Variation in seed morpho-physiological traits of Nauclea orientalis L. in a seedling seed orchard and the effect of seed sortation on their physiological parameters

The aim of this study was to identify the morpho-physiological diversity of seeds in several mother trees of N. orientalis at 6-year-old in a seedling seed orchard in Parungpanjang, Bogor and to identify the effectiveness of seed sortation in improving the seed germination. A total of 17 mother trees were collected for their fruit, measured the morphology of the seeds and tested their germination. The results showed that fruit and seed dimensions and weight, germination capacity were strongly influenced by the mother tree. Germination capacity and the highest germination capacity was resulted by family no. 22-BT (90%) and 48-MJ (92%). The highest coefficient of variation was given by seed weight per fruit (83.03), followed by germination capacity (66.66) and fruit weight (65.95). Seed sortation had a significant effect on germination capacity, germination rate, and mean germination time, but had no significant effect on germination value. In general, a sieve size of 300-420 μm gave the best results for increasing germination capacity (59.5%), germination rate (1.67% day-1), and mean germination time (18 days). Other sieve sizes, i.e., 420-600 μm was also able to improve seed germination so that for seed sortation applications 300-600 μm sieve sizes can be used.


Introduction
Restoration of degraded forest and land is a priority program for forestry development in Indonesia.Based on data from the Ministry of Environment and Forestry, the area of degraded forest and land in Indonesia reaches 14.01 million ha [1] with a deforestation rate of 4.4 million ha per year [2].By 2021, the Government of Indonesia has committed to rehabilitating 12 million ha of degraded land and 2 million ha of peatland by 2030 as part of the commitment to the United Nations Framework Convention on Climate Change through the Nationally Determined Contribution (NDC) [3].The program certainly requires technological and innovation supports, including selecting the appropriate species to reforestation of degraded forests and lands.
One of the forest tree species that has high adaptability for rehabilitation of degraded forests and land is Nauclea orientalis L. [4,5].N. orientalis has a wide distribution which naturally can be found in swamp areas [6], peat swamps [1], forests along rivers [7,8], savannas and grasslands [6], as well as in coastal areas up to 1,400 m asl [9,2].Apart from land rehabilitation, N. orientalis also has the potential to be developed in plantation forest development because it is a fast-growing species.N. orientalis can be used as carpentry wood, pulp and paper raw material, furniture industry, flooring, molding, packaging boxes, sculpture, carving [10] and has the potential as an ingredient for medicines, such as antimalarials 1315 (2024) 012070 IOP Publishing doi:10.1088/1755-1315/1315/1/012070 2 [11], antibacterial [12], and inflammation [13].
The success of planting N. orientalis in the context of forest and land restorations requires the support of providing quality seeds which can be carried out with a breeding program that involves a relatively wide genetic diversity.The wide genetic diversity of seed sources will increase the adaptation and tolerance to stress of the seeds and seedlings, which will affect the success of the degraded forest and land restoration [14,15].The establishment of progeny tests followed by selection, both individual selection within families and family selection, is needed to obtain seeds with superior genetic quality [5].Apart from genetics, seed quality is also greatly influenced by its physical and physiological quality [16] which is the result of the seed handling process [17].However, the physical and physiological qualities of seed are also influenced by the genetic character of the mother tree [18] so that it allows for variations in the physical and genetic characteristics of seeds between mother trees.The physical and physiological diversity of seeds between mother trees will affect the effectiveness of seed handling techniques such as seed sortation.Seed cleaning and sortation are also important to improve the quality of N. orientalis seeds because the seeds have very small sizes (fine seeds) so that the level of seed purity is generally very low.The progeny test of N. orientalis has been established at Parungpanjang Forest Research Station in Bogor, West Java involving 49 families.Currently, the progeny test has been selected and determined as a seedling orchard by the Forest Tree Seed and Certification Institute, the West Java Provincial Forestry Service with Certificate No. 522/051/S.SB-5/SPTH/2019.To improve the quality of the seeds produced from the seedling orchard, a study of the diversity of seeds and their handling techniques is needed.The aim of this study was to identify the morpho-physiological diversity of seeds in several mother trees at 6-year-old in a N. orientalis seedling seed orchard in Parungpanjang, Bogor and to identify the effectiveness of seed sorting treatment in improving the physiological quality of the seeds.

Materials
Seeds were collected from the N. orientalis seedling seed orchard at Parungpanjang, Bogor, West Java.Geographically the location is located at 0623'08.8"S and 10631'14.4"E with an altitude of 52 m asl (Figure 1a).The seedling seed orchard was the result of a progeny test conversion that was established involving 49 families from two provenances, i.e., Banten and Majalengka, West Java (Figure 1b).The initial design of progeny test establishment used a completely randomized block design with 4-tree-row plots planted in 10 blocks.Seed collection was carried out by climbing and picking ripe fruit that was physiologically ripe.Seeds were collected from 17 mother trees (Table 2) which ripened at the same time.The height and diameter (diameter at breast height) of mother tree were measured using a measuring stick and a diameter measuring tape.The fruit from each parent tree was separated and given an identity and then brought to the Seed Testing Laboratory, Center for Application of Environmental and Forestry Standards and Instruments, Bogor.

Measurement of seed morpho-physiological traits
A completely randomized design was used to test differences in fruit and seed characteristics from different mother trees.The fruit of N. orientalis has a round shape and some of it is slightly oval so that the measurement of fruit morphology was carried out on the parameters of fruit length and diameter.
Fruit length and diameter were measured using a digital caliper which is the largest diameter (fruit length) and the smallest fruit diameter (fruit diameter or width).In addition, fruit weight was measured using a digital scale.Measurement of fruit dimensions was carried out with 30 replicates (fruit) for each mother tree.
Fruit extraction was carried out separately for each fruit group from each mother tree using the wet extraction method [18].To calculate the number of seeds per fruit was carried out manually by extracting 10 seeds per mother tree separately and counting the number of seeds per fruit.
The extracted seeds are then air-dried for two days at room temperature (25-30° C and 70-80% relative humidity).Observation of the morphology and testing of the physiological quality of the seeds was carried out separately on groups of seeds originating from different mother trees.Weight was measured with a digital scale, while the characteristics of the seeds measured were seed length, seed width, seed weight per fruit and number of seeds per fruit.The method used is the test method on paper in a germinator at a temperature of 24-30° C and 90-95% relative humidity.The test samples from each group of seeds totaled 4 replicates with 100 seeds each.Observations and calculations of seed germination were carried out every day until no more seeds germinated.The germination parameter calculated was the seed germination capacity [19].

Effectiveness of seed sortation on improving seed quality
The N. orientalis seeds from 17 mother trees were composited, i.e., by mixing the seeds in the same weight proportion for each parent tree.Seed sortation was carried out using multilevel sieves using 3 sieve sizes (600 m, 420 m, and 300 m) starting from the largest to the smallest sieve, so that 5 seed size classes were obtained, namely seeds with sizes >600 μm, 600-420 μm, 420-300 μm, <300 μm, and control (without sifting).A completely randomized design was used to test the 5 seed sortation treatments with 8 replications and each replication consisted of 100 seeds.Testing the physiological quality of seeds was carried out in the germinator (temperature 24-30° C and relative humidity 90-95%) using the top paper test [16].Seeds are declared to have germinated when a pair of leaves has emerged completely.Seed germination was observed every day until no more seeds germinated.Germination parameters that were calculated were germination capacity, germination rate, mean germination time, and germination value [20,21].

Data analysis
Data were analyzed by analysis of variance to test the significance of the effect of different mother trees of N. orientalis on the morphophysiological characteristics of fruit and seeds and to test the effect of the effectiveness of seed sortation using several sieve sizes on their physiological quality.Duncan's Multiple Range Test (DMRT) was used to test differences between treatments (differences in seed characteristics and effectiveness of seed sorting).A simple correlation test (Pearson) was used to determine the relationship between the parameters of the parent tree and the morpho-physiological characteristics of N. orientalis seeds.

Result
3.1.1.Variation of mother trees and seed characteristics.In general, the seventeen mother trees whose seeds were tested had no significant differences in growth, both in height and tree diameter.Meanwhile, the dimensions and weight of the fruit and seeds are greatly influenced by the mother tree.For germination capacity, differences in mother trees affect the germination capacity (Table 1).The highest coefficient of variation was given by seed weight per fruit (83.03), followed by germination capacity (66.66) and fruit weight (65.95).The lowest coefficient of variation is shown by mother tree diameter (14.79) and seed width (14.13).Table 2 shows that all the parameters tested gave a significant difference, except the height and diameter of mother trees.Based on the fruit and seed morphological traits, family no.17-BT (Banten) gave the best results for fruit length, fruit diameter, fruit weight, seed weight per fruit, while for seed length and seed width, family no.20-BT (Banten) gives the best results.For seed physiological trait or gemination capacity, family no.22-BT (Banten) and 48-MJ (Majalengka) gave the highest gemination capacity, i.e., 90% and 92% respectively.If we look at the correlation between the parameters tested in Table 3, almost all parameters of mother tree growth (height and diameter) are not correlated, except for the seed length.Morphology and fruit weight were only correlated with seed length and not significantly correlated with other seed morphophysiological parameters.Notes: ** = significant at P < 0.05, * = Significant at P > 0.05

Effectiveness of sortation on seed germination.
Seed sortation using different sieve sizes had a significant effect on germination capacity, germination rate, and mean germination time, but had no significant effect on germination value (Table 4).In general, a sieve size of 300-420 m gave the best results for increasing germination capacity (59.5%), germination rate (1.67% day -1 ), and mean germination time (18 days).Other sieve sizes, i.e., 420-600 m, were also able to improve seed germination of N. orientalis when viewed from the increase in their germination capacity compared to control (Figure 2).

Variation of mother trees and seed characteristics
The fruits and seeds of N. orientalis produced from the seedling seed orchard at Parungpanjang, Bogor have various fruit and seed morphophysiological traits.Differences in fruit, seed and seed germination characteristics of N. orientalis were also reported by [22] by examining its four natural habitats, i.e., swamps, temporal swamps, dry land, and temporal swamp savanna.Several studies have reported that differences in the morphophysiological characteristics of fruit and seeds from different population or provenance are strongly influenced by genetic factors [18,23,24,25].Differences in mother tree of N. orientalis also affect the growth of seedlings in nurseries [26].The germination capacity range of N. orientalis seeds from the seventeen mother trees was very diverse (14-95%) with an average of 52.29%.The germination capacity was lower when compared to N. orientalis from Southeast Sulawesi (74-91%) [22] or white jabon from Pomalaa (82.25%) [27].The low germination capacity of N. orientalis seeds was suspected because the seedling seed orchard stands are still young (6 years old).A similar thing was reported by [4] on Acacia mangium, Gmelina arborea, and Eucalyptus deglupta, which stated that at a relatively young tree age, seed germination tends to be low and increases with increasing age of the tree stand, then decreases again when the tree stand is old.The diversity of seed and fruit morphophysiological characteristics indicates that the seedling seed orchards that have been established have a fairly wide diversity considering that the differences in fruit and seed characters are more influenced by genetic factors.According to [15] and [28], seeds originating from seed sources with high diversity will increase adaptation to various suitable sites and increase the success of forest and landscape restoration.
Meanwhile, the seventeen mother trees that were selected to collect the seeds did not have a significant difference in growth (height and diameter).This is presumably because the seedling seed orchard is the result of conversion from progeny testing with a 4-row-tree plot planting system so that after thinning (selection), only 1 tree is left for each family and 3 other trees are cut down, leaving only the tree with the best performance for each family.The height of the mother tree correlates with its diameter, but the height and diameter of the mother tree do not correlate with most of the characteristics of the fruit and seeds.The morphophysiological characteristics of N. orientalis seeds in this study were also not significantly correlated with most of the fruit and seed characters.Likewise, the character dimensions and weight of fruit and seeds are not significantly correlated with seed germination.Basically, larger seeds produce better seed viability and germination vigor [29], but in this case the results are not significantly correlated.Similar results with this study were also reported for Toona sinensis [30], Fagraea fragrans [31] and Sterculia foetida [25], that the morphological characteristics of the fruit and seeds were not significantly correlated with the germination capacity of the seeds.

Effectiveness of sortation on seed germination
N. orientalis seeds are very small seeds (fine seeds) so it is difficult to distinguish between seeds and other impurities.To increase the purity as well as the germination of seeds, seed sortation treatment is necessary.Several studies have stated that sifting using sieves of various sizes is quite effective in increasing the purity and germination of small or very small seeds [32] such as the seeds of Calliandra calothyrsus [33] and Eucalyptus pellita [31].
Seed sortation of N. orientalis using three different sieve sizes (300 m, 420 m, and 600 m) resulted in differences in germination capacity, germination rate, and mean germination time.The use of a 300-420 m sieve, i.e., seeds that passed through a 420 m sieve and were retained on a 300 m sieve resulted in the highest seed germination capacity (59.5%).However, sieve size >420-600 µm also resulted in a relatively high germination capacity (55%).Seed sortation is an activity that aims to improve the purity and quality of seeds.Operationally, these activities often accompany or coincide with seed cleaning and selection activities [32].Seed cleaning is an activity of separating seeds from other plant parts and nonseed parts, such as fruit parts, leaves or twigs.Selection is done to separate filled seeds from empty seeds, other types of seeds and dirt.Seed weight and size in general are the basis for sorting seeds [34].Operationally, seed sortation of N. orientalis can use a 300-600 µm sieve which allows not too much seed to be wasted and the physiological quality of the seeds can be improved.

Conclusion
The fruit and seed morphophysiological traits from several mother trees of N. orientalis seedling seed orchard at Parungpanjang, Bogor have a fairly high diversity with a coefficient of variation ranging from 14.13 (seed length) -83.03 (seed weight per fruit).This diversity indicates that the seedling orchard has sufficiently high genetic diversity to make it suitable for use in forest and landscape restoration.To improve the physiological quality of N. orientalis seeds, seed sieving can be done by sieving using a sieve size of 300-600 m.This sortation technique can increase germination capacity, germination rate, and accelerate germination (shorter mean germination time), but not lose too much seed so that it is operationally more effective.

Figure 1 .
Figure 1.Location (1a) and tree stand performance (1b) of seedling seed orchard of Nauclea orientalis at the Parungpanjang Forest Research Station, Bogor, West Java, Indonesia

Table 2 .
Mother tree height and diameter, fruit and seed morphophysiological traits of Nauclea orientalis

Table 3 .
Simple (Person) correlation between height and diameter of mother trees with fruit and seed morpho-physiological traits of Nauclea orientalis

Table 4 .
Summary of analysis of variance of seed sortation treatment by sieving using different sieve sizes on the Nauclea orientalis physiological seed quality