Structure and diversity of earthworm communities in long-term irrigated soils with raw effluent and treated wastewater

Abstract

This study was conducted in two natural meadows: first, soils were irrigated with raw wastewater (SIRWW) and in the second, soils were irrigated with treated wastewater (SITWW). Earthworms were sampled in eight soil blocks spaced 10 m apart at each site. Earthworm community was characterized and compared using density, biomass, composition, structure, species richness, and diversity parameters. At both meadows, 459 earthworm individuals from two families and seven species were collected. The highest earthworm density and species richness were recorded at SIRWW. Nicodrilus caligenus, was the most abundant species. Most of earthworm community parameters decreased significantly at SITWW. Only two species (N. caligenus and Octodrilus complanatus) were common between the two grasslands. Among the seven species identified at both meadows, four (Allolobophora longa, Easinia fetida, Allolobophora rosea, Allolobophora chlorotica) were exclusively present in SIRWW, whereas a single species (Amynthas sp.) was characterized in SITWW. Three ecological earthworm groups (epigeic, endogeic, and anectic) were represented in SIRWW, with the dominance of endogeics. Further studies are needed to quantify pollution in this soils and the accumulation of pollutant load in earthworms. It is also important to highlight the relationship between the abundance and diversity of earthworms in these two ecosystems with soil biological activity.

Full text

Structure and diversity of earthworm communities in long-term irrigated soils with raw
effluent and treated wastewater
Nawal Ababsa a
,
b, Sihem Fellah b
,
c, Haroun Chenchouni b
,
d,*, Rania Lallaouna a, Khaled Bouchama a,
Mounia Bahaeand Mohamed Kribaa b
a
Department of Ecology and Environment, Faculty of Nature and Life Sciences, University of Khenchela, El-Hamma 40016, Khenchela, Algeria
b
Laboratory of Natural Resources and Management of Sensitive Environments ‘RNAMS’, University of Oum-El-Bouaghi, Oum-El-Bouaghi 04000, Algeria
c
Département de Médicine Dentaire, Faculté de Médicine, University of Salah Boubnider Constantine 3, Constantine 25000, Algeria
d
Higher National School of Forests, Khenchela 40000, Algeria
e
The Animal Eco-Biology Laboratory (LEBA), École Normale Supérieure de Kouba Bachir El Ibrahimi, Kouba 16050, Algeria
*Corresponding author. E-mail: chenchouni@gmail.com
NA, 0000-0002-4052-6619;SF,0000-0002-0296-6701; HC, 0000-0001-9077-2706; RL, 0009-0008-7970-9835; KB, 0000-0001-7007-7327;
MK, 0000-0003-0925-4640
ABSTRACT
This study was conducted in two natural meadows: first, soils were irrigated with raw wastewater (SIRWW) and in the second, soils were
irrigated with treated wastewater (SITWW). Earthworms were sampled in eight soil blocks spaced 10 m apart at each site. Earthworm com-
munity was characterized and compared using density, biomass, composition, structure, species richness, and diversity parameters. At both
meadows, 459 earthworm individuals from two families and seven species were collected. The highest earthworm density and species rich-
ness were recorded at SIRWW. Nicodrilus caligenus was the most abundant species. Most of earthworm community parameters decreased
significantly at SITWW. Only two species (N. caligenus and Octodrilus complanatus) were common between the two grasslands. Among the
seven species identified at both meadows, four (Allolobophora longa, Eisenia foetida, Allolobophora rosea, Allolobophora chlorotica) were
exclusively present in SIRWW, whereas a single species (Amynthas sp.) was characterized in SITWW. Three ecological earthworm groups
(epigeic, endogeic, and anectic) were represented in SIRWW, with the dominance of endogeics. Further studies are needed to quantify
pollution in this soils and the accumulation of pollutant load in earthworms. It is also important to highlight the relationship between the
abundance and diversity of earthworms in these two ecosystems with soil biological activity.
Key words: biodiversity, earthworm community, natural meadows, soil bioindicators, vermifiltration, wastewater irrigation
HIGHLIGHTS
•Diversity and composition of earthworm community were studied in meadows receiving wastewater.
•Species richness and abundance varied between sites irrigated with raw and treated wastewaters.
•Grassland meadows irrigated with raw wastewater created a favorable environment for earthworms.
•Endogeic species tolerate large variations of environmental conditions in wastewater-irrigated lands.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (CC BY 4.0), which permits copying, adaptation and
redistribution, provided the original work is properly cited (http://creativecommons.org/licenses/by/4.0/).
© 2023 The Authors Water Science & Technology Vol 88 No 9, 2473 doi: 10.2166/wst.2023.345
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GRAPHICAL ABSTRACT
1. INTRODUCTION
Soils are complex ecosystems that sustain diverse organisms, including bacteria and pedofauna, which play an important
role in maintaining their functions (Byrne 2022;Ding & Eldridge 2022). Macrofauna has a significant influence on the
entire soil fauna community and, therefore, on ecosystem functions (Ohgushi et al. 2018). Among the soil fauna, the
earthworms, invertebrates belonging to the phylum Annelida, are ecosystem engineers capable of modifying the soil
environment they inhabit (Sizmur & Richardson 2020;Bora et al. 2021). Their incredible architectural heel contributes
to the modification of the soil structure. This biological reworking of soil is accomplished through the movement of
materials known as bioturbation. Thus, earthworms play a key beneficial role in soil structure, function, and productivity
(Liu et al. 2019). Dhiman & Pant (2022) consider them as indicators of soil biological activity. They can withstand high
concentrations of soil pollutants and play a vital role in their effective removal (Zeb et al. 2020). Earthworms have also
shown considerable potential for the remediation of soils polluted by heavy metals such as Pb, Cd, and Ni (Ahadi et al.
2020) and organic contaminants including polychlorinated biphenyls, polybrominated diphenyl ethers, polycyclic aro-
matic hydrocarbons, and pesticides (Zeb et al. 2020). The decontamination mechanisms include the accumulation of
high concentration of heavy metals in the bodies of earthworms, and/or absorption of the water soluble and extractable
fractions of heavy metals by direct contact with earth skin or by gut absorption after ingestion of contaminated organic
matter.
Earthworms drive multiple soil processes, but their specific impact on soil functions differs among species and ecological
categories (Arrázola-Vásquez et al. 2022). They play an important role in organic matter decomposition and nutrient recy-
cling (Coleman 2013;Singh et al. 2019;Yang et al. 2020;Cheng et al. 2021). This gives them a critical role in terrestrial
ecosystems as an important biomarker for toxicology research (Bart et al. 2018;Chen et al. 2020;Zhou et al. 2020;
Dhiman & Pant 2022;Edwards & Arancon 2022). Depending on the source of the contaminants, earthworms can be
used to rehabilitate contaminated soils (Al-Maliki et al. 2021). They are relatively tolerant of toxic elements in soils
(Fründ et al. 2011;Richardson et al. 2020). Sizmur & Richardson (2020) found that both endogeic and epigeic earthworms
increase the mobility of potentially toxic elements in soil and in casts, and all ecological groups (epigeic, endogeic, and
anecic) mobilize potentially toxic elements during soil passage through the earthworm gut. Through the vermifiltration pro-
cess, earthworms are used to treat organic pollutants in wastewater (Kanaujia et al. 2020;Arora et al. 2021,2022) as well
as chemical and pathogenic contaminants (Mohan et al. 2022) and even better and more stable performance has been
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found in the removal of antibiotics from hospital wastewater (Shokouhi et al. 2020). To survive in such an environment,
earthworms have developed effective immune defense mechanisms against invading microorganisms existing in wastewater
(Romo et al. 2016). Furthermore, earthworms play a vital role in shaping and maintaining healthy soil ecosystems, making
them valuable indicators of soil pollution and the effects of receiving wastewaters (Ahadi et al. 2020). Earthworms are a
diverse group of soil-dwelling organisms that contribute to soil fertility and structure through their feeding and burrowing
activities. They consume organic matter, such as dead plant material, and excrete nutrient-rich casts, which enhance soil
nutrient cycling and increase its water-holding capacity. As such, earthworms are considered ecosystem engineers, influen-
cing soil physical, chemical, and biological properties (Liu et al. 2019;Sizmur & Richardson 2020;Arrázola-Vásquez et al.
2022). In the context of soil pollution, earthworms act as sensitive bioindicators. Their abundance, diversity, and behavior
can be significantly affected by pollutants present in the soil (Ouahrani 2003;Sekhara-Baha 2008). Various studies have
demonstrated the adverse effects of contaminants, such as heavy metals, pesticides, and organic pollutants, on earthworm
populations (Nahmani et al. 2003;Zeb et al. 2020). Changes in earthworm community composition and reduced popu-
lation densities can indicate soil degradation and the presence of pollutants (Dlamini & Haynes 2004;Singh et al.
2021;Ahmed et al. 2022). Moreover, the physiological responses of earthworms to pollutants, such as alterations in
enzyme activities or reproductive patterns, provide additional insights into the severity of soil contamination (Pelosi
et al. 2014;Romo et al. 2016;Chen et al. 2020;Cheng et al. 2021).
The use of reclaimed wastewater for irrigation is increasingly recognized as a way to reduce water consumption by promot-
ing the reuse of treated wastewater (Boudjabi et al. 2023). The drawback is that reclaimed wastewater can be a vector of soil
contamination, and its use is expected to alter soil properties, especially the microbial community (Guedes et al. 2022). In
recent years, the potential of using earthworms to treat sewage sludge, domestic wastewater, and human feces is increasing
(Arora et al. 2020). Sinha et al. (2008) suggest that the bodies of earthworms behave as biofilters and they remove contami-
nants from wastewater at a rate of 80–90%. According to Zeb et al. (2020), earthworms can withstand high concentrations of
soil pollutants and play a key role in removing them effectively.
From the point of view of diversity, it has been reported by European Commission (2016) that more than 7,000 earthworm
species have been described worldwide, but the expected number of species is much higher. Earthworm abundance tends to
decline during the dry or extremely cold season and reaches the highest densities and biomass when climatic and soil con-
ditions are more favorable (Singh et al. 2019). The diversity of the earthworm community in a given environment is
influenced by the characteristics of climate, soil, vegetation, and organic resources, as well as the history of land use and
soil disturbance (Edwards & Arancon 2022). Based on the overall strategy of biodiversity conservation and ecosystem ser-
vices provided by organisms, it is of current and future interest to assess the functional and structural biodiversity of
arable soils (van Capelle et al. 2012). Biological diversity has been suggested to be linked to ecosystem functioning
(Scherer-Lorenzen et al. 2022). It is in this context that earthworms are regularly used as bioindicators for management
changes or soil contamination (Fründ et al. 2011;Pelosi et al. 2014).
Wastewater is a common source of soil pollution, as it often contains various contaminants that can adversely affect
soil quality (Ababsa et al., 2020; Boudjabi et al. 2023). Wastewater, whether from domestic, industrial, or agricultural
sources, may contain high levels of organic matter, nutrients, heavy metals, and harmful chemicals (Shokouhi et al.
2020). When wastewater is discharged onto land or used for irrigation, it can significantly impact soil ecosystems
(Chenchouni et al. 2022; Guedes et al. 2022). In this context, this study assumes that earthworms can act as reliable indi-
cators of the effects of wastewater on soil health. Accordingly, monitoring earthworm populations and assessing their
responses to wastewater application can provide valuable information about the potential risks and impacts of this prac-
tice. By studying earthworm communities in soil receiving wastewaters, this study aimed at gaining insights into the
pollution levels and the overall health of the soil, facilitating the development of effective soil management strategies
and pollution mitigation measures. This paper investigates the effect of irrigation with treated and raw wastewater on
the abundance and diversity of earthworms in natural meadow grasslands. In fact, we have evaluated the impact of
the reuse of these two ‘non-conventional waters’on the biological component of the soil characterized by earthworms,
and we hypothesize that the soil irrigated with treated water will be more abundant in earthworms than the soil irrigated
with raw sewage. The site irrigated with raw wastewater (SIRWW) can be considered as a natural bio-filter that uses the
biological component of the soil as a decontamination agent that protects the soil against the polluting components of
raw wastewater.
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2. MATERIALS AND METHODS
2.1. Study area
The two sites that were the subject of this study are located in the province ‘wilaya’of Setif, situated in the high plains of
eastern Algeria. The studied sites are irrigated meadow grasslands. The first site is located at the peri-urban area of the
city of Setif (36°1103.66″N, 05°22050.66″E, elevation ¼984 m a.s.l., https://goo.gl/maps/4qLUePJR61LFsFWA7). It is a
large area of natural meadow grassland that has been irrigated for a long time with raw wastewater discharged from the
city of Setif. The soil of this meadow is sub-permanently submerged by water (i.e. gley soil). The second site is another grass-
land located 1 km east of Beni Fouda city at Setif (36°17098.00″N, 05°36056.00″E, elevation ¼833 m a.s.l., https://goo.gl/
maps/6WWZ7evjKtY8EPCL9). The irrigation of this meadow was ensured by treated wastewater generated from the city’s
wastewater plant. The irrigation of the second meadow has been practiced since the installation of the wastewater treatment
plant in 2006.
Landscapes of the region of Setif include mainly plains used for rainfed cereal cropping and mountains occupied by Med-
iterranean sclerophyllous forests. The climate is Mediterranean semi-arid with hot and dry summers (maximum temperature
of July ¼32.5 °C) and cold winters (minimum temperature of January ¼0.6 °C). Precipitation is low and erratic, ranging from
228.1 to 503.8 mm/year (Ababsa et al. 2020). Soil characteristics of the two meadows are represented in Table 1.
2.2. Earthworm sampling and counting
Sampling was conducted in April 2021, when earthworm activity was at its maximum. We used the hand-sorting standard
sampling method, which consists of hand-sorting all earthworms included in a block of soil 50 cm square and 30 cm deep.
Separation of earthworms from the soil matrix was performed onsite. For each site, i.e. meadow soils irrigated with raw waste-
water (SIRWW) and meadow soils irrigated with treated wastewater (SITWW), earthworms were sampled in eight soil blocks
spaced 10 m apart. The sampling points were chosen along the diagonal in each meadow. All collected earthworms were
placed in plastic jars (300 mL) containing ethanol, and upon arrival at the laboratory, the earthworms were rinsed and
dried on paper, after which they were weighed and counted. Then the specimens were preserved in 70% ethanol before
being identified. The identification of earthworm species was based on morphological characteristics that involves carefully
observing and analyzing various physical traits and features of the earthworm’s body, including: the external morphology
such as color, size and shape of the body, the number of segments (annuli), and any distinctive markings or structures on
the skin; segmentation and clitellum (its location, shape, and coloration); arrangement, number, and distribution of setae
on each segment; and shape and structure of the earthworm’s head, as well as its mouthparts. Some features might require
microscopic examination. This could involve looking at structures like the prostomium, mouthparts, and reproductive organs
in finer detail. To accurately identify a species, the observed characteristics are compared to established taxonomic keys,
identification guides, and existing literature on the available species descriptions of the region (Bouché 1972;Omodeo
et al. 2003;Sekhara-Baha 2008). The identified earthworm species were classified into three ecological groups: epigeic
(litter dwelling), endogeic (shallow burrowing), and anecic (deep burrowing) (Bouché 1972).
Table 1 |Soil characteristics of the two meadow sites: SIRWW and SITWW
Soil properties SITWW SIRWW
pH 8.0 7.5
Electrical conductivity (mS/cm) 0.592 1.231
P
2
O
5
(ppm) 181.7 253.2
Total nitrogen (%) 0.26 0.26
Organic matter (%) 3.78 7.52
Total CaCO
3
(%) 33.88 17.94
Soil moisture (%) 20 90
Soil texture Clayey Clayey
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2.3. Data analysis
The assessment of earthworm biodiversity was determined using several descriptors based on the absolute abundance (n
i
)
measured for each earthworm species at each sample in the two meadows. The relative abundance (RA) was determined
as the ratio of the number of individuals of a species (n
i
) to the total number of individuals (N) of all species (RA ¼n
i
/
N100). Frequency of occurrence (Occ) was calculated for each species as the ratio of the number of samples where the
species was present (m
i
) to the total number of samples (M) realized (Occ ¼m
i
/M100). Based on occurrence values,
species were classified as: very accidental species (VA) when Occ ,10%, accidental species (AC) when Occ ¼[10–25%],
common species (CM) when Occ ¼[25–50%], and constant species (CN) when Occ 50%.
All earthworm individuals of all species pooled were weighed to determine total fresh biomass (B) per sample. Species rich-
ness (S) was defined as the total number of species present at a given sample or a batch of samples (meadow). At the sample
level, abundance-to-species richness ratio (N/S) and biomass-to-species richness ratio (B/S) were computed to standardize
abundances and biomasses to species richness.
Earthworm alpha diversity was explored using Shannon diversity index (H), with H¼Σ((n
i
/N)log
2
(n
i
/N)); Pielou’s
evenness index (E), E¼H/log
2
(S); Simpson reciprocal index (SRI) diversity, where SRI ¼(N(N1))/Σ(n
i
(n
i
1)); and
SRI/S ratio. The latter ratio and Evary from 0 (indicating uneven distribution of species densities within the community,
often related to the dominance of 1–2 species) to 1 (indicating high balance between densities of populations). The range
of SRI values included between 1 (indicating low diversity) and S(indicating high diversity) (Chenchouni 2017).
Using the software EstimateS version 9.1 (Colwell 2013), species richness was estimated and interpolated in each meadow
separately and in both meadows combined. In each case, estimates of S was carried out using four asymptotic species richness
estimators: first-(S
Jack1
)-order and second-(S
Jack2
)-order Jackknife estimators, Chao1 (S
Chao1
), and Chao2 (S
Chao1
) estimators.
We employed sample-based rarefaction curves to compare species richness in sampled meadows, and accumulation curves
for species richness interpolations to a sample size of 100, which represented a sampling effort higher by 12.5 times than the
reference sample size in each meadow (M¼8).
2.4. Statistical analysis
The R software (version 4.2.1) was used for statistical analyzes of the data (R Core Team 2022). Sample-based data of earth-
worm species and community were summarized using basic statistics (mean, standard deviation, minimum, maximum, and
coefficient of variation) for each sampling meadow. Community parameters for SIRWW and SITWW were plotted using the
R package {ggplot2}. Prior testing the variation of earthworm community parameters, i.e. those related to density, biomass,
composition, structure, species richness, and diversity; Shapiro-Wilk test was used for verifying normality. Then differences
of these parameters among SIRWW and SITWW were analyzed using Kruskal–Wallis rank sum tests. The variation of each
species abundance per sample between SIRWW and SITWW was also tested using the same procedure. The interrelation-
ships between earthworm community parameters were analyzed Pearson correlation tests separately for each sampled
meadow. The obtained correlation matrices were mapped in an interactive plot using the R package {corrplot}. The statistical
significance of the applied tests was set at the threshold (p,0.05).
3. RESULTS
3.1. Community composition and systematics
The results including all earthworm species collected at the two sites studied (SIRWW and SITWW) are reported in Table 2.
For both meadows sampled, a total of 459 individuals of earthworms were collected, of which 90.2% were collected from
SIRWW. The three ecological groups: epigeic, endogeic, and anecic were represented, with a dominance of endogiec earth-
worms. Only seven species, attached to two families, were identified based on all earthworm individuals collected (Table 2).
The meadow with SIRWW enclosed the highest earthworm abundance, with a total of 414 individuals classified into six
species, five genera and one family (Lumbricidae). Nicodrilus caligenus was the most abundant species (62.32% of the
total abundance), whereas Octodrilus complanatus represented 18.12% of individuals, and the remaining four species had
abundances less than 10%, namely Allolobophora chlorotica (RA ¼9%), Allolobophora longa (RA ¼4.59%), Eisenia fetida
(RA ¼3.14%), and Allolobophora rosea (RA ¼2.17%). With only 45 individuals (RA ¼9.2% of the total), the abundance of
earthworms collected in SITWW was much lower compared to SIRWW. The community included three species belonging
to three genera and two families (Lumbricidae and Megascolecidae), of which O. complanatus was the most abundant
(RA ¼95.6%) and the other two species (N. caligenus and Amynthas sp.) represented each 2.2% of the total abundance
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Table 2 |Systematic list, ecological groups, absolute abundance (n
i
and N), relative abundance (RA, %) and occurrence (occ, %) of earthworm species collected in the two meadow
soils irrigated with raw wastewater (SIRWW) and treated wastewater (SITWW)
Family Species
Ecological
groups
SITWW SIRWW Overall
ni +SD N RA Occ Scale ni +SD N RA Occ Scale ni +SD N RA Occ Scale
Lumbricidae Octodrilus complanatus (Dugés, 1828) En–An 5.4 +8.9 43 95.6 50.0 CN 9.4 +15.7 75.0 18.1 50.0 CN 7.4 +12.5 118 25.7 50.0 CN
Lumbricidae Nicodrilus caligenus (Bouché, (1972) En 0.1 +0.4 1 2.2 12.5 AC 32.3 +38.6 258.0 62.3 75.0 CN 16.2 +31.2 259 56.4 43.8 CM
Lumbricidae Allolobophora longa (Ude, 1885) An — ———— 2.4 +4.8 19.0 4.6 25 CM 1.2 +3.5 19 4.1 12.5 AC
Lumbricidae Allolobophora rosea (Bouché, (1972) En — ———— 1.1 +2.2 9.0 2.2 25 CM 0.6 +1.6 9 2.0 12.5 AC
Lumbricidae Allolobophora chlorotica (Savigny, 1826) En — ———— 5.0 +14.1 40.0 9.7 12.5 AC 2.5 +10.0 40 8.7 6.3 VA
Lumbricidae Eisenia fetida (Savigny, 1826) Ep — ———— 1.6 +4.6 13.0 3.1 12.5 AC 0.8 +3.3 13 2.8 6.3 VA
Megascolecidae Amynthas sp. —0.1 +0.4 1 2.2 12.5 AC —————0.1 +0.3 1 0.2 6.3 VA
Total 5.6 +8.7 45 100 51.8 +33.3 414 100 28.7 +33.5 459 100
Species abundances are given in mean +standard deviation (n
i
+SD) the total at all samples (N). Ecological groups (Ep: epigeic, En: endogeic, En–An: endogeic–anecic, An: anecic). Scales of occurrence (CN: constant species,
CM: common species, AC: accidental species, VA: very accidental species).
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(Table 2). The Kruskal–Wallis rank sum test showed that only N. caligenus species were site-dependent (χ²¼7.08, p¼0.008).
While for the other species, no statistically significant difference (p.0.05) was recorded between the two meadows.
According to specific values of species occurrence frequency (Occ) at soil samples collected at SIRWW (Table 2), E. fetida
and A. chlorotica were accidental and rare (Occ 12.5%), A. longa and A. rosea were considered common species (12.5 ,
Occ ,50) and O. complanatus and N. caligenus were constant species (Occ .50%). At SITWW, O. complanatus was classi-
fied as a constant species as it occurred in 50% of samples, while N. caligenus and Amynthas sp. occurred as accidental
species.
3.2. Density, biomass and Alfa diversity estimates of earthworms
Figure 1 summarizes the values of earthworm community parameters (density, biomass, composition, structure, species rich-
ness, and diversity) obtained for the two meadows. The boxplots highlighted a large variability in data between and within
SIRWW and SITWW. The SIRWW was more abundant and richer in earthworms with an N/S ratio of 34 +31.52. On aver-
age it had 51.75 +33.28 individuals and 2 +0.76 species, while SITWW showed an abundance of 5.63 +8.73 individuals
and 0.75 +0.71 species. The edaphic conditions of the meadow with SIRWW allowed the development of earthworm bio-
mass 12 times more important than the biomass recorded in SITWW (22.23 +17.6 g vs. 2.73 +5.73 g, respectively). The
biomass-to-species richness ratio was also higher at SIRWW. Earthworm diversity and community structure were character-
ized using Shannon index, Pielou’s evenness index, Simpson reciprocal index, and SRI/S ratio, which denoted the highest
scores in SIRWW with 0.8 +0.59, 0.66 +0.44, 1.8 +0.7, and 0.91 +0.13, respectively (Figure 1); compared to the low
values recorded in SITWW (0.13 +0.35, 0.13 +0.35, 0.75 +0.71 and 0.63 +0.52, respectively).
3.3. Variations of density, biomass and diversity parameters
The results of the Kruskal–Wallis rank sum test (Figure 1) revealed significant differences (K².3.87, p,0.05) among the two
studied meadows in almost all the parameters characterizing the community of earthworms. These significant variations con-
cerned earthworm abundance (number of individuals), species richness, abundance-to-richness ratio, biomass, biomass-to-
richness ratio, Shannon diversity index, evenness, and SRI. Only values of SRI/S ratio did not show a significant difference
(K²¼0.11, p¼0.741) between SIRWW and SITWW according to this test.
3.4. Interrelationships between diversity parameters
Out of the 36 correlation tests realized between the studied parameters in each meadow (Figure 2), 22 were significant (p,0.05)
in SIRWW and 14 in SITWW. Overall and for both meadows, non-significant correlations ( p.0.05) were recorded between the
pairs N–(S,H,E, SRI, SRI/S), B–(S,E), and SRI/S–(N/S,B,B/S,H, and E). The significant correlations observed in N(with N/S,
B, and B/S) were all positive, these four parameters were all positively correlated. The parameters of density and biomass
(N,N/S,B, and B/S) were negatively correlated with diversity and structure indices (H,E, and SRI); however, these negative
correlations were significant only in SIRWW. Species richness and community diversity parameters, i.e. S,H,E, and SRI,
were all significantly and positively auto-correlated. The majority of negative correlations among the studied parameters was
observed in SIRWW, in which S revealed significant correlations with all parameters except Band SRI/S, and N/S with all par-
ameters except SRI/S. The significant correlations observed with SRI/S were established with S (p¼0.004) and SRI ( p¼0.004).
3.5. Traits of earthworm ecological groups
Data on abundance, biomass, species richness, occurrence parameters of earthworm ecological groups recorded at the two
studied meadows are reported in Table 3. In SITWW, the endogeic–anecic group was the most abundant (N ¼43 individuals).
Endogeics were marked by only one individual, while no earthworms belonging to the epigeic and anecic groups were col-
lected. Moreover, all ecological groups were present in SIRWW, with a dominance of endogeics (307 individuals), followed
by endogeic–anecic, anecic and epigeic with abundance values of 75, 19, and 13, respectively. The highest value of RA was
recorded by the endogeic–anecic group (RA ¼95.6%) in SITWW, while RA was higher in the endogeic group (RA ¼74.2%) of
SIRWW. The endogeic group was represented by four species which had highest species richness score in SIRWW, while the
other groups, when present, were represented by only one species. The highest value of N/S ratio was recorded by the endo-
geic–anecic group in SITWW, whereas in SIRWW, the two groups endogeic–anecic and endogeic had almost similar N/S
ratios, 75 and 76.75, respectively. Earthworms of the endogeic–anecic ecological group occurred regularly in both meadows
(Occ ¼50%), which assigned them to the class of constant groups. The same occurrence class was distinguished in SIRWW
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Figure 1 |Boxplots displaying statistics and values of earthworm density, biomass, composition, and diversity parameters in meadow soils
irrigated with raw (SIRWW) and treated wastewater (SITWW). Chi-squared statistics (χ²) and p-values are results of Kruskal–Wallis rank sum
tests of differences among the two meadows.
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for the endogeic group (Occ ¼87.5%), which was classified as incidental (Occ ¼12.5%) in SITWW. For the latter meadow,
epigeic and anecic groups occurred as accidental and common earthworms, respectively.
3.6. Species richness rarefication
The values of the four estimators of species richness applied in this study are summarized in Figure 3. The first-order Jackknife
species richness estimator (S
Jack1
) showed that rarified species richness was expected to increase by 34.8% in SITWW and
23.1% in SIRWW. For both meadows combined, estimated species richness was 9.8 species exceeding the observed species
richness (S¼7), which indicated that the overall completeness of earthworm inventory was 72%. The application of the
second-order Jackknife estimator (S
Jack2
) revealed that species richness was 48.3% (S
Jack2
¼5.8 species) and 24.6%
(S
Jack2
¼7.96) higher than the observed richness in SITWW and SIRWW, respectively. For the samples of two meadows
pooled, estimated species richness was expected to stretch to 10.8 species (S
obs
¼7 species). Chao1 species richness estimator
demonstrated that inventory completeness was 100% in SIRWW and in the two meadows combined (S
Chao1
¼S
obs
). In
SITWW, inventory completeness was estimated to 75.4% as S
Chao1
was 3.98 species and S
obs
¼3 species. Furthermore,
the application of the Chao2 estimator revealed slight to moderate increases in species richness, with S
Chao2
¼3.8,
S
Chao2
¼6.29 and S
Chao2
¼9.11, which matched up inventory completeness levels of 78.9, 95.4, and 76.8% in SITWW,
SIRWW, and both meadows combined, respectively (Figure 3).
Figure 2 |Probability values of correlation coefficients of Pearson’s correlation tests between density, biomass, and diversity parameters of
earthworms collected in soils of the two meadows (SITWW above the diagonal and SIRWW below the diagonal). P-values are superimposed
over the solid circles whose size and color are proportional to Pearson correlation coefficient scores. Correlations marked with a cross are
non-significant (p.0.05). (Variable abbreviations: N: abundance (number of individuals), S: species richness, N/S: abundance-to-species
richness ratio, B: total biomass, B/S: biomass-to-species richness ratio, H: Shannon diversity index, E: Pielou’s evenness index, SRI: Simpson
reciprocal index, and SRI/S ratio.)
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3.7. Species richness interpolation
Overall, the rarefaction curves increased with the increase in number of samples and then reached a plateau (Figure 4). The
interpolation curves of species richness showed that S
est
appeared to reach stability from the 12th sample in SITWW and from
the 20th sample in SIRWW. The predicted values of S
est
(mean +SD [lower 95%–upper 95% of confidential intervals]) at 100
samples were 3.8 +1.38 species (CI: 1.09–6.51), 6.87 +1.64 species (CI: 3.66–10.09) and 9.11 +3.15 species (CI: 2.93–
15.28) for SITWW, SIRWW, and both meadows combined, respectively. These projected species richness scores were
Table 3 |Number of individuals, relative abundance, species richness, N/S ratio, and occurrence frequency for the ecological groups of
earthworms collected in meadow soils irrigated with raw wastewater (SIRWW) and treated wastewater (SITWW) in northeastern
Algeria
Sampled meadows
Ecological groups
Epigeic Endogeic Endogeic–Anecic Anecic Not determined
Total density ‘N’
SITWW —143 —1
SIRWW 13 307 75 19 —
Relative abundance (%)
SITWW —2.2 95.6 —2.2
SIRWW 3.1 74.2 18.1 4.6 —
Species richness ‘S’
SITWW —11 —1
SIRWW 1 4 1 1 —
N/S ratio
SITWW —143 —1
SIRWW 13 76.75 75 19 —
Occurrence frequency (%)
SITWW —12.5 50 —12.5
SIRWW 12.5 87.5 50 25 —
Figure 3 |Observed (S
est
¼analytical) and estimated (S
Jack1
,S
Jack2
,S
Chao1
, and S
Chao2
) earthworm species richness recorded at meadows
irrigated with treated and untreated wastewater, and for both meadows combined (Overall). Rectangular bars and error bars represent mean
values and standard deviations, respectively, of species richness estimates averaged over 100 randomizations.
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associated to specific densities of 900; 5,175; and 2,869 individuals, respectively. Accordingly, with the increase if number of
samples to 100, the number of earthworm species was expected to increase by 26.6, 14.5, and 30.14% for SITWW, SIRWW,
and both meadows combined.
3.8. Analysis of similarity (beta diversity)
The Venn diagram indicated that only two species (N. caligenus and O. complanatus) were common between the two mea-
dows (Figure 5). Out of the seven species identified in this study, four were exclusively present for SIRWW (A. longa,E. fetida,
A. rosea,A. chlorotica). In contrast, SITWW was characterized by the exclusivity of a single species (Amynthas sp.). The
application of qualitative similarity indices (Jaccard and Sørensen) showed that two meadows had a low similarity of 28.6
and 44.4%, respectively. Similarly, quantitative similarity estimated using Morisita-Horn index and Bray–Curtis index indi-
cated low resemblance values with 28.8 and 20.8%, respectively. However, both raw and estimated metrics of either
Chao-Jaccard and Chao-Sørensen showed high similarity scores with 86.4, 97.8, 92.7, and 98.9%, respectively (Figure 5).
These high scores were related to the abundance-based coverage estimations (ACE) of species richness in SITWW
(ACE ≈4 species), which was greater than the observed species richness (three species).
4. DISCUSSION
This study analyzed and compared various parameters (density, biomass, composition, structure, species richness, and diver-
sity) of the earthworm community living in long-term irrigated soils with wastewater in meadow grasslands in a semiarid region
of Algeria (North Africa). In this study, we collected a total of 459 individuals classified into two families (Lumbricidae and
Megascolecidae), five genera and seven species, with the family Lumbricidae being the most represented with 458 individuals
and six species. According to Edwards & Arancon (2022), these two families are the most ecologically important in Europe,
North America, Australia, and Asia. Mainly endemic in the Palearctic region, including Europe and the north region of North
Africa, the family Lumbricidae is spread all over the world, mainly by humans. Earthworms naturally disperse through various
Figure 4 |Sample-based interpolation curves of species richness estimated for earthworms at meadows irrigated with treated and
untreated wastewater, and for both meadows combined (overall). Light grey-shaded areas represent lower and upper bounds of 95% con-
fidence intervals for S
(est)
. Color-shaded areas indicate +standard deviations of S
(est)
determined over 100 randomizations. The black solid
circle refers to the reference sample size used in rarefication.
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mechanisms. One way is through natural events like heavy rainfall, where earthworms can be carried in streams or run-off.
Additionally, accidental transportation by humans, known as anthropochore transport, can occur when earthworms are unin-
tentionally carried along with plants or other materials. Earthworm cocoons can also be transported by birds and other animals
(zoochory), clinging to their feet or bodies. The accidental introduction of earthworms into soil where plants grow can signifi-
cantly impact their dispersal patterns. Moreover, passive dispersal can happen through activities such as tractor usage, where
earthworms can be inadvertently spread, influencing the rate of population expansion in newly colonized agricultural fields
(Mathieu et al. 2018). According to Reynolds (2018), a total of 35 species belonging to six families were identified in Algeria,
including: Acanthodrilidae: 3 species, Criodrilidae: 2 species, Glossoscolecidae: 1 species, Haplotaxidae: 1 species, Hormogas-
tridae: 1 species, and Lumbricidae: 27 species. Earthworms are found worldwide in soils with sufficient moisture to support
and sustain them, except in arid lands such as deserts. Their abundance is extremely variable, ranging from only a few individ-
uals to more than 2,000 per m² (Bora et al. 2021;Edwards & Arancon 2022). They are common worldwide in forests and
natural grasslands as well as in agrosystems (Philips et al. 2019;Zerrouki et al. 2022).
In total, we recorded the presence of seven earthworm species in both irrigated meadow, with a specific richness of six
species in SIRWW and only three species in SITWW. Of the 35 species identified in Algeria, Bazri et al. (2013), determined
18 species in eastern Algeria at 62 sites distributed over different bioclimatic zones; whereas Sekhara-Baha (2008), recorded
11 species in the Mitidja plain; and Ouahrani (2003), determined 11 species in the Constantinois region. In meadows irri-
gated with raw treated and agricultural effluents, Ababsa et al. (2020), determined three species at Setif. Following the
study on the effects of grassland management on earthworm communities under ambient and future climatic conditions
(Singh et al. 2021), earthworm communities were significantly impacted by different grassland types, with earthworm abun-
dance and biomass decreased in grasslands with high intensification use and low plant diversity.
The distribution of earthworm densities per studied meadow revealed that SIRWW was more abundant (N¼414 individ-
uals) in earthworms than SITWW (N¼45 individuals). According to Ababsa et al. (2020), the irrigation with raw urban
wastewater, despite its richness in organic and particulate matter, resulted in increased earthworm density, higher soil por-
osity and water transfer.
Environmental conditions directly affect the abundance and diversity of earthworms (Ahmed et al. 2022;Ding & Eldridge
2022). In the present study, despite the commonality between the two meadows, as an irrigated grassland ecosystem, it
Figure 5 |Venn diagram displaying observed (S) and estimated (ACE: abundance-based coverage estimation) species richness of earth-
worms recorded in meadow soils irrigated with raw wastewater (SIRWW) and treated wastewater (SITWW) in northeastern Algeria. Figures in
black are number of exclusive species of each meadow, whereas the white number between square brackets represents the number of
shared species among these meadows. Binomials of earthworm species are given in italics, with only initials of genera. Similarity statistics (in
%) are represented within the overlapped area of the diagram (J: the classic Jaccard index, Sø: the classic Sørensen index, C–J
(raw)
: raw Chao’s
abundance-based Jaccard index, C–J
(est)
: estimated Chao’s abundance-based Jaccard index, C–Sø
(raw)
: raw Chao’s abundance-based
Sørensen index, C–Sø
(est)
: estimated Chao’s abundance-based Sørensen index, MH: Morisita–Horn index, BC: Bray–Curtis index).
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appears that SIRWW created favorable conditions for the development of earthworm populations. Earthworms can with-
stand high concentrations of soil pollutants and play a vital role in their effective removal. They can remove contaminants
from the soil or help degrade non-recyclable chemicals; these are the basis for vermifiltration, which has proven to be an
alternative and inexpensive technology to treat contaminated soils (Zeb et al. 2020).
In SIRWW, earthworms are an integrated component of the pedofauna that participates in the remediation of pollution,
especially in meadows and grasslands, which cover about 40% of global inland surface and harbor the greatest number of
earthworms (Edwards & Bohlen 1996). These ecosystems have high economic, ecological and biodiversity values due to
their role in providing fodder for livestock and retaining high levels of carbon in the soil (Lenhart et al. 2015). The ecological
conditions at SIRWW allowed for the development of a high abundance and richness of earthworms, with an earthworm
biomass 12 times greater than the biomass at SITWW. The reason for this high abundance and richness could be attributed
to the adaptation of earthworms to the quality of the irrigation wastewater. Rodriguez-Campos et al. (2014), reported that the
presence, and in some cases the high abundance, of earthworms in contaminated environments suggests their high eco-phys-
iological tolerance and ecological plasticity.
Apart from soil moisture that was considered in this study, and which is dependent on air and soil temperature as well as
other physicochemical soil properties, soil factors are reported to influence the abundance and diversity for earthworms.
According to Mishra et al. (2020) and Zerrouki et al. (2022), soil temperature, moisture, and other edaphic factors are key
regulators of earthworm abundance and activity in nature. Earthworms are most sensitive to the hydraulic properties of
the soil, and in times of drought, they move to deeper soil layers for protection (Johnston et al. 2014). The studies of Philips
et al. (2019),Singh et al. (2019), and Ahmed et al. (2022), reported the presence of a significant positive relationship between
earthworm density with soil moisture and rainfall. Thakur et al. (2018) reported that soil moisture is a major factor determin-
ing soil biological activity, while Al-Maliki et al. (2021) concluded that earthworms are more sensitive to changes in soil
temperature than to the effects of moisture content and thus can be used as a bio-indicator of soil quality. It is true that earth-
worm activity depends on adequate soil moisture availability, but not all species have the same moisture requirements, and
within a species, the moisture requirements of earthworm populations in different parts of the world can be very different. For
instance, A. chlorotica,A. longa, and A. caliginosa are among the earthworm species that can survive long periods submerged
in water (Edwards & Arancon 2022).
The soil of the studied meadow with the highest earthworm abundance was submerged in raw wastewater (gley soil), while
the grassland with the lowest earthworm abundance was characterized by low soil moisture compared to SIRWW. Further-
more, a high-performance of treatment against chemical oxygen demand (COD), total nitrogen (TN), total phosphorus (TP),
total suspended solids and even heavy metals has been approved by the vermifiltration of domestic and industrial wastewater
(Namaldi & Azgin 2023). Removal rates were 82.1, 92.8, 96.4, and 96.3%, respectively, for COD, NH
4
þ
-N, NT and TP as
reported by Das & Paul (2023). Studies reported the presence of significant positive relationships between earthworms
and the content of soil organic carbon, where high soil organic carbon levels favored high earthworm abundance (Dlamini
& Haynes 2004;Johnston et al. 2014;Ahmed et al. 2022).
The ecological categories of earthworms help us understand the associated ecological processes and establish mechanistic
links between earthworm community structure and ecosystem function (Hsu et al. 2023). Despite the dominance of endo-
geics (four species), all three ecological groups are represented in this study (Table 2). The results of the study conducted
by Singh et al. (2021) reveal the dominance of endogeic species in terms of total abundance and biomass. Our findings
are in agreement with previous studies, where endogeic species dominance in grasslands is clearly described (Didden
2001;Butt et al. 2022). This may be related to the fact that endogeic species tolerate large variations in environmental con-
ditions. Similar findings were reported by Mantoani et al. (2022), where a dominance of the endogenic earthworms
(A. chlorotica,A. caliginosa, and A. rosea) was observed in all sampled plots across five grassland sites.
Some earthworm species are widely distributed and are not characteristic of any particular site/habitat/ecosystem. In our
study, the two species, O. complanatus and N. caligenus were ubiquitous regardless of the quality of irrigation water.
Nahmani et al. (2003) reported that A. caliginosa was dominant in unpolluted grasslands (density ¼45 ind./m²). This species
consumes high proportions of soil organic matter and microorganisms in its diet (Potapov et al. 2019). Fonte et al. (2009)
reported that it is a cosmopolitan species and commonly found in temperate agroecosystems worldwide. A large-bodied
anecic worm, O. complanatus is present from North Africa and Spain through southern European countries and Cyprus
to Turkey and the Levant region (Pavlícek & Csuzdi 2016). It is typically restricted to wet meadows (Chenchouni 2017),
and common in grasslands and pastures, with a wide distribution in Europe and North Africa (Monroy et al. 2007). Because
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A. caliginosa is sensitive to metal toxicity and has high ecological plasticity and adaptability in agroecosystems, it is con-
sidered an excellent bioindicator model of pollution and environmental status (Bouché 1972;Otmani et al. 2018). Adults
of this endogeic species were dominant in all agricultural systems sampled by Lemtiri et al. (2018). In our study, A. chlorotica
was present only at SIRWW (in gley soil) with an AR of 8.7%. According to Plum & Filser (2005), this species was dominant I
gley soil of wet grasslands. It is a species known for its tolerance to wet soils (Zorn et al. 2008). A. chlorotica and A. rosea
species are probably the most widespread earthworm species in the world (Szederjesi 2017). The taxon Amynthas sp. exhib-
ited a more restricted distribution where we collected only a single individual from SITWW. Eisenia fetida worms are widely
used to test the toxicity of pollutants ( Jiang et al. 2020;Chenchouni et al. 2022). Weight change and cocoon production of this
earthworm as well as mortality were significantly affected when salinity increases (Owojori et al. 2009;Yang et al. 2022).
Raiesi et al. (2020) reported that salinity increased lead (Pb) toxicity to the life cycle and activity of E. fetida. In our study,
this species was recorded only at SIRWW where soil electrical conductivity is twice higher than of SITWW.
5. CONCLUSION AND RECOMMENDATION
This study explored the diversity of earthworms in the rhizosphere of two meadow grassland ecosystems, SIRWW and SITWW.
Our results revealed a good abundance and specific richness, as well as a higher biomass of earthworms in SIRWW compared to
SITWW. Given the high abundance of N. caligenus, we suggest to set up an experimental study aiming at using this species in a
vermifiltration system for urban wastewater treatment. Thus, we suggest carrying out other studies on the chemical quality of
soils and quantifying the accumulation of pollutant load in the body of earthworms. Therefore, it is important to study the bio-
logical activity of soils under irrigation with both types of water to consider the relationship of abundance and biodiversity of
earthworms with microorganisms, soil respiration, as well as the enzymatic activity of the soils.
The findings of this study hold significant relevance for regions beyond its immediate context. By investigating earthworm
communities under contrasting wastewater irrigation practices, the study provides insights applicable to diverse global set-
tings facing similar challenges. As wastewater utilization for irrigation gains prominence worldwide, understanding its
impact on soil ecosystems is crucial. The observed higher earthworm abundance, species richness, and biomass in
SIRWW highlight the potential benefits of such practices. The suggestion to explore the utilization of abundant, well-adapted
and resilient species in vermifiltration systems for urban wastewater treatment introduces an innovative solution with broader
implications. Furthermore, the call for studies on soil chemical quality, pollutant accumulation in earthworms, and the inter-
play between earthworms, microorganisms, soil respiration, and enzymatic activity underscores the need for comprehensive
assessments of soil health. These findings resonate across regions grappling with sustainable agricultural practices, environ-
mental conservation, and efficient wastewater management, making this study’s insights invaluable for shaping informed
decision-making globally.
AUTHORS’CONTRIBUTIONS
N.A. was involved in conceptualization, methodology, resources, investigation, writing –original draft, writing –review and
editing. S.F. was involved in visualization, methodology, resources, investigation. H.C. was involved in formal analysis, visu-
alization, writing –original draft, writing –review and editing. R.L. was involved in investigation. K.B. was involved in writing
–review and editing. M.B. was involved in investigation. M.K. was involved in conceptualization and investigation. Please
refer the CRediT taxonomy for the term explanation.
DATA AVAILABILITY STATEMENT
All relevant data are included in the paper or its Supplementary Information.
CONFLICT OF INTEREST
The authors declare there is no conflict.
REFERENCES
Ababsa, N., Kribaa, M., Tamrabet, L., Addad, D., Hallaire, V. & Ouldjaoui, A. 2020 Long-term effects of wastewater reuse on hydro physicals
characteristics of grassland grown soil in Semi-Arid Algeria.J. King Saud Univ. Sci. 32 (1), 1004–1013. https://doi.org/10.1016/j.jksus.
2019.09.007.
Water Science & Technology Vol 88 No 9, 2486
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Ahadi, N., Sharifi, Z., Hossaini, S. M. T., Rostami, A. & Renella, G. 2020 Remediation of heavy metals and enhancement of fertilizing
potential of a sewage sludge by the synergistic interaction of woodlice and earthworms.J. Hazard. Mater. 385, 121573. https://doi.org/
10.1016/j.jhazmat.2019.121573.
Ahmed,S., Marimuthu, N., Tripathy, B.,Julka, J. M. & Chandra, K. 2022Earthworm community structure and diversityin different land-usesystems
along an elevation gradient in the Western Himalaya, India.Appl. Soil Ecol. 176, 104468. https://doi.org/10.1016/j.apsoil.2022.104468.
Al-Maliki, S., Al-Taey, D. K. & Al-Mammori, H. Z. 2021 Earthworms and eco-consequences: Considerations to soil biological indicators and
plant function: A review.Acta Ecol. Sin. 41 (6), 512–523. https://doi.org/10.1016/j.chnaes.2021.02.003.
Arora, S., Saraswat, S., Mishra, R., Rajvanshi, J., Sethi, J., Verma, A., Nag, A. & Saxena, S. 2020 Design, performance evaluation and
investigation of the dynamic mechanisms of earthworm-microorganisms interactions for wastewater treatment through vermifiltration
technology.Bioresour. Technol. Rep. 12, 100603. https://doi.org/10.1016/j.biteb.2020.100603.
Arora, S., Saraswat, S., Rajpal, A., Shringi, H., Mishra, R., Sethi, J., Rajvanshi, J., Nag, A., Saxena, S. & Kazmi, A. A. 2021 Effect of earthworms
in reduction and fate of antibiotic resistant bacteria (ARB) and antibiotic resistant genes (ARGs) during clinical laboratory wastewater
treatment by vermifiltration.Sci. Total Environ. 773, 145152. https://doi.org/10.1016/j.scitotenv.2021.145152.
Arora, S., Saraswat, S., Kazmi, A. A., 2022 Vermifilter: A biofilter with earthworms for wastewater treatment. In: An Innovative Role of
Biofiltration in Wastewater Treatment Plants (WWTPs) (Shah, M., Rodriguez-Couto, S. & Biswas, J., eds). Elsevier, London, UK,
pp 87–112. https://doi.org/10.1016/b978-0-12-823946-9.00013-9.
Arrázola-Vásquez, E., Larsbo, M., Capowiez, Y., Taylor, A., Sandin, M., Iseskog, D. & Keller, T. 2022 Earthworm burrowing modes and rates
depend on earthworm species and soil mechanical resistance.Appl. Soil Ecol. 178, 104568. https://doi.org/10.1016/j.apsoil.2022.104568.
Bart, S., Amossé, J., Lowe, C. N., Mougin, C., Péry, A. R. & Pelosi, C. 2018 Aporrectodea caliginosa, a relevant earthworm species for a
posteriori pesticide risk assessment: current knowledge and recommendations for culture and experimental design.Environ. Sci. Pollut.
Res. 25 (34), 33867–33881. https://doi.org/10.1007/s11356-018-2579-9.
Bazri, K., Ouahrani, G., Gheribi-Aoulmi, Z. & Díaz Cosín, D. J. 2013 La diversité des lombriciens dans l’Est algérien depuis la côte jusqu’au
désert.Ecol. Mediterr. 39 (2), 5–17. https://doi.org/10.3406/ecmed.2013.1276.
Bora, S., Bisht, S. S. & Reynolds, J. W. 2021 Global diversity of earthworms in various countries and continents: A short review.
Megadrilogica 26 (9), 127–154.
Bouché, M. B. 1972 Lombriciens de France. Ecologie et Systématique. Doctoral Thesis, Univ. Paris-Sud, Orsay, France.
Boudjabi, S., Ababsa, N. & Chenchouni, H., 2023 Sewage and sewage treatment. In: The Palgrave Handbook of Global Sustainability
(Brinkmann, R., ed.). Palgrave Macmillan, Cham, Switzerland, pp 719–745. https://doi.org/10.1007/978-3-030-38948-2_50-1.
Butt, K. R., Gilbert, J. A., Kostecka, J., Lowe, C. N., Quigg, S. M. & Euteneuer, P. 2022 Two decades of monitoring earthworms in translocated
grasslands at Manchester Airport.Eur. J. Soil Biol. 113, 103443. https://doi.org/10.1016/j.ejsobi.2022.103443.
Byrne, L. B. 2022 The essence of soil biodiversity.Conserv. Lett. 15, 412900. https://doi.org/10.1111/conl.12900.
Chen, Y., Liu, X., Leng, Y. & Wang, J. 2020 Defense responses in earthworms (Eisenia fetida) exposed to low-density polyethylene
microplastics in soils.Ecotoxicol. Environ. Saf. 187, 109788. https://doi.org/10.1016/j.ecoenv.2019.109788.
Chenchouni, H. 2017 Variation in White Stork (Ciconia ciconia.) diet along a climatic gradient and across rural-to-urban landscapes in North
Africa.Int. J. Biometeorol. 61, 549–564. https://doi.org/10.1007/s00484-016-1232-x.
Chenchouni, H., Chaminé, H. I., Khan, M. F., Merkel, B. J., Zhang, Z., Li, P., Kallel, A. & Khélifi, N. 2022 New Prospects in Environmental
Geosciences and Hydrogeosciences. Springer, Cham, Switzerland. https://doi.org/10.1007/978-3-030-72543-3.
Cheng, Y., Song, W., Tian, H., Zhang, K., Li, B., Du, Z., Zhang, W., Wang, J., Wang, J. & Zhu, L. 2021 The effects of high-density polyethylene
and polypropylene microplastics on the soil and earthworm Metaphire guillelmi gut microbiota.Chemosphere 267, 129219. https://doi.
org/10.1016/j.chemosphere.2020.129219.
Coleman, D. C. 2013 Soil biota, soil systems, and processes.Encycl. Biodivers. 6. https://doi.org/10.1016/B978-0-12-384719-5.00128-3.
Colwell, R. K. 2013 EstimateS: statistical estimation of species richness and shared species from samples. Version 9. Available from: https://
www.robertkcolwell.org/pages/1407-estimates.
Das, P. & Paul, K. 2023 A review on integrated vermifiltration as a sustainable treatment method for wastewater.J. Environ. Manage. 328,
116974. https://doi.org/10.1016/j.jenvman.2022.116974.
Dhiman, V. & Pant, D., 2022 Bioindication and biomarker responses of earthworms: A tool for soil pollution assessment. In: Advances in
Bioremediation and Phytoremediation for Sustainable Soil Management (Malik, J. A., ed.). Springer, Cham, Switzerland, pp 365–378.
https://doi.org/10.1007/978-3-030-89984-4_23.
Didden, W. A. M. 2001 Earthworm communities in grasslands and horticultural soils.Biol. Fertil. Soils 33, 111–117. https://doi.org/10.1007/
s003740000297.
Ding, J. & Eldridge, D. J. 2022 Drivers of soil biodiversity vary with organism type along an extensive aridity gradient.Appl. Soil Ecol. 170,
104271. https://doi.org/10.1016/j.apsoil.2021.104271.
Dlamini, T. C. & Haynes, R. J. 2004 Influence of agricultural land use on the size and composition of earthworm communities in northern
KwaZulu-Natal, South Africa.Appl. Soil Ecol. 27 (1), 77–88. https://doi.org/10.1016/j.apsoil.2004.02.003.
Edwards, C. A. & Arancon, N. Q. 2022 Biology and Ecology of Earthworms, 4th edn. Springer, New York, NY, USA. https://doi.org/10.1007/
978-0-387-74943-3.
Edwards, C. A. & Bohlen, P. J. 1996 Biology and Ecology of Earthworms, 3rd edn. Chapman and Hall, London, UK, p. 426.
Water Science & Technology Vol 88 No 9, 2487
Downloaded from http://iwaponline.com/wst/article-pdf/doi/10.2166/wst.2023.345/1323509/wst2023345.pdf
by guest
on 17 November 2023

European Commission, Joint Research Centre, Johnson, N., Scheu, S., Ramirez, K., Lemanceau, P., Eggleton, P., Jones, A., Moreira, F.,
Barrios, E., De Deyn, G., Briones, M., Kaneko, N., Kandeler, E., Wall, D., Six, J., Fierer, N., Jeffery, S., Lavelle, P., Putten, W., Singh, B.,
Miko, L., Hedlund, K., Orgiazzi, A., Chotte, J., Bardgett, R., Behan-Pelletier, V., Fraser, T. & Montanarella, L. 2016 Global Soil
Biodiversity Atlas. Publications Office of the European Union, Luxembourg. https://data.europa.eu/doi/10.2788/2613.
Fonte, S. J., Winsome, T. & Six, J. 2009 Earthworm populations in relation to soil organic matter dynamics and management in California
tomato cropping systems.Appl. Soil Ecol. 41, 206–214. https://doi.org/10.1016/j.apsoil.2008.10.010.
Fründ, H. C., Graefe, U. & Tischer, S., 2011 Earthworms as bioindicators of soil quality. In: Biology of Earthworms. Soil Biology, Vol. 24
(Karaca, A., ed.), Springer, Cham, Switzerland, pp 261–278. https://doi.org/10.1007/978-3-642-14636-7_16.
Guedes, P., Martins, C. & Couto, N. 2022 Irrigation of soil with reclaimed wastewater acts as a buffer of microbial taxonomic and functional
biodiversity.Sci. Total Environ. 802, 149671. https://doi.org/10.1016/j.scitotenv.2021.149671.
Hsu, G. C., Szlavecz, K. & Csuzdi, C. 2023 Ecological groups and isotopic niches of earthworms.Appl. Soil Ecol. 181, 104655. https://doi.
org/10.1016/j.apsoil.2022.104655.
Jiang, X., Chang, Y., Zhang, T., Qiao, Y., Klobucar, G. & Li, M. 2020 Toxicological effects of polystyrene microplastics on earthworm (Eisenia
fetida).Environ. Pollut. 259, 113896. https://doi.org/10.1016/j.envpol.2019.113896.
Johnston, A. S., Holmstrup, M., Hodson, M. E., Thorbek, P., Alvarez, T. & Sibly, R. M. 2014 Earthworm distribution and abundance predicted
by a process-based model.Appl. Soil Ecol. 84, 112–123. https://doi.org/10.1016/j.apsoil.2014.06.001.
Kanaujia, K., Trivedi, A., Hait, S., 2020 Applicability of vermifiltration for wastewater treatment and recycling. In: Earthworm Assisted
Remediation of Effluents and Wastes (Bhat, S., Vig, A., Li, F. & Ravindran, B., eds). Springer, Singapore, pp 3–17. https://doi.org/
10.1007/978-981-15-4522-1.
Lemtiri, A., Colinet, G., Alabi, T., 2018 Short-term effects of tillage practices and crop residue exportation on soil organic matter and
earthworm communities in silt loam arable soil. In: Soil Management and Climate Change (Munoz, M. & Zornoza, R., eds), Elsevier,
London, UK, pp 53–71, https://doi.org/10.1016/B978-0-12-812128-3.00005-7.
Lenhart, P. A., Eubanks, M. D. & Behmer, S. T. 2015 Water stress in grasslands: Dynamic responses of plants and insect herbivores.Oikos
124, 381–390. https://doi.org/10.1111/oik.01370.
Liu, T., Chen, X., Gong, X., Lubbers, I. M., Jiang, Y., Feng, W., Li, X., Whalen, J. K., Bonkowski, M., Griffiths, B. S. & Hu, F. 2019 Earthworms
coordinate soil biota to improve multiple ecosystem functions.Curr. Biol. 29 (20), 3420–3429. https://doi.org/10.1016/j.cub.2019.08.045.
Mantoani, M. C., Alhakami, F. T. & Fearon, H. 2022 Gunnera tinctoria invasions increase, not decrease, earthworm abundance and diversity.
Biol. Invasions 24 (12), 3721–3734. https://doi.org/10.1007/s10530-022-02873-9.
Mathieu, J., Caro, G. & Dupont, L. 2018 Methods for studying earthworm dispersal.Appl. Soil Ecol. 123, 339–344. https://doi.org/10.1016/
j.apsoil.2017.09.006.
Mishra, C. S. K., Samal, S., Rout, A., Pattanayak, A. & Acharya , P. 2020 Evaluating the implications of moisture deprivation on certain
biochemical parameters of the earthworm Eudrilus eugeniae with microbial population and exoenzyme activities of the organic
substrate.Invertebr. Surviv. J. 17 (1), 1–8. https://doi.org/10.25431/1824-307x/isj.v0i0.1-8.
Mohan, M., Manohar, M., Muthuraj, S., 2022 Vermifiltration: A novel sustainable and innovative technology for wastewater treatment. In:
Innovations in Environmental Biotechnology (Arora, S., Kumar, A., Ogita, S. & Yau, Y. Y., eds), Springer, Cham, Switzerland, pp 597–
611. https://doi.org/10.1007/978-981-16-4445-0_24.
Monroy, F., Aira, M., Gago, J. Á. & Domínguez, J. 2007 Life cycle of the earthworm Octodrilus complanatus (Oligochaeta, Lumbricidae).CR
Biol. 330 (5), 389–391. https://doi.org/10.1016/j.crvi.2007.03.016.
Nahmani, J., Lavelle, P., Lapied, E. & van Oort, F. 2003 Effects of heavy metal soil pollution on earthworm communities in the north of
France.Pedobiologia 47 (5–6), 663–669. https://doi.org/10.1078/0031-4056-00243.
Namaldi, O. & Azgin, S. T. 2023 Evaluation of the treatment performance and reuse potential in agriculture of organized industrial zone (OIZ)
wastewater through an innovative vermifiltration approach.J. Environ. Manage. 327, 116865. https://doi.org/10.1016/j.jenvman.2022.116865.
Ohgushi, T., Wurst, S., Johnson, S. N., 2018 Current knowledge and future challenges of aboveground and belowground community ecology.
In: Aboveground–Belowground Community Ecology. Ecological Studies, Vol. 234 (Ohgushi, T., Wurst, S. & Johnson, S., eds). Springer,
Cham, Switzerland, pp 345–361. https://doi.org/10.1007/978-3-319-91614-9_15.
Omodeo, P., Rota, E. & Baha, M. 2003 The megadrile fauna (Annelida: Oligochaeta) of Maghreb: A biogeographical and ecological
characterization.Pedobiologia 47(5–6), 458–465. https://doi.org/10.1078/0031-4056-00213.
Otmani, H., Tadjine, A., Moumeni, O., Zeriri, I., Amamra, R., Samira, D. B., Djebar, M. R. & Berrebbah, H. 2018 Biochemical responses of
the earthworm Allolobophora caliginosa exposed to cadmium contaminated soil in the Northeast of Algeria.Bull. Soc. Roy. Sci. Liège.
87,1–12. https://doi.org/10.25518/0037-9565.7331.
Ouahrani, G. 2003 Lombritechniques appliquées aux évaluations et aux solutions environnementales. [Lombritechniques Applied to
Environmental Assessments and Solutions]. Doctoral Thesis, Univ Constantine, Algeria.
Owojori, O. J., Reinecke, A. J. & Rozanov, A. B. 2009 The combined stress effects of salinity and copper on the earthworm Eisenia fetida.
Appl. Soil Ecol. 41 (3), 277–285. https://doi.org/10.1016/j.apsoil.2008.11.006.
Pavlícek, T., Csuzdi, C., 2016 Clitellata: Oligochaeta: Earthworms, Chapter 23. In: An Introduction to the Wildlife of Cyprus (Sparrow, D. J. &
Eddie, J., eds). Terra Cypria, Limassol, Cyprus, pp 587–599.
Pelosi, C., Barot, S., Capowiez, Y., Hedde, M. & Vandenbulcke, F. 2014 Pesticides and earthworms. A review.Agron. Sustainable Dev. 34,
199–228. https://doi.org/10.1007/s13593-013-0151-z.
Water Science & Technology Vol 88 No 9, 2488
Downloaded from http://iwaponline.com/wst/article-pdf/doi/10.2166/wst.2023.345/1323509/wst2023345.pdf
by guest
on 17 November 2023

Philips, H.R. P., Guerra, C. A., Bartz, M. L. C., Briones, M. J., Brown, G., Crowther,T. W., Ferlian, O., Gongalsky, K. B., Van Den Hoogen, J., Krebs,
J. & Orgiazzi, A. 2019 Global distribution of earthworm diversity.Science 366 (6464), 480–485. https://doi.org/1 0.1126/science.aax4851.
Plum, N. M. & Filser, J. 2005 Floods and drought: Response of earthworms and potworms (Oligochaeta: Lumbricidae, Enchytraeidae) to
hydrological extremes in wet grassland.Pedobiologia 49 (5), 443–453. https://doi.org/10.1016/j.pedobi.2005.05.004.
Potapov, A. M., Tiunov, A. V., Scheu, S., Larsen, T. & Pollierer, M. M. 2019 Combining bulk and amino acid stable isotope analyses to
quantify trophic level and basal resources of detritivores: A case study on earthworms.Oecologia 189 (2), 447–460. https://doi.org/10.
1007/s00442-018-04335-3.
Raiesi, F., Motaghian, H. R. & Nazarizadeh, M. 2020 The sublethal lead (Pb) toxicity to the earthworm Eisenia fetida (Annelida, Oligochaeta)
as affected by NaCl salinity and manure addition in a calcareous clay loam soil during an indoor mesocosm experiment.Ecotoxicol.
Environ. Saf. 190, 110083. https://doi.org/10.1016/j.ecoenv.2019.110083.
R Core Team 2022 R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing, Vienna, Austria.
Available from: https://www.R-project.org
Reynolds, J. W. 2018 Preliminary key to Algerian Megadriles (Annelida, Clitellata, Oligochaeta), based on external characters, insofar as
possible. Megadrilogica 2(4), 1–16.
Richardson, J. B., Görres, J. H. & Sizmur, T. 2020Synthesis of earthworm trace metal uptake and bioaccumulationdata: Role of soil concentration,
earthworm ecophysiology, and experimental design.Environ. Pollut. 262, 114126. https://doi.org/10.1016/j.envpol.2020.114126.
Rodriguez-Campos, J., Dendooven, L. & Alvarez-Bernal, D. 2014 Potential of earthworms to accelerate removal of organic contaminants
from soil: A review.Appl. Soil Ecol. 79,10–25. https://doi.org/10.1016/j.apsoil.2014.02.010.
Romo, M. R., Pérez-Martínez, D. & Castillo Ferrer, C. 2016 Innate immunity in vertebrates: An overview.Immunology 148 (2), 125–139.
https://doi.org/10.1111/imm.12597.
Scherer-Lorenzen, M., Gessner, M. O., Beisner, B. E.,Messier, C., Paquette, A.,Petermann, J. S., Soininen, J.& Nock, C. A. 2022 Pathwaysfor cross-
boundary effects of biodiversity on ecosystem functioning.Trends Ecol. Evol. 37 (5), 454–467. https://doi.org/10.1016/j.tree.2021.12.009.
Sekhara-Baha,M.2008Étude bioécologique desoligochètes du Nordde . Doctoral Thesis,INA, El Harrach,Algiers, Algeria. Availablefrom: http://
dspace.ensa.dz:8080/jspui/bitstream/123456789/486/1/Sekhara_M.
Shokouhi, R., Ghobadi, N., Godini, K., Hadi, M. & Atashzaban, Z. 2020 Antibiotic detection in a hospital wastewater and comparison of
their removal rate by activated sludge and earthworm-based vermifilteration: Environmental risk assessment.Process Saf. Environ. Prot.
134, 169–177. https://doi.org/10.1016/j.psep.2019.10.020.
Singh, J., Schädler, M., Demetrio, W., Brown, G. G. & Eisenhauer, N. 2019 Climate change effects on earthworms-a review. Soil Org. 91 (3),
114–138. https://doi.org/10.25674/so91iss3pp114.
Singh, J., Cameron, E. & Reitz, T. 2021 Grassland management effects on earthworm communities under ambient and future climatic
conditions.Eur. J. Soil Sci. 72, 343–355. https://doi.org/10.1111/ejss.12942.
Sinha, R. K., Bharambe, G. & Chaudhari, U. 2008 Sewage treatment by vermifiltration with synchronous treatment of sludge by earthworms:
A low-cost sustainable technology over conventional systems with potential for decentralization.The Environmentalist. 28 (4), 409–420.
https://doi.org/10.1007/s10669-008-9162-8.
Sizmur, T. & Richardson, J. 2020 Earthworms accelerate the biogeochemical cycling of potentially toxic elements: Results of a meta-analysis.
Soil. Biol Biochem. 148, 107865. https://doi.org/10.1016/j.soilbio.2020.107865.
Szederjesi, T. 2017 Earthworms of Crete (Oligochaeta: Lumbricidae, Acanthodrilidae): New records, remarks and biogeographical review.
North-Western J. Zool. 13 (1), 128–135.
Thakur, M. P., Reich, P. B., Hobbie, S. E., Stefanski, A., Rich, R., Rice, K. E., Eddy, W. C. & Eisenhauer, N. 2018 Reduced feeding activity of
soil detritivores under warmer and drier conditions.Nat. Clim. Change 8,75–78. https://doi.org/10.1038/s41558-017-0032-6.
van Capelle, C., Schrader, S. & Brunotte, J. 2012 Tillage-induced changes in the functional diversity of soil biota–A review with a focus on
German data.Eur. J. Soil Biol. 50, 165–181. https://doi.org/10.1016/j.ejsobi.2012.02.005.
Yang, H., Zhou, J., Weih, M., Li , Y., Zhai, S., Zhang, Q., Chen, W., Liu, J., Liu, L. & Hu, S. 2020 Mycorrhizal nitrogen uptake of wheat is
increased by earthworm activity only under no-till and straw removal conditions.Appl. Soil Ecol. 155, 103672. https://doi.org/10.1016/
j.apsoil.2020.103672.
Yang, X., Shang, G. & Wang, X. 2022 Biochemical, transcriptomic, gut microbiome responses and defense mechanisms of the earthworm
Eisenia fetida to salt stress.Ecotoxicol. Environ. Saf. 239, 113684. https://doi.org/10.1016/j.ecoenv.2022.113684.
Zeb, A., Li, S., Wu, J., Lian, J., Liu, W. & Sun, Y. 2020 Insights into the mechanisms underlying the remediation potential of earthworms in
contaminated soil: A critical review of research progress and prospects.Sci. Total Environ. 740, 140145. https://doi.org/10.1016/j.
scitotenv.2020.140145.
Zerrouki, H., Hamil, S., Alili, M., Isserhane, W. & Baha, M. 2022 Contribution to the knowledge of earthworm fauna of Chrea National Park
(Algeria).Ukr. J. Ecol. 12,1–10. https://doi.org/10.15421/2022_348.
Zhou, Y., Liu, X. & Wang, J. 2020 Ecotoxicological effects of microplastics and cadmium on the earthworm Eisenia foetida.J. Hazard. Mater.
392, 122273. https://doi.org/10.1016/j.jhazmat.2020.122273.
Zorn, M. I., Van Gestel, C. A. M. & Morrien, E. 2008 Flooding responses of three earthworm species, Allolobophora chlorotica,Aporrectodea
caliginosa and Lumbricus rubellus, in a laboratory-controlled environment.Soil Biol. Biochem. 40 (3), 587–593. https://doi.org/
10.1016/j.soilbio.2007.06.028.
First received 10 June 2023; accepted in revised form 14 September 2023. Available online 30 October 2023
Water Science & Technology Vol 88 No 9, 2489
Downloaded from http://iwaponline.com/wst/article-pdf/doi/10.2166/wst.2023.345/1323509/wst2023345.pdf
by guest
on 17 November 2023