GIS-Facilitated Germination of Stored Seeds from Four Wild-Growing Populations of Petromarula pinnata (L.) A. DC.—A Valuable, yet Vulnerable Local Endemic Plant of Crete (Greece)
by
, , , , and
Ioannis Anestis
1,
Elias Pipinis
2,
Stefanos Kostas
1,
Eleftherios Karapatzak
3
,
Eleftherios Dariotis
3,
Veroniki Paradeisopoulou
1,
Vasileios Greveniotis
4,
Georgios Tsoktouridis
3,
Stefanos Hatzilazarou
1,* and
Nikos Krigas
3,*
1
Laboratory of Floriculture, School of Agriculture, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece
2
Laboratory of Silviculture, School of Forestry and Natural Environment, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece
3
Institute of Plant Breeding and Genetic Resources, Hellenic Agricultural Organization Demeter, P.O. Box 60458, 57001 Thermi, Greece
4
Institute of Industrial and Forage Crops, Hellenic Agricultural Organization Demeter, 41335 Larissa, Greece
*
Authors to whom correspondence should be addressed.
Agronomy 2024, 14(2), 274; https://doi.org/10.3390/agronomy14020274
Submission received: 30 December 2023
/
Revised: 17 January 2024
/
Accepted: 24 January 2024
/
Published: 26 January 2024
(This article belongs to the Special Issue Effect of Agronomic Treatment on Seed Germination and Dormancy)
Abstract
:The ex situ conservation and sustainable exploitation of neglected or underutilized plant species (NUPs) is an urgent and vital endeavor. To this end, we focused on Petromarula pinnata (Campanulaceae), a vulnerable local plant endemic to Crete (Greece) that has been garnering interest for its agro-alimentary, medicinal, and ornamental value. A GIS ecological profile was established herein based on the natural distribution of this species in Crete. This profile contains detailed information on the climatic conditions (minimum, maximum, and mean temperatures; precipitation), as well as information on 19 bioclimatic variables that shape its natural adaptations. This profiling contributed to a better understanding of the species’ ecological requirements and facilitated germination trials employing stored seeds from four distinct populations (two from lowlands and two from semi-mountainous areas) at four temperatures (10, 15, 20, and 25 °C) and two light conditions. The results presented here show that both incubation temperature and population of origin, as well as the interaction between these variables, significantly affected seed germination rates. Incubation temperatures of 10 and 15 °C were the most appropriate for the successful germination of this species (>81.25% for both temperatures in three out of four populations), with light conditions having no effect on seed germination (86% in light and 80% in darkness). The establishment of a protocol for the successful germination of P. pinnata seeds opens avenues for further sustainable exploitation of this valuable yet vulnerable NUP as a new Greek native crop.
1. Introduction
Several members of Campanulaceae are commonly appreciated as ornamental plants worldwide, and many of them are praised for their attractive flowers, which can vary in color from deep violet to the palest milky blue [1]. Consequently, many members of Campanulaceae are often used as garden plants, potted plants, and possibly as cut flowers. These plants thus represent a growing trend in the development of new ornamental plants [2,3]. Within Campanulaceae, some genera and/or species are unique, such as the monotypic genus Petromarula, with P. pinnata (L.) A. DC., which is exclusively found on the island of Crete, Greece [4,5].
Beyond the ornamental appreciation of Campanulaceae species, people in France, Italy, and Spain are reported to consume raw roots of Campanula rapunculus L. in salads or in hot soups, while Chinese people also traditionally use dried roots of Codonopsis pilosula (Franch.) Nannf. in soups. Plants for all of these uses are sourced directly from the wild due to their nutritional benefits [6,7]. Previous studies have shown that the aerial parts (mainly leaf rosettes) of several Campanula spp. sourced from the wild, such as C. rapunculus in Spain [8,9], C. pelviformis L. in Crete [10] and P. pinnata in Crete [11], are used as wild edible greens in salads. Current research has shown that such ethnobotanical wild food sources can also provide particular nutritional value and may have medicinal properties, as illustrated in the case of C. pelviformis [10] or P. pinnata [12]. The latter local Cretan species is a key ingredient in small traditional pastries filled with a mixture of wild greens, cultivated vegetables, and local cheeses, called ‘kaltsounia’ [11]. Consuming just two pieces of Cretan ‘kaltsounia’ can provide nearly 40% of the estimated daily intake of strong antioxidants (flavonols and flavones) in the local Mediterranean diet in only 100 g of food [11].
Sustainable exploitation of neglected and underutilized plant species (NUPs) in agricultural settings [13] may alleviate the pressure on wild populations of threatened local endemic plants created by over-harvesting [3]. In response to this need, propagation and cultivation practices for NUPs must first be developed and established [3,14]. To this end, sexual propagation constitutes a highly effective and low-cost solution for the production of many plant species [15], as well as an appropriate propagation method for conservation-oriented research due to maintenance of high genetic diversity in the species of concern [16,17]. Seed-germination tests provide detailed and species-specific information on the range of environmental conditions required for the germination of fresh or stored mature seeds, thus shedding light on how target species adapt to their natural environment during this critical phase of their life cycle, e.g., [3,16,17,18,19,20,21].
As a support for seed-germination studies, species-specific ecological profiles provide comprehensive information on the autecology of the species of concern, including insight into the prevailing climatic conditions at the natural distribution sites of the species throughout the year, which may facilitate the design of seed-germination experiments [16,17]. To this end, identifying and confirming through experimentation the temperature range that favors high percentages of seed germination of a certain species can effectively enhance their reproduction potential, thus facilitating conservation actions or ensuring the production of seedlings for further development of cultivation protocols. Such protocols can then be used to create new value chains even for rare and/or threatened NUPs [3,20,22]. Moreover, the species-specific germination protocols derived from such studies are also very useful for large-scale production of newly introduced plant species by commercial nurseries, thus representing basic knowledge that can be exploited in the ornamental-floriculture sector or the medicinal and agro-alimentary industries [3,18,23].
In this context, an analytical seed-germination protocol has been developed for a threatened, range-restricted, local Cretan endemic NUP, C. pelviformis, which is of ornamental, medicinal, and agro-alimentary interest [17]. Additional research has also confirmed the presence of various isolated compounds with notable pharmaceutical properties in C. pelviformis [10], paving the way for the sustainable exploitation of this NUP as part of a newly launched national research project employing pilot cultivation in Crete (N. Krigas, pers. comm.). This study is focused on Petromarula pinnata (L.) A.DC., which has the above-described uses and ranks high among other Mediterranean NUPs, making it potentially highly valuable in the context of sustainable exploitation. This study aims to enhance the limited data on seed germination of P. pinnata [19,24] and builds upon previous studies that showed this species to have significant potential and value in different economic sectors [3], acting in parallel with undertaken pharmacognostic investigations (D. Lazari, pers. comm. and manuscript in preparation). Such applied research can also contribute to the conservation of this vulnerable species through targeted collection in the natural environment, which will allow for enhanced ex situ conservation in seed banks, and species-specific propagation and cultivation. The latter will enhance our practical knowledge of these plants, which can then be applied to re-introduction when needed or to pilot production of plant material for novel cultivations [19,25]. Considering all of the points above, the present investigation aimed to achieve the following objectives: (i) using Geographical Information Systems (GIS), to identify the climate range and the ideal conditions for seed germination in P. pinnata based on the ecological profiles of the sites where P. pinnata grows in the wild, and (ii) using the results of the ecological profiling, to test the germination of stored seeds from four populations collected at different altitudes under different temperature and light conditions.
2. Materials and Methods
2.1. Characteristics of the Focal Plant Species
The monotypic plant Petromarula pinnata (Figure 1) of Campanulaceae thrives as a chasmophyte in crevices of limestone cliffs, usually in semi-shaded places across the island of Crete, and it may also be found frequently on the stone walls of old buildings and fortresses [4,5]. Its distribution ranges from sea level up to 800 or even occasionally up to 1300 m a.s.l. It flowers during April and May or later, depending on altitude and slope [5]. Hence, P. pinnata is considered a widely scattered hemicryptophyte in Crete [4,5], occurring in almost all of the island’s regions (http://www.cretanflora.com/petromarula_pinnata.html, accessed 17 January 2024). It is a local single-island endemic plant that is currently assessed as ‘Vulnerable’ [26].
2.2. GIS Ecological Profiling
The ecological profile of P. pinnata was developed in GIS, following the methodology applied previously to C. pelviformis, a species with similar characteristics and conservation concerns e.g., [17]. In summary, the natural distribution points of P. pinnata (n = 51) (http://www.cretanflora.com/petromarula_pinnata.html, accessed 17 January 2024) as derived from a previous study [27] were herein linked with climate and precipitation data (minimum, maximum, and average values), as well as with 19 bioclimatic variables, all based on 30 years of historical climate data (1970–2000), with a pixel size of 30 s and a spatial resolution of 1 km2 (https://www.worldclim.org/data/worldclim21.html, accessed on 15 June 2023).
2.3. Seed Collection and Storage
All mature seeds of P. pinnata were collected on the same day (29 August 2018) from up to five individuals in each of four wild-growing populations found in different locations in Crete (Table 1). The seeds were collected using a special collection permit (182336/879 of 16 May 2019 and 64886/2959 of 6 July 2020) issued by the Greek Ministry of Environment and Energy. Seed storage conditions (three years’ storage) followed those described in previous studies [17].
2.4. Germination Tests
Prior to the germination experiments, morphological measurements such as weight of 1000 seeds and average length/width of 30 seeds were obtained for every population of P. pinnata [17]. The weight of 1000 seeds was 0.041 g for GR-BBGK-1-19,97; 0.044 g for GR-BBGK-1-19,126; 0.034 g for GR-BBGK-1-19,124; and 0.043 g for GR-BBGK-1-19,130. The average length/width ranged from 0.206/0.107 to 0.231/0.114 mm for lowland populations, while semi-mountainous populations had relatively larger seeds, ranging from 0.217/0.117 to 0.258/0.126 mm.
Germination tests were conducted following the germination protocol described in a previous study [17], and all trials took place at the Laboratory of Floriculture, School of Agriculture, Aristotle University of Thessaloniki (Thermi, Greece) in April 2022. In brief, seed responses as a function of incubation temperature were examined at four different temperatures (10, 15, 20, and 25 °C) using four replicates of 20 seeds from populations GR-BBGK-1-19,97 and GR-BBGK-1-19,126, and four replications of 25 seeds from populations GR-BBGK-1-19,124 and GR-BBGK-1-19,130. Germinated seeds were counted and removed every five days for a period of 60 days.
The light requirements for P. pinnata seed germination were investigated after the end of the germination experiments according to a protocol described in a previous study [17]. Only seeds from population GR-BBGK-1-19,124 were used for this experiment, which was conducted in a controlled-temperature chamber set to 15 °C. In this experiment, the mean germination time (MGT) was calculated [17].
2.5. Statistical Analysis
In the germination tests, the experimental design was a completely randomized factorial design, with the population and incubation temperature as separate factors. The germination data for the incubation temperature of 25 °C were not analyzed, as either none of the seeds germinated or the percentages of germinated seeds were very low (<3%). Therefore, in the statistical analysis, there were four levels for the population factor and three levels for the incubation-temperature factor (4 × 3 factorial design). The data were analyzed using ANOVA in the frame of the GLM (General Linear Model) [28]. The germination rate data were transformed into arc-sine square-root values before analysis [29]. The transformed data were checked for normality and homogeneity of variances and then were analyzed by ANOVA, while the comparisons of the means were performed using the LSD test at a significance level of p ≤ 0.05. In the comparisons between the two light conditions (alternating light/dark and continuous dark conditions), Student’s t-test was used [30]. All statistical analyses were carried out using SPSS 27.0 (SPSS, IΒΜ, Inc., Armonk, NY, USA).
3. Results
3.1. Ecological Profiling
The ecological profile of P. pinnata was generated in GIS based on its natural distribution in Crete, which was used to determine favorable climate conditions for wild-growing populations (Figure 2). According to historical climate data, the lowest average temperatures are recorded during the first annual quarter (9.49 ± 1.11 °C, 9.52 ± 1.12 °C, 10.95 ± 1.08 °C in January, February, and March, respectively). When such data were combined with the mean temperature of the coldest quarter (9.98 ± 1.10 °C) and the minimum temperature of the coldest month (6.48 ± 1.06 °C), it was concluded that P. pinnata wild-growing populations thrive in areas with relatively ‘mild’ winters (that rarely experience temperatures below 0 °C). Between April (14.05 ± 1.03 °C) and June (22.25 ± 0.93 °C), the average temperature in these areas gradually rose, with July (24.28 ± 0.94 °C) and August (24.09 ± 0.94 °C) having the highest average temperatures. This period coincides with the natural flowering and fruiting of wild-growing populations of P. pinnata. Even during fruiting, in the warmest period for P. pinnata, the average temperature was close to 25 °C (maximum temperature of warmest month = 28.39 ± 0.87 °C). After summer ended, the average temperature in these areas decreased from September (21.64 ± 0.97 °C) to December (11.05 ± 1.10 °C). Considering all the above-mentioned data as well as the lowest and highest temperatures (Tmin of Tmin = 4.15 °C in February, Tmax of Tmax = 29.90 °C in July) and the mean diurnal range (7.22 ± 0.13 °C), P. pinnata wild-growing populations thrive in environments with no extreme climate conditions. Such temperature limits may illustrate the natural adaptations of wild-growing populations of P. pinnata.
Wild-growing populations of P. pinnata thrive in habitats with a distinct rainy season, as indicated by the precipitation data shown in Figure 2. The data indicate a rainy season starting in mid-October (73.16 ± 8.88 mm) and lasting until mid-March (86.72 ± 8.63 mm), with January being the wettest month (142.40 ± 14.44 mm). From April (37.77 ± 5.43 mm) onwards, while P. pinnata populations are in flower [5], the precipitation values decreased significantly until June (5.75 ± 1.63 mm). This period marks the onset of the dry season (precipitation of driest quarter = 10.81 ± 3.31 mm) and the fruit-setting period, during which wild-growing plant individuals dry out, completing their role in the biological cycle. Starting in September (18.06 ± 1.77 mm), rainfall patterns increased again until the onset of the rainy season. Similar to the temperature limits described above, these precipitation patterns may illustrate the natural adaptations of wild-growing populations of P. pinnata.
3.2. Seed Germination Success
3.2.1. Effect of Incubation Temperature
According to the results of the statistical analysis, the main effects, population and incubation temperature, significantly affected the germination of P. pinnata seeds, as did their interaction (Table 2). In detail, the seeds of population GR-BBGK-1-19,97, which was collected from the highest altitude, exhibited the lowest germination rate (Figure 3). Additionally, the germination rate of population GR-BBGK-1-19,124 was higher than that of population GR-BBGK-1-19,126 (both collected at lower altitudes). The germination rate of seeds incubated at 20 °C was the lowest, while no statistical difference was observed between the germination rates of seeds incubated at 10 and 15 °C (Figure 4).
Significant differences were observed among the germination rates of seeds incubated at 10, 15, and 20 °C (Figure 5). Specifically, the seeds of populations GR-1-BBGK-19,126 and GR-1-BBGK-19,130 germinated at higher rates when they were incubated at 10 and 15 °C (BBGK-19,126: 81.25% and 85%; GR-1-BBGK-19,130: 90% and 89%, respectively) compared to when they were incubated at 20 °C (65% and 79%, respectively). In population GR-1-BBGK-19,97, the highest germination rate was observed after incubation at 15 °C (75%), whereas in population GR-1-BBGK-19,124, the highest germination rate was observed after incubation at 10 °C (91%, versus 84% at 15 °C), with fewer seeds germinating after incubation at 20 °C (79%) (Figure 5, p < 0.05). As shown in Figure 5, in all populations, the seeds incubated at 15 °C germinated earlier than those incubated at 10 °C. Specifically, germination of the seeds began on the 10th day and completed on the 30th day.
Furthermore, the incubation temperature had a significant impact on the germination of seeds from four wild populations of P. pinnata (Table 3, p < 0.05). Specifically, at 10 °C, the seeds from population GR-1-BBGK-19,97 germinated at the lowest rate (48.75%), while no statistically significant difference was observed among the remaining three populations. At an incubation temperature of 15 °C, the seeds from population GR-1-BBGK-19,130 germinated at a higher rate (89%) than those from population GR-1-BBGK-19,97 (75%). An incubation temperature of 20 °C generally reduced the germination rates. In general, the seeds of population GR-1-BBGK-19,97 exhibited the lowest germination rate (48.75% at 10 °C), whereas the seeds of population GR-1-BBGK-19,124 germinated at higher rates than did those of population GR-1-BBGK-19,126 (Table 3).
3.2.2. Effects of Light Treatments
Light requirements for germination of P. pinnata seeds at 15 °C were also tested. No statistically significant results were observed in terms of mean germination percentage (MGP) between seeds germinated under alternating light/dark and continuous dark conditions (Table 4). In both treatments, the germination rates were high (86% in alternating light/dark conditions and 80% in continuous dark conditions). Furthermore, the seeds showed similar germination speeds under different light conditions. Specifically, the mean germination time (MGT) was 18.59 days for seeds exposed to light/dark conditions and 20.05 days for seeds exposed to dark conditions (Table 4).
4. Discussion
In general, the development of seed-propagation protocols for threatened species is necessary both for effective conservation, whether ex situ or in situ [3,31,32], and for the sustainable utilization of NUPs. Such plants may be in demand due to their significant value in different economic sectors such as the ornamental sector [3] or the agro-alimentary sector, as means of contributing to food security [33].
Although seed gemination of several members of the family Campanulaceae has been previously investigated, the seed biology of this large family generally remains understudied [24]. Petromarula pinnata has been previously subjected to multi-species in vivo [19] and in vitro seed trials [24], with promising general results. This study, however, investigated for the first time the light and temperature dependance of P. pinnata seed germination. The investigation was enhanced by GIS-derived ecological profiling, which furnished insight into the most appropriate treatments in terms of temperature and light requirements. The GIS-derived ecological profile of P. pinnata indicated the temperature limits and precipitation patterns to which its wild-growing populations are exposed.
Several members of the genus Campanula appear to have specific temperature requirements for their seed germination, as suggested by previous studies [24]; this finding was also confirmed in the current study, which a strong temperature effect (p = 0.0001) was found in P. pinnata seed-germination trials. Overall, incubation temperatures of 10 and 15 °C resulted in the highest rates of germination of P. pinnata seeds. The exception was population GR-1-BBGK-19,97, for which the highest germination rate was observed after incubation at 15 °C (75%). These incubation temperatures also improved the germination of endemic Cretan members of genus Campanula, i.e., C. saxatilis L. subsp. saxatilis [31] and C. pelviformis [17]. An incubation temperature of 20 °C resulted in relatively high germination rates (>50%), while an incubation temperature of 25 °C dramatically reduced germination rates (<3%). However, previous studies on many members of the family Campanulaceae have shown that temperatures of 25 °C can benefit seed germination when appropriate pretreatment methods are applied, as in Asyneuma chinense D.Y.Hong (88%), Cyananthus inflatus Hook f. & Thompson (85%) or Lobelia oahuensis Rock (95%) [24].
This study investigated for the first time the germination of P. pinnata seeds collected from lowland and semi-mountainous populations of Central and Eastern Crete, Greece. The examination of the population effect (p = 0.0001) investigated whether origination from populations at different altitudes, in conjunction with four incubation temperatures, can affect the seed-germination rates of wild-growing P. pinnata from two semi-mountainous and two lowland areas in Central and Eastern Crete. Previous studies have reported that some endemic Balkan Campanula species may have significantly different germination responses to temperatures, depending on the altitude at which the original collections were found [34]. In this study, however, no significant variation was observed in the seed-germination rates of P. pinnata populations from different altitudes. In particular, the lowland populations were similar to each other, with the highest germination rates at 10 and 15 °C, while the same was true for the semi-mountainous populations, which had the highest germination rates at 15 °C. However, it is well known that differences are to be expected across different populations of the same species [16]. Moreover, variability in germination has been detected between populations originating from similar altitudes for C. pelviformis, as shown in a recent study [17]. Undoubtedly, experiments with freshly collected seeds (seeds that have not been subjected to storage) will yield a more complete picture of the germination behaviors and preferences of the focal species, including the identification of any after-ripening effects and possible dormancy types. Further research is therefore recommended, with the aim, among others, to verify previously published in vivo germination results [19].
P. pinnata was shown here to have no light preference for germination, with 86% germination in alternating light/dark conditions and 80% germination in darkness at 15 °C. Our results contrast with those of a previous study that observed different germination rates in seeds of this species maintained under constant light (76%) and constant darkness (3%) at 20 °C, probably because that trial was conducted with fresh seeds at 20 °C [24] rather than with stored seeds at 15 °C, as in this study. However, it is worth noting that many members of the Campanulaceae family may exhibit light preferences, as observed in other local Greek endemics of genus Campanula such as C. cretica (A.DC.) A.Dietr. (99% germination in light and 24% in darkness) and C. goulimyi Turrill (96% germination in light and 0% in darkness) [24]. In terms of seed dormancy, temperatures above 15 °C are known to trigger embryo development in the seeds of several Campanulaceae species that undergo morphological or morphophysiological dormancy [35].
When the ecological profile of P. pinnata is combined with the germination results obtained herein, it can be concluded that in a conservation project, three-year stored seeds of P. pinnata could be directly sowed in a field during the last quarter of the year, as prevailing mean temperatures are 17.98, 14.09, and 11.05 °C during October, November, and December, respectively. During this period, temperatures were shown to reach 10 and 15 °C in the natural habitats of this species in Crete. Simultaneously, moisture levels in these periods increase due to the increase in precipitation, which starts at 73.16 mm in October and peaks at 142.40 mm in January (Figure 2). Most probably, April temperatures (14.05 °C) and the relatively low precipitation (37.77 mm) in early spring would not favor seed germination (Figure 2).
The results of this study of this valuable, yet vulnerable local endemic plant of Crete can be used either for conservation purposes or to facilitate the sustainable exploitation of P. pinnata [3]. Notably, this species is traditionally used across Crete as an ingredient of a widely consumed local everyday snack known as ‘kaltsounia’ in western Crete and ‘chortopitakia’ in central and eastern Crete, a savory dough stuffed with wild-sourced and cultivated greens that have strong antioxidant potential [11]. A newly launched research project will apply this knowledge to produce enough plant material for a pilot cultivation project, which will be established in the next growing season in this species’ place of origin, Crete (Greece).
5. Conclusions
The development of seed-propagation protocols for threatened species is necessary for effective ex situ conservation and enables their sustainable utilization. The investigation described here provides for the first time a detailed profiling of the non-biotic environmental conditions experienced evolutionarily by wild-growing P. pinnata populations based on the natural distribution of this plant in rocky habitats of Crete, Greece. Subsequently, the germination trials performed in this investigation using three-year-old stored seeds sourced from four wild-growing populations revealed significant variation in seed germination among populations and across temperatures, as well as an interaction between population (genotype) and temperature. This work also identified the most appropriate incubation temperature for the effective germination of this species at the population level. The combination of the GIS ecological profile developed herein with the germination results for the three-year-old stored seeds may facilitate the successful ex situ conservation and further cultivation of this vulnerable local Cretan endemic species. These results also offer guidance for its cultivation in man-made settings for purposes of conservation or sustainable exploitation. These results can pave the way for the successful ex situ conservation of natural species diversity in seed banks and may further aid in current attempts towards the sustainable exploitation of this valuable, yet vulnerable plant, which possesses ornamental, medicinal, and agro-alimentary potential.
Author Contributions
Conceptualization, S.H., N.K., I.A. and G.T.; data curation, N.K., I.A., S.K., E.P., E.D. and G.T.; formal analysis, I.A., E.P., N.K., S.K. and S.H.; investigation, I.A., N.K., S.H., E.D., E.P., E.K., S.K. and G.T.; methodology, I.A., N.K., S.H., E.P., S.K., E.D., E.K., V.P., V.G. and G.T.; project administration, S.H., N.K. and G.T.; resources, N.K., G.T., S.H., V.G. and S.K.; software, I.A.; supervision, N.K., S.H. and G.T.; validation, N.K., I.A., E.K., E.D., V.G. and G.T.; visualization, I.A., E.P., V.G. and N.K.; writing—original draft, I.A., E.P., S.H., N.K., E.K. and G.T.; writing—review and editing, all authors. All authors have read and agreed to the published version of the manuscript.
Funding
This research received no external funding.
Data Availability Statement
All data supporting the results of this study are included in the manuscript and the datasets are available upon request.
Acknowledgments
This investigation was performed as part of the PhD research of Ioannis Anestis. The research of Nikos Krigas and Georgios Tsoktouridis was supported by the project “Indigenous edible plants of Crete as alternative new crops contributing to biodiversity preservation, protection from soil degradation and mitigation of climate change impacts” (acronym: Cretan Greens 4 Clima Pro, Μ16ΣΥΝ-01106) co-funded under Measure 16—Cooperation (16.1–16.5) by Greece and the European Union (European Agricultural Fund for Rural Development 2014–2020).
Conflicts of Interest
The authors declare no conflicts of interest.
References
- Seglie, L.; Scariot, V.; Larcher, F.; Devecchi, M.; Chiavazza, P.M. In vitro seed germination and seedling propagation in Campanula spp. Plant Biosyst. 2011, 146, 15–23. [Google Scholar] [CrossRef]
- Scariot, V.; Seglie, L.; Caser, M.; Devecchi, M. Evaluation of ethylene sensitivity and postharvest treatments to improve the vase life of four Campanula species. Eur. J. Horticult. Sci. 2008, 73, 166–170. Available online: https://www.pubhort.org/ejhs/2008/file_723254.pdf (accessed on 15 January 2024).
- Krigas, N.; Tsoktouridis, G.; Anestis, I.; Khabbach, A.; Libiad, M.; Megdiche-Ksouri, W.; Ghrabi-Gammar, Z.; Lamchouri, F.; Tsiripidis, I.; Tsiafouli, M.A.; et al. Exploring the potential of neglected local endemic plants of three Mediterranean regions in the ornamental sector: Value chain feasibility and readiness timescale for their sustainable exploitation. Sustainability 2021, 13, 2539. [Google Scholar] [CrossRef]
- Cellinese, N.; Smith, S.A.; Edwards, E.J.; Kim, S.T.; Haberle, R.C.; Avramakis, M.; Donoghue, M.J. Historical biogeography of the endemic Campanulaceae of Crete. J. Biogeogr. 2009, 36, 1253–1269. [Google Scholar] [CrossRef]
- Strid, A. Atlas of the Aegean Flora Part 1 (Text & Plates) & Part 2 (Maps), 1st ed.; Englera 33 (1 & 2); Botanic Garden and Botanical Museum: Berlin, Germany; Freie Universität: Berlin, Germany, 2016. [Google Scholar]
- González-Tejero, M.R.; Casares-Porcel, M.; Sánchez-Rojas, C.P.; Ramiro-Gutiérrez, J.M.; Molero-Mesa, J.; Pieroni, A.; Giusti, M.E.; Censorii, E.; de Pasquale, C.; Della, A.; et al. Medicinal plants in the Mediterranean area: Synthesis of the results of the project Rubia. J. Ethnopharmacol. 2008, 116, 341–357. [Google Scholar] [CrossRef] [PubMed]
- Lim, T.K. Edible Medicinal and Non-Medicinal Plants; Springer International Publishing: Gewerbestr, Switzerland, 2016; ISBN 9789401772761. [Google Scholar]
- Heinrich, M.; Müller, W.E.; Galli, C. Local Mediterranean Food Plants and Nutraceuticals; Karger Medical and Scientific Publishers: Basel, Switzerland, 2006; Volume 59. [Google Scholar]
- Hadjichambis, A.C.H.; Hadjichambi, D.P.; Della, A.; Giusti, M.E.; De Pasquale, C.; Lenzarini, C.; Censorii, E.; Reyes Gonzales Tejero, M.; Sanchez-Rojas, C.P.; Ramiro Gutierrez, J.M.; et al. Wild and semi-domesticated food plant consumption in seven circum-Mediterranean areas. Int. J. Food Sci. Nutr. 2008, 59, 383–414. [Google Scholar] [CrossRef] [PubMed]
- Tsiftsoglou, O.S.; Lagogiannis, G.; Psaroudaki, A.; Vantsioti, A.; Mitić, M.N.; Mrmošanin, J.M.; Lazari, D. Phytochemical analysis of the aerial parts of Campanula pelviformis Lam. (Campanulaceae): Documenting the dietary value of a local endemic plant of Crete (Greece) traditionally used as wild edible green. Sustainability 2023, 15, 7404. [Google Scholar] [CrossRef]
- Vasilopoulou, E.; Trichopoulou, A. Green pies: The flavonoid rich Greek snack. Food Chem. 2011, 126, 855–858. [Google Scholar] [CrossRef]
- Kalpoutzakis, E.; Chatzimitakos, T.; Athanasiadis, V.; Mitakou, S.; Aligiannis, N.; Bozinou, E.; Gortzi, O.; Skaltsounis, L.A.; Lalas, S.I. Determination of the total phenolics content and antioxidant activity of extracts from parts of plants from the Greek Island of Crete. Plants 2023, 12, 1092. [Google Scholar] [CrossRef]
- Padulosi, S.; Thompson, J.; Rudebjer, P. Fighting Poverty, Hunger and Malnutrition with Neglected and Underutilized Species (NUS): Needs, Challenges and the Way Forward; Biodiversity International: Rome, Italy, 2013. [Google Scholar]
- Yucel, G. Seed propagation, adaptation to cultivation conditions, determination of ornamental plant properties, and ex situ conservation of the endemic species Centaurea hermannii F. hermann. BioResources 2022, 17, 616–633. [Google Scholar] [CrossRef]
- Macdonald, B. Practical Woody Plant Propagation for Nursery Growers; Timber Press Inc.: Portland, OR, USA, 2006; p. 669. ISBN 9780881920628. [Google Scholar]
- Baskin, C.C.; Baskin, J.M. Seeds: Ecology, Biogeography, and Evolution of Dormancy and Germination, 2nd ed.; Academic Press: San Diego, CA, USA, 2014; ISBN 978-0-12-416677-6. [Google Scholar]
- Anestis, I.; Pipinis, E.; Kostas, S.; Papaioannou, E.; Karapatzak, E.; Dariotis, E.; Tsoulpha, P.; Koundourakis, E.; Chatzileontari, E.; Tsoktouridis, G.; et al. GIS-facilitated germination of stored seeds from five wild-growing populations of Campanula pelviformis Lam. and fertilization effects on growth, nutrients, phenol content and antioxidant potential. Horticulturae 2023, 9, 877. [Google Scholar] [CrossRef]
- O’Reill, C.; Arnott, J.T.; Owens, J.N. Effects of photoperiod and moisture availability on shoot growth, seedling morphology, and cuticle and epicuticular wax features of container-grown western hemlock seedlings. Can. J. For. Res. 1989, 19, 122–131. [Google Scholar] [CrossRef]
- Sarropoulou, V.; Krigas, N.; Tsoktouridis, G.; Maloupa, E.; Grigoriadou, K. Seed germination trials and ex situ conservation of local prioritized endemic plants of Crete (Greece) with commercial interest. Seeds 2022, 1, 279–302. [Google Scholar] [CrossRef]
- Varsamis, G.; Merou, T.; Tseniklidou, K.; Goula, K.; Tsiftsis, S. How different reproduction protocols can affect the germination of seeds: The case of three stenoendemic species on Mt. Olympus (NC Greece). Eur. J. Environ. Sci. 2023, 13, 23–30. [Google Scholar] [CrossRef]
- Varsamis, G.; Merou, T.; Takos, I.; Malesios, C.; Manolis, A.; Papageorgiou, A.C. Seed adaptive traits of Fagus sylvatica populations in northeastern Greece. For. Sci. 2020, 66, 403–415. [Google Scholar] [CrossRef]
- Fernandes, M.P.; Pinto-Cruz, C.; Almeida, E.; Emídio, M.; Simões, M.P.; Gazarini, L.; Belo, A.D. Seed germination of six Iberian endemic species—A contribution to enhance plant conservation. Plant Biosyst. 2021, 155, 1146–1152. [Google Scholar] [CrossRef]
- Alp, S.; Ipek, A.; Arslan, N. The effect of gibberellic acid on germination of rosehip seeds (Rosa canina L.). Acta Hortic. 2008, 885, 33–37. [Google Scholar] [CrossRef]
- Koutsovoulou, K.; Daws, M.I.; Thanos, C.A. Campanulaceae: A family with small seeds that require light for germination. Ann. Bot. 2014, 113, 135–143. [Google Scholar] [CrossRef]
- Fanourakis, D.; Paschalidis, K.; Tsaniklidis, G.; Tzanakakis, V.A.; Bilias, F.; Samara, E.; Liapaki, E.; Jouini, M.; Ipsilantis, I.; Maloupa, E.; et al. Pilot cultivation of the local endemic Cretan marjoram Origanum microphyllum (Benth.) Vogel (Lamiaceae): Effect of fertilizers on growth and herbal quality features. Agronomy 2021, 12, 94. [Google Scholar] [CrossRef]
- Kougioumoutzis, K.; Kokkoris, I.P.; Panitsa, M.; Strid, A.; Dimopoulos, P. Extinction risk assessment of the Greek endemic flora. Biology 2021, 10, 195. [Google Scholar] [CrossRef]
- Lazarina, M.; Kallimanis, A.S.; Dimopoulos, P.; Psaralexi, M.; Michailidou, D.E.; Sgardelis, S.P. Patterns and drivers of species richness and turnover of neo-endemic and palaeo-endemic vascular plants in a Mediterranean hotspot: The case of Crete, Greece. J. Biol. Res.-Thessaloniki 2019, 26, 12. [Google Scholar] [CrossRef] [PubMed]
- Gomez, K.A.; Gomez, A.A. Statistical Procedures in Agricultural Research, 2nd ed.; Wiley: New York, NY, USA, 1984; ISBN 0-471-87092-7. [Google Scholar]
- Snedecor, G.W.; Cochran, W.C. Statistical Methods, 7th ed.; The Iowa State University Press: Ames, IA, USA, 1980. [Google Scholar]
- Klockars, A.; Sax, G. Multiple Comparisons; Sage Publications: Newbury Park, CA, USA, 1986; p. 87. ISBN 0-8039-2051-2. [Google Scholar]
- Hatzilazarou, S.; Anestis, I.; Pipinis, E.; Kostas, S.; Avramakis, M.; Greveniotis, V.; Dariotis, E.; Tsoktrouridis, G.; Krigas, N. GIS-facilitated seed germination of six local endemic plants of Crete (Greece) and multifaceted evaluation in three economic sectors. J. Biol. Res. 2023, 30, 2241–5793. [Google Scholar] [CrossRef]
- Maunder, M.; Higgens, S.; Culham, A. The effectiveness of botanic garden collections in supporting plant conservation: A European case study. Biodivers. Conserv. 2001, 10, 383–401. [Google Scholar] [CrossRef]
- Singh, A.; Dubey, P.K.; Chaurasia, R.; Dubey, R.K.; Pandey, K.K.; Singh, G.S.; Abhilash, P.C. Domesticating the Undomesticated for Global Food and Nutritional Security: Four Steps. Agronomy 2019, 9, 491. [Google Scholar] [CrossRef]
- Blionis, G.; Vokou, D. Reproductive attributes of Campanula populations from Mt Olympos, Greece. Plant Ecol. 2005, 178, 77–88. [Google Scholar] [CrossRef]
- Baskin, C.C.; Baskin, J.M.; Yoshinaga, A.; Wolkis, D. Seed dormancy in Campanulaceae: Morphological and morphophysiological dormancy in six species of Hawaiian lobelioids. Botany 2020, 98, 327–332. [Google Scholar] [CrossRef]
Figure 1.
Wild-growing Petromarula pinnata in full flower during spring in its natural rocky habitat in Crete (A), bearing ripe fruits during the summer months, (B) mature seeds, (C) germinated seeds with evident radicle protrusion (D).
Figure 1.
Wild-growing Petromarula pinnata in full flower during spring in its natural rocky habitat in Crete (A), bearing ripe fruits during the summer months, (B) mature seeds, (C) germinated seeds with evident radicle protrusion (D).
Figure 2.
Ecological profile across the natural distribution range (http://www.cretanflora.com/petromarula_pinnata.html, accessed 17 January 2024) of wild-growing Petromarula pinnata populations in Crete (n = 51) linked in GIS with geodatabases (WorldClim version 2.1) to provide values for the following: (A) minimum temperature per month (°C), (B) maximum temperature per month (°C), (C) average temperature per month (°C), (D) precipitation per month (mm), and (E) calculated values for 19 bioclimatic variables. For (A–E), the minimum, maximum, average, and standard deviation are shown based on data from 1970–2000. The colors of the plotted lines illustrate the minimum (blue), maximum (gray), and mean (orange) monthly values for temperature (°C) and precipitation (mm).
Figure 2.
Ecological profile across the natural distribution range (http://www.cretanflora.com/petromarula_pinnata.html, accessed 17 January 2024) of wild-growing Petromarula pinnata populations in Crete (n = 51) linked in GIS with geodatabases (WorldClim version 2.1) to provide values for the following: (A) minimum temperature per month (°C), (B) maximum temperature per month (°C), (C) average temperature per month (°C), (D) precipitation per month (mm), and (E) calculated values for 19 bioclimatic variables. For (A–E), the minimum, maximum, average, and standard deviation are shown based on data from 1970–2000. The colors of the plotted lines illustrate the minimum (blue), maximum (gray), and mean (orange) monthly values for temperature (°C) and precipitation (mm).
Figure 3.
Effect of population on germination rate (±standard deviation) of Petromarula pinnata seeds, regardless of incubation temperature. Columns accompanied by the same letter do not differ significantly. The comparisons were made using the LSD test.
Figure 3.
Effect of population on germination rate (±standard deviation) of Petromarula pinnata seeds, regardless of incubation temperature. Columns accompanied by the same letter do not differ significantly. The comparisons were made using the LSD test.
Figure 4.
Effect of incubation temperature on germination rate (±standard deviation) of Petromarula pinnata seeds from four wild-growing populations. Columns accompanied by the same letter do not differ significantly. The comparisons were made using the LSD test.
Figure 4.
Effect of incubation temperature on germination rate (±standard deviation) of Petromarula pinnata seeds from four wild-growing populations. Columns accompanied by the same letter do not differ significantly. The comparisons were made using the LSD test.
Figure 5.
Cumulative percent germination (±standard deviation) of Petromarula pinnata seeds sourced from four populations incubated at 10 °C (●), 15 °C (■) and 20 °C (□). 1 In each population, means are statistically different at p ≤ 0.05, and statistically significant groups do not share a common letter. The comparisons were carried out using the LSD test (n = 4).
Figure 5.
Cumulative percent germination (±standard deviation) of Petromarula pinnata seeds sourced from four populations incubated at 10 °C (●), 15 °C (■) and 20 °C (□). 1 In each population, means are statistically different at p ≤ 0.05, and statistically significant groups do not share a common letter. The comparisons were carried out using the LSD test (n = 4).
Table 1.
Collection data with geographical information and IPEN (International Plant Exchange Network) accession numbers of the investigated Petromarula pinnata wild-growing populations.
Table 1.
Collection data with geographical information and IPEN (International Plant Exchange Network) accession numbers of the investigated Petromarula pinnata wild-growing populations.
| IPEN Accession Number | Collection Site | Latitude (North) | Longitude (East) | Altitude (m) |
|---|---|---|---|---|
| GR-BBGK-1-19,97 | Viannos, Heraklion | 35.0471 | 25.4174 | 632 |
| GR-BBGK-1-19,126 | Agios Georgios Selinaris, Lasithi | 35.2753 | 25.5549 | 225 |
| GR-BBGK-1-19,124 | Agia Eirini gorge, Heraklion | 35.2844 | 25.1653 | 140 |
| GR-BBGK-1-19,130 | Tylissos gorge, Heraklion | 35.2874 | 24.9716 | 395 |
Table 2.
Significance of the effects of individual factors (population, temperature) and their interaction on germination rate of Petromarula pinnata seeds from different wild-growing populations, as estimated by ANOVA.
Table 2.
Significance of the effects of individual factors (population, temperature) and their interaction on germination rate of Petromarula pinnata seeds from different wild-growing populations, as estimated by ANOVA.
| Source | Sum of Squares | df | Mean Square | F | Sig. |
|---|---|---|---|---|---|
| Population | 2314.13 | 3 | 771.38 | 20.36 | 0.000 |
| Temperature | 1240.85 | 2 | 620.43 | 16.38 | 0.000 |
| Population × Temperature | 701.35 | 6 | 116.89 | 3.09 | 0.015 |
Table 3.
Effect of three incubation temperatures on the germination rates of four wild-growing populations of Petromarula pinnata. Values are given as means ± standard deviation.
Table 3.
Effect of three incubation temperatures on the germination rates of four wild-growing populations of Petromarula pinnata. Values are given as means ± standard deviation.
| Area | Population | Incubation Temperature | ||
|---|---|---|---|---|
| 10 °C | 15 °C | 20 °C | ||
| Semi-mountainous | GR-1-BBGK-19,97 | 48.75 b 1 ± 8.54 | 75.00 b ± 10.80 | 50.00 c ± 8.16 |
| GR-1-BBGK-19,130 | 90.00 a ± 7.66 | 89.00 a ± 6.83 | 70.00 ab ± 6.93 | |
| Lowland | GR-1-BBGK-19,126 | 81.25 a ± 11.09 | 85.00 ab ± 9.13 | 65.00 b ± 7.07 |
| GR-1-BBGK-19,124 | 91.00 a ± 2.00 | 84.00 ab ± 5.66 | 79.00 a ± 6.83 | |
1 Values in the same column followed by the same letter are not significantly different (p ≤ 0.05) according to the LSD test (n = 4).
Table 4.
Effect of light conditions on germination of Petromarula pinata seeds. Means and standard deviation values are provided.
Table 4.
Effect of light conditions on germination of Petromarula pinata seeds. Means and standard deviation values are provided.
| Germination Rate (%) | Mean Germination Time (Days) | |
|---|---|---|
| Light/dark | 86.00 ± 6.93 a 1 | 18.59 a ± 1.02 a |
| Dark | 80.00 ± 8.64 a | 20.06 a ± 1.72 a |
1 In each column, the means are statistically different at p ≤ 0.05 (n = 4), shown as values that do not share a common small letter. The comparisons were made using Student’s t-tests.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content. |
© 2024 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
MDPI and ACS Style
Anestis, I.; Pipinis, E.; Kostas, S.; Karapatzak, E.; Dariotis, E.; Paradeisopoulou, V.; Greveniotis, V.; Tsoktouridis, G.; Hatzilazarou, S.; Krigas, N. GIS-Facilitated Germination of Stored Seeds from Four Wild-Growing Populations of Petromarula pinnata (L.) A. DC.—A Valuable, yet Vulnerable Local Endemic Plant of Crete (Greece). Agronomy 2024, 14, 274. https://doi.org/10.3390/agronomy14020274
AMA Style
Anestis I, Pipinis E, Kostas S, Karapatzak E, Dariotis E, Paradeisopoulou V, Greveniotis V, Tsoktouridis G, Hatzilazarou S, Krigas N. GIS-Facilitated Germination of Stored Seeds from Four Wild-Growing Populations of Petromarula pinnata (L.) A. DC.—A Valuable, yet Vulnerable Local Endemic Plant of Crete (Greece). Agronomy. 2024; 14(2):274. https://doi.org/10.3390/agronomy14020274
Chicago/Turabian StyleAnestis, Ioannis, Elias Pipinis, Stefanos Kostas, Eleftherios Karapatzak, Eleftherios Dariotis, Veroniki Paradeisopoulou, Vasileios Greveniotis, Georgios Tsoktouridis, Stefanos Hatzilazarou, and Nikos Krigas. 2024. "GIS-Facilitated Germination of Stored Seeds from Four Wild-Growing Populations of Petromarula pinnata (L.) A. DC.—A Valuable, yet Vulnerable Local Endemic Plant of Crete (Greece)" Agronomy 14, no. 2: 274. https://doi.org/10.3390/agronomy14020274
Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.
