Vermifiltration and sustainable agriculture: unveiling the soil health-boosting potential of liquid waste vermicompost

Abstract

Vermifiltration is a promising technique that can help recover nutrients from wastewater for further use in agriculture. We conducted a field experiment to assess the effectiveness of vermicompost produced from the vermifiltration of liquid waste (manure and food production waste) and how it can affect the soil health and yield of a squash crop. We tested the effect of three rates of vermicompost (low, medium, and high) applied over two consecutive years and measured physical, chemical, and biological soil health indicators, squash yield, and nutritional status. The results showed that the use of vermicompost, especially at a high rate, increased total soil carbon, total nitrogen, potentially mineralizable nitrogen, and particulate organic matter, as well as the activity of C-N-P cycling enzymes, as compared to a control with only inorganic fertilization. The yield of the squash crop remained stable, while the crop nutritional value improved as the levels of boron and copper in the treated squash increased. These findings indicate an improvement in soil health after the use of vermicompost. Overall, results strongly support using this type of vermicompost as a sustainable management approach to recycle nutrients and enhance soil health.

Full text

Frontiers in Sustainable Food Systems 01 frontiersin.org
Vermifiltration and sustainable
agriculture: unveiling the soil
health-boosting potential of
liquid waste vermicompost
HalimaMalal
1,2*, VeronicaSuarezRomero
2,
WilliamR.Horwath
2, SabinaDore
3, PatrickBeckett
3,
MohamedAitHamza
4, HichamLakhtar
1 and CristinaLazcano
2
1 Microbial Biotechnology and Plant Protection Laboratory, Faculty of Science, Ibn Zohr University,
Agadir, Morocco, 2 Department of Land, Air and Water Resources, University of California, Davis, Davis,
CA, United States, 3 Biofiltro, Inc., Davis, CA, United States, 4 Laboratory of Biotechnology and
Valorization of Natural Resources, Faculty of Science, Ibn Zohr University, Agadir, Morocco
Vermifiltration is a promising technique that can help recover nutrients from
wastewater for further use in agriculture. Weconducted a field experiment to
assess the eectiveness of vermicompost produced from the vermifiltration
of liquid waste (manure and food production waste) and how it can aect the
soil health and yield of a squash crop. Wetested the eect of three rates of
vermicompost (low, medium, and high) applied over two consecutive years
and measured physical, chemical, and biological soil health indicators, squash
yield, and nutritional status. The results showed that the use of vermicompost,
especially at a high rate, increased total soil carbon, total nitrogen, potentially
mineralizable nitrogen, and particulate organic matter, as well as the activity
of C-N-P cycling enzymes, as compared to a control with only inorganic
fertilization. The yield of the squash crop remained stable, while the crop
nutritional value improved as the levels of boron and copper in the treated
squash increased. These findings indicate an improvement in soil health after
the use of vermicompost. Overall, results strongly support using this type of
vermicompost as a sustainable management approach to recycle nutrients and
enhance soil health.
KEYWORDS
agricultural wastewater, vermifiltration, vermicompost, soil organic matter, microbial
activity, micro and macro nutrients
1 Introduction
Agricultural and agro-industrial activities have grown considerably over the last few
decades, leading to substantial environmental pollution (Martinez-Burgos etal., 2021; De
Rooij et al., 2024). ese activities generate large amounts of wastewater with high
concentrations of organic matter and nutrients, especially nitrogen compounds (Simas etal.,
2019). Inappropriate disposal of this liquid waste can cause soil acidication, nitrate leaching
to groundwater, eutrophication of surface waters, emissions of greenhouse gasses, and
unpleasant odors (Lazcano etal., 2016; Köninger et al., 2021; Zhang etal., 2022; Rocha
etal., 2024).
OPEN ACCESS
EDITED BY
Çağrı Akyol,
Ghent University, Belgium
REVIEWED BY
Ana Robles Aguilar,
University of Vic- Central University of
Catalonia, Spain
Hongzhen Luo,
Ghent University, Belgium
*CORRESPONDENCE
Halima Malal
hmalal@ucdavis.edu
RECEIVED 07 February 2024
ACCEPTED 09 April 2024
PUBLISHED 24 April 2024
CITATION
Malal H, Romero VS, Horwath WR, Dore S,
Beckett P, Ait Hamza M, Lakhtar H and
Lazcano C (2024) Vermifiltration and
sustainable agriculture: unveiling the soil
health-boosting potential of liquid waste
vermicompost.
Front. Sustain. Food Syst. 8:1383715.
doi: 10.3389/fsufs.2024.1383715
COPYRIGHT
© 2024 Malal, Romero, Horwath, Dore,
Beckett, Ait Hamza, Lakhtar and Lazcano. This
is an open-access article distributed under
the terms of the Creative Commons
Attribution License (CC BY). The use,
distribution or reproduction in other forums is
permitted, provided the original author(s) and
the copyright owner(s) are credited and that
the original publication in this journal is cited,
in accordance with accepted academic
practice. No use, distribution or reproduction
is permitted which does not comply with
these terms.
TYPE Original Research
PUBLISHED 24 April 2024
DOI 10.3389/fsufs.2024.1383715

Malal et al. 10.3389/fsufs.2024.1383715
Frontiers in Sustainable Food Systems 02 frontiersin.org
By treating these wastes through vermiltration process, farmers
can reduce their demand for high-quality water (Sarkar etal., 2006;
Lahlou etal., 2021), recover nutrients from wastewater that can beused
for crop production (Kenneth etal., 2023), and reduce greenhouse gas
emissions (Dore etal., 2022). Indeed, vermiltration is an eco-friendly
and cost-eective biological aerobic process that can be used for
secondary wastewater treatment using epigenic earthworms (Pasha
etal., 2018; Singh etal., 2019). Earthworms ingest and burrow through
the lter media, producing mucus and casting that alter the properties
of the biolm. is, in turn, increases the microbial population and
activity, resulting in a signicant reduction in biological oxygen
demand (up to 90%), chemical oxygen demand (between 80 and 90%),
and total dissolved solids (up to 90%) (Sinha etal., 2008).
Using vermicompost from vermiltration treatment in agriculture
could increase the attractiveness and ecacy of vermiltration as a
recycling strategy, reducing the environmental impact of liquid
organic wastes in the livestock farming industry and agroindustries.
is approach aligns with the principles of a circular economy,
developing eective wastewater recycling processes and sustainable
soil management strategies.
Vermicompost produced aer vermiltration is rich in plant-
available nutrients (Xing et al., 2005; Kumar et al., 2015; Dey
Chowdhury etal., 2022), indicating its potential use as an organic
amendment to improve soil health and crop production. Additionally,
vermicomposting and vermiltration represent two distinct methods
of earthworm-mediated waste management, each with its own unique
processes and applications (Enebe and Erasmus, 2023; Saapi etal.,
2024). ese distinctions may introduce specic physico-chemical
and microbial characteristics, ultimately impacting the nal
composition and functionality of the vermicompost produced (A’ali
etal., 2017; Rynk etal., 2022; Stehouwer etal., 2022; Luo etal., 2023).
erefore, it is crucial to investigate whether these distinctions lead to
dierential impacts on soil health, nutrient availability, and
plant growth.
e current body of literature primarily focuses on applying
vermicompost derived from treating solid organic waste materials in
agricultural practices. is type of vermicompost has previously been
observed to enhance plant growth (Lazcano etal., 2009; Song etal.,
2022), increase soil microbial biomass and diversity (Danish Toor
etal., 2024), act as a source of macro-and micro-nutrients (Nurhidayati
etal., 2016; Raza etal., 2021), and support soil carbon sequestration
(Ngo etal., 2012, 2014; Naikwade, 2019). However, there is a need for
more information about the use of vermicompost produced from
liquid waste treatment in agriculture.
Furthermore, soils in arid and semi-arid regions are characterized
by poor soil health, low nutrient content, high soil pH, low microbial
biomass and activity, and limited vegetation cover. e intense use of
chemical fertilizers, monoculture farming, and overall expansion of
land use worsens the state of soil health in those regions (Ayangbenro
and Babalola, 2021), aecting negatively agricultural production and
favoring land degradation process (Fernández-Ugalde etal., 2011).
Hence, using nutrient-rich organic amendments such as
vermicompost can bea sustainable soil management practice that can
help improve soil health and restore degraded lands. To our
knowledge, there is a lack of studies on the impact of vermicompost
resulting from vermiltration on soil health. Specically, carbon
pools, enzymatic activities, macro and micronutrients on soil
and crops.
In this context, this study aimed to investigate the impact of
vermicompost obtained from the vermiltration of wastewater on
physicochemical soil properties, nutrient cycling, enzymatic activities,
and carbon sequestration potential, as well as on the yield and nutrient
status of summer squash, a widely cultivated crop in nearly all regions
of California giving its adaptability to arid and semi-arid climate.
Wehypothesized that applying the vermiltration byproduct to the
soil would stimulate microbial activity and nutrient cycling, providing
valuable soil nutrients and enriching soil organic matter. Furthermore,
we expected that its application could increase particulate and
mineral-associated organic matter, ultimately contributing to carbon
sequestration and improved crop yield and quality.
2 Materials and methods
2.1 Study site
We conducted a eld experiment on a commercial farm in
Winters, California, UnitedStates, in 2021 and 2022. e area has a
semi-arid climate with an average annual precipitation of 557 mm, an
average maximum temperature of 24.6°C, and an average minimum
temperature of 9.3°C (Western Regional Climate Center, n.d.). e
soil in the eld is classied as Rincon silty clay loam, and the baseline
top15 cm soil has a pH of 6.47 ± 0.05, total %N of 0.15 ± 0.01, total %C
of 1.28 ± 0.07, and EC of 759.42 ± 19.82 μS cm−1. e eld is subjected
to a crop-rotating system, with tomatoes planted in 2021 and summer
squash (Cucurbita pepo), the ‘Ananashyi’ variety planted in 2022.
2.2 Characterization of the organic
amendment
We employed vermicompost generated as a by-product of the
vermiltration system. e vermiltration system is operated by
Bioltro Inc. e system consisted of a rectangular concrete enclosure
(49 m × 11 m × 1.5 m) lled with 1 meter of ltering material
(woodchips) and earthworms (Eisenia andrei) in the top30 cm. It has
a 30 cm deep drainage area at the bottom vented to the outside by PVC
exhaust pipes (15 cm in diameter) for passive air exchange (Dore etal.,
2022). e sprinkler system applied inuent for 2 min to the
vermilter surface every 30 min. e applied inuent percolated
through the vermilter to the underlying drainage space and drained
under gravity in about 4h. e vermilter material, comprising
mature vermicompost and the residual ltering material, is typically
extracted from the system every 18 months. It was stored in piles and
used for soil application.
Due to the availability of the organic waste feedstock and output
rates of the vermiltration system, it was not possible to harvest
enough of the same material for two consecutive years. erefore,
we used vermicompost produced from two dierent feedstock
materials in each year. In the initial year of the study (2021), pepper
plant processing wastewater served as the feedstock, while dairy
manure wastewater was utilized in the subsequent year (2022). In both
cases, woodchips were utilized as the ltering material. Characteristics
of the vermicompost used in the study are outlined in (Table1). To
uphold the experiment’s consistency, application rates to the soil were

Malal et al. 10.3389/fsufs.2024.1383715
Frontiers in Sustainable Food Systems 03 frontiersin.org
determined based on the available nitrogen content of the
vermicompost, maintaining uniformity across both study years.
2.3 Experimental design and management
We conducted a eld experiment to test the impact of
vermicompost, used as an organic amendment, on both soil health
and crop yield. e eld experiment consisted of four treatments
arranged in a randomized complete block design with three
replications (Figure 1). e treatments were (1) control with no
vermicompost applied, (2) low rate of vermicompost application (LV,
18 kg ha−1N), (3) medium rate of vermicompost application (MV, 28.8
Kg ha
−1
N), and (4) high rate of vermicompost application (HV,
54 kg ha
−1
N). e application rate was based on the available nitrogen
content of the vermicompost applied and summarized in Table2.
e application of vermicompost was performed manually aer
the beds were established and prior to spring planting. We evenly
raked the soil to mix in the material within each bed while the areas
between the beds remained free of any amendment. Each treatment
was applied to three 8 m × 8 m plots containing eight beds, resulting
in 12 experimental plots distributed in the eld. Buer zones of six
beds were set between plots to avoid disturbances. All treatments were
provided with standard mineral fertilizer to supplement the plants
with essential nutrients during their growth. is mineral fertilization
was applied through subsurface drip irrigation (20 cm), consisting of
100 kg ha−1 of 32% N of urea and ammonium nitrate.
2.4 Laboratory analyses
2.4.1 Determination of soil health indicators
is study aimed to investigate the cumulative eect of
vermicompost application on soil health. erefore, wecollected soil
samples at the end of the second year. In September 2022, aer the
summer squash harvest, we collected three soil cores from each
experimental plot at 0–15 cm depth. e samples were homogenized
and composited into one sample per plot and then moist sieved to
8 mm to remove plant matter and organic debris. e soil samples
were then stored at 4°C until further use.
Nitrate (NO
3
−
-N) and ammonium (NH
4+
-N) concentrations in
the soil samples were determined colorimetrically in a soil extract
prepared with 8 g of fresh soil using 0.5 M of K
2
SO
4
(Miranda etal.,
2001; Doane and Horwath, 2003). Another 8 g subsample was taken
to measure potentially mineralizable nitrogen (PMN) in the soil
samples. Briey, 10 mL of water was added to the soil, and then the
solution was purged with N2 gas and incubated for 7days at 37°C. A
soil extract was subsequently prepared using 0.67 M of K
2
SO
4
, and
NH
4+
-N was determined colorimetrically in the soil extracts. e
dierence between NH
4+
-N in the incubated and non-incubated
samples was the PMN.
To measure the microbial biomass carbon (MBC), a 6 g subsample
was exposed to chloroform for 24 h and then extracted using a
solution of 30 mL of 0.5K
2
SO
4
. Another 6 g subsample was used to
prepare a non-fumigated extract. Dissolved organic carbon
concentrations were determined by UV-persulfate oxidation
(Teledyne-Tekmar Fusion), and the MBC was calculated as the
dierence between the fumigated and non-fumigated samples
(Horwath and Paul, 1994). e soil pH and electrical conductivity
(EC) were determined in soil slurries [1:2 soil to Deionized water (DI)
water] using a pH/EC meter (Mettler Toledo, Columbus, OH,
UnitedStates).
A high-throughput microplate assay method was used to
measure the potential activity of carbon cycle related enzymes
(𝛂-Glucosidase, β-Glucosidase, Cellulase, Xylosidase) and nitrogen
cycle-related enzymes (Leucine aminopeptidase, N-Acetyl-
glucosaminidase) and phosphate cycle enzyme (Phosphatase) as
described in Bell etal. (2013). Briey, 2.75 g of moist soil was mixed
with 91 milliliters of a 50-millimolar solution of Sodium Acetate
buer to prepare the soil slurry. e 4-methylumbelliferone (MUB)
standard was used for all enzymes except the LAP enzyme, where
weused the 7-Amino-4-methylcoumarin (MUC) standard. Two
deep96-well plates were utilized for the two standards, while one
plate was dedicated to the enzyme substrates. Each corresponding
well in the plates was pipetted with 200 μL of the standards or the
substrates, followed by adding 800 μL of soil slurry to all wells in the
three plates. Next, the three plates were placed in a dark incubator
for 3h. Aerward, they were centrifuged for 3min at approximately
2,900 x g. en, 250 μL was carefully transferred from each well of
the incubated deep well plates to the corresponding wells in clear
96-well plates to beread using a microplate reader (BioTek synergy
HTX multi-mode reader) (Excitation Wavelength = 365 nm,
Emission Wavelength = 450 nm).
Soil respiration was determined aer 24 h of soil incubation in
227.3 mL glass jars. Aer determining soil water holding capacity, DI
water was added to 10 g of 2 mm sieved air-dried soil to reach 60% of
water holding capacity. e jars were airtight and incubated in the
dark for 24 h. Another set of jars was used to set up standards with
known concentrations of CO
2
. e soil respiration was estimated
through the CO
2
evolved from the samples measured with a
continuous ow LICOR 850 IRGA CO
2
/H
2
O analyzer (LI-COR
Environmental, Lincoln Nebraska, UnitedStates). To determine the
concentration of active C in soil, wemeasured Pyruvate oxidase/
carboxylase (POXC) using the colorimetric protocol described in Weil
etal. (2003). Briey, wemixed 2.5 g of air-dried soil samples with
0.2 M potassium permanganate solution and measured the resulting
solution’s absorbance at 550 nm using a microplate reader (BioTek
synergy HTX multi-mode reader).
TABLE1 Physicochemical characteristics of the vermicompost extracted
from the vermifiltration system analyzed before application to the field.
Vermicompost
(pepper plant)
vermicompost
(dairy manure)
N-NO3
− (mg g−1 dw) 0.378 0.237
N-NH4+ (mg g−1 dw) 0.170 0.406
Inorganic N (mg g−1 dw) 0.548 0.643
Total N (%) dw 2.02 1.31
Organic N (mg g−1 dw) 19.65 12.50
C (%) 42 38.63
C/N 20.79 29.39
BD1 Wet (g cm−3)0.48 0.51
BD1 Dry (g cm−3 dw) 0.17 0.15
1BD, bulk density.

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Total soil C (%) and N (%) were determined by dry combustion
in an elemental analyzer (Costech analytical technologies Inc. model
ECS 4010).
To measure particulate organic matter (POM) and mineral-
associated organic matter (MAOM), 10 g subsamples were taken from
each plot and mixed with 30 mL of 5 g/L sodium hexametaphosphate.
e mixture was shaken for 15 h on a reciprocal shaker and then
passed through a 53-μm sieve. e material that remained on the sieve
aer rinsing with water several times was identied as POM, whereas
the soil slurry that passed through the sieve was identied as
FIGURE1
Aerial view of the experimental design, LV stands for low rate of vermicompost, MV stands for vermicompost at a medium rate, and HV stands for
vermicompost at a high rate.
TABLE2 The rate of vermicompost applied to the soil each year.
Target available N (kg ha−1)vermicompost applied (dry ton ha−1)
2021 Low rate (LV) 18 2,8
Medium rate (MV) 28.8 4.48
High rate (HV) 54.0 8.39
2022 Low rate (LV) 18 4.22
Medium rate (MV) 28.8 6.56
High rate (HV) 54.0 12.20

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MAOM. Both parts were then dried overnight at 50°C and ground
with a mortar and pestle. Aerward, the samples were analyzed for
total organic C and total Kjeldahl N (Cambardella and Elliott, 1992).
Soil available phosphorus was extracted by adding 50.0 mL of
0.5 M NaHCO3 (pH = 8.5) to 2.50 g soil. e mixture was shaken for
30 min and ltered. 40 μL aliquot of soil NaHCO
3
extract was mixed
with 20 μL MA reagent in each well of the 96-well microplate and
shaken for 1 min, then 140 μL aliquot of deionized water was added.
Absorbance was read at 700 nm using a microplate reader, and soil
Olsen-P concentrations were calculated based on the standard curve
(Song etal., 2019).
Soil samples were sent to the UC DAVIS Analytical Lab to
measure the potential availability of soil micronutrients; Zn, Mn, Cu,
and Fe using the diethylenetriaminepentaacetic acid (DTPA)
extraction method (Lindsay and Norvell, 1978).
2.4.2 Crop performance indicators
At harvest, the yield of the squash crop (kg ha
−1
) was measured by
randomly harvesting 6 healthy plants from each plot. All the squash
resulting from the harvest of six plants was weighed in the eld and
then taken to the lab. Squashes from each batch were cut horizontally,
and one-half of each squash was mixed into a puree and frozen; the
samples were then freeze-dried and ground to a powder. Samples were
sent to the UC Davis analytical lab to analyze macro and
micronutrients. e concentration of B, Ca, Cu, Fe, Mg, Mn, P, K, Na,
S, and Zn were analyzed in a 5 g sample using the nitric acid/hydrogen
peroxide microwave digestion and determination by Inductively
Coupled Plasma Atomic Emission Spectrometry (ICP-AES) (Meyer
and Keliher, 1992; Sah and Miller, 1992). e total nitrogen in the
squash was measured using the combustion method (AOAC Ocial
Method 972.43, 2006; AOAC Ocial Method 990.03, 2005).
2.5 Data analysis
All statistical analyses were done using R studio statistical soware
(v.2023.03.0+386). Weconducted a one-way ANOVA to determine if
there was a signicant treatment eect on the soil and plant variables
measured in this experiment. To ensure the accuracy of our analysis,
wetransformed any variables that did not follow a normal distribution
using box-cox transformations. In the case of signicant treatment
eects, wefollowed up with a Tukey Post Hoc test to evaluate the
treatment eects further. Wereported our data as mean values or
treatment dierences, followed by the standard error. Weused a
p-value of less than 0.05 to determine statistical signicance.
3 Results
3.1 Eects of the vermicompost treatments
on soil health indicators
According to Table3, the soil pH ranged between 7.29 and 7.54
and generally increased with vermicompost application, but the
change was not signicant (p > 0.05). e soil EC ranged between 259
and 345 μs cm−1, with a tendency to decrease when vermicompost was
applied. However, the variation was not signicant (p > 0.05). As for
the soil bulk density, its values ranged between 1.04 and 1.14 g cm3 and
did not change signicantly aer applying the vermicompost
treatments compared to the control (p > 0.05). Applying vermicompost
with dierent rates did not aect the total inorganic nitrogen in the
soil (p > 0.05). However, the amount of PMN increased signicantly
in the HV treatment compared to the control and LV (p < 0.05).
e total carbon content in the soil at harvest was 37.14% higher
in the HV treatment than in the control (Figure2A). e total carbon
was 21.42 and 15.00% higher in the MV and LV treatments,
respectively, compared to the control, but these dierences were not
signicant (Figure 2A). e total nitrogen in the HV treatment
increased signicantly by 24.66% compared to the control. e total
nitrogen also increased in the two other treatments (MV, LV)
compared to the control, but the dierence was not signicant
(Figure2B).
e vermicompost application had a signicant eect on
particulate organic carbon (POC) (p < 0.05). e low vermicompost
treatment showed no signicant change in POC compared to the
control. In contrast, the MV and the HV treatment increased the POC
by 40.82 and 93.02%, respectively, compared to the control, with the
increase in HV treatment being signicant (Figure2C). e particulate
organic nitrogen (PON) exhibited the same trend as the POC;
According to ANOVA, the use of vermicompost had a signicant
eect on PON (p < 0.05). e LV treatment maintained the same level
of PON as the control, whereas the MV and HV treatments increased
the PON by 31.67 and 93.12% compared to the control. However, the
Tukey test revealed that the dierence was signicant only when
comparing the HV treatment to the control (Figure 2D).
Vermicompost application did not have any signicant eect (p > 0.05)
on the mineral-associated organic carbon (MAOC) and nitrogen
(MAON) (p > 0.05) (Figures2E,F).
According to Table 4, the available P on soil samples varied
between 0.57 and 0.90 mg kg
−1
. Similarly, the concentration of Zn
varied between 2.40 and 2.93 mg kg
−1
across treatments, while the
amount of Cu on soil samples ranged between 6.09 and 7.03 mg kg−1.
TABLE3 Variation in soil physicochemical properties after vermicompost application.
Control LV MV HV P-value
pH 7.29 ± 0.16 7.42 ± 0.21 7.43 ± 0.26 7.54 ± 0.15 0.5
EC (μs cm-1) 345 ± 115 259 ± 62.5 280 ± 60.4 276 ± 83.3 0.6
Bulk density (g cm −3)1.07 ± 0.05 1.04 ± 0.07 1.14 ± 0.14 1.09 ± 0.08 0.6
TIN (kg ha−1)47.1 ± 29.2 38.1 ± 18 38.1 ± 18 27.3 ± 8.39 0.4
PMN (kg ha−1)18.1 ± 4.37 b 20.1 ± 7.68 b 27.8 ± 12.8 ab 51.6 ± 17.5 a 0.02
LV stands for vermicompost at a low rate, MV stands for vermicompost at a medium rate, and HV stands for vermicompost at a high rate. TIN stands for total inorganic nitrogen, PMN stands
for potentially mineralizable nitrogen. Dierent letters indicate a signicant dierence between treatments according to the Tukey test results.

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Additionally, the available Fe concentration fell between 10.6 and
11.7 mg kg−1.
Although the MV treatment tended to exhibit higher nutrient
content for all measured nutrients in the soil samples, these
dierences were not found to bestatistically signicant according
to ANOVA tests.
3.2 Microbial biomass and activity
Applying vermicompost with dierent rates did not aect POXC
concentration (Figure2G), the soil microbial biomass carbon, nor the
mineralizable carbon (p > 0.05) (Figures3A,B).
e ANOVA analysis indicated that the treatment type had a
signicant eect on the activity of β-glucosidase (Figure4B) and
cellulase (Figure 4C), where the HV treatment had the highest
enzymatic activity among the four treatments. e activity of
𝛂-Glucosidase and Xylosidase showed the same trend where the
enzymatic activity increased with an increasing application rate of the
vermicompost, with the HV treatment having the highest enzymatic
activity. However, the dierence was not signicant (p > 0.05 for both
enzymes) (Figures4A,D). e enzymatic activity related to the N cycle
tended to behigher on the HV treatment, especially for the N-Acetyl-
glucosaminidase enzymes (p > 0.05) (Figures 4E,F). e ANOVA
analysis revealed that the treatment type signicantly aected the
phosphatase activity (p < 0.05), with the HV treatment having the
FIGURE2
(A) Total carbon, (B) total nitrogen, (C) particulate organic carbon (POC), (D) particulate organic N (PON), (E) mineral associated organic C (MAOC),
(F) mineral associated organic N (MAN), (G) active carbon POXC, under dierent treatments. LV stands for low rate of vermicompost, MV stands for
vermicompost at a medium rate, and HV stands for vermicompost at a high rate. NS stands for non-significant when the p-value is higher than 0.05,
and S means a significant eect when the p < 0.05. Dierent letters indicate a significant dierence between treatments according to the Tukey test
results.

Malal et al. 10.3389/fsufs.2024.1383715
Frontiers in Sustainable Food Systems 07 frontiersin.org
highest enzymatic activity compared to the other treatments
(389 ± 30.2 nmol activity/g dry soil/h) (Figure4G).
3.3 Eects of the treatments on crop
performance
e squash yield ranged between 6.78 and 8.25 tons ha−1 and did
not change signicantly aer the application of the vermicompost
treatments as compared to the control without vermicompost
(p > 0.05) (Figure5).
Table 5 summarizes the changes in squash macro and
micronutrient content across treatments. e concentration of
macronutrients (N, K, Ca, Mg, P, and S) remained consistent across
dierent treatments. Similarly, the application of vermicompost did
not aect the concentration of micronutrients Zn, Fe, and Mn.
However, the treatments did have a signicant impact on the B
concentration (p = 0.05). Specically, the B concentration increased by
14.07, 31.98, and 6.6% in LV, MV, and HV treatments compared to the
control. e use of vermicompost also had a noticeable impact on the
concentration of Cu (p < 0.05). In the HV treatment, the concentration
of copper decreased by 9.59% compared to the control. In contrast, in
the LV and MV treatments, the concentration of copper increased by
36.52 and 47.90%, respectively, compared to the control.
4 Discussion
In contrast to existing research primarily centered on evaluating
the eciency of vermiltration, our study breaks new ground by
exploring the potential of its byproduct, vermicompost. Focusing on
the impact of vermicompost on soil health and crop performance,
wehave established compelling evidence supporting its use as an
ecient organic amendment. When applied at a rate equivalent to
54 kg N ha−1 it resulted in improved soil health compared to lower rates
and control groups. is was manifested by increased carbon and
nitrogen content, (mostly as POM), and enhanced enzymatic activity.
It also supported similar crop yields as the control treatment with only
inorganic N fertilizer. Our ndings demonstrate the validity of the
vermiltration process to recycle agro-industrial wastewater to close
the cycle of nutrients in agricultural systems while supporting soil
health and sustainable soil management in arid and semi-arid regions.
TABLE4 Variation in soil nutrient concentrations across treatments at harvest time.
Control LV MV HV P-value
P (mg kg−1)0.59 ± 0.23 0.57 ± 0.21 0.90 ± 0.22 0.75 ± 0.22 0.3
Zn (mg kg−1)2.63 ± 0.1 2.8 ± 0.34 2.93 ± 0.05 2.4 ± 0.7 0.5
Mn (mg kg−1)30.4 ± 0.63 31.5 ± 1.4 30.3 ± 1.16 26.4 ± 7.42 0.4
Cu (mg kg−1)6.93 ± 0.32 6.56 ± 0.32 7.03 ± 0.20 6.09 ± 1.15 0.3
Fe (mg kg−1)10.9 ± 1.63 11.7 ± 2.66 11.6 ± 1.28 10.6 ± 3.16 0.9
LV stands for vermicompost at low rate, MV stands for vermicompost at a medium rate, and HV stands for vermicompost at a high rate.
FIGURE3
(A) microbial biomass carbon, (B); respiration in dierent treatments at harvest time. LV stands for low rate of vermicompost, MV stands for
vermicompost at a medium rate, and HV stands for vermicompost at a high rate. NS stands for non-significant when the p-value is higher than 0.05,
and S means a significant eect when the p < 0.05.

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Frontiers in Sustainable Food Systems 08 frontiersin.org
4.1 Applying vermicompost improved soil
health
Healthy soils play a pivotal role in fostering both environmental
resilience and sustainable agriculture (Salomon and Cavagnaro, 2022).
Our ndings reveal notable improvements in key indicators such as
total carbon, total nitrogen, PMN, POC, PON, and enzymatic activity
following vermicompost application for two consecutive years.
Importantly, our approach of basing application rates on the available
nitrogen content of each vermicompost type ensured consistency in
nutrient inputs across both years of the study, regardless of the
feedstock material used.
Soil organic carbon (SOC) constitutes the largest terrestrial
carbon pool and serves as a vital indicator of soil health, and carbon
sequestration, (Just etal., 2023; Sun etal., 2023; Tang etal., 2023).
e increase in carbon content following the application of
vermicompost observed in our study is consistent with previous
research on organic amendments and suggests that vermicompost
improved soil health (Rahman etal., 2020; Wu etal., 2021; Xu etal.,
2022; Bai etal., 2023; Cooper and DeMarco, 2023; Zhang etal.,
2023). However not all soil C fractions contributes equality to soil
health. To further evaluate the impact of vermicompost use on
SOC, weassessed alterations in various SOC fractions, including
POC and MAOC.
FIGURE4
Potential enzymatic activities in soil after vermicompost application at dierent rates. (A) Alpha-Glucosidase, (B) Beta-Glucosidase, (C) Cellulase,
(D) Xylosidase, (E) N-Acetyl-Glucosaminidase, (F) Leucine-Amino-Peptidase, (G) phosphatase. LV stands for vermicompost applied at a low rate, MV
stands for vermicompost at a medium rate, and HV stands for vermicompost at a high rate. NS stands for non-significant when the p value is higher
than 0.05, and S means a significant eect when the p<0.05. dierent letters indicate a significant dierence between treatments according to the
Tukey test results.

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Particulate organic carbon is the labile fraction of SOC composed
of lightweight fragments of organic matter and characterized by a low
turnover time (Sun etal., 2023). e addition of vermicompost in this
study introduced labile organic matter into the soil, increasing
POC. is is consistent with other studies, such as Plaza etal. (2016)
and Giannetta etal. (2024), which also conrm the positive impact of
organic amendments on the POC fraction of soil organic matter.
Increasing POC can address soil health by improving soil structure,
water inltration, aeration, root growth, and cation exchange capacity
(Carter etal., 2003; Fronning etal., 2008). Furthermore, an increase
in POC is oen associated with a rise in enzymatic activity (Tang etal.,
2023). Our experiment observed that the activity of C, N, and P
cycling enzymes increased with vermicompost application, in
agreement with Li et al. (2021) and Yan et al. (2023). e soil
microbiome produces extracellular enzymes that break down the
labile fraction of soil organic matter, releasing carbon and nutrients
available to microbes and plants (Verrone etal., 2024).
Despite the increase in enzymatic activity following the
vermicompost, it was not associated with any changes in microbial
biomass carbon and respiration. e rise in soil enzyme activity can
FIGURE5
Squash yield variation after the application of three rates of vermicompost. LV, low; MV, medium; HV, high. NS stands for non-significant when the
p-value is higher than 0.05.
TABLE5 The concentration of squash macro and micronutrients across treatments.
Control LV MV HV P-value
N (%) 4.04 ± 0.57 3.64 ± 0.53 3.86 ± 0.51 3.96 ± 0.59 0.84
P (%) 0.49 ± 0.05 0.49 ± 0.02 0.47 ± 0.01 0.43 ± 0.02 0.19
K (%) 3.69 ± 0.62 3.62 ± 0.42 3.75 ± 0.19 3.41 ± 0.24 0.77
Ca (%) 0.33 ± 0.08 0.34 ± 0.03 0.35 ± 0.05 0.31 ± 0.03 0.83
Mg (%) 0.41 ± 0.06 0.38 ± 0.06 0.36 ± 0.04 0.39 ± 0.02 0.73
S (ppm) 2,517 ± 482 2,403 ± 431 2,323 ± 225 2,377 ± 238 0.93
B (ppm) 46.9 ± 4.62 b 53.5 ± 6.61 ab 61.9 ± 5.26 a 50 ± 5.90 ab 0.05
Zn (ppm) 59.2 ± 13.3 55.1 ± 9.75 52.5 ± 8.85 56.6 ± 5.24 0.86
Mn (ppm) 14.1 ± 3.70 13.3 ± 1.59 12.8 ± 1.36 12.4 ± 0.69 0.8
Fe (ppm) 122 ± 23.1 119 ± 21.8 113 ± 52.6 108 ± 17.3 0.96
Cu (ppm) 16.7 ± 1.67 ab 22.8 ± 6.54 a 24.7 ± 4.68 a 15.1 ± 0.56 b 0.008
LV stands for vermicompost at a low rate, MV stands for vermicompost at a medium rate, and HV stands for vermicompost at a high rate. Dierent letters indicate a signicant dierence
between treatments according to the Tukey test results.

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Frontiers in Sustainable Food Systems 10 frontiersin.org
bedue to the growth of microbes, modications in the microbial
community, or the stimulation of microbial activity through the
addition of organic matter (Yan etal., 2023). erefore, it is crucial to
analyze the eects of vermicompost on microbial community’s
dynamics, composition, and variation in further experiments to gain
further insights into the soil’s C and N dynamics.
e MAOC fraction of SOC primarily consists of small molecules
formed through microbial processing of C inputs. ese molecules are
subsequently protected through organo-mineral associations or
encapsulated within microaggregates. Consequently, MAOC exhibits
long turnover time, stability, and greater resistance to microbial
degradation (Tang etal., 2023). e MAOC fraction did not change
during the study despite the application of vermicompost, probably
because the MAOC unlike POC forms slowly over the years. is slow
accumulation process makes it dicult to detect signicant shis in
MAOC content in short-term studies. erefore, more time is needed
to detect any changes in this fraction resulting from stabilizing the
supplemented organic matter (Sokol etal., 2019), or the transformation
of POC to MAOC with time (Püspök etal., 2023).
Nitrogen is an essential macronutrient for plant growth, soil
health, climate change regulation and carbon sequestration. e
application of vermicompost increased soil total nitrogen, particularly
in the HV treatment, which is consistent with previous studies
demonstrating the positive impact of organic amendments on soil
nitrogen levels (Ryals etal., 2014; Wang etal., 2018). e increase in
PMN in the vermicompost treatments, combined with no signicant
dierence in inorganic nitrogen (nitrate and ammonium)
concentration between the control and the vermicompost-treated
groups, suggests that vermicompost addition has aected the organic
nitrogen pool without substantially impacting the readily available
mineral nitrogen pool. In fact, the use of vermicompost especially the
HV treatment also increased the PON fraction.
e organic nitrogen pool is an important indicator of soil health
(Lehmann etal., 2020), and considered as a sensitive indicator of
changes in agricultural management practices (Hossain etal., 2021).
Studies by Hossain etal. (2021) and Yao etal. (2021) indicated that the
application of organic amendments increases the PON fraction. is
fraction contributes to aggregate formation water holding capacity,
nutrient cycling (Hossain etal., 2021) and it may pave the way for
gradual nutrient release, which may not beimmediately apparent but
can benet soil health in the long run. Similar to the MAOC fraction,
the use of vermicompost did not increase the MOAN, needing more
time to depict changes in this fraction of soil organic nitrogen.
4.2 The vermicompost application did not
enhance the yield but it changed the
nutrient status of summer squash
e vermicompost used was rich in nutrients and organic matter,
and its application was expected to increase both the yield and
nutrient content of the squash. However, there were no dierences in
yield compared to the control. is may beattributed to many factors;
It is possible that the plants were already receiving all the required
nutrients through the fertigation system, or that more time was
needed to observe any changes in crop performance. According to Ma
etal. (2022), repeated application of organic amendment over the
years can improve soil conditions and lead to visible eects on crop
yield. Additionally, the nature of the irrigation system might explain
the limited impact of vermicompost on squash yield. Using a
subsurface drip irrigation system could restrict the benets of applying
vermicompost at the soil surface because the moisture is concentrated
in the root zone, and the nutrients contained in vermicompost may
not move eectively through the soil prole. Liquid fertilizers like
vermicompost tea can beutilized to overcome this limitation.
On the other hand, the use of vermicompost, especially at a
medium rate, enhanced the nutritional value of squash as it boosted
the concentration of B and Cu – two crucial plant micronutrients
(Khaliq etal., 2018; Lafuente etal., 2023). Copper, for example, is
crucial for brain development and immune system functioning
(Chiou and Hsu, 2019). However, excessive amounts of copper can
pose a risk to human health (Yang etal., 2002). In this study, the levels
of Cu in summer squash samples remained below the established
toxicity threshold of 40 ppm as recommended by Codex Alimentarius
Commission (1984) and FAO/WHO (1988). As for B, it is an essential
micronutrient for forming cell walls and membrane structure and
functioning (Läuchli and Grattan, 2014). It is also an essential
micronutrient in many physiological activities, including bone growth
and nervous system functioning (Nielsen, 2016). e B concentration
in squash did not exceed the toxicity threshold of 200 ppm (Vitosh
etal., 2006).
e increase in Cu and B concentrations following the application
of vermicompost at a medium rate may beattributed to the diverse
responses of plants to nutrient uptake, which vary depending on the
level of organic amendment applied (Bar-Tal etal., 2004; Chang etal.,
2007). Moreover, the same trend is reected in soil and squash total
nutrients, where the medium-rate application of vermicompost
resulted in higher concentrations of nutrients. Interestingly, the
relationship between nutrient concentration and vermicompost rate
does not follow a linear trend. is phenomenon might beelucidated
by a synergistic eect between nutrients and organic matter, a
phenomenon documented by Bar-Tal et al. (2004). A plausible
mechanism could involve the moderate application rate optimizing
cation exchange capacity (CEC), thereby enhancing the retention and
availability of copper and boron to plants. Importantly, this
optimization occurs without the risk of excessive metal xation that
could arise from overly high organic matter application rates
(Clemente etal., 2006).
5 Conclusion
e utilization of vermicompost derived from vermiltration of
wastewater as an organic amendment has shown promise as a
sustainable means of addressing soil health. e incorporation of
vermiltration-derived vermicompost, especially at a rate of
54 kg N ha
−1
, improved the nutritional status of the summer squash
and improved soil health by increasing the total carbon, nitrogen,
particulate organic matter, and microbial activity. ese encouraging
results, obtained in the context of a short-term investigation,
highlighted that vermicompost enabled prompt soil quality recovery
and heightened crop nutritional status. However, the long-term
evaluation of this practice remains a subject of interest. It is crucial to
conduct long-term studies to determine the fate of the added organic
carbon and whether it will endure stable sequestration within the soil
matrix. Additionally, it is important to investigate whether repeated

Malal et al. 10.3389/fsufs.2024.1383715
Frontiers in Sustainable Food Systems 11 frontiersin.org
applications would increase yields and promote overall
agricultural sustainability.
Data availability statement
e original contributions presented in the study are included in
the article/supplementary material, further inquiries can bedirected
to the corresponding author.
Author contributions
HM: Writing – review & editing, Writing – original dra,
Visualization, Investigation, Data curation, Conceptualization. VR:
Writing – review & editing, Investigation, Funding acquisition, Data
curation, Conceptualization. WH: Writing – review & editing,
Resources, Funding acquisition. SD: Writing – review & editing,
Conceptualization. PB: Writing – review & editing, Conceptualization.
MA: Writing – review & editing, Visualization. HL: Writing – review
& editing, Validation, Supervision. CL: Writing – review & editing,
Validation, Supervision, Resources.
Funding
e author(s) declare that nancial support was received for the
research, authorship, and/or publication of this article. is research
was supported by the California Climate investments program and the
California department of Food and Agriculture Healthy soils program
(Grant agreement 18700800). HM acknowledges support from a
Fulbright Fellowship.
Acknowledgments
We thank Bioltro Inc. for providing the vermicompost for
this study.
Conflict of interest
SD and PB were employed by Bioltro Inc.
e remaining authors declare that the research was conducted in
the absence of any commercial or nancial relationships that could
beconstrued as a potential conict of interest.
Publisher’s note
All claims expressed in this article are solely those of the authors
and do not necessarily represent those of their aliated organizations,
or those of the publisher, the editors and the reviewers. Any product
that may be evaluated in this article, or claim that may be made by its
manufacturer, is not guaranteed or endorsed by the publisher.
References
A’ali, R., Jafarpour, M., Kazemi, E., and Pessarakli, M. (2017). Eects of raw materials on
vermicompost qualities. J. Plant Nutr. 40, 1635–1643. doi: 10.1080/01904167.2016.1270319
Ayangbenro, A. S., and Babalola, O. O. (2021). Reclamation of arid and semi-arid soils:
the role of plant growth-promoting archaea and bacteria. Curr. Plant Biol. 25:100173.
doi: 10.1016/j.cpb.2020.100173
Bai, X., Tang, J., Wang, W., Ma, J., Shi, J., and Ren, W. (2023). Organic amendment
eects on cropland soil organic carbon and its implications: a global synthesis. Catena
231:107343. doi: 10.1016/j.catena.2023.107343
Bar-Tal, A., Yermiyahu, U., Beraud, J., Keinan, M., Rosenberg, R., Zohar, D., et al.
(2004). Nitrogen, phosphorus, and potassium uptake by wheat and their distribution in
soil following successive, annual compost applications. J. Env iron. Qual. 33, 1855–1865.
doi: 10.2134/jeq2004.1855
Bell, C. W., Fricks, B. E., Rocca, J. D., Steinweg, J. M., McMahon, S. K., and
Wallenstein, M. D. (2013). High-throughput uorometric measurement of potential soil
extracellular enzyme activities. J. Vis. Exp. 81:e50961. doi: 10.3791/50961
Cambardella, C. A., and Elliott, E. T. (1992). Particulate soil organic-matter changes
across a grassland cultivation sequence. Soil Sci. Soc. Am. J. 56, 777–783. doi: 10.2136/
sssaj1992.03615995005600030017x
Carter, M. R., Angers, D. A., Gregorich, E. G., and Bolinder, M. A. (2003).
Characterizing organic matter retention for surface soils in eastern Canada using density
and particle size fractions. Can. J. Soil Sci. 83, 11–23. doi: 10.4141/S01-087
Chang, E.-H., Chung, R.-S., and Ts ai, Y.-H. (2007). Eect of dierent application rates
of organic fertilizer on soil enzyme activity and microbial population. Soil Sci. Plant
Nut r. 53, 132–140. doi: 10.1111/j.1747-0765.2007.00122.x
Chiou, W.-Y., and Hsu, F.-C. (2019). Copper toxicity and prediction models of copper
content in leafy vegetables. Sustain. For. 11:6215. doi: 10.3390/su11226215
Clemente, R., Escolar, Á., and Bernal, M. P. (2006). Heavy metals fractionation and
organic matter mineralisation in contaminated calcareous soil amended with organic
materials. Bioresour. Technol. 97, 1894–1901. doi: 10.1016/j.biortech.2005.08.018
Codex Alimentarius Commission (1984). Contaminants, joint FAO/WHO food
standards program, vol. XVII. 1st Edn Codex Alimentarius.
Cooper, A., and DeMarco, J. (2023). Composted biosolids amendments for enhanced
soil organic carbon and water storage in perennial pastures in Colorado. Agric. Ecosyst.
Environ. 347:108401. doi: 10.1016/j.agee.2023.108401
Danish Toor, M., Kizilkaya, R., Anwar, A., Koleva, L., and Eldesoky, G. E. (2024).
Eects of vermicompost on soil microbiological properties in lettuce rhizosphere: an
environmentally friendly approach for sustainable green future. Environ. Res.
243:117737. doi: 10.1016/j.envres.2023.117737
De Rooij, M. M. T., Erbrink, H. J., Smit, L. A. M., Wouters, I. M., Hoek, G., and
Heederik, D. J. J. (2024). Short-term residential exposure to endotoxin emitted from
livestock farms in relation to lung function in non-farming residents. Environ. Res.
243:117821. doi: 10.1016/j.envres.2023.117821
Dey Chowdhury, S., Bhunia, P., and Surampalli, R. Y. (2022). Sustainability assessment
of vermiltration technology for treating domestic sewage: a review. J. Water Proc. Eng.
50:103266. doi: 10.1016/j.jwpe.2022.103266
Doane, T. A., and Horwath, W. R. (2003). Spectrophotometric determination of nitrate
with a single reagent. Anal. Lett. 36, 2713–2722. doi: 10.1081/AL-120024647
Dore, S., Deverel, S. J., and Christen, N. (2022). A vermiltration system for low
methane emissions and high nutrient removal at a California dairy. Bioresour. Technol.
Rep. 18:101044. doi: 10.1016/j.biteb.2022.101044
Enebe, M. C., and Erasmus, M. (2023). Vermicomposting technology—a perspective
on vermicompost production technologies, limitations and prospects. J. Environ. Manag.
345:118585. doi: 10.1016/j.jenvman.2023.118585
FAO/WHO. (1988). Report of 33rd meeting, joint FAO/WHO joint expert committee
on food additives, toxicological evaluation of certain food additives and contami- nants no.
24. International Programme on Chemical Safety, WHO, Geneva.
Fernández-Ugalde, O., Virto, I., Barré, P., Gartzia-Bengoetxea, N., Enrique, A.,
Imaz, M. J., et al. (2011). Eect of carbonates on the hierarchical model of aggregation
in calcareous semi-arid Mediterranean soils. Geoderma 164, 203–214. doi: 10.1016/j.
geoderma.2011.06.008
Fronning, B. E., elen, K. D., and Min, D.-H. (2008). Use of manure, compost, and
cover crops to supplant crop residue carbon in corn Stover removed cropping systems.
Agron. J. 100, 1703–1710. doi: 10.2134/agronj2008.0052
Giannetta, B., Plaza, C., Galluzzi, G., Benavente-Ferraces, I., García-Gil, J. C.,
Panettieri, M., et al. (2024). Distribution of soil organic carbon between particulate
and mineral-associated fractions as affected by biochar and its co-application with
other amendments. Agric. Ecosyst. Environ. 360:108777. doi: 10.1016/j.
agee.2023.108777

Malal et al. 10.3389/fsufs.2024.1383715
Frontiers in Sustainable Food Systems 12 frontiersin.org
Horwath, W. R., and Paul, E. A. (1994). “Microbial biomass” In: Methods of Soil
Analysi s. R. W. Weaver, J. S. Angle, P. J. Bottomley, D. F. Bezdicek, M. S. Smith and M.
A. Tabatabaiet al. Eds (Hoboken: John Wiley & Sons, Ltd.), 753–774.
Hossain, M. E., Mei, X., Zhang, W., Dong, W., Yan, Z., Liu, X., et al. (2021). Substitution
of chemical fertilizer with organic fertilizer aects soil Total nitrogen and its fractions
in northern China. Int. J. Environ. Res. Public Health 18:12848. doi: 10.3390/
ijerph182312848
Just, C., Armbruster, M., Barkusky, D., Baumecker, M., Diepolder, M., Döring, T. F.,
et al. (2023). Soil organic carbon sequestration in agricultural long-term eld
experiments as derived from particulate and mineral-associated organic matter.
Geoderma 434:116472. doi: 10.1016/j.geoderma.2023.116472
Kenneth, M. J., Koner, S., Hsu, G.-J., Chen, J.-S., and Hsu, B.-M. (2023). A review on
the eects of discharging conventionally treated livestock waste to the environmental
resistome. Environ. Pollut. 338:122643. doi: 10.1016/j.envpol.2023.122643
Khaliq, H., Juming, Z., and Ke-Mei, P. (2018). e physiological role of boron on
health. Biol. Trace Elem. Res. 186, 31–51. doi: 10.1007/s12011-018-1284-3
Köninger, J., Lugato, E., Panagos, P., Kochupillai, M., Orgiazzi, A., and Briones, M. J.
I. (2021). Manure management and soil biodiversity: towards more sustainable food
systems in the EU. Agric. Syst. 194:103251. doi: 10.1016/j.agsy.2021.103251
Kumar, T., Bhargava, R., Prasad, K. S. H., and Pruthi, V. (2015). Evaluation of
vermiltration process using natural ingredients for eective wastewater treatment. Ecol.
Eng. 75, 370–377. doi: 10.1016/j.ecoleng.2014.11.044
Lafuente, M. T., Sampedro, R., Vélez, D., and Romero, P. (2023). Decient copper
availability on organoleptic and nutritional quality of tomato fruit. Plant Sci. 326:111537.
doi: 10.1016/j.plantsci.2022.111537
Lahlou, F.-Z., Mackey, H. R., and Al-Ansari, T. (2021). Wastewater reuse for livestock
feed irrigation as a sustainable practice: a socio-environmental-economic review. J.
Clean. Prod. 294:126331. doi: 10.1016/j.jclepro.2021.126331
Läuchli, A., and Grattan, S. R. (2014). Plant abiotic stress: salt. In AlfenN. K. Van (Ed.),
Encyclopedia of agriculture and food systems (313–329). Oxford: Academic Press.
Lazcano, C., Arnold, J., Tato, A., Zaller, J. G., and Dominguez, J. (2009). Compost and
vermicompost as nursery pot components: eects on tomato plant growth and
morphology. Span. J. Agric. Res. 7, 944–951. doi: 10.5424/sjar/2009074-1107
Lazcano, C., Tsang, A., Doane, T. A., Pettygrove, G. S., Horwath, W. R ., and Burger, M.
(2016). Soil nitrous oxide emissions in forage systems fertilized with liquid dairy manure
and inorganic fertilizers. Agric. Ecosyst. Environ. 225, 160–172. doi: 10.1016/j.
agee.2016.04.009
Lehmann, J., Bossio, D. A., Kögel-Knabner, I., and Rillig, M. C. (2020). e concept
and future prospects of soil health. Nat. Rev. Earth Environ. 1, 544–553. doi: 10.1038/
s43017-020-0080-8
Li, X., Zhu, W., Xu, F., Du, J., Tian, X., Shi, J., et al. (2021). Organic amendments aect
soil organic carbon sequestration and fractions in elds with long-term contrasting
nitrogen applications. Agric. Ecosyst. Environ. 322:107643. doi: 10.1016/j.
agee.2021.107643
Lindsay, W. L., and Norvell, W. A. (1978). Development of a DTPA soil test for Zinc,
Iron, Manganese, and Copper. Soil Sci. Soc. Am. J. 42, 421–428. doi: 10.2136/sssaj197
8.03615995004200030009x
Luo, Y., Chavez-Rico, V. S., Sechi, V., Bezemer, T. M., Buisman, C. J. N., and Ter
Heijne, A. (2023). Eect of organic amendments obtained from dierent pretreatment
technologies on soil microbial community. Environ. Res. 232:116346. doi: 10.1016/j.
envres.2023.116346
Ma, B., Ma, B.-L., McLaughlin, N. B., Li, M., and Liu, J. (2022). Improvement in
dryland crop performance and soil properties with multiple annual applications of
lignite-derived humic amendment. Soil Tillage Res. 218:105306. doi: 10.1016/j.
still.2021.105306
Martinez-Burgos, W. J., Bittencourt Sydney, E., Bianchi Pedroni Medeiros, A.,
Magalhães, A. I., de Carvalho, J. C., Karp, S. G., et al. (2021). Agro-industrial wastewater
in a circular economy: characteristics, impacts and applications for bioenergy and
biochemicals. Bioresour. Technol. 341:125795. doi: 10.1016/j.biortech.2021.125795
Meyer, G. A., and Keliher, P. N. (1992). An over view of analysis by inductively coupled
plasma-atomic emission spectrometry. In: Inductively coupled plasmas in analytical
atomic spectrometry, A. Montaser and D. W. Golightly, Eds (New York, NY: VCH
Publishers) 473–516.
Miranda, K. M., Espey, M. G., and Wink, D. A. (2001). A rapid, simple
spectrophotometric method for simultaneous detection of nitrate and nitrite. Nitric
Oxide Biol. Chem. 5, 62–71. doi: 10.1006/niox.2000.0319
Naikwade, P. V. (2019). Soil organic carbon sequestration by Long-term application
of manures prepared from Trianthema portulacastrum Linn. Commun. Soil Sci. Plant
Anal. 50, 2579–2592. doi: 10.1080/00103624.2019.1671442
Ngo, P.-T., Rumpel, C., Doan, T.-T., and Jouquet, P. (2012). e eect of earthworms on
carbon storage and soil organic matter composition in tropical soil amended with compost
and vermicompost. Soil Biol. Biochem. 50, 214–220. doi: 10.1016/j.soilbio.2012.02.037
Ngo, P.-T., Rumpel, C., Doan u, T., Henry-des-Tureaux, T., Dang, D.-K., and
Jouquet, P. (2014). Use of organic substrates for increasing soil organic matter quality
and carbon sequestration of tropical degraded soil: a 3-year mesocosms experiment.
Carbon Manag. 5, 155–168. doi: 10.1080/17583004.2014.912868
Nielsen, F. H. (2016). “Boron” in Encyclopedia of food and health. eds. B.
Caballero, P. M. Finglas and F. Toldrá (Oxford: Academic Press), 451–455.
Nurhidayati, N., Ali, U., and Murwani, I. (2016). Yield and quality of cabbage (Brassica
oleracea L. var. Capitata) under organic growing media using Vermicompost and
earthworm Pontoscolex corethrurus inoculation. Agric. Agric. Sci. Proc. 11, 5–13. doi:
10.1016/j.aaspro.2016.12.002
Pasha, M. F. K., Yeasmin, D., Zoldoske, D., Kc, B., and Hernandez, J. (2018).
Performance of an earthworm-based biological wastewater-treatment Plant for a Dairy
Farm: case study. J. Environ. Eng. 144:04017086. doi: 10.1061/(ASCE)
EE.1943-7870.0001290
Plaza, C., Giannetta, B., Fernández, J. M., López-de-Sá, E. G., Polo, A., Gascó, G., et al.
(2016). Response of dierent soil organic matter pools to biochar and organic fertilizers.
Agric. Ecosyst. Environ. 225, 150–159. doi: 10.1016/j.agee.2016.04.014
Püspök, J. F., Zhao, S., Calma, A. D., Vourlitis, G. L., Allison, S. D., Aronson, E. L.,
et al. (2023). Effects of experimental nitrogen deposition on soil organic carbon
storage in Southern California drylands. Glob. Chang. Biol. 29, 1660–1679. doi:
10.1111/gcb.16563
Rahman, M. L., Rahman, M. M., Rahman, G. K. M. M., Solaiman, A. R. M.,
Karim, M. A., Bokhtiar, S. M., et al. (2020). Inuence of tillage and organic amendments
on terrace soil bio-physico-chemical properties and carbon sequestration index.
Commun. Soil Sci. Plant Anal. 51, 2683–2700. doi: 10.1080/00103624.2020.1845355
Raza, S. T., Wu, J., Ali, Z., Anjum, R., Bazai, N. A., Feyissa, A., et al. (2021). Dierential
eects of organic amendments on maize biomass and nutrient availability in upland
calcareous soil. Atmos. 12:1304. doi: 10.3390/atmos12081034
Rocha, D. H. D., Freitas, F. R. S., Sakamoto, I. K., Silva, E. L., and Varesche, M. B. A.
(2024). Co-digestion of solid-liquid waste from citrus agroindustrial: eect of hydraulic
retention time and organic loading rate on H2 production in a long-term continuous
operation leach bed reactor. Int. J. Hydrog. Energy 49, 554–571. doi: 10.1016/j.
ijhydene.2023.10.289
Ryals, R., Kaiser, M., Torn, M. S., Berhe, A. A., and Silver, W. L. (2014). Impacts of
organic matter amendments on carbon and nitrogen dynamics in grassland soils. Soil
Biol. Biochem. 68, 52–61. doi: 10.1016/j.soilbio.2013.09.011
Rynk, R., Schwarz, M., Richard, T. L., Cotton, M., Halbach, T., and Siebert, S. (2022).
“Chapter 4—compost feedstocks” in e composting handbook. ed. R. Rynk (Oxford:
Academic Press), 103–157.
Sah, R. N., and Miller, R. O. (1992). Spontaneous reaction for acid dissolution of biological
tissues in closed vessels. Anal. Chem. 64, 230–233. doi: 10.1021/ac00026a026
Saapi, S. S. Y., Andrianisa, H. A., Zorom, M., Mounirou, L. A., Kouassi, H. A. A.,
and Ahossouhe, M. S. (2024). New developments on vermifiltration as a bio-
ecological wastewater treatment technology: mechanism, application, performance,
modelling, optimization, and sustainability. Heliy on 10:e25795. doi: 10.1016/j.
heliyon.2024.e25795
Salomon, M. J., and Cavagnaro, T. R . (2022). Healthy soils: the backbone of productive,
safe and sustainable urban agriculture. J. Clean. Prod. 341:130808. doi: 10.1016/j.
jclepro.2022.130808
Sarkar, B., Chakrabarti, P. P., Vijaykumar, A., and Kale, V. (2006). Wastewater
treatment in dairy industries—possibility of reuse. Desalination 195, 141–152. doi:
10.1016/j.desal.2005.11.015
Simas, A., Mores, R., Steens, J., Da llago, R . M., Kunz, A., Michelon, W., et al. (2019).
Electrodisinfection of real swine wastewater for water reuse. Environ. Chem. Lett. 17,
495–499. doi: 10.1007/s10311-018-0782-z
Singh, R., Samal, K., Dash, R. R., and Bhunia, P. (2019). Vermiltration as a sustainable
natural treatment technology for the treatment and reuse of wastewater: a review. J.
Environ. Manag. 247, 140–151. doi: 10.1016/j.jenvman.2019.06.075
Sinha, R., Nair, J., Bharambe, G. R., Patil, S. S., and Bapat, P. (2008). Vermiculture
revolution: A low-cost and sustainable technology for management of municipal and
industrial organic wastes (solid and liquid) by earthworms with signicantly low
greenhouse gas emissions. In: Progress in waste management research (159–227). Nova
Science Publishers.
Sokol, N. W., Sanderman, J., and Bradford, M. A. (2019). Pathways of mineral-
associated soil organic matter formation: integrating the role of plant carbon source,
chemistry, and point of entry. Glob. Chang. Biol. 25, 12–24. doi: 10.1111/gcb.14482
Song, X., Li, H., Song, J., Chen, W., and Shi, L. (2022). Biochar/vermicompost
promotes hybrid Pennisetum plant growth and soil enzyme activity in saline soils. Plant
Physiol. Biochem. 183, 96–110. doi: 10.1016/j.plaphy.2022.05.008
Song, S., Wen, Y., Zhang, J., and Wang, H. (2019). Rapid spectrophotometric
measurement with a microplate reader for determining phosphorus in NaHCO3 soil
extracts. Microchem. J. 146, 210–213. doi: 10.1016/j.microc.2019.01.002
Stehouwer, R., Cooperband, L., Rynk, R., Biala, J., Bonhotal, J., Antler, S., et al. (2022).
“Chapter 15—compost characteristics and quality” in e composting handbook. ed. R.
Rynk (Oxford: Academic Press), 737–775.
Sun, T., Mao, X., Han, K., Wang, X., Cheng, Q., Liu, X., et al. (2023). Nitrogen addition
increased soil particulate organic carbon via plant carbon input whereas reduced
mineral−associated organic carbon through attenuating mineral protection in
agroecosystem. Sci. Total Environ. 899:165705. doi: 10.1016/j.scitotenv.2023.165705

Malal et al. 10.3389/fsufs.2024.1383715
Frontiers in Sustainable Food Systems 13 frontiersin.org
Tang, B., Rocci, K. S., Lehmann, A., and Rillig, M. C. (2023). Nitrogen increases soil
organic carbon accrual and alters its functionality. Glob. Chang. Biol. 29, 1971–1983. doi:
10.1111/gcb.16588
Verrone, V., Gupta, A., Laloo, A. E., Dubey, R. K., Hamid, N. A. A., and Swarup, S. (2024).
Organic matter stability and lability in terrestrial and aquatic ecosystems: a chemical and
microbial perspective. Sci. Total Environ. 906:167757. doi: 10.1016/j.scitotenv.2023.167757
Vitosh, M. L., Warncke, D. D., and Lucas, E. R.. (2006). Secondary and micronutrients for
vegetables and eld crops. East Lansing, Michigan: Michigan State University Extension.
Wang, L., Zheng, X., Tian, F., Xin, J., and Nai, H. (2018). Soluble organic nitrogen
cycling in soils aer application of chemical/organic amendments and groundwater
pollution implications. J. Contam. Hydrol. 217, 43–51. doi: 10.1016/j.jconhyd.2018.08.003
Weil, R., Stine, M., Gruver, J., and Samson-Liebig, S. (2003). Estimating active carbon
for soil quality assessment: a simplied method for laboratory and eld use. Am. J.
Altern. Agric. 18, 3–17. doi: 10.1079/AJAA200228
Western Regional Climate Center. (n.d.). Available at: https://wrcc.dri.edu
Wu, L., Zhang, S., Ma, R., Chen, M., Wei, W., and Ding, X. (2021). Carbon
sequestration under dierent organic amendments in saline-alkaline soils. Catena
196:104882. doi: 10.1016/j.catena.2020.104882
Xing, M., Yang, J., and Lu, Z. (2005). Microorganism-earthworm integrated biological
treatment process—A sewage treatment option for rural settlements. In: ICID 21st
European regional conference, Frankfurt (pp. 15–19).
Xu, C., Wang, J., Wu, D., Li, C., Wang, L., Ji, C., et al. (2022). Optimizing organic
amendment applications to enhance carbon sequestration and economic benefits
in an infertile sandy soil. J. Environ. Manag. 303:114129. doi: 10.1016/j.
jenvman.2021.114129
Yan, B., Zhang, Y., Wang, Y., Rong, X., Peng, J., Fei, J., et al. (2023). Biochar
amendments combined with organic fertilizer improve maize productivity andmitigate
nutrient loss by regulating the C-N-P stoichiometry of soil, microbiome, and enzymes.
Chemosphere 324:138293. doi: 10.1016/j.chemosphere.2023.138293
Yang, X.-E., Long, X.-X., Ni, W.-Z., Ye, Z.-Q., He, Z.-L., Stoella, P. J., et al. (2002).
Assessing copper thresholds for phytotoxicity and potential dietary toxicity in selected
vegetable crops. J. Environ. Sci. Health B 37, 625–635. doi: 10.1081/PFC-120015443
Yao, Z., Xu, Q., Chen, Y., Liu, N., Li, Y., Zhang, S., et al. (2021). Leguminous green
manure enhances the soil organic nitrogen pool of cropland via disproportionate
increase of nitrogen in particulate organic matter fractions. Catena 207:105574. doi:
10.1016/j.catena.2021.105574
Zhang, Y., Li, F., Lu, Z., Pei, Z., Zhao, H., Shen, Q., et al. (2023). Organic amendments
eects on soil aggregation and carbon sequestration in saline-alkaline croplands in
China. Agron. J. 115, 2083–2095. doi: 10.1002/agj2.21332
Zhang, M., Tashiro, Y., Ishida, N., and Sakai, K. (2022). Application of autothermal
thermophilic aerobic digestion as a sustainable recycling process of organic liquid waste:
recent advances and prospects. Sci. Total Environ. 828:154187. doi: 10.1016/j.
scitotenv.2022.154187