Exploring pea soaking water as alternative to synthetic fertilizer: growth and microbial analysis of pea and tomato plants

Abstract

Synthetic fertilizers deplete soil microbiome. Organic fertilizers lack specific nutrients for plant growth. A fertilizer that delivers essential nutrients to plants without deteriorating soil is lacking. This study investigated the viability of using pea soaking water (PSW) as substitute for synthetic fertilizers to enhance the growth of pea and tomato plants. Traits included plant growth stages, shoot and root weight, estimated chlorophyll content, and soil microbial populations. Pea plants exhibited consistent growth stages and rates of development across treatments, whereas tomato plants displayed treatment-dependent growth variations and differences in rates of development. Synthetic fertilizer (NPK) and PSW treatments increased shoot weight and chlorophyll content in both pea and tomato plants, compared with their controls. Interestingly, PSW produced comparable shoot growth to synthetic fertilizer in both crops. Root weights were similar in response to both fertilizer treatments in tomato but only increased in response to synthetic fertilizer in pea plants. Soil microbial analysis highlighted differences in Lactobacillus amount with soil supporting pea plants having higher bacteria counts. Notably, Lactobacillus amounts were reduced by 48% in the synthetic fertilizer treatment, but not by PSW, compared with the control, for tomatoes only. These findings suggest that PSW is a potential alternative to synthetic fertilizer to sustainably support plant growth. Replacement of synthetic fertilizer with PSW could reduce the environmental impact of agriculture by promoting healthy soil microbiota and preventing eutrophication, as well as reducing reliance on fertilizers. Further research is needed to explore its effect on crop yield, and applicability across crop species, field, and environmental conditions.

Introduction

Application of mineral fertilizers such as nitrogen (N), phosphorous (P) and potassium (K) is the primary method to maintain or restore soil nutrients and increase crop yields¹. NPK fertilizers not only can increase crop yields, but also directly or indirectly affect the chemical, physical and biological characteristics of the soil².

Increasing food productivity means increasing the use of fertilizers, but this may have harmful environmental consequences because fertilizers are a source of pollution, along with compost, manure and plant residue. This and excess use of fertilizers may have a major negative impact on food safety and human health³, for example through eutrophication, nitrate (NO₃) contamination of groundwater, and the accumulation of heavy metals in soil⁴. Eutrophication is characterized by increased availability of one or more restrictive growth factors required for photosynthesis such as carbon dioxide and nutrient fertilizers, which is leading to excessive growth of plants and algae⁵. In intensive agricultural regions, the excess use of NPK fertilizer resulted in an excessive accumulation of nutrients in soils, ground water and surface water, causing eutrophication. Moreover, about 70% of waterways are polluted by N and 40% by P because of agricultural practices in the UK, and in the USA, eutrophication from agriculture is one of the main sources of water pollution⁶. As such, many agricultural systems are focusing on increasing the organic matter content in soil as a measure of increased fertility, improved soil health and carbon isolation for greenhouse gas mitigation.

Soil microbial biomass and the number of bacteria and fungi are affected by long-term fertilizer use². Lactobacilli microorganisms are beneficial to soil health, and consequently plant growth. Numerous species of Lactobacillus plantarum (ONU 991, ONU 12, ONU 313, and ONU 316) have been shown to increase shoot length and root growth of tomatoes⁷. Such effects were attributed to the metabolites produced by lactic acid microorganisms. Other lactic acid bacteria, such as Lactobacillus strain BB6 and Enterococcus sp. strain BB3, were found to increase photochemical ability in tomato plants by preventing oxidative damage. Lactic acid bacteria metabolize sugar, synthesizing thiols and antioxidant enzymes that protect plants⁸. Similarly, pea plants were shown to benefit from the presence of lactic acid bacteria in the soil. Both direct and indirect effects can be observed: lactic acid bacteria directly support plant growth, with mechanisms such as synthesis of phytohormones (compounds that stimulate plant growth) and siderophores (iron chelating compounds), solubilization of phosphate and fixation of nitrogen. Indirect support of plant health by lactic acid bacteria includes competition with pathogens for nutrients, synthesis of antimicrobial compounds (organic acids and bacteriocins) and synthesis of exopolysaccharides (water retention)⁹.

Processing of food legumes generates wastewater that contains nutrients useful for plant growth. For example, in a study by Huang and collaborators¹⁰, soaking water of legumes contained 1.89 g/100 mL of dry matter, consisting of 0.69 g/100 mL of soluble carbohydrates, 0.34 g/100 mL of insoluble carbohydrates, 0.60 g/100 mL of protein, and 0.26 g/100 mL of minerals. Legume-derived wastewater has been applied to food to improve taste and nutrition due to its content of soluble fibre and protein, as well as minerals, saponins and phytochemicals^11,12. Tofu whey has been used successfully in replacement of urea in cherry tomatoes¹³. Partial replacement of urea with tofu whey (50:50 ratio) resulted in less ammonia volatilization from the soil since nitrogen in tofu whey is bound (amino acids, peptides, protein). Improved soil fertility was attributed to prebiotic fibre which favoured the growth of beneficial microorganisms and improved nutrient solubility in the soil. Higher fruit weight was observed and attributed to organic compounds such proteins, reducing sugars and polyphenols. In addition, it was estimated to have a cost saving output of 26–52%, although processing costs of tofu whey (drying, etc.) were not considered. Tofu whey was also used in replacement of synthetic fertilizer NPK in komatsuna (Japanese mustard spinach) and potatoes¹⁴. Plant growth was comparable between NPK and tofu whey, as a result of soluble protein, sugars and minerals. Unlike compost, manure or tofu whey, pea soaking water contains soluble fibre, oligosaccharides and saponins, on top of soluble protein. This composition allows for both prebiotic activity (oligosaccharides, soluble fibre) and antimicrobial (saponins).

In the present study, it was hypothesized that the soaking water of peas could replace synthetic fertilizer (NPK) and successfully support plant growth. Consequently, this study investigated the potential of pea soaking water (PSW) as a fertilizer. A leguminous (pea) and a non-leguminous crop species (tomato) were chosen as representative crops. Growth stages, shoot and root weight, estimated chlorophyll content and microbial load of the soil were measured. Tomatoes were chosen because they do not fix nitrogen and require N, P, and K. Peas were chosen to investigate the possibility of returning the nutrients from the soaking water back to the next crop of peas¹⁵. To the best of our knowledge, this is the first study investigating the potential of legume wastewater as an alternative to NPK (Table 1).

Table 1.

Mineral composition of the two fertilisers tested: Synthetic (Osmocote Exact) and pea soaking water (PSW)^10,16,38.

	Mineral content (% dry matter)
	Synthetic fertilizer	Pea soaking water
Nitrogen	16	5.9
Phosphorous	3.9	0.9
Potassium	10	6.4
Magnesium	1.2	0.2
Iron	0.45	0.004
Manganese	0.06	0.002
Boron	0.02	Not detected
Copper	0.05	0.002
Molybdenum	0.02	0.0001
Zinc	0.015	0.014

The main aims of this study were to:

1. Evaluate pea soaking water as organic fertilizer to grow pea and tomato plants.

2. Assess the impact of pea soaking water on soil Lactobacilli.

Results

Growth stages

In pea plants, there were no significant differences among growth stages in the three treatments (control, fertilizer, and PSW). Sprouts appeared after 7–8 days, followed by seedlings with leaves after 12 days. Longer time was required for pea flowers (40–42 days) and pea pods (46 days) to develop (Table 2). Statistically significant shorter times were required for flowers to appear in the treated pea plants: 40 vs. 42. (Table 2). In contrast, the appearance of pea pods occurred at the same time (46 days) regardless of treatment (Table 2). These findings indicate that nutrient availability does not influence the timing of vegetative and reproductive development in this species.

Table 2.

Growing stages of the plant treatments: PC (Pea, Control), PF (Pea, Fertilizer), PS (Pea, Soaking water), TC (Tomato, Control), TF (Tomato, Fertilizer), and TS (Tomato, Soaking water). Results are indicated in days. Different letters represent statistically significant differences withing each growing stage (Tukey, p < 0.05).

Treatment	Sprouts	Leaves	Flowers	Fruits
PC	7 ± 1a	12 ± 1a	42 ± 76a	46 ± 1a
PF	8 ± 1a	12 ± 1a	40 ± 76b	46 ± 1a
PS	8 ± 1a	12 ± 1a	40 ± 76b	46 ± 1a
TC	13 ± 1a	−	-	-
TF	14 ± 1a	22 ± 1a	-	-
TS	13 ± 0a	24 ± 1a	-	-

In contrast, treatment-specific differences were observed among the tomato plants. While tomato sprouts appeared after 13–14 days for all treatments, no further growth was observed in the tomato control plants (Table 2). Leaves appeared only in tomato plants treated with NPK and with PSW after 22–24 days (Table 2). No flowers or fruits appeared in the tomato plants during the 49-day trial, suggesting that a longer trial period would have been required for the formation of reproductive organs.

Shoot and root weight

As expected, shoot fresh weights and dry weights were significantly higher (3.5- to 4-fold) upon use of fertilizer in pea plants than the control plants. Interestingly, the treatment with PSW produced comparable shoot growth to the NPK treatment in this species (Fig. 1). While control tomato plants did not grow further after sprouting without fertilizer, the tomato shoots were comparable between NPK and PSW, ranging from 22 to 26 g fresh weight and 2.6 to 2.8 g dry weight (Fig. 1). NPK was shown to yield larger leaves in peas, resulting in significantly higher fresh weight and dry weight of pea leaves. As for the belowground growth of pea plants, the NPK treatment resulted in roots with 70% higher fresh weights, and more than 2-fold higher dry weights than control plants (Fig. 2). No significant root weight differences were observed between control pea plants and those treated with PSW. This contrasted with tomato, where both fertilizer treatments (synthetic and PSW) had comparable fresh and dry root weights, both higher than the untreated plant (Fig. 2).

Fig. 1.

Wet weight (fresh) and dry weight (dried) of shoots harvested for all treatments: PC (Pea, Control), PF (Pea, Fertilizer), PS (Pea, Soaking water), TC (Tomato, Control), TF (Tomato, Fertilizer), and TS (Tomato, Soaking water). No data shown for TC (Tomato, Control) because the plant had stopped growing. Results are indicated in grams. Different letters represent statistically significant differences within each plant species (peas, tomatoes) (Tukey, p < 0.05).

Fig. 2.

Wet weight (fresh) and dry weight (dried) of roots harvested for all treatments: PC (Pea, Control), PF (Pea, Fertilizer), PS (Pea, Soaking water), TC (Tomato, Control), TF (Tomato, Fertilizer), and TS (Tomato, Soaking water). No data shown for TC (Tomato, Control) due to the plant halted growth prior to collection. Results are indicated in grams. Different letters represent statistically significant differences within each plant species (peas, tomatoes) (Tukey, p < 0.05).

The NPK-treated pea roots showed more and larger nodules, likely due to nutrient accumulation, whereas the PSW-treated counterpart (PS) showed a denser network of roots with fewer and smaller nodules, comparable to that of the control. The tomato control plants were not able to develop roots because of the lack of nutrients. Enrichment of soil with NPK and with PSW resulted in dense and comparable tomato root networks (Fig. 3).

Fig. 3.

Representative images of the roots from the plant treatments: TC (Tomato, Control), TF (Tomato, Fertilizer), TS (Tomato, Soaking water).

SPAD

The readings acquired by the SPAD meter revealed that addition of either NPK or PSW resulted in significantly higher chlorophyll/nitrogen concentration estimates than the controls (Fig. 4). In the case of peas, the SPAD readings increased by more than 50% in both fertilizer treatments, when compared with the control. Values increased from 27.1 to 43.6 and 40.5 for NPK and PSW, respectively (Fig. 4). In tomatoes, SPAD readings more than doubled on average in the fertilizer treatments, from 14.3 to 36.0 in the case of NPK, reaching 45.0 with PSW (Fig. 4).

Fig. 4.

Chlorophyl content of leaves measured with the Soil Plant Analysis Development SPAD meter for all treatments: PC (Pea, Control), PF (Pea, Fertilizer), PS (Pea, Soaking water), TC (Tomato, Control), TF (Tomato, Fertilizer), and TS (Tomato, Soaking water). Results are indicated in SPAD units. Different letters represent statistically significant differences within each plant species (peas, tomatoes) (Tukey, p < 0.05).

Microbial enumeration of the soil

Two-way ANOVA was used to evaluate the effects of plant species and fertilizer treatment on Lactobacillus abundance. Plant species had a clear influence on bacterial counts, with pea soils consistently showing higher values than tomato soils (Table 3). Fertilizer effects were more pronounced in tomatoes than in peas, indicating that treatment responses differed between species. The potting mix in which pea plants were grown was richer in Lactobacilli than the tomato-growing soil, approximately twice as much: 370 vs. 180 CFU/g (Table 3). The difference induced by the fertiliser treatment was marginally significant (p = 0.077).

Table 3.

Microbial enumeration of Lactobacillus in the soil after 49 days for the treatments PC (Pea, Control), PF (Pea, Fertilizer), PS (Pea, Soaking water), TC (Tomato, Control), TF (Tomato, Fertilizer), and TS (Tomato, Soaking water). Results are indicated in colony forming units (CFU/g). Different letters represent statistically significant differences within each plant species (Tukey, p < 0.05).

Microbial enumeration (CFU/g)	Peas	Tomatoes
Control	358 ± 76a	249 ± 45a
Fertilizer	348 ± 74a	131 ± 8b
Pea soaking water	408 ± 209a	174 ± 11ab

The second observation was a significant reduction by 48% of Lactobacilli counts in the NPK treatment when compared with the control for tomatoes. No statistically significant difference was observed between control and PSW for tomato soil. The microbial count of lactic acid bacteria in the PSW-treated soil was between the control and NPK amounts (Table 3).

Discussion

Growth stages

In agreement with previous literature this study showed no significant change in sprouting time or leaf appearance time in response to different fertilizer treatments in pea. These results agree with other findings, showing that NPK fertilizers reduced flowering time in pea plants from 53 to 49 days¹⁶. Comparable fruiting times agreed with previously published data on the effect of fertilizers on peas, both inorganic (NPK) and organic (biogas)^19,20.

The lack of growth in the control group of tomato plants highlights the higher nutrient demand of tomato plants when compared with the nitrogen-fixing peas and shows that nutrient availability and genetics can affect different plant growth stages¹.

It must be noted that the two fertilizers tested have different compositions. The PSW contained 33% N, 25% P and 60% K, with lower amounts of trace minerals, relatively to the NPK product (Table 1). Nitrogen is a component of chlorophyll, responsible for plant growth²¹. Phosphorous is particularly important in energy conversion and early rot development, whereas it does not affect the flowering time²². Potassium affects both plant development and yield²³.

Shoot and root weight

NPK was shown to boost plant growth, particularly during leaf appearance and fruiting^24,25. Differences were particularly large for tomato plants with a five-fold increase in root weight²⁶. The comparable effect of inorganic and organic fertilizers on shoot and root weights was demonstrated previously: NPK (solid form) and a Trichoderma-enriched biofertilizer (in liquid form) resulted in statistically comparable weights of shoots and roots in tomatoes²⁷. In our study, root data revealed that NPK yielded heavier roots, both in terms of fresh and dry weight, but only in the case of peas (Fig. 2). Nitrogen in the soil is directly correlated with plant yield²⁸; therefore, PSW would be expected to support bigger pea and tomato plant weights due to its nitrogen content (Table 1). Nonetheless, other minerals are equally important. It is possible that the difference in phosphorous content (3.9 vs. 0.9% in NPK and PSW, respectively) (Table 1) played a role here. Lower phosphorus availability can limit root growth, even when nitrogen is sufficient. The lower phosphorus concentration in the presence of nitrogen in PSW may therefore have supported shoot growth while restricting root biomass in peas. The lower the phosphorous content in the soil, the lower is the root weight in pea²⁹ and tomato³⁰.

These findings demonstrate that nutrient availability and nitrogen transformations significantly affect tomato shoot and root growth³¹, with both fertilizer treatments boosting above- and below-ground development.

SPAD

Chlorophyll content

In tomatoes, SPAD readings highlighted the benefits of nitrogen-containing fertilizers to supply nitrogen to non-leguminous crops³². Despite no significant difference observed among fertilizer- and PSW-treated tomato plants, it is noteworthy to point out that PSW lifted chlorophyll content more than NPK. Similarly, manure and biogas fertilizer with low concentrations of these key minerals (0.8–1.5% nitrogen, 0.7–0.9% phosphorous, and 0.89–1.2% potassium) were less effective than NPK at supporting chlorophyll synthesis in pea plants¹⁹. In the current experiment, the organic fertilizer, PSW, contained twice as much nitrogen as NPK, with nutritionally relevant concentrations of phosphorous and potassium (Table 1). Therefore, PSW can be used as a viable alternative to NPK to support growth and chlorophyll synthesis in pea and tomato plants.

Studies on peas highlighted the beneficial effect of inorganic fertilizers on chlorophyll concentrations^33,34. Other studies reveal the crucial role of nitrogen, phosphorous and potassium for this attribute. For example, an organic manure containing 4% nitrogen, 4% phosphorous and 4% potassium failed to enhance chlorophyll content in peas³³. Similarly, manure and biogas fertilizer with low concentrations of these key minerals (0.8–1.5% nitrogen, 0.7–0.9% phosphorous, and 0.89–1.2% potassium) were less effective than NPK at supporting chlorophyll synthesis in pea plants¹⁹.

Nutrient release

Another important factor in evaluating fertilizers is release time. The synthetic product used in this trial had a 3-month release time (data from manufacturer). However, data on mineral release from the PSW were not available. The nitrogen found in PSW was attributed mostly to protein (albumin, globulin, lipoxygenase), peptides (7s globulin subunit) and free amino acids^21,35. Rapid release of plant-available nitrogen from proteins applied to soils is well documented²².

Microbial enumeration of the soil

Release of nitrogen from organic compounds depends on soil conditions. Two intrinsic factors in PSW may affect such release: emulsifying activity and prebiotic properties. Firstly, PSW expresses excellent emulsifying activity in oil-water emulsions, reflecting a capacity to homogeneously hold water-soluble and fat-soluble molecules. Such characteristics might allow water to be held in the soil for longer periods of time, thus allowing superior hydration to the roots. Such effects could be beneficial in dry soil, but potentially deleterious in wet soils. Secondly, prebiotic properties indicate faster growth of Lactobacilli, which contribute to the digestion of organic fertilizers. In the unintended case of fertilizer leaching into waterways, such as rivers and lakes, eutrophication by PSW would likely be far less pronounced compared with the NPK due to: (i) no free nitrate available; (ii) prebiotic properties; and (iii) antimicrobial activity.

Peas are legumes, which are known to promote the growth of beneficial microorganisms in the soil²³. Legumes such as pea can enrich the rhizosphere through nitrogen fixation and root exudates, which often support higher microbial activity. In contrast, tomato relies entirely on external nitrogen sources and may deplete nutrients more rapidly in pot systems, which can contribute to lower bacterial counts. Therefore, the consistently high amount of lactic acid bacteria found in the three pea soils (control, fertilizer, and PSW) was expected. PSW-grown Lactobacilli numbers were not different from the control, nor from the NPK treatment. When performing one-way ANOVA, significant differences emerged within the tomato study, with higher Lactobacillus count in sample-treated and control soil, compared with the fertilizer-treated soil (p-value 0.048). These results warrant further research in the impact of PSW on soil microbes. Tomato plants require high nutrient concentrations, which are depleted from the soil³⁶. Thus, tomato growing can result in altered bacterial consortium in the soil, with loss of beneficial microorganisms such as Lactobacilli. Recent studies have shown that replacement of NPK with organic waste (cattle manure, spent mushroom liquid) resulted in larger bacterial consortium^37,38. Specifically, use of a liquid organic fertilizer based on spent mushrooms resulted in a more diverse bacterial community³⁸.

The second observation was a significant reduction by 48% of Lactobacilli counts in the NPK treatment when compared with the control for tomatoes. Tomato is highly demanding in terms of nutrients, depleting the soil of its minerals. Consequently, microbial populations diminish upon tomato plant growth, with loss of lactic acid bacteria in favour of other bacteria (Nocardioides spp. and Bradyrhizobium) and fungi (Pseudogymnoascus, Acremonium, Oidiodendron, Phialemonium, Penicillium, Phialosimplex)³⁹. This is in line with other findings demonstrating negative impacts of chemical fertilizer application on soil microbial communities⁴⁰. Mineral fertilizers supply readily available nutrients but little organic carbon, which can limit some microbial groups. In contrast, PSW provides soluble carbohydrates and peptides that may help maintain Lactobacillus populations. No statistically significant difference was observed among control and PSW for tomato soil. These results suggest maintenance of microbial soil health under PSW application. The PSW contained oligosaccharides such as raffinose and stachyose, which selectively promote probiotic growth over pathogen growth. It must be noted that the observed prebiotic activity was moderate, lower than the soaking water of other legumes, such as haricot beans and yellow soybeans⁴¹. It has been hypothesized that the peptide defensin could partially hinder microbial growth. Defensin is a peptide found mostly in peas and lentils, which acts as antimicrobial over a wide range of microorganisms. In addition, 3 mg/g saponins were quantified in PSW¹⁰. Saponins are known to limit the growth of a variety of microorganisms. This result is even more interesting, since it may potentially control microbial growth. This hypothesis, if confirmed, would define PSW as a low-impact fertilizer. It is noteworthy that the microbial count of lactic acid bacteria in the PSW-treated soil was between the control and NPK concentrations (Table 3). Prebiotic activity of PSW was established to be moderate⁴¹. Consequently, other processing water from legumes such as haricot beans and yellow soybeans might offer more pronounced benefits in terms of Lactobacilli growth. Prebiotic activity of PSW was established to be moderate⁴¹. Consequently, other processing water from legumes such as haricot beans and yellow soybeans might offer more pronounced benefits in terms of Lactobacilli growth.

Practical implications

PSW in the liquid state must be stored frozen to prevent microbial spoilage. A study on spray-drying of PSW yielded highly functional powder but with some loss during drying due to stickiness and low solid content of the starting material³⁵, possibly requiring pre-concentration or other treatments. This limitation adds costs and technological requirements such as freezing storage or drying. In addition, pasteurization should be implemented in order to eliminate microbial contamination.

This study was limited to vegetative plant growth. Future work should also consider the reproductive and fruiting stages for the interpretation of fertiliser equivalence. The objective was to determine whether plant can grow fully in absence of synthetic fertilizer, with the sole use of PSW. In addition to photographical depiction of root architecture, image analysis could be used in future studies to quantify root measurements such as root length density and branching intensity under the different treatments.

Methods

Material preparation

Split yellow peas (Pams, New Zealand) were soaked for 16 h at room temperature in a 1:3.3 ratio of seed to water (w/v). PSW was drained from the peas, collected, frozen, and thawed 1 day prior to use. The liquid was not sterilized or filtered. The freezing step was necessary to store the soaking water throughout the length of the study (49 days). Pea seeds, variety ‘Dwarf Massey’ (Oderings, New Zealand) and tomato seeds, variety ‘Grosse Lisse’ (Mr Fothergill’s, New Zealand) were planted in individual pots filled with dry soil, with holes at the bottom and saucers. Seeds were planted following the suppliers’ instructions.

Experimental design and growing conditions

Plants were arranged in a randomized design with three fertilizer treatments and six replicates. Control treatments for peas (PC) and for tomatoes (TC) consisted of seeded pots filled with bark-pumice that did not receive fertilizer. Fertilizer treatment pots for peas (PF) and tomatoes (TF) were additionally filled with Osmocote Exact (16-3.9-10 TE, 3-month release) (Daltons, New Zealand). This NPK was chosen as a representative of commonly used fertilizers for potted plants. It contained 16% N, 3.9% P, 10% K and trace elements such as magnesium, iron, manganese, boron, copper, molybdenum, and zinc (Table 1). In the third treatment, peas (PS) and tomatoes (TS) were treated with PSW. The PSW contained 1.89 g of solids per 100 mL. One-third of the solution (32%) consisted of soluble protein, followed by 36.5% of soluble carbohydrates, 18% of insoluble carbohydrates and 14% of minerals^10,38. The mineral profile of PSW consisted of 5.9% N, 0.9% P and 6.4% K³⁸ (Table 1). In comparison, PSW contained twice as much nitrogen as the NPK fertilizer (yet in the bound form of amino acids and protein), less phosphorous, potassium and trace minerals, except for zinc. Irrigation of all treatments was performed with 150 mL of water per pot on day 1, followed by 50 mL daily for the rest of the experiment (49 days total). For PS and TS, tap water was replaced with equal volumes of PSW. The experiment took place in a greenhouse at Lincoln University (Lincoln, New Zealand) from 20 July 2020 until 4 September 2020 with natural light and an average temperature of 18 °C.

Growth stages

Key growth stages of plants were observed and recorded as the number of days needed for sprouting, leaf development, flowering and fruit appearance.

Shoot and root weight

Whole plants were harvested after 49 days, separating roots (below ground) from shoots (above ground). Harvesting took place manually, separating plants from their soil and shoots from their roots. Plant material was washed under running water to remove soil. Subsequently, washed plant material was dried in an oven at 80 °C for 24 h before weighing. Representative pictures of washed roots were taken before drying.

SPAD (soil plant analysis development)

A SPAD meter (SPAD-502Plus, Konica Minolta, Japan) was used to estimate leaf chlorophyll and nitrogen status on the leaves at harvest. Measurements were conducted in the center of fully unfolded leaves, testing three leaves per plant.

Microbial load of the soil

Soil samples were tested for microbial load. Specifically, Lactobacilli were the focus of this experiment since they represent beneficial microorganisms for soil health. Other microbial groups were not assessed for the scope of this study. Lactobacillus was selected as an indicator group because of its well-documented role in plant growth promotion and organic matter transformation 439]. Petri dishes were used as the matrix. The growing media chosen was MRS Agar (de Man, Rogosa, Sharpe). Samples were incubated at 35 °C for 2 days before microbial enumeration.

Statistical analysis

Treatment effects were analyzed using one-way ANOVA in Minitab (Minitab, 2021). As for the microbial enumeration, 2-way ANOVA was performed to test for two variables: treatment (control, fertilizer, PSW) and species (peas, tomatoes). Before ANOVA, all data were tested for the ANOVA assumption of homogeneity of variances to examine the potential need for data transformation. Statistical analysis was run with a Tukey post-hoc test at p < 0.05.

Conclusions

The findings presented here are the first comparison of the effects of NPK and PSW on edible crop plants. Results in a legume and in a non-leguminous species showed that PSW may be a viable and more sustainable alternative for NPK to promote crop growth. This was reflected in morphological and physiological attributes and beneficial effects on soil microbial populations. The study contributes towards the development of circular sustainable economies by proposing an innovative, more sustainable strategy to optimize plant growth. Further research is needed to validate these results across different crop yields, and across different crops and soil types and environmental conditions. In addition, plant growth should be allowed to reach full maturity to test the effects on fruit yield and quality. Application of PSW and other legume wastewater to agriculture could significantly reduce the use of synthetic fertilizers, whilst preserving healthy soil microbiome and preventing eutrophication. Soluble proteins, sugars, minerals, phenolics and saponins allow superior nutrient solubility in the soil and promote the growth of microorganisms useful to plants and the ecosystem. Plant growth and estimated chlorophyll content was comparable to that of NPK, whilst soil Lactobacilli were preserved. Techno-economic analysis is warranted to establish feasibility on large scale. Specifically, in order to have a stable product, PSW must be pasteurised, then either stored frozen or dried, thus increasing processing costs. PSW as organic fertilizer can be a novel opportunity to grow high quantity and quality of produce whilst preserving the environment.

Author contributions

L.S. planned the experimental design, wrote part of the manuscript, and performed the final editing. C.H. performed the experiments and wrote part of the manuscript. R.H. planned the experimental design, wrote part of the manuscript and performed the final editing.

Data availability

The authors declare that the data supporting the findings of this study are available within the paper and its Supplementary Information files. Should any raw data files be needed in another format they are available from the corresponding author upon reasonable request.

Declarations

Competing interests

The authors declare no competing interests.