The Yellow River Delta wetland (118°33′-119°207′E, 37°35 ′-38°12′N) is the best preserved, youngest, and widest wetland ecosystem in the warm temperature zone of China (Chen et al., 2016; Yu et al., 2016). With a temperate continental monsoon climate, this area has an average annual temperature of 11.7-12.6°C and average annual precipitation of 540-600mm, which primarily concentrates in Summer. Also, loam is the dominant soil in this area (Wang et al., 2015). Its coastal area has a high level of salinization, with few plant species and vegetation varieties.

In January 2022, samples of soil seed banks were collected in the nascent wetland of the Yellow River Delta. Along the silting direction of the nascent wetland of the Yellow River Delta, two parallel belt transects ( Figure 1 belt transect A and B) with representative vegetation communities and lower frequency of anthropogenic disturbances perpendicular to the direction of the Yellow River entering the sea were set up on the north bank of the Yellow River, and samples were collected in a way that combines belt transect and sample point method. There was a space range of 500-700m between the two adjacent belt transects. Within each belt transect, there was an interval of 50-150m between two adjacent sample points. With a soil collection tool, a soil sample of 10 cm×10cm was collected at each point with a sampling depth of 20cm. Soil samples were collected at four layers with depths of 0-5cm, 5-10cm, 10-15cm, and 15-20cm, respectively. Three repetition sampling points were set at each sample point, with a distance of 5-10m between two adjacent repetition sampling points. Then, soil samples collected from the same layer of the repetition sampling points were mixed into a sample.

A method of soil sample concentration was applied in this experiment (Heerdt et al., 1996). First, the collected seed bank samples were dried in sunlight. And then the large plant fragments and stones were removed using a 4 mm sieve, and then the soil was filtered with a 2 mm sieve (Ma et al., 2017b). After the residual plant tissues and stones in the samples were carefully removed, the seed bank samples were placed in a pot (30cm×20cm×10cm) for germination in a greenhouse in LuDong University. First, a 5cm layer of vermiculites were placed at the bottom of the experimental soil to hold water (Li, 2022). Second, prepared seed bank samples were evenly spread on the vermiculites layer. During the germination process, all pots were watered regularly to ensure adequate moisture. When seedlings appeared in the pots, they were identified. The species name and number of seedlings per pot were recorded. The identified seedlings were removed, meanwhile, unknown seedlings were transplanted to separately pots to grow until it could be identified (Gao et al., 2021). After all seedlings were identified, the soil samples were turned over to continue germination until no seedlings appeared for 2 weeks (Ma et al., 2011).

Three soil cores (d = 3.6 cm, h = 10 cm) were randomly taken next to seed samples quadrats and mixed into a sample to determine soil properties. Soil electrical conductivity and pH value were measured with a conductivity meter with glass electrodes and a pH meter, respectively. Soil total nitrogen content was measured with the Kjeldahl method (Stanley et al., 2019), soil total organic matter content was measured with the potassium dichromate external heating method (Kopáček et al., 2001), and soil total phosphorus content was measured with the ultraviolet spectrophotometric method (Kopáček et al., 2001; Miller and Arai, 2016; Mikajlo et al., 2023).

In this paper, the Shannon-Wiener diversity index (SWI) (Zou et al., 2021), Patrick richness index (PRI) (Courkamp et al., 2022), Jaccard similarity index (JCI) (Zhang et al., 2017), and Total seed density (TSD) were used to reflect the diversity of seed banks.

Patrick richness index:

Shannon-Wiener diversity index:

Jaccard similarity index:

Where, S represents the number of plant species; p_i refers to the ratio of the number of individual plant species to the total number of the plant community; A represents the composite set of ground vegetation species; and B is the composite set of the soil seed bank.

Microsoft Excel 2023, Origin 9.2, and GraphPad 9.3 software were used to perform the data sorting, statistical analysis, and chart drawing. One-way ANOVA by Duncan’s multiple range test (P< 0.05) was used to assess the differences in seed composition, distribution and diversity along transect A and B. In addition, a correlation analysis method was applied to analyze the relationship between the soil seed bank and its influencing factors, with all applied data being averages of three values repeatedly measured.

The results of diversity indices show that, from the sea to the river, the PRI, SWI, and JCI of the soil seed bank present similar trends of variation, all showing a trend of increasing first and decreasing then ( Figure 5 ). The diversity index of belt transect A has a higher value than that of belt transect B. In addition, the diversity of soil seed bank at different depth are also different. The SWI, PRI, and JCI indices of belt transect A at a depth of 5-15cm have a highest value, while the indices of belt transect B have a highest value at a depth of 0-5cm. Also, these three indices of belt transect A and B have a lowest value at a depth of 15-20cm.

In order to ensure the succession of species, plants often propagate through seeds to reduce their survival risks, thus maintaining the long-term existence of species (Childs et al., 2010; Metzner et al., 2017). Annual and perennial plants usually adopt different reproductive strategies. Annual plants mainly regenerate by seeds, while perennial plants can reproduce multiple times depending on their underground roots (Ida et al., 2013). Therefore, the species composition and density of seed banks are closely related to the species composition of surface aboveground vegetation (Capers, 2003; Liu, 2005; Chu et al., 2019). In the study of urban soil seed bank, herb plant species accounted for most of the soil seed banks, herb species were about one third of herb species in the aboveground vegetation communities, while woody species only were tenth of the woody plant species in the aboveground vegetation communities (Zhao et al., 2023). In alpine wetland, species and diversity of soil seed bank were significantly correlated with aboveground vegetation species, but seed bank dynamics do not change with aboveground vegetation during wetland degradation (Zhao et al., 2021). In order to ensure the continuation of species, plants often propagate through seeds to reduce their survival risks, thus maintaining the long-term existence of species (Childs et al., 2010; Childs et al., 2010; Metzner et al., 2017). In our study, 5 plant species of the soil seed bank were all herbs without woody plant, including one annual herb and four perennial herbs. S. salsa is annual herb which only regenerate by seeds. T P. australis, S. mariqueter, C. album and S. brachyotus are perennial herbs most depending on asexual reproduction around their aboveground vegetation communities. There are 11 species in aboveground vegetation, including 9 herb species and 2 woody species ( Table 3 ). Species and diversity of soil seed bank did not have close relationship with aboveground vegetation ( Figures 4E, F ). Tiny seeds of herb species nearby the river could be not spread over the woody plant to establish a new population by wind or water (Wang et al., 2015), which mainly resulted in species composition of soil seed bank of the nascent wetland in the YRD was significant different with the aboveground vegetation. The formation of a seed bank is influenced by various factors (Baldwin et al., 2010; Ruano et al., 2015; Benvenuti and Mazzoncini, 2021).

Many factors could affect species and density of soil seed bank, such as surface configuration (Havrdová et al., 2015), climate (Andrea et al., 2015), vegetation characteristics (Ma et al., 2017a), soil conditions (Ma et al., 2017b; Seibert et al., 2018), water regime (Anneke et al., 2018), and so on. In areas with good conditions, soil seed banks have a complex species composition and a higher density. On the contrary, in areas with disturbed factors, soil seed banks have a relatively simple composition and lower density (Zhao et al., 2022). To a certain extent, the fluctuation of soil water and salt contents will interfere with the balance of density and species composition of the soil seed bank. Also, the soil nutrient content will influence the seed variety and density of the soil seed bank with different influencing degrees. Some studies have shown that the species richness and seed density of wetland seed banks can be slightly affected by soil nutrient enrichment (Miao and Zou, 2009). Meanwhile, it is argued that the density condition of soil seed banks has a close relationship with the soil nutrient contents of nitrogen, phosphorus, and potassium (He et al., 2016).

In the estuary wetland, the initial vehicle of dispersal is wind (Chen et al., 2019; Zhu et al., 2022). When seeds fall to the ground, they will dispersal again faraway under the action of tides and surface runoff (Kim et al., 2022; Wang et al., 2022). Consequently, the seed dispersal distance and the consequent changes in the relevant factors have greatly impacted density and diversity of the seed bank. Existing studies have shown that dispersal distance has a significant influence on density of soil seed bank, but they did not show significant impact on the species composition (Wei et al., 2017). In the Tianjin coastal wetland, soil seed bank size is huge, but it contains relatively few species, which are primarily saline-alkaline tolerant plant, such as P. australis, S. salsa, and Aeluropus littoralis (He et al., 2014). Results in this paper are consistent with above results. In our study, density of soil seed bank in the new-formed wetland is great, but the species is only 5 herbs.

Among these factors, environmental factors can affect the compositional structure and density of soil seed banks. In areas with good environmental factors, soil seed banks have a complex compositional structure and a high richness of species. On the contrary, in areas with bad environmental factors, soil seed banks have a simple compositional structure and low richness of species (Zhao et al., 2022). To a certain extent, the fluctuation of soil water and salt contents will interfere with the balance of density and species composition of the soil seed bank. Also, the soil nutrient content will influence the seed variety and density of the soil seed bank, with different influencing degrees. Some studies have shown that the species richness and seed density of wetland seed banks can be slightly affected by soil nutrient enrichment (Miao and Zou, 2009). Meanwhile, it is argued that the density condition of soil seed banks has a close relationship with the soil nutrient (He et al., 2016). The results of this research show that EC has a certain influence on TSD, while there is no significant relationship between soil nutrients and TSD. The results of this study are slightly different from the results reported in the previous literature. The primary reason is that the salt content can exert a main inhibiting effect on the growth and manifestation of the plant community, thus further affecting the formation of the seed bank (Li and Xu, 2009).

In this study, an interesting finding is distance to the sea or river determining the density of soil seed bank, and salinity greatly affecting the diversity of soil seed bank(p ≤ 0.01) ( Figure 7 ). But there is no significant relationship between soil nutrients and density of seed bank. This are halfway similar to the results reported in the previous literature (Li and Xu, 2009; Erfanzadeh et al., 2010; Bai et al., 2014; Gao et al., 2018). The primary reason is that the salinity can exert a main inhibiting effect on the growth and manifestation of the plant community in the estuary wetlands, thus further affecting the formation of the seed bank. In this paper, diversity of soil seed bank with different buried depth are also affected by soil salt ( Figure 6 ). Our results have fully verified the influence of salt water and freshwater interactions on the spatial heterogeneity of soil seed banks.

Spartina alterniflora is an invasive species with great negative effects in the YRD. However, in this study, no S. alterniflora seeds were found in the soil bank in the nascent wetland. This is probably due to the fact that samples were obtained from areas with few disturbances, and distance to S. alterniflora community is long enough, limiting the dispersal and establishment of invasive species.