ± 3.34 b6.241 ± 1.89 c
Natural dispersal timeSeptember–DecemberSeptember–DecemberOctober–December
Data are means ± standard deviation (SD). Different lower-case letters indicate a significant difference among the three Castanopsis in each basic characteristic (p < 0.05).
Table 2. Relevant parameters of the desiccation sensitivity model curve in Figure 2c,d.
Table 2. Relevant parameters of the desiccation sensitivity model curve in Figure 2c,d.
ViR2WC50 (FWB)b
Fresh acornsC. chinensis94.67122.627 ± 0.0010.87509
C. purpurella85.330.9977627.848 ± 0.2140.50127
C. sclerophylla82.670.9995228.594 ± 0.0900.44457
CS acornsC. chinensis85.330.9988024.413 ± 0.0770.73186
C. purpurella61.330.7939827.347 ± 1.3330.44289
C. sclerophylla94.670.9679127.450 ± 0.6250.43443
Vi: the initial viability (fresh acorns) or biggest viability (CS acorns); R2: proportion of variance explained by the desiccation sensitivity model; WC50: the moisture content at which half of the initial viability was lost; b: specific parameter in Formula (1); FWB: the moisture content on a fresh weight basis (%); b is acorn lot-specific parameters.
Table 3. Mean (±SD) germination percentages and germination time for Castanopsis chinensis (a), Castanopsis purpurella (b), and Castanopsis sclerophylla (c) acorns dried for different periods after CS. Different letters indicate significant differences in germination percentages at p < 0.05 for the same incubation time (lower-case) and drying period (upper-case) for each species based on Tukey’s post hoc test. SD, standard deviation.
Table 3. Mean (±SD) germination percentages and germination time for Castanopsis chinensis (a), Castanopsis purpurella (b), and Castanopsis sclerophylla (c) acorns dried for different periods after CS. Different letters indicate significant differences in germination percentages at p < 0.05 for the same incubation time (lower-case) and drying period (upper-case) for each species based on Tukey’s post hoc test. SD, standard deviation.
SpeciesGermination Time (Days)Drying PeriodF Ratiop-Value
0 (Day)1 (Days)3 (Days)5 (Days)7 (Days)
C. chinensis0–3030.67 ± 0.47 Aa45.33 ± 1.70 Ab14.67 ± 1.25 Ac4.00 ± 0.00 Acd0.00 ± 0.00 Ad48.0240.000
30–6036.00 ± 1.25 ABa10.67 ± 0.00 Bb0.00 ± 0.00 Bb0.00 ± 0.00 Ab0.00 ± 0.00 Ab42.8130.000
60–9018.67 ± 1.25 Ba2.67 ± 0.00 Bb0.00 ± 0.00 Bb0.00 ± 0.00 Ab0.00 ± 0.00 Ab9.3000.002
F ratio6.65028.93317.2861.750
p-Level0.0300.0010.0030.252
C. purpurella0–309.33 ± 1.25 Aa37.33 ± 1.25Ab9.33 ± 2.05 Aa4.00 ± 0.82 Aa0.00 ± 0.00 Aa16.8750.000
30–6010.67 ± 2.45 Aab14.67 ± 1.41 Ba8.00 ± 1.89 Aab1.33 ± 1.25 Ab0.00 ± 0.00 Ab7.2330.005
60–9017.33 ± 1.25 Aa9.33 ± 0.47 Bab2.67 ± 2.16 Ab0.00 ± 0.00 Ab0.00 ± 0.00 Ab10.4330.001
F ratio1.47620.7220.9131.750
p-Level0.3010.0020.4510.252
C. sclerophylla0–3041.33 ± 0.94 Aa50.67 ± 2.49 Aa28.00 ± 2.83 Aab16.00 ± 1.41 Abc0.00 ± 0.00 Ac14.7600.000
30–6032.00 ± 0.94 Ba29.33 ± 1.63 ABa22.67 ± 1.70 Aa1.33 ± 1.70 Bb2.67 ± 1.70 Ab46.4230.000
60–9012.00 ± 0.94 Cab14.67 ± 0.47 Ba12.00 ± 0.47 Aab1.33 ± 1.70 Bab0.00 ± 0.00 Ab5.0000.018
F ratio94.75012.2892.03611.0004.000
p-Level0.0000.0080.2110.0100.079
Table 4. Castanopsis species known to have germination characteristics and dormancy type.
Table 4. Castanopsis species known to have germination characteristics and dormancy type.
SpeciesTemperature (°C)Germination Time (days)Germination (%)Seed Collection TimeTime of Initiation of GerminationIncubation EnvironmentPre-Sowing TreatmentDormancy TypeGermination JudgmentCollection Site (Province)Reference
C. carlesii >15055DecemberMay–JuneSoil burial test PD 3 + ePDRadicle/seedling emergenceChongqing[39]
C. chinensis 90–12024–66DecemberMid MarchNursery (shade house) PD 3 + ePDSeedling emergenceGuangdong[40]
C. chinensis25/2027 ChamberSoak in water for 12 hND Guangdong[41]
C. chinensis20/35, 18~323–7 in incubator, 60-180 in field>60November Chamber/soil burial test ePDRadicle/seedling emergenceGuangdong[32]
C. fargesii Late October–early NovemberApril–MayNurseryCS in wet sand; Soak in water for 3–5 dPD *Seedling emergenceChongqing[42]
C. fargesii23/134–2085–64NovemberMarchChamberStorage in wet sand for 4 monthsPD 3Radicle emergenceChongqing[43]
C. fargesii256–2095DecemberFebruaryChamberStorage for winter and soak in cold water for 24 hPD *Radicle emergenceJiangxi[44]
C. fargesii 60–9011 seeds, 45 seedlingsDecemberFebruarySoil seed bank PD 2Seedling emergenceGuizhou[45]
C. fargesii >15059DecemberMay–JuneSoil burial test PD 3 + ePDRadicle/seedling emergenceChongqing[39]
C. fargesii 51.9–82.566.6–81.0NovemberMaySoil burial testStorage in wet sand, fridge (3 °C), and wet soil for 3 monthsPD 3 Chongqing[46]
C. fargesii 12031.7–33.3November–DecemberAprilSoil burial test PD 3 + ePDRadicle/seedling emergenceZhejiang[47]
C. fargesii >150 germination, >210 seedling October–November Soil seed bank PD 3 + ePDSeedling emergenceChongqing[28]
C. fissa31.54–6030–35DecemberMarchGreenhouseStorage at 4 °C for 3 monthsPD 2, PD 3 Guangdong[48]
C. fissa25/2010 ChamberSoak in water for 12 hND Guangdong[41]
C. purpurella306–2097 in incubator, 25–59 in the fieldSeptember–November Chamber/shade house ND Yunan[30]
C. purpurella 1207.2 seedlings May–JuneSoil burial test PD 3 + ePDSeedling emergenceFujian[49]
C. purpurella A. DC 68–98 NurserySoak in 98% concentrated sulphuric acid for 15 to 30 minPD * Guangdong[50]
C. indica 174.625–67AugustOctoberNurseryControl; sandpaper rubbingPD 3 + ePDSeedling emergenceBangladesh[51]
C. kawakamii25/15 57November–December ChamberStorage at 5 °C for 60–80 d, 50 mg/L GA3 24 h, manual scratching of seed coatPD 2 Fujian[52]
C. kawakamii15~2020 September–November Soil burial testStorage in wet sand for 60–80 d, 300–500 mg/L GA3 24 hPD 2 Guangdong[53]
C. kawakamii Hayata2513.5463–78October Artificial climate chamberStorage in wet sand at 5 °C for 60–80 d; 10–50 mg/L GA3 24 h; remove the seed coat; incubation at 40–50 °C for 2.5 hPD 2Radicle emergenceFujian[54]
C. lamontii30–20 82October Artificial climate chamberSoak in water at 40 °C for 24 h, 50 mg/L GA3 24 hPD 1 Fujian[55]
C. sclerophylla15–3516100 without pericarpOctober ChamberStorage at 4 °C, remove the pericarpPD * Jiangxi[56]
C. tibetana Hance28/20 39late OctoberAprilArtificial climate chamberStorage in wet sand; soak in water for 24 hPD * Zhejiang, Anhui[57]
PD 1, non-deep physiological dormancy; PD 2, intermediate physiological dormancy; PD 3, deep physiological dormancy; ePD, epicotyl physiological dormancy; PD *, levels of physiological dormancy cannot be determined; ND, non-dormant. The blanks indicate missing information.
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Li, J.; Jaganathan, G.K.; Kang, H.; Liu, B. The Co-Occurrence of Physiological and Epicotyl Physiological Dormancy in Three Desiccation-Sensitive Castanopsis (Fagaceae) Acorns from China with Specific Reference to the Embryonic Axis Position. Forests 2023, 14, 2330. https://doi.org/10.3390/f14122330

AMA Style

Li J, Jaganathan GK, Kang H, Liu B. The Co-Occurrence of Physiological and Epicotyl Physiological Dormancy in Three Desiccation-Sensitive Castanopsis (Fagaceae) Acorns from China with Specific Reference to the Embryonic Axis Position. Forests. 2023; 14(12):2330. https://doi.org/10.3390/f14122330

Chicago/Turabian Style

Li, Jiajin, Ganesh K. Jaganathan, Han Kang, and Baolin Liu. 2023. "The Co-Occurrence of Physiological and Epicotyl Physiological Dormancy in Three Desiccation-Sensitive Castanopsis (Fagaceae) Acorns from China with Specific Reference to the Embryonic Axis Position" Forests 14, no. 12: 2330. https://doi.org/10.3390/f14122330

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