Enhanced Sweet Sorghum Growth and Soil Quality in Coastal Saline–Alkali Soils Through Organic Acid-Containing Bio-Based Materials and Microbial Synergy

Abstract

Coastal mudflats are characterized by high salinity and alkalinity, along with low mineral nutrient availability, making it challenging to achieve high biomass or effective yields when directly cultivating food or fodder crops. Exogenous complex saline soil amendments can enhance forage production, but their effects on soil salinity reduction and nutrient activation remain unclear. This study used pot experiments and laboratory analyses to investigate these effects. A 0.3% saline–alkali soil was treated with a combination of organic acids (fulvic acid and citric acid), bio-based materials (cow dung and pine needles), and beneficial microbial mixtures (Priestia megaterium + Trichoderma harzianum, Bacillus subtilis + Aspergillus niger, and Bacillus pumilus + Paecilomyces lilacinus). The organic acid bio-modifier significantly alleviated salinity stress in sweet sorghum, reducing soil salinity, increasing soil nutrient levels, enhancing root vigor and photosynthesis, and improving plant morphology, resulting in higher biomass yields. Among the factors tested, bio-based materials had the most pronounced effect. Citric acid, pine needles, Priestia megaterium, and Trichoderma harzianum enhanced sweet sorghum growth during the seedling stage, whereas fulvic acid, pine needles, Bacillus pumilus, and Paecilomyces lilacinus were more beneficial during the elongation stage.

1. Introduction

Jiangsu Province (30°45′–35°08′ N, 116°21′–121°56′ E) is situated in the central part of China’s coastal region, downstream of the Yangtze and Huai Rivers. Including an area of 68.7 × 10⁴ hm², mudflats are widely distributed throughout Jiangsu Province, with Yancheng alone covering approximately 45.3 × 10⁴ hm², accounting for over 70% of the province’s total. These mudflats continue to expand landward each year [1,2]. The prudent development of mudflats, which represent a vital reserve of land resources, can alleviate pressure on food production and arable land [3,4]. However, plant metabolism and growth are significantly hindered by salt stress due to the accumulation of ions such as Na⁺ and K⁺. Elevated concentrations of these ions, along with Cl⁻, are found in the soil [5]. High levels of Na⁺ and K⁺ ions can affect soil properties, including porosity, aeration, and hydraulic conductivity [6], ultimately reducing nutrient mobilization, microbial diversity, and crop yield [7]. Consequently, crops grown for seed harvest are often ill-suited for saline soils. In contrast, forages, which are cultivated for their nutritional constituents such as stems and leaves, can more effectively leverage climatic, land, and biological resources, resulting in a significant increase in biomass per unit area [8]. Additionally, the cultivation of forages enhances soil nutrient content and modifies soil bacterial and fungal communities [9]. Therefore, the establishment of salt-tolerant forage grasses on saline and alkaline coastal land not only provides economic benefits but also serves as an effective strategy for soil improvement.

Due to the complexity of the coastal mudflat development process, meeting planting requirements through a single-treatment approach is challenging. Currently, a prominent area of research focuses on the application of efficient chemical enhancement techniques in treatment processes, complemented by biological treatment methods. Exogenous organic acids can effectively activate or transform insoluble nutrients in the soil [10,11]. Furthermore, these organic acids promote interactions between crops and soil microbes, thereby enhancing enzyme activity and microbial vitality within the soil [12]. However, since exogenous organic acids are primarily available in granular or powdered form and are highly soluble, their direct application in coastal soils is susceptible to losses due to natural rainfall and soil hydrological processes [13]. To enhance the durability of organic acid utilization, it is essential to incorporate bio-based materials with good absorptive and buffering capacities for composite applications [14,15]. Previous studies demonstrate that the porosity and large specific surface area of decomposed bio-based materials [16] facilitate soil water retention [17], increase soil organic carbon content [18], reduce nutrient leaching, and improve nitrogen utilization [19]. Additionally, these materials create an optimal environment for the growth, reproduction, and metabolism of microorganisms [20].

Constrained by high salt ion concentrations, poor hydrothermal conditions, and limited carbon and nitrogen sources, the total number of soil microorganisms in coastal saline soils is lower than that in non-saline soils within the same ecological region [21]. In terms of microbial community composition, bacteria are the predominant microorganisms in saline–alkaline soils, exhibiting significantly higher abundance compared to fungi and actinomycetes [22,23]. Beneficial microorganisms serve as auxiliary additives, creating a favorable living environment for crop growth [24] and enhancing the availability of effective nutrients in the soil [25]. The microbial community structure of saline–alkali soils may also be altered during the remediation of saline soils through microbial amendments [26]. Kumar et al. [27] demonstrated that the Bacillus pumilus strain JPVS11 promotes plant growth, enhances soil enzyme activity, and mitigates the adverse effects of salinity in rice. Similarly, López-Bucio et al. [28] found that Trichoderma releases a specific biostimulant that fosters crop growth and yield. Additionally, it can reduce soil salinity and pH while increasing soil organic matter, quickly available phosphorus, and total nitrogen (TN) [29]. Li et al. [30] showed that Trichoderma, when combined with low concentrations of nitrogen fertilizers, can improve soil nutrient status and enhance maize production potential. Given the diversity of microbial functions, inoculated strains can enhance the soil environment and facilitate nutrient cycling. For example, the inoculation of nitrogen-fixing microorganisms can immobilize atmospheric nitrogen, thereby meeting the nitrogen demands of plants [31]. Phosphorus-solubilizing microorganisms can break down a variety of organic acids that react with insoluble phosphates, converting them into bioavailable phosphorus [32]. Despite the effectiveness of many microorganisms in improving saline soils, their treatment effects are often low and unstable, largely because a single strain may not adequately cope with multiple environmental factors, such as soil pH, osmotic pressure, organic matter content, and salt concentration. Composite microbial agents (CMAs), which consist of multiple functional strains, have been shown to enhance crop growth and improve management efficacy in saline soils. The combination of Aspergillus niger MJ1 with Pseudomonas stutzeri DSM4166 or mutant Pseudomonas fluorescens CHA0-nif has been shown to affect the diversity and abundance of bacterial communities in soils cultivated with lettuce [33]. Co-inoculation of Trichoderma with various Bacillus spp. of plant growth-promoting rhizobacteria (PGPR) plays a crucial role in enhancing plant growth, increasing nutrient uptake, and boosting the yields of different crops. Under greenhouse conditions, the application of a cell suspension of T. atroviride with Bacillus sp. and Pseudomonas sp. resulted in a 37% increase in the biomass of banana plants, outperforming the effects of each microorganism applied individually [34]. Hansen et al. demonstrated that co-inoculation of Penicillium bilaiae and Bacillus simplex enhances phosphorus uptake in low-phosphorus soils [35]. Likewise, Escobar et al. found that co-inoculation of Aspergillus and Bacillus promotes cotton growth [36]. Therefore, the combined application of organic acids, bio-based materials, and CMAs can improve soil properties and crop yields with greater efficiency.

However, the internal mechanisms of organic acids and the physicochemical properties of bio-based materials exhibit considerable variability. The effects of microorganisms also differ significantly. Their combined influence impacts both the growth of salt-tolerant forage grasses in saline soils and the enhancement of soil quality itself. Sun et al. [37] demonstrated that the composite application of biochar and xanthate can markedly reduce SS, regulate soil physical properties, and enhance nutrient availability, ultimately leading to increased crop yields. Xing et al. [38] developed a composite soil conditioner by combining biochar, salt-tolerant bacteria, alkaline soil conditioner, and earthworms. This amendment was applied to saline soils to decrease salt levels, thereby improving both the physical and chemical properties as well as enzyme activity. Farhadian et al. [39] found that the combination of humic acid and cow dung promotes optimal growth and development of coneflowers by directing photosynthetic products toward osmoregulatory compounds, such as proline (PRO) and soluble carbohydrates, which help plants withstand drought stress. Nacoon et al. [40] demonstrated that the inoculation of sandy soil with Ascomycetes mycorrhizal fungi (AMF) and phosphate-solubilizing bacteria (PSB) effectively promotes the growth and yield of sunflowers. Zai et al. [41] reported that the dual application of AMF and phosphorus-solubilizing fungi (PSF) mitigated the effects of salt stress on various growth parameters and nutrient uptake, with significant effects observed on root and PSF populations. Zhu et al. [42] indicated that the application of Bacillus sphaericus complex biofertilizers increased soil bacterial community diversity and significantly reduced bioavailable cadmium levels in the soil. Karimi et al. [43] pointed out that combining bacterial and fungal amendments demonstrated a stable and effective growth-promoting regulatory mechanism to enhance quinoa growth under salt stress. Although both national and international studies have explored the addition of various types of organic acids, bio-based materials, or CMAs, there is a notably limited body of research focused on the integration of biological amendments in composite formulations.

In order to identify the optimal combination of well-stabilized organic acids and bio-based materials inoculated with CMAs, this study utilized salt-tolerant sweet sorghum as the experimental subject. The sweet sorghum was cultivated in 0.3% high saline beach soil, and various organic acid bio-based materials were introduced alongside CMAs (Figure 1). By measuring parameters such as sweet sorghum biomass, antioxidant enzyme activity, chlorophyll (Chl) content, root activity (RA), and soil soluble salt (SS) content indices, we investigated the effects of organic acids and bio-based materials inoculated with CMAs on the growth of sweet sorghum and the quality of seashore saline soil. The aim was to provide a theoretical basis for the application of biofertilizers in the sustainable agricultural development of Jiangsu’s seaside saline soil.

2. Materials and Methods

2.1. Experimental Material

The experimental soil was collected from a coastal saline area in Snapping Town, Dongtai City, Jiangsu Province (32°86′ N, 120°93′ E). The classification and grading of the saline soil were determined in accordance with the national standard of the People’s Republic of China, “Classification and grade of saline–alkali soil for agricultural use”. Light saline soils (Ece: 2–4) and very heavy saline soils (Ece > 16) were used in this experiment. These soils were combined in a ratio of 7:3 and subsequently measured using a salinometer. If the salinity was too high, low salinity soil was added until the salinity was adjusted to 3 g·kg⁻¹. The basic physical and chemical properties of the soil were as follows: pH 8.47, soil organic matter (SOM) 2.54 g·kg⁻¹, TN 0.628 g·kg⁻¹, alkali-hydrolyzed nitrogen (AH-N) 11.17 mg·kg⁻¹, available phosphorus (AP) 16.1 mg·kg⁻¹, and available kalium (AK) 2.6 mg·kg⁻¹.

The variety of sweet sorghum tested was the forage sweet sorghum “Hunnigreen”, which is an interspecific hybrid with a late maturation period of approximately 180 d. This variety is characterized by high yield, strong drought resistance, good disease resistance, waterlogging resistance, and salt alkali resistance [44,45].

The tested strains included Priestia megaterium CICC no. 20665 (Pr. megaterium 20065), Trichoderma harzianum CICC no. 13010 (T. harzianum 13010), Bacillus subtilis CICC no. 10089 (B. subtilis 10089), Aspergillus niger CICC no. 2432 (A. niger 2432), Bacillus pumilus CICC no. 22096 (B. pumilus 22096), and Purpureocillium lilacinum CICC no. 40210 (P. lilacinum 40210), all sourced from Hubei Qiming Biological Engineering Co., Ltd (Yichang City, Hubei Province, China). Luria–Bertani medium (LB medium) were utilized to cultivate B. subtilis 10089, Pr. megaterium 20065, and B. pumilus 22096. Potato Dextrose Agar medium (PDA medium) were employed for the cultivation of A. niger 2432, T. harzianum 13010, and P. lilacinum 40210. The organic acids used in this study were citric acid and fulvic acid. The bio-based materials consisted of cow dung and pine needles, the latter supplied by Shijiazhuang Nongyou Biotechnology Co., Ltd (Shijiazhuang City, Hebei Province, China). The basic physical and chemical properties of pine needles were as follows: TN 14.2 g·kg⁻¹, total phosphorus (TP) 0.67 g·kg⁻¹, and total kalium (TK) 2.73 g·kg⁻¹. Pine needles underwent a natural drying process; they were spread in a shaded, ventilated area at a thickness of 5 to 8 cm before being crushed for subsequent use. Cow dung was sourced from Jurong Blue Sky Water Biotechnology Co., Ltd (Zhenjiang City, Jiangsu Province, China). with the following basic physical and chemical properties: TN 15.00 g·kg⁻¹, TP 3.79 g·kg⁻¹, and TK 14.00 g·kg⁻¹. The cow dung was dried to ensure moisture content remained below 85%. It was then mixed with straw, crushed, and placed in plastic pots with water to mature for over one month before being sun-dried.

2.2. Experimental Design

The potting experiment was conducted on 29 July 2023, in the net room of Nanjing Agricultural University’s Pailou Experimental Base. Each pot used for this experiment had a diameter of 20 cm and a height of 19 cm. We began by filling each pot with 4 kg of dry soil and thoroughly mixing in the organic acid, bio-based materials, CMAs, and fertilizer. The soil was added to a height of approximately 3 to 5 cm from the rim of the pot. After loading each treatment, pots were watered daily to stabilize the soil for a duration of one month. Prior to planting, we ensured the soil was at its maximum field moisture capacity. To sanitize the seeds before planting, they were soaked in a 5% sodium hypochlorite (NaClO) solution for ten minutes. After removing contaminants and debris, the seeds were rinsed five times with distilled water. Each pot was seeded with three seeds. Once the first true leaves emerged, we selected strong, uniformly growing plants, leaving two plants per pot. Throughout the testing period, soil water content was maintained at 60% ± 5% of the field moisture capacity.

The experimental setup and distribution were organized in a completely randomized design with three factors. Two types of organic acids were utilized: citric acid and fulvic acid, denoted as H and F, respectively. Additionally, two types of bio-based materials were employed: pine needles and cow dung, referred to as PN and CM. Three combinations of CMAs were included: Pr. megaterium 20065 + T. harzianum 13010, B. subtilis 10089 + A. niger 2432, and B. pumilus 22096 + P. lilacinum 40210, abbreviated as MT, SA, and PP. In total, there were 13 distinct treatments (the specific distribution of these treatments is illustrated in Figure 2 and

Table 1. Treatment of different organic acid bio-modifier additives.

Treatment
Citric acid (C)	Pine needle (PN)	Pr. megaterium 20065 + T. harzianum 13010 (MT)
		B. subtilis 10089 + A. niger 2432 (SA)
		B. pumilus 22096 + P. lilacinum 40210 (PP)
	Cow dung (CD)	Pr. megaterium 20065 + T. harzianum 13010 (MT)
		B. subtilis 10089 + A. niger 2432 (SA)
		B. pumilus 22096 + Pa. lilacinus 40210 (PP)
Fulvic acid (F)	Pine needle (PN)	Pr. megaterium 20065 + T. harzianum 13010 (MT)
		B. subtilis 10089 + A. niger 2432 (SA)
		B. pumilus 22096 + Pa. lilacinus 40210 (PP)
	Cow dung (CD)	Pr. megaterium 20065 + T. harzianum 13010 (MT)
		B. subtilis 10089 + A. niger 2432 (SA)
		B. pumilus 22096 + Pa. lilacinus 40210 (PP)

Control group (CK): no treatments were applied; only sweet sorghum was planted.

). Each treatment was replicated with five pots, and two additional pots were planted as backups in the event that any of the plants did not thrive, resulting in a total of 91 pots. The amount of organic acid applied was 0.44 g·pot⁻¹, whereas the substrate quantity was 17.76 g·pot⁻¹, and the volume of CMA administered was 200 mL·pot⁻¹ (with functional total microorganisms in the soil measured at 5 × 10⁶ CFU·g⁻¹ and a mixing ratio of strains established at 1:1). The conventional fertilizer application rate in the field was 300 kg/hm²; however, in the pot test, this rate was adjusted to be double that of conventional nitrogen and phosphorus application rates.

2.3. Measurement Items and Methods

2.3.1. Plant and Soil Sampling

Samples were collected at 20, 40, and 60 d into the growth of sweet sorghum. Sampling involved the random selection of three uniformly growing sweet sorghum plants from different pots within the same treatment group. Each index was repeated three times to assess its morphological, physiological, and biochemical characteristics [46].

Soil sampling occurred at 30, 60, and 90 d during the growth of sweet sorghum. The soil sampling point was positioned at the inner point of the equilateral triangle formed by the test pot, with soil collected to a depth of 10 cm. Inter-root soil was extracted by gently shaking the roots after carefully excavating the root system of the sweet sorghum plants. The soil samples were then placed into sterile Ziploc bags after removing debris and stones. Each sealed bag was labeled with the date of sampling and the treatment name [47].

2.3.2. Measurement of Plant Growth Indicators

The following morphological and biomass parameters were measured: Stem diameter (SD) was assessed using a vernier caliper, whereas stem height (SH) and root length (RL) were recorded with a tape measure. The fresh weight (FW) of a single plant was determined using an electronic balance. Subsequently, the samples were killed green in an electric thermostatic drying oven at 105 °C and then dried at 65 °C until a constant weight was reached. The single plant biomass (BP) was calculated accordingly.

2.3.3. Measurement of Plant Physiological and Ecological Indicators

The triphenyltetrazolium chloride method [48] was employed to assess RA. For the measurement of Chl, the acetone–ethanol extraction method [49] was utilized. Proline (PRO) levels were determined using the ninhydrin method [50], whereas malondialdehyde (MDA) content was measured through the thiobarbituric acid colorimetric method [51]. The methodology proposed by Zhang et al. [52] was followed to evaluate the superoxide anion radical content (${\mathrm{O}}_2\stackrel{-}{·}$).

The activity of superoxide dismutase (SOD) was analyzed using the nitrotetrazolium blue chloride reduction method, whereas catalase (CAT) activity was assessed via the ultraviolet absorption method. The O-methoxy-phenol oxidation method was employed to determine peroxidase (POD) activity. For comprehensive details on the methodologies and calculation procedures, please refer to Zahir et al. [53].

2.3.4. Determination of Soil Physical and Chemical Properties

Soil pH was measured utilizing a conductivity meter (Eutech pH 700, New York, ES, USA) with a soil-to-water ratio of 1:5. SS was determined through the mass method, maintaining a soil-to-water ratio of 1:5. Soil electrical conductivity (EC) was also assessed using a conductivity meter (DDS-307A, Shanghai INESA Scientific Instrument CO., LTD, Shanghai, China) with the same soil-to-water ratio of 1:5. Additionally, SS was evaluated via the residue drying mass method, again using a 1:5 soil-to-water ratio [54]. Following the establishment of the association between SS and EC, the SS of all samples was calculated using the formula:

$$\mathrm{S}\mathrm{S}=0.0028\mathrm{E}\mathrm{C}+0.7085,$$

where SS means soil soluble salt content (g∙kg⁻¹), and EC represents soil electrical conductivity (μs∙cm⁻¹).

2.4. Data Processing and Statistical Analysis

IBM SPSS Statistics 23.0 (SPSS Inc., Chicago, IL, USA) software was employed for one-way and three-way analysis of variance (ANOVA), whereas preliminary statistics were conducted using Microsoft Excel 2021. The interactions among the three components were evaluated through a three-way ANOVA, and multiple comparisons were performed using Duncan’s new multiple-range test. Data representation in charts was expressed as “mean ± standard error”, with an alphabetical method employed to denote the significance of differences. Graph construction was facilitated by GraphPad Prism 9.0.

3. Results

3.1. Impact of Organic Acid Bio-Modifier on Morphological Characteristics of Sweet Sorghum

Exogenous organic acids, bio-based materials, and CMAs had varying effects on sweet sorghum at different growth stages (Figure 3 and Figure 4). From 20 to 60 d, SH, SD, FW, RL, and BP generally increased in sweet sorghum, with highly significant differences (p < 0.01) in their responses to the bio-based materials and CMAs. The responses to organic acids were also highly significant (p < 0.01), except for SH and SD at 20 d and RL at 60 d, which showed significant differences (p < 0.05). At 20 and 40 d, SH, SD, FW, RL, and BP were significantly higher (p < 0.05) in all treatments compared to the CK, with increases ranging from 4.21% to 213.22%, 13.83% to 145.69%, 2.57% to 341.06%, 3.03% to 424.18%, and 2.58% to 341.06%, respectively. The highest increases were observed in the CPN + MT treatment. At 60 d, sweet sorghum SH, SD, FW, RL, and BP in the FPN + PP treatment were higher than in the other treatments, reaching their highest values. Compared to the CK, SH, SD, FW, RL, and BP increased by 48.28%, 177.07%, 386.27%, 204.35%, and 387.45%, respectively. In conclusion, the CPN + MT and FPN + PP treatments were most beneficial for stem elongation and thickening during the nutritive growth period of sweet sorghum.

3.2. Impact of Organic Acid Bio-Modifier on Photosynthesis and Root Physiology of Sweet Sorghum

The responses of CMAs, bio-based materials, and exogenous organic acids to Chl and RA in sweet sorghum leaves varied between 20 d and 60 d (Figure 5). The effects of CMAs, organic acids, and bio-based materials yielded highly significant differences (p < 0.01), with the exception of Chl at 20 d, where the differences were not significant (p ≥ 0.05). At 20 d, CPN + MT exhibited the highest Chl content, followed by FPN + PP, with a significant difference between the two treatments (p < 0.05). Similarly, RA was highest in the CPN + MT treatment, significantly differing from the other treatments (p < 0.05). Compared to the CK, increases in Chl and RA were observed at 60.56% and 72.94%, respectively. At 40 d, CPN + MT again displayed the highest Chl content, whereas CPN + PP followed closely, showing no significant difference (p ≥ 0.05) between these treatments. However, RA was highest in the CPN + SA treatment, which significantly differed from the others (p < 0.05). Compared to the CK, Chl and RA increased by 160.01% and 321.62%, respectively. By 60 d, Chl and RA values for FPN + PP were significantly greater than those of all other treatments (p < 0.05), increasing by 208.94% and 129.41%, respectively, in comparison to the CK.

3.3. Impact of Organic Acid Bio-Modifier on Antioxidant Enzyme Systems of Sweet Sorghum

The application of CMAs, organic acids, and bio-based materials increased the antioxidant enzyme activities (SOD, POD, and CAT) in sweet sorghum leaves at different growth stages compared to the CK (Figure 6). From 40 d to 60 d, bio-based materials and CMAs significantly enhanced POD and CAT activities (p < 0.01). In contrast, organic acids showed no significant effects (p ≥ 0.05), except for a highly significant increase in SOD and CAT activities at 60 d (p < 0.01). At 20 d, CMAs did not significantly affect SOD, POD, or CAT (p ≥ 0.05), whereas bio-based materials significantly improved all three enzyme activities (p < 0.01). Organic acids significantly increased CAT activity (p < 0.01) but had no significant effects on SOD and POD (p ≥ 0.05). SOD, POD, and CAT activities peaked at 20 d, decreased slightly at 40 d, and increased again at 60 d. At 20 d, the highest SOD and CAT activities were observed in the CPN + MT treatment, which was not significantly different from FPN + PP and CPN + PP treatments (p ≥ 0.05) but was significantly higher than other treatments (p < 0.05). Similarly, POD and CAT activities were highest in CPN + MT and significantly differed from other treatments (p < 0.05). At 40 d, SOD activity was highest in the CPN + MT treatment, although it did not significantly differ from FPN + MT, FPN + PP, CPN + PP, or CPN + SA treatments (p ≥ 0.05). CAT activity, however, was significantly higher in CPN + MT than in other treatments (p < 0.05). At 60 d, SOD activity peaked in the FPN + PP and CPN + PP treatments, significantly exceeding other treatments (p < 0.05). FPN + PP treatment also exhibited the highest POD activity, which was significantly different from other treatments (p < 0.05). CAT activity was highest in FPN + PP but did not differ significantly from CPN + MT and CPN + PP treatments.

3.4. Impact of Organic Acid Bio-Modifier on Osmoregulation in Sweet Sorghum

The results demonstrated that the application of CMAs, organic acids, and bio-based materials exerted a significant influence on the ${\mathrm{O}}_2\stackrel{-}{·}$ production rate, MDA content, and PRO content of sweet sorghum at various growth stages (Figure 7). The sweet sorghum ${\mathrm{O}}_2\stackrel{-}{·}$ production rate, MDA content, and PRO content were highly significant (p < 0.01) in response to bio-based materials from 20 d to 60 d, CMAs at 40 d, and organic acids at 60 d. In addition, CMAs also had a highly significant (p < 0.01) effect on the ${\mathrm{O}}_2\stackrel{-}{·}$ production rate as well as the MDA content at 60 d, and organic acids had a significant effect on the ${\mathrm{O}}_2\stackrel{-}{·}$ production rate of 20 d (p < 0.01). The PRO content was highest at 20 d, particularly in the CPN + MT treatment, followed by FPN + PP, with no significant differences between these two treatments but significant differences with the other treatments (p ≥ 0.05). The lowest MDA content was observed in the CPN + MT, CPN + PP, FPN + PP, and CPN + SA treatments, with no significant differences among them (p ≥ 0.05). The ${\mathrm{O}}_2\stackrel{-}{·}$ production rate was the lowest in CPN + MT, significantly differing from other treatments (p < 0.05). At 40 d, the PRO content was significantly higher in the CPN + MT treatment compared to other treatments (p < 0.05). The MDA content and the production rate were the lowest in CPN + MT and significantly different from other treatments (p < 0.05). At 60 d, FPN + PP, CPN + PP, CPN + MT, FPN + MT, and CPN + SA treatments exhibited the highest PRO content, with no significant differences among these treatments (p < 0.05).

3.5. Impact of Organic Acid Bio-Modifier on Saline and Alkaline Characteristics of Beach Soisl

The application of CMAs, organic acids, and bio-based materials affected the pH, EC, and SS values of beach saline soils at different growth stages (Figure 8). Organic acids significantly reduced soil pH from 30 to 90 d (p < 0.01) and soil EC at 90 d, with no significant effects on other parameters (p ≥ 0.05). Bio-based materials significantly affected pH, EC, and SS from 30 to 90 d (p < 0.01) and had significant effects on EC and SS values at 40 d (p < 0.05). At 30 d, the CPN + MT treatment exhibited the lowest soil pH, EC, and SS values, though these did not significantly differ from FPN + PP and CPN + PP treatments (p ≥ 0.05). At 60 d, the soil pH was the lowest in the CPN + MT treatment but did not differ significantly from other treatments (p ≥ 0.05). Soil EC and SS values were also lowest in CPN + MT and FPN + PP treatments, with no significant differences from CPN + PP (p ≥ 0.05). At 90 d, the lowest soil pH was observed in the FPN + PP treatment, which did not significantly differ from CPN + PP (p ≥ 0.05). EC and SS values were also lowest in FPN + PP and significantly differed from other treatments (p < 0.05).

3.6. Correlation Analysis Between Growth Physiology of Sweet Sorghum and Soil Saline–Alkali Indexes in Different Growth Periods

Correlation analysis revealed that the morphological indicators and biomass (SH, SD, FW, RL, and BP) of sweet sorghum were significantly positively correlated (p < 0.01) with physiological and biochemical indicators of sweet sorghum (Chl, RA, PRO, SOD, CAT and POD) and significantly negatively correlated with MDA and ${\mathrm{O}}_2\stackrel{-}{·}$ in the three periods of time when a CMA was inoculated with the organic acid-based material (Figure 9).

As illustrated in

Table 2. The magnitude of change in each index for the most relevant treatment compared with CK.

The Maximum Change Amplitude of Each Index Compared with CK (%)	CPN + MT	FPN + MT
SH	213.22%	48.28%
BP	341.06%	387.45%
RL	424.18%	204.35%
SD	145.69%	177.07%
FW	341.06%	386.27%
Chl	160.01%	208.94%
RA	321.62%	129.41%
SOD	225.13%	86.53%
POD	110.89%	116.69%
CAT	90.86%	115.23%
${\mathrm{O}}_2\stackrel{-}{·}$	82.43%	89.23%
MDA	52.95%	54.73%
PRO	238.36%	37.70%
pH	10.31%	12.68%
EC	19.08%	14.06%
SS	19.08%	14.06%

, the CPN + MT treatment enhanced the growth of morphological indices such as SH and SD of sweet sorghum during the seedling stage, yielding highly significant differences from the control treatment (CK). The CPN + MT treatment effectively regulated the saline conditions and nutrient levels of the saline soil, facilitating the early growth of sweet sorghum during seedling establishment. Meanwhile, the FPN + PP treatment improved ground strength over the long term and promoted stable development and biomass accumulation of sweet sorghum in the middle and late stages. In coastal beach areas, the CPN + MT treatment was employed to enhance the fresh grass yield of green-mowed forage using CPN treatment or to increase the biological yield of silage and seed forage via the FPN + PP treatment, in alignment with the production characteristics of salt-tolerant forage grasses.

4. Discussion

4.1. Mechanisms of Organic Acid Bio-Modifier Formulated to Regulate the Growth of Sweet Sorghum in Saline Soil

Under saline and alkaline stress conditions, salt-damaging ions infiltrate and accumulate within the plant [55], significantly impacting its ability to absorb water and maintain cellular equilibrium [56]. This accumulation adversely affects various growth metrics, including SH and physiological and biochemical indicators such as Chl content in sweet sorghum. In this study, each treatment exhibited a notable positive effect on stalk thickening, elongation growth, and biomass when compared with the control group. This improvement may be attributed to the neutralization of exogenous organic acids with alkaline substances and the complexation of soluble base ions within the root zone of the soil [57]. The resultant complexation products were further stabilized by bio-based materials, thereby enhancing the growth environment and effectively reducing the overall concentration of soil salts and alkali components [58]. Additionally, under salt and alkaline stress, exogenous microorganisms contributed to the maintenance of root integrity, regulation of root architecture, and enhancement of sweet sorghum vitality [59]. Moreover, a range of plant hormones and signaling molecules were generated during metabolic processes, which play a role in regulating root development, stimulating plant growth, and assisting sweet sorghum in adapting to saline–alkaline conditions [60].

When plants experience salt–alkali stress, they produce a significant amount of reactive oxygen species (ROS), which leads to the peroxidation of lipids and alters the activity of antioxidant enzymes. To mitigate oxidative damage, plants that are persistently subjected to oxidative stress activate their own antioxidant defense mechanisms [61]. The results indicated that the application of organic acids, bio-based materials, and CMAs enhanced the content of Chl, increased the activities of SOD, POD, and CAT, slowed down the production rate of ${\mathrm{O}}_2\stackrel{-}{·}$, increased the content of PRO and decreased the content of MDA. This suggests that the complex additive can facilitate the accumulation of osmoregulatory substances in sweet sorghum leaves, enhance antioxidant enzyme activity, and maintain cellular osmotic homeostasis under saline and alkaline stress. Furthermore, it may serve as a biomolecule protector and ROS scavenger. This finding aligns with the experimental results of Wang et al. [62], which concluded that salinity stress prompts the accumulation of PRO and other compounds, thus improving tolerance to salt and heat stress in rice. The highest activity of antioxidant enzymes across all treatments was observed around 20 d, likely due to the accumulation of ROS in sweet sorghum leaves shortly after seedling emergence, which triggers antioxidant enzyme production. Our experimental results also demonstrated that the addition of various bio-amendments had differential effects on sweet sorghum metrics across the three observation periods. The most pronounced effects were noted at 20 d and 40 d with CPN + MT and at 60 d with FPN + PP. This variation may be attributed to sweet sorghum being exposed to different concentrations and types of saline and alkaline chemicals at different growth stages. Exogenous organic acids and bio-based materials exhibit varying complexation abilities and physicochemical properties, whereas the CMA has specific functional effects. Consequently, sweet sorghum’s growth and physiological and biochemical indices display distinct response patterns over time.

4.2. The Dynamic Change Pattern of Salinity and Alkalinity of Beach Soil by Organic Acid Bio-Amendments

The physicochemical characteristics of saline soil were effectively enhanced by organic acids and bio-based materials, which also promoted the amelioration of salt stress [63]. In this study, the CPN treatment exhibited a lower pH, EC, and SS at both 30 and 60 d compared to the other treatments. This phenomenon can be attributed to the application of citric acid, which significantly increases the solubility of calcium sulfate and calcium carbonate in the soil [64,65]. The resultant liberated Ca²⁺ ions displace a considerable amount of Na⁺ that is adsorbed to soil colloids. Consequently, the displaced Na⁺ interacts with carboxyl groups to form relatively stable complexes that do not compromise crop growth. Furthermore, this process aids in complexation solubilization, enhancing the soil environment by lowering the soil pH and efficiently neutralizing CO₃²⁻, HCO₃⁻, and other anions [66]. Although decomposing pine needles possess a significant adsorption capacity and can stabilize the adsorption of citric acid, thereby slowing the leaching and volatilization processes, this effect is challenging to maintain over time due to the higher solubility and crystallization potential of citric acid. In contrast, fulvic acid is more acidic and soluble than citric acid due to its higher concentration of acidic functional groups (−COOH) [67]. This translates to an increased cation exchange capacity, which may further promote the substitution of Na⁺ by soil colloids. Naturally, the effects of fulvic acid are more enduring than those of citric acid [68,69].

Alkaline conditions can severely damage the structural integrity and functionality of bacterial cell membranes [70]. Bacillus spp. strains counter this by maintaining a neutral pH within their cells through increased concentrations of proteins on the exterior [71]. They also adapt to alkaline environments by regulating Na⁺ ion concentrations within their cell membranes. When subjected to high salinity and alkaline environmental stresses, Bacillus spp. strains can induce the production of a significant quantity of general stress proteins (GSPs), which are capable of sensing surrounding pH levels and activating specific signaling pathways. These pathways can regulate the expression of the clpC gene, facilitating adaptation to saline environments. Furthermore, T. harzianum is able to colonize the root system, thereby stimulating the expression of defense response genes and enhancing antioxidant enzyme activities [72]. This organism also produces various organic acids during metabolism [73], which combine with Ca²⁺ and Mg²⁺, displacing Na⁺ and effectively regulating soil acidity and alkalinity, thus creating a more favorable environment for survival.

The results of this experiment indicated that the saline control, characterized by high soluble salt content, along with the application of Pr. megaterium 20065 and T. harzianum 13010, significantly reduced SS and pH. This reduction in salinity and pH mitigated the transfer of high concentrations of Na⁺ ions from roots to shoots, enabling sweet sorghum to preserve its antioxidant enzyme activities. This preservation occurs by stabilizing the cytoplasm against the toxic effects of Na⁺ ion accumulation and enhancing the activity of defense-related antioxidants. Among the three microbial agents tested, the combination of Pr. megaterium 20065 and T. harzianum 13010 exhibited the most substantial effects at 30 and 60 d. This observation can be attributed to the distinct mechanisms through which Pr. megaterium and T. harzianum operate in saline environments, as each has unique influences in these conditions. T. harzianum enhances plant resilience by upregulating genes associated with osmoprotection before salt stress occurs and activating the stress response system in sweet sorghum. Additionally, it synthesizes stress-resistant biochemicals and scavenges ROS, thus aiding the plant in enduring salt stress during the seedling phase, a finding that aligns with the results of Viterbo et al. [74]. Furthermore, it regulates the community structure and functions of rhizosphere microorganisms, promoting root growth by providing space and nutrients for microbial activity. Conversely, Pr. Megaterium serves as a PGPR. It enhances the abundance of phosphorus cycling genes [75], leading to the production of organic acids that solubilize otherwise unavailable inorganic and organic phosphorus. It also releases phosphatases, which mineralize organic phosphorus, converting it into plant-available orthophosphate and improving overall soil nutrient conditions [76].

The effects of B. pumilus 22096 and Pa. lilacinus 40210 were most pronounced at 90 d, likely attributed to their sustained effects. B. pumilus produces phytase, which degrades phytic acid and enhances phosphorus availability in the soil [77]. It also generates non-enzymatic fractions that scavenge excess reactive oxygen species [78], thereby improving the plant’s salt tolerance and counteracting soil re-salting processes in the later stages of growth. Pa. lilacinus acts as an effective bioremediating agent and phytostimulant, producing secondary metabolites and phytohormones (such as indole-3-acetic acid, IAA) that promote plant and root growth following the later growth stages of sweet sorghum [79,80]. Additionally, it releases organic acids that chelate mineral ions or lower pH levels to solubilize insoluble phosphates for plant uptake [81]. Its strong environmental adaptability and wider acid–base tolerance enable it to thrive in saline and alkaline conditions alongside B. pumilus.

4.3. The Feedback Mechanism of Organic Acid Bio-Amendment Formulation on Plants and Soil Microorganisms

PSF refer to a plant’s ability to influence its own growth, development, metabolism, defense, and tolerance to biotic and abiotic stresses by altering the soil environment surrounding it or affecting subsequent generations of plants growing in that environment [82]. As integral components of the soil microbiological landscape, soil microorganisms play a pivotal role in soil functionality, impacting nutrient availability, plant growth, and overall ecosystem function. Their physiological traits, including symbiosis, resistance, and decomposition, enhance the efficiency of plant nutrients within the soil, thereby promoting plant colonization, growth, and development [83,84]. When the structure of the soil microbial community shifts, it provides feedback to plants, resulting in corresponding changes in plant indicators [85]. Typically, salt-stressed situations harbor fewer microorganisms compared to normal soils, with soil salinization inversely correlated with microorganism abundance, as salt stress diminishes microbial activity [86]. In general, microbial populations are smaller in salt-stressed environments than in their non-stressed counterparts. The degree of soil salinization has a negative correlation with microbial activity, which is decreased by salt stress [87].

The distribution and community structure of soil microbial diversity is also impacted by physiological changes in plant root litter and inter-root secretions [88]. Under salt stress, both exogenously supplied organic acids and those released by the root system act as signaling molecules to regulate the plants’ tolerance to saline conditions. Moreover, these organic acids help maintain intracellular homeostasis within root cells and impact the inter-root environment [89]. Among the two organic acids examined in this study, citric acid demonstrated a more pronounced effect on the soil through plant interaction [90]. This is largely due to the significantly increased concentration of citric acid secreted by the root system in response to exogenous application. Such an increase establishes a new ionic equilibrium by reducing intracellular Na⁺ concentration and impairing the transfer of Na⁺ from roots to the soil [91]. Additionally, citric acid provides microorganisms with a carbon source, promoting microbial growth and reproduction while altering the composition of the soil microbial community [92]. The interactions between plants and soil microorganisms represent a burgeoning area of research. Future studies should further explore the role of soil microbial diversity in regulating the reciprocal interactions of “salt-tolerant forage—coastal saline soil”.

5. Conclusions

The growth performance of sweet sorghum cultivated in saline–alkali soil was influenced by the interaction among CMAs, organic acids, and bio-based materials. In this study, the slow-release properties of fulvic acid contributed to long-term enhancements in soil quality. Decayed pine needles provided stability to the adsorption of organic acids, effectively reducing their leaching and volatilization. B. pumilus 22096 and Pa. lilacinus 40210 facilitated the growth of both plant roots and vegetative organs. Additionally, Pr. megaterium 20065 demonstrated the ability to solubilize insoluble phosphates in the soil, thereby creating a more favorable growth environment for plants. T. harzianum 13010 secretes substances that deliver essential nutrients to the plant. In conclusion, the combination of citric acid, pine needles, Pr. megaterium 20065, and T. harzianum 13010 was most beneficial for the growth of saline sweet sorghum during the seedling stage. Conversely, the combination of fulvic acid, pine needles, B. pumilus 22096, and Pa. lilacinus 40210 proved to be more suitable for the elongation stage of saline sweet sorghum.