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Journal of the Marine
Biological Association of the
United Kingdom
cambridge.org/mbi
Research Article
Cite this article: Rekik A, Pagano M, Ayadi H,
Guermazi W, Elloumi J (2023). Spatial and
seasonal changes in microphytoplankton and
ciliate communities in a stressed area of the
southeastern Mediterranean coast (Tunisia).
Journal of the Marine Biological Association of
the United Kingdom 1–15. https://doi.org/
10.1017/S0025315423000462
Received: 27 January 2023
Revised: 14 June 2023
Accepted: 16 June 2023
Keywords:
Ciliates; microphytoplankton; physico-
chemical parameters; seasonal distribution;
southern coast of Sfax
Corresponding author:
Jannet Elloumi;
Email: jannetelloumi@yahoo.fr
© The Author(s), 2023. Published by
Cambridge University Press on behalf of
Marine Biological Association of the United
Kingdom
Spatial and seasonal changes in
microphytoplankton and ciliate communities in
a stressed area of the southeastern
Mediterranean coast (Tunisia)
Amira Rekik1, Marc Pagano2, Habib Ayadi1, Wassim Guermazi1
and Jannet Elloumi1
1
Faculty of Sciences of Sfax, Department of Sciences of Life, Laboratory LR/18ES30 Marine Biodiversity and
Environment, University of Sfax, Street Soukra Km 3.5 –BP 1171 –CP 3000 Sfax, Tunisia and
2
Aix Marseille
University, CNRS/INSU, University of Toulon, IRD, Mediterranean Institute of Oceanography (MIO) UM 110, 13288
Marseille, France
Abstract
The spatial and seasonal variability of the microphytoplankton and ciliates communities in
relation to the environmental factors were studied in the southern coastal area of Sfax.
Results revealed a striking difference between seasons regarding pH, with strong acidification
in autumn generated by industrial activity. Spatial distribution of pH in autumn impacted the
microorganisms in different ways: acidic stations to the south showed significant correlations
with Cyanobacteria, dinoflagellates and loricate ciliates whereas higher pH values in spring
(pH > 8) were linked to diatoms richness. The high availability of inorganic phosphate is
associated with the high release of phosphate due to residue from a phosphate treatment
manufacture along the coast; consequently, N/P ratios were low (1.34–13.43) suggesting
nitrogen limitation. Microphytoplankton abundance shifted from dinoflagellates dominance
in autumn to dominance of diatoms during winter and of Euglenophyceae in summer.
Loricate ciliates accounted for the largest proportion of the ciliates community while aloricate
ciliates were relatively scarce during all seasons. Variability of ciliate community appeared not
directly linked to environmental conditions, but significant positive relationships between
abundance of loricate ciliates and microphytoplankton suggest that these ciliates may feed
on microphytoplankton.
Introduction
Due to their abundance and vital roles, microphytoplankton communities are fundamental to
the functioning and evolution of marine ecosystems. They are the primary producers in the
pelagic marine food web, representing the main pathway for transferring matter and energy
to the higher trophic levels (Ben Salem et al., 2015). Hence the diversity and fluctuations of
microphytoplankton can affect the food web and the ecological functions and thus explicit
knowledge of the structure of this component is major for identifying trophic regimes and
investigating the general functioning of the marine ecosystems (Lagaria et al., 2016).
Moreover, the analysis of the composition, abundance and changes in the frequency of micro-
phytoplankton are informative of the actual and future changes in water quality (Belén Sathicq
et al., 2016). Simultaneously of the most important components of plankton, ciliates are
trophic link between the traditional food chain and microbial food web (Elloumi et al.,
2015). Marine planktonic ciliates are a major, ubiquitous and varied group of protozooplank-
ton (Ying et al., 2013). Their dynamics are closely related to variations in environmental para-
meters (Küppers and Claps, 2012), particularly in coastal ecosystems due to the combination
of marine and land influences, making ciliates useful as indicators of ecosystem health status
(Kchaou et al., 2009; Elloumi et al., 2015). With their fast growth, ciliates react more rapidly to
environmental variations than most other microorganisms (Gong et al., 2005).
Several studies have been undertaken in the southern coast of Sfax regarding the spatial dis-
tribution of plankton assemblages (Rekik et al., 2013a,2015a,2016a,2016b; Ben Salem et al.,
2015,2016; Drira et al., 2016) to compare the spatial and seasonal distribution of dinoflagel-
lates and diatoms along Sfax northern and southern coasts (Rekik et al., 2017a,2017b). The
present study is the first examining the distribution of microphytoplankton and ciliates assem-
blage through sampling these communities simultaneously at high spatial resolution sampling
in the shallow coastal waters south of Sfax during four seasons. It is therefore of interest to
assess to the high impact of human pressure, chiefly by phosphogypsum, on plankton assem-
blages in a stressed ecosystem (Rekik et al., 2015a). Our objectives are (1) to study the spatial
and seasonal distribution of ciliates in relation to microphytoplankton, that constitute one of
their potential prey, in the shallow coastal waters of Sfax, (2) to determine their potential rela-
tionship with environmental factors by using statistical analyses and (3) to determine marine
water quality based on biological parameters as a bioindicator.
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Material and methods
Study site
The southern coast of Sfax, the second largest city in Tunisia
(Figure 1) is marked by salt extraction ponds from solar salter
located over an area of about 1500 ha (COTUSAL) (Kobbi-
Rebai et al., 2013). In addition, phosphogypsum, the residue of
phosphate treatment, has been stored along the coastline at an
uncontrolled dumpsite from the manufacture which produces
phosphoric acid (SIAPE) (Rekik et al., 2012). This coast is subject
to degradation of water quality (Drira et al., 2016), increasing
eutrophication (Kobbi-Rebai et al., 2013), green tides caused by
coastal Ulva rigida replacing the Posidonia oceanica seagrass
beds (Ben Brahim et al., 2013) and thus degrading benthic habi-
tats (Turki et al., 2006). It also suffered over the last two decades
from an important decrease in fish resources that might have
resulted from industrial and urban activities, menacing Tunisia’s
socio-economic resources (Abdennadher et al., 2012). Many stud-
ies have reported the high level of atmospheric pollution (Azri
et al., 2010), marine pollution such as hydrocarbon (Zaghden
et al., 2014), and heavy metal contamination (Serbaji et al.,
2012; Naifar et al., 2018).
Field sampling
Samples for nutrients, microphytoplankton and ciliates were
taken during four one-day campaigns in winter (16 February),
spring (22 May), autumn (11 October), and summer (15 July)
2011 along the southern coast of Sfax. During each campaign,
water samples were collected in 20 stations, divided in to five
transects from coast to open water (Figure 1). The stations were
located at different depths due to different distances off the
coast: S1, S5, S9, S13, and S17 with depth < 0.5 m; S2, S6, S10,
S14, and S18 with depth varying between 0.5 and 3 m; S3, S7,
S11, S15, and S19 with depth varying between 3 and 5 m; S4,
S8, S12, S16, and S20 with depth > 5 m. A total of 80 samples
were collected with a Van Dorn-type closing bottle that was
deployed horizontally and at a depth ranging from 0.5 to 7 m.
Nutriment samples (120 ml) were kept immediately upon collec-
tion at −20°C in the dark. Samples for microphytoplankton were
preserved with acid Lugol solution (at 3%; Parsons et al., 1984)
and alkaline Lugol solution was used for fixation of ciliate samples
(at 5%; Sherr and Sherr, 1993). Samples for microphytoplankton
and ciliates were placed at 4°C in the dark for enumeration. Water
samples for Chlorophyll-a(1 l) and suspended matter (0.5 l) ana-
lyses were filtered by vacuum filtration onto Whatman GF/F and
Whatman GF/C glass fibre filters, respectively, which were then
immediately stored at −20°C.
Physico-chemical variables
Physical parameters (temperature, salinity, and pH) were mea-
sured using a multi-parameter kit (Multi 340 i/SET) immediately
after sampling. Subsamples for the nutrients (nitrite, nitrate,
ammonium, orthophosphate, silicate, total nitrogen, and total
phosphate) were collected in plastic containers of 4.5 ml previ-
ously washed with distilled water. They were analysed with a
Bran and Luebbe type 3 autoanalyzer and concentrations were
determined colourimetrically using a UV-visible (6400/6405)
spectrophotometer (Grasshof, 1983). Analyses were independent.
The automatic analysis system provides fast and accurate analysis
of these nutrients. Although each nutrient is determined in a dif-
ferent way, but the method remains similar. It is used colourime-
try to determine the dosage of each nutrient. Percentages of
dissolved inorganic nitrogen were calculated from [(NO
3
−
+
NO
2
−
+NH
4
+
)/T-N] × 100. Percentages of dissolved inorganic
phosphate were calculated from [PO
4
3−
/T-P] × 100. Suspended
matter concentrations were measured using the dry weight of
the residue after filtration of 0.5 l of seawater onto Whatman
GF/C membrane filters and drying at 60°C during 24 h.
Fig. 1 - B/W online, B/W in print
Figure 1. Location of sampling stations (1–20) in the south coast of Sfax.
2 Amira Rekik et al.
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Ciliates and microphytoplankton enumeration
Sub-samples (50 ml) for microphytoplankton and ciliates count-
ing to estimate the abundance were analysed under an inverted
microscope (Leica) using the Utermöhl method (1958) after
24 h settling. Microphytoplankton and ciliates species counts
were carried out on the entire sedimentation chamber with 40×
magnified. Identification of microphytoplankton species was
made according to various keys (Balech, 1959; Tomas et al.,
1996). Ciliates were identified to genus or species level after the
works of Alder (1999), Petz (1999) and Strüder-Kypke and
Montagnes (2002). The importance value for the different species
was determined by their relative frequency.
Chlorophyll-a
Chlorophyll-awas estimated by spectrometry, after extraction of
the pigments in acetone (90%). The concentrations were then esti-
mated using the equations of SCOR-UNESCO (SCOR-UNESCO,
1966). This method consists of filtering 1 l of sea water by vacuum
filtration onto Whatman GF/F glass fibre filters, without exceed-
ing 400 mmHg to prevent cell breakdown. A pinch of carbonate
magnesium is added to avoid the degradation of pigments in
pheopigments at the end of filtration. Filters are kept in alumin-
ium paper and are dried under vacuum on silica gel during 24 h
and were then conserved at −20°C until the time of extraction.
The pigments extraction is carried out in 90% acetone in the
dark and cold for 5 h. After 10 min of centrifugation at 3500g,
the absorbance is measured using a Jenway spectrophotometer
at 630, 645 and 663 nm.
Data analyses
Means and standard deviations (SD) were reported when appro-
priate. The potential relationships between variables were tested
with Pearson’s coefficient correlation. One-way ANOVA followed
by a post hoc comparison using Tukey’s test was applied to iden-
tify significant differences between seasons.
The variations of phytoplankton and ciliate communities were
investigated using multivariate analysis, specifically Nonmetric
Multidimensional Scaling (NMDS). The mean percentage abun-
dance of the taxa per transect and per seasonal period were square
root transformed before estimation of resemblance using the Bray
Curtis metric. The similarity matrix was then ordinated using
NMDS. A SIMPER (percentage of similarity) analysis was per-
formed to identify the species contributing most to similarity
within and dissimilarity between clusters.
The physico-chemical and biological parameters assessed at 20
stations during four seasons were submitted to a normalized prin-
cipal component analysis (PCA) (Dolédec and Chessel, 1989).
Table 1. Seasonal variation of physical-chemical and biological parameters in the south coast of Sfax (Mean + SD; n= 20)
Variables Autumn Winter Spring Summer Fvalues Pvalues
Physical variables
Temperature (°C) 21.71 ± 0.49 15.55 ± 0.87 26.97 ± 3.25 31.75 ± 0.85 316.45 7.73 × 10
−43
***
Salinity (p.s.u.) 39.43 ± 0.47 36.50 ± 1.98 38.25 ± 1.40 37.05 ± 0.95 19.44 1.88 × 10
−9
***
pH 7.17 ± 0.08 7.80 ± 0.10 8.13 ± 0.29 7.91 ± 0.19 97.14 6.20 × 10
−26
***
Suspended matter (mg l
−1
) 30.08 ± 3.38 38.60 ± 11.09 49.47 ± 11.86 34.58 ± 23.14 6.78 0.00***
Chemical variables
NO
3
−
(μM) 10.39 ± 7.87 7.77 ± 2.77 7.37 ± 2.86 3.35 ± 2.34 8.12 9.28 × 10
−5
***
NO
2
−
(μM) 3.58 ± 2.40 1.54 ± 1.59 0.26 ± 0.09 0.31 ± 0.33 22.98 1.08 × 10
−10
***
NH
4
+
(μM) 6.43 ± 6.12 6.10 ± 1.55 4.53 ± 2.08 3.74 ± 5.10 1.85 0.14
T-N (μM) 34.14 ± 13.51 23.05 ± 5.28 21.12 ± 4.55 18.18 ± 9.62 12.01 1.61 × 10
−6
***
PO
4
3−
(μM) 3.42 ± 1.10 2.51 ± 1.09 11.35 ± 4.75 3.45 ± 3.13 39.31 1.92 × 10
−15
***
T-P (μM) 17.02 ± 4.62 10.61 ± 3.16 29.39 ± 13.37 13.53 ± 9.79 17.84 7.35 × 10
−9
***
N/P ratio 6.24 ± 3.23 13.43 ± 24.72 1.34 ± 0.80 3.00 ± 2.05 3.66 0.01*
Si (OH)
4
(μM) 24.69 ± 21.62 5.90 ± 3.14 30.92 ± 12.00 23.58 ± 21.98 8.37 7.10 × 10
−5
***
Biological variables
Chlorophyll a(mg l
−1
) 11.08 ± 11.67 0.42 ± 0.87 6.95 ± 4.27 0.58 ± 1.66 13.68 3.19 × 10
−7
***
Total microphytoplankton (× 10
2
cells l
−1
) 44.10 ± 41.92 32.75 ± 23.56 50.85 ± 48.45 84.10 ± 57.91 4.86 0.00**
Bacillariophyceae (× 10
2
cells l
−1
) 16.25 ± 22.23 25.35 ± 22.27 26.05 ± 26.86 21.45 ± 14.99 0.84 0.48
Dinophyceae (× 10
2
cells l
−1
) 21.60 ± 26.86 5.95 ± 8.00 16.45 ± 33.28 24.40 ± 32.17 1.81 0.15
Cyanobacteria (× 10
2
cells l
−1
) 1.27 ± 1.14 0.90 ± 1.74 3.03 ± 6.34 0.65 ± 1.22 1.99 0.12
Euglenophyceae (× 10
2
cells l
−1
) 2.05 ± 3.50 0.55 ± 1.31 2.30 ± 4.29 37.60 ± 27.31 33.29 7.61 × 10
−14
***
Dictyochophyceae (× 10
2
cells l
−1
) 0.05 ± 0.22 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 1.00 0.40
Chlorophyceae (× 10
2
cells l
−1
) 0.35 ± 1.34 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 1.35 0.27
Total ciliates (× 10
2
cells l
−1
) 7.15 ± 4.39 9.70 ± 4.99 4.15 ± 3.51 11.00 ± 8.89 5.39 0.00**
Loricate ciliates (× 10
2
cells l
−1
) 5.35 ± 4.31 7.30 ± 4.05 3.05 ± 3.54 9.00 ± 9.15 3.99 0.01*
Naked ciliates (× 10
2
cells l
−1
) 1.80 ± 2.06 2.40 ± 2.13 1.10 ± 1.55 2.00 ± 2.38 1.39 0.25
In the last column, results of one-way ANOVA analysis. *P< 0.05, **P< 0.01, ***P< 0.001, show significant differences among sampled levels.
Journal of the Marine Biological Association of the United Kingdom 3
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Simple log (x+ 1) transformation was applied to data in order to
correctly stabilize variance (Frontier, 1973). These statistical ana-
lyses were performed using Primer 7 software.
Results
Hydrological features
The mean values of physical variables recorded at the 20 sampled
stations are summarized in Table 1. Temperature varied among
stations and seasons (Figure 2 and Table 1). The temperature
was in the range 14–33°C, the lowest values being observed at sta-
tions 6 in winter and the highest at stations 5, 6, 8, and 13 in sum-
mer. At each station, temperature exhibited increasing values
from winter to summer and a slight decline in spring compared
to summer. In winter, the observed temperatures were at their
lowest (Figure 2 and Table 1). Thermal stratification did not
develop because of the shallowness at the sampled stations
(<7 m). Salinity varied from 35.2 in winter (stations 9, 16, and
20) to 40 in autumn (stations 1, 2, 3, 7, 11, 13, and 20) and spring
(stations 11, 14, 16, 18, and 19). The pH values ranged from 7.01
(autumn, station 6) to 8.51 (spring, station 17). Concentrations of
suspended matter varied between 30.08 ± 3.38 mg l
−1
during
autumn and 49.47 ± 11.86 mg l
−1
during spring (Table 1).
Nutrients
NO
3
−
concentration varied between 1.31 and 39.66 μM in the
study area, with the lowest concentration observed in summer
at station 11 and the highest in autumn at station 17 (Figure 3).
Mean values were also higher in autumn (10.39 ± 7.87 μM) than
in summer (3.35 ± 2.34 μM), whereas winter and spring were
intermediate (7.77 ± 2.77 and 7.37 ± 2.86 μM respectively;
Table 1). NO
2
−
,NH
4
+
and total nitrogen (T-N) concentrations
were higher in autumn and winter than that in spring and sum-
mer. Nitrogen appeared mainly in its dissolved inorganic form
(DIN, NO
3
−
+NO
2
−
+NH
4
+
) representing 57.35% of the total nitro-
gen. Orthophosphate and total phosphate concentrations had
almost the same distribution pattern (Figure 3), with low concen-
trations during winter and maximum values during spring
(Table 1). The N/P ratio (dissolved inorganic nitrogen (NO
2
−
+
NO
3
−
+NH
4
+
) to dissolved inorganic phosphate (PO
3
4−
) ratio), ran-
ged from 1.34 in spring to 13.43 in winter (Figure 3). These values
were less than the Redfield ratio (16), suggesting a potential N
limitation. Silicate concentrations ranged from 5.90 ± 3.14 μM
(winter) to 30.92 ± 12.00 μM (spring) (Table 1).
Chlorophyll-a
Average Chl aconcentrations remained <12 mg l
−1
(Table 1), but
exhibited higher values like the maximum (39.40 mg l
−1
) observed
at station 9 in autumn. Meanwhile, Chl awas very low and some-
times undetected in some samples during winter and summer
(Figure 4).
Microphytoplankton
Mean microphytoplankton abundance was the highest in summer
(84.10 ± 57.91 × 10
2
cells l
−1
) and the lowest in winter (32.75 ±
23.56 × 10
2
cells l
−1
)(Figure 5 and Table 1), and displayed signifi-
cant differences from season to season (F= 4.86; df = 80; P< 0.01).
In the present study, 65 microphytoplankton taxa were observed, 25
among them were identified to the species level (Table 2). Diatoms
werethe most species-richgroup with 30 taxa,followed by dinoflagel-
lates with 29 taxa and Cyanobacteria with 3 taxa. Other groups
such as Dictyochophyceae (Dictyocha sp.), Euglenophyceae
(Euglena acusformis) and Chlorophyceae (Merismopedia sp.) were
represented by only one species each. The genus Protoperidinium
(9 taxa) was the most diverse among dinoflagellates and the genera
Lithodesmium,Skeletonema and Synedra (2 taxa) among diatoms
(Table 2). Diatoms were, on average, the most abundant group
throughout the survey period (Table 1), but dinoflagellates and
Euglenophyceae were punctually more abundant in autumn and
summer respectively (Figure 6). Microphytoplankton diversity chan-
ged significantly throughout our study, shifting from the predomin-
ance of diatoms, particularly Grammatophora sp., Navicula sp.,
Coscinodiscus sp., Pinnularia sp., and Bellarochea sp. during the win-
ter and spring, to thatof dinoflagellates represented by Gymnodinium
sp., Prorocentrum gracile,andProtoperidinium steinii in autumn
(Table 2). The highest microphytoplankton abundance observed in
summer (84.10 ± 57.91 × 10
2
cells l
−1
,Table 1), was associated with
Fig. 2 - B/W online, B/W in print
Figure 2. Spatial and seasonal variations of physical variables. Stations (1–20) and
transects (1–5).
4 Amira Rekik et al.
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an important proliferation of Euglenophyceae (37.60 ± 27.31 × 10
2
cells l
−1
,Table 1), with Euglena acusformis accounting for 44.71%
of total microphytoplankton abundance. The dominance of E. acus-
formis was coupled with a low number of microphytoplankton taxa
(only 25 taxa, Figure 5), but no significant correlation was found
between this species and physico-chemical variables.
Ciliates
Ciliate abundance ranged from 0 (stations 1, 7, 12 (spring), and 20
(summer)) to 32 × 10
2
cells l
−1
(station 3, summer) (mean =
8.00 × 10
2
± 3.02 × 10
2
cells l
−1
). The highest ciliate abundance
was recorded in summer and the highest number of ciliate taxa
was observed in winter (34 taxa) (Figure 5). The ciliate commu-
nity consisted of 64 taxa (33 taxa in autumn, 34 taxa in winter,
25 taxa in spring, and 20 taxa in summer) belonging to 32 genera
and 2 groups: loricate ciliates and naked ciliates (Table 3). Loricate
ciliates were the most diversified with 43 taxa and representing
73–82% of total ciliates abundance. The genus Tintinnopsis was
dominant among loricate ciliates (13 taxa), followed by
Codonellopsis and Undella (4 taxa) (Table 3). Loricate ciliates
and total ciliate abundance showed the same temporal and spatial
distribution patterns (Figure 7). Loricate ciliate abundance varied
from 0 to 32 × 10
2
cells l
−1
, with the highest abundance at station
3 in summer, associated with an important reproduction of
Poroecus apiculatus and Tintinnopsis beroidea. High abundances
were also recorded at the same season at station 2 (26 × 10
2
cells
l
−1
,Tintinnopsis aperta) and station 4 (27 × 10
2
cells l
−1
,
Tintinnopsis parvula and Tintinnopsis complex)(Figure 7). Some
loricate ciliates species (among which Tintinnidium balechi,
Fig. 3 - B/W online, B/W in print
Figure 3. Spatial and seasonal variations of chemical parameters. Stations (1–20) and transects (1–5).
Journal of the Marine Biological Association of the United Kingdom 5
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Tintinnopsis beroidea, and Tintinnopsis nana) were omnipresent at
all seasons (Table 3). Naked ciliates abundance varied from 0 to 10.
10
2
cells l
−1
(maximum in summer at station 9), and showed its
highest mean value (2.40 ± 2.13 × 10
2
cells l
−1
) in winter and its low-
est 1.10 ± 1.55 × 10
2
cells l
−1
in spring (Figure 7;Table 1).
Statistical analysis
Non-metric dimensional scaling (NMDS) and similarity (SIMPER)
analyses on microphytoplankton and ciliate species
The NMDS ordination of relative abundances of the microphyto-
plankton species (stress value of 0.16 indicating a strong ordin-
ation) roughly identified four clusters corresponding to the four
seasons (Figure 8A). However, in winter one transect (T5) clearly
distinguished from the four other clusters, mainly due to some
species (Gonyaulax sp., Grammatophora sp., Licmophora sp.,
Bellarochea sp., Gymnodinium sp., and Prorocentrum triestinium)
that explained 62% of the dissimilarity with the four other trans-
ects (T1–T4). Also, in spring two transects differentiate from the
three others: T3 due to Anabeana sp., Coscinodiscus sp., Nitschia
longissimi,Prorocentrum micans,Prorocentrum triestinium,
Navicula sp., and Polykrikos sp. (50% cumulated dissimilarity) and
T5 due to Prorocentrum lima,Navicula sp., Anabeana sp.,
Coscinodiscus sp., P. micans,andProrocentrum triestinium (42%
cumulated dissimilarity). The autumn group (60.45 average similar-
ity) was mainly explained by Gymnodinium sp., Grammatophora sp.,
Navicula sp., Achnanthes sp., and Anabeana sp. that explained 72%
cumulative similarity. The main winter group (70.73 similarity, with-
out T5) was explained by Navicula sp., Grammatophora sp.,
Bellarochea sp., Gymnodinium sp., and Pinnularia sp. (72% cumu-
lated), the main spring group (46.78 similarity, without T3 and T5)
by Navicula sp., Coscinodiscus sp., Euglena sp., and Prorocentrum
Triestinium (62% cumulated), and the summer group (69.23 similar-
ity) by Euglena sp., Navicula sp., Grammatophora sp., Gymnodinium
sp., and Prorocentrum triestinium (75% cumulated).
The NMDS ordination of the relative abundances of ciliate
species (stress value of 0.16 indicating a strong ordination) clearly
identified three clusters corresponding to autumn, winter and
summer transects, showing a relative spatial homogeneity of the
ciliate communities at these three periods, whereas the five trans-
ects of spring were scattered (low similarity: 17.99), suggesting
high spatial variability at this period (Figure 8B). The summer
group had the highest similarity (60.87) mostly explained by
Tintinnopsis aperta,Tintinnopsis beroidea,Poroecus apiculatus,
and Euplotes Charon (72% cumulative). Winter and autumn
groups were less homogeneous (38.52 and 35.07 similarity, respect-
ively) and both highly explained by Tintinnopsis beroidea (>50%).
Relationships between biological and environmental variables
Simple correlation analyses between biological and environmental
variables and between microphytoplankton and ciliate variables
are detailed in Tables S1 and S2, respectively.
The PCA on the mean values per transect of the four seasonal
sets of hydrological (temperature, salinity, pH, suspended matter,
nutrients) and biological (Chlorophyll a, microphytoplankton
groups’abundance and ciliates groups’abundance) variables
(Figure 9) allowed clear discrimination of the four seasonal sam-
pling groups around the F1 and F2 components. The F1 compo-
nent axis (26% of the variance) opposed the autumn sampling
points to the summer sampling points. The formers were charac-
terized by high concentrations of N-nutrients and Chlorophyll a
and by the presence of Dictyochophyceae and Chlorophyceaea,
and the latter by high temperature and pH and by high
Euglenophyceae, diatoms and ciliate densities. The F2 component
axis (23% of the variance) opposed spring points correlated with
pH, temperature, P and Si nutrients to winter points correlated
with loricate and naked ciliates.
Discussion
The current study is the first report concerning the distribution of
microphytoplankton and ciliates assemblage through high spatial
resolution sampling in the shallow coastal waters south of Sfax
during four seasons.
The south coast of Sfax, a typical stressed Mediterranean
coastal zone
Our results allow characterizing the environmental context of a
typical stressed area of the southeastern Mediterranean coast.
The high values of temperature and salinity are in agreement
with other studies performed in arid to semi-arid
Mediterranean areas (Elloumi et al., 2015). A strong acidification
of seawater was observed in autumn with pH values down to 7
(mean = 7.17 ± 0.08), contrasting with the highest pH levels in
spring (8.13 ± 0.29). Such low pH values could reasonably be
attributed to the industrial activity still in operation along the
Fig. 4 - B/W online, B/W in print
Figure 4. Spatial and seasonal variations of chlorophyll-aconcentration. Stations
(1–20) and transects (1–5).
Fig. 5 - B/W online, B/W in print
Figure 5. Seasonal variations of average values of microphytoplankton abundance,
number of microphytoplankton taxa, ciliates abundance, and number of ciliates taxa.
6 Amira Rekik et al.
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Table 2. List and frequency of microphytoplankton species found in the southern coast of Sfax during the study conducted at four successive seasons
Microphytoplankton species (cells l
−1
) Autumn Winter Spring Summer
Cyanophyceae
Anabaena sp. C –C–
Oscillatoria sp. R R R R
Spirulina sp. R –––
Bacillariophyceae
Achnanthes sp. C C –C
Amphiprora sp. ––R–
Amphora sp. R R R –
Bellerochea sp. R C C
Biddulphia sp. R C R R
Climacosphenia sp. R R R –
Cocconeis sp. –RRR
Coscinodiscus sp. –RCR
Diploneis sp. R –R–
Epithemia sp. –R–
Grammatophora sp. C C R C
Gyrosigma sp. ––R–
Leptocylindrus danicus (Cleve, 1889) ––C–
Leptocylindrus sp. ––R–
Licmophora sp. R C R –
Lithodesmium sp. ––R–
Lithodesmium undulatum (Ehrenberg, 1839) R –R–
Navicula sp. C C A C
Nitschia longissima (Ralf, 1861) R –R–
Pinnularia sp. CRR
Plagiotropis sp. ––R–
Pleurosigma sp. R C R R
Rhabdonema sp. C R ––
Rhizosolenia sp. R R ––
Rhizosolenia stolforthii (Cupp, 1943) R ––
Skeletonema costatum (Cleve, 1873) ––R–
Skeletonema sp. ––R–
Striatella unipunctata (Agardh, 1832) –R–R
Synedra sp. R –––
Synedra ulna (Ehrenberg, 1832) R –––
Thalassiosira sp. R –R–
Dinophyceae
Alexandrium sp. ––R–
Amphidinium sp. R R R R
Tripos lineatus (Gomez, 2013) R –––
Dinophysis caudata (Saville-Kent, 1881) R –––
Dinophysis sp. –––R
Gonyaulax sp. –CRR
Gymnodinium marinum (Saville-Kent, 1880) R –––
Gymnodinium sp. A C C C
Gyrodinium sp. R R –R
(Continued)
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south coast (Rekik et al., 2013a). In particular, the phosphate pro-
cessing industries (SIAPE-Sfax) generate very acidic residues
(phosphogypsum) that can result in very low pH values of coastal
marine water: in the Ghannouch-Gabes zone, values lower than
3.5 have been recorded close to the discharge (Ben Amor and
Gueddari, 2016), and values close to or even lower than 7 can
be observed at several kilometres off the coastline (El Kateb
et al., 2018). The high concentration of suspended matter may
be the result of the shallow depth of the sampled area and the
intensity of the dominant winds (southwest and north-east),
which usually provoke not only sediment mixing but also remo-
bilization from the surface deposits (Ben Salem et al., 2015).
The high concentrations of orthophosphate and total phosphate
are associated with an important release of phosphate due to resi-
due from the phosphate processing industries (SIAPE-Sfax) (Ben
Brahim et al., 2010). Additionally, the N/P–DIN to DIP ratio was
highly variable and in average lower than the Redfield ratio (16)
during the four periods. Strong variability in the N/P ratio charac-
terizes coastal ecosystems, particularly under eutrophication condi-
tions, where the high terrestrial inputs of nutrients, tide and
turbulence-driven resuspension cause situations far from the relative
equilibrium found in the open ocean (Ryther and Dunstan, 1971).
Low N/P ratio during our study agrees with the results in the north
Sfax coast before the restoration process (Rekiket al., 2012), also sug-
gesting an overall nitrogen limitation in this stressed coastal zone.
Microphytoplankton community of the south coast of Sfax and
its environmental drivers
Withis the oligotrophic Eastern Mediterranean Sea, the southern
coast of Sfax stands out as a highly productive ecosystem (Rekik
et al., 2015a). The high productivity has been further confirmed
by compiling satellite observations and biogeochemical data,
which reinforce the contrast with the Eastern Mediterranean
Sea (D’Ortenzio and Riberad’Alcalà, 2009;Ayataet al., 2017).
Microphytoplankton assemblages recorded in our study in the
southern coast of Sfax showed some similarities compared to
other coastal environments (Rekik et al., 2013b). A high number
of microphytoplankton taxa (65 species), with a prevalence of dia-
toms species was observed in agreement with previous studies
conducted in the north Sfax coast, during the 2009–2010 period,
showing a comparable number of taxa (70 taxa/90 taxa) at the
surface and the water-sediment interface, respectively (Rekik
et al., 2013b,2015b,2016a). Microphytoplankton abundance
shifted from dinoflagellates dominance in autumn to diatoms
dominance in winter and spring and dominance of
Table 2. (Continued.)
Microphytoplankton species (cells l
−1
) Autumn Winter Spring Summer
Noctiluca sp. ––R–
Peridinium sp. R –RC
Polykrikos sp. R –RR
Prorocentrum compressum (Dodge, 1975) ––R–
Prorocentrum gracile (Schütt, 1895) C –––
Prorocentrum lima (Stein, 1878) R –CR
Prorocentrum micans (Ehrenberg, 1834) ––CR
Prorocentrum triestinium (Schiller, 1918) –CCC
Protoperidinium bipes (Balech, 1974) R –R–
Protoperidinium cerasus (Balech, 1973) ––R–
Protoperidinium conicoides (Balech, 1973) ––R–
Protoperidinium conicum (Balech, 1974) R –––
Protoperidinium depressum (Balech, 1974) ––R–
Protoperidinium globulum (Balech, 1974) ––R–
Protoperidinium minutum (Loeblich III, 1970) –RRR
Protoperidinium sp. R R R R
Protoperidinium steinii (Jorgensen, 1899) C –––
Pyrophacus sp. R –––
Scrippsiella trochoidea (Stein, 1883) R R R R
Euglenophyceae
Euglena acusformis (Schiller, 1925) C R C A
Dictyochophyceae
Dictyocha sp. R –––
Chlorophyceae
Merismopedia sp. R –––
(–) en dash means not detected.
(R) Rare means 0–100 cells l
−1
.
(C) Common means 100–300 cells l
−1
.
(A) Abundant means > 300 cells l
−1
.
8 Amira Rekik et al.
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Euglenophyceae in summer in the southern coast. On the north
coast of Sfax, the microphytoplankton community consisted
mainly of diatoms in autumn and winter, dinoflagellates in spring
and Cyanobacteriae in summer (Rekik et al., 2013b). The varia-
tions of microphytoplankton community were mainly related to
nutrient and environmental parameters. Dinoflagellates were
positively correlated to pH, TN and N/P ratio in autumn. The
important abundance of dinoflagellates in autumn may be
explained by their cell motility allowing them to explore different
depths (Rekik et al., 2017a). The abundance of dinoflagellates in
such polluted situation (low pH, high nutrients) agrees with their
cosmopolitan and less demanding character in terms of environ-
mental conditions compared with other groups (Ben Salem et al.,
2015). In our study, dinoflagellates species composition showed
similarity between the southern coast of Sfax and the Gulf of
Gabes. Some dinoflagellates, such as Gymnodinium,Gonyaulax,
Fig. 6 - B/W online, B/W in print
Figure 6. Spatial and seasonal variations of the abundance of microphytoplankton,
diatoms (diat), dinoflagellates (dino), and other microphytoplankton (other micro).
Table 3. List and frequency of ciliate species found in the southern coast of
Sfax during the study conducted at four successive seasons
Ciliates species (cells l
−1
) Autumn Winter Spring Summer
Loricate ciliates
Acanthostomella norvegica
(Kofoid and Campbell, 1929)
–R––
Amphorellopsis sp. R –R
Ascampbeliella armilla (Kafoid
and Campbell, 1929)
R–––
Ascampbeliella urceolata
(Ostenfeld, 1899)
–RR–
Codonellopsis cylindroconica
(Alder, 1999)
––R–
Codonellopsis obesa (Balech,
1948)
R–––
Codonellopsis pusilla
(Jörgensen, 1924)
R–––
Codonellopsis sp. –R––
Cyttarocylis sp. ––RR
Favella errhenbergii (Claparède
and Lachmann, 1858)
R–R–
Favella serrata (Möbius, 1887) R –––
Favella sp. –R––
Helicostomella sp. R –––
Helicostomella subulata
(Ehrenberg, 1833)
R––R
Metacylis jorgenseni (Cleve,
1902)
R–––
Metacylis sp. R R R –
Ormosella acantharus (Kofoid
and Campbell, 1929)
R––
Ormosella cormicopia
(Campbell, 1929)
–R–R
Petalotricha ampulla (Fol, 1881) –RR–
Petalotricha sp. R –––
Poroecus apiculatus (Cleve,
1899)
R–RC
Proplectella ovata (Jörgensen,
1924)
R––
Rhabdonella amor (Cleve, 1900) R –––
Rhabdonella spiralis (Fol, 1881) –R––
Steenstrupiella steenstrupii
(Claparède and Lachmann,
1858)
––R–
Tintinnidium balechi (Barra de
Cao, 1981)
CRRR
Tintinniopsis campanula
(Ehrenberg, 1840)
–R–R
Tintinnopsis lobiancoi (Daday,
1887)
RRRR
Tintinnopsis amphora (Kofoid
and Campbell, 1929)
––R–
Tintinnopsis aperta (Brandt,
1906)
RRC
Tintinnopsis beroidea (Stein,
1867)
CCRC
Tintinnopsis butschlii (Daday,
1887)
R–––
(Continued)
Journal of the Marine Biological Association of the United Kingdom 9
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Protoperidinium, and Prorocentrum attained high abundance in
stressed areas like in the northern coast of Sfax (Rekik et al.,
2012), the southern coast of the Kerkennah islands (Rekik et al.,
2018), the Kneiss island (Rekik et al., 2017b), and the Gulf of
Gabès (Drira et al., 2008), indicating their tolerance to local envir-
onmental conditions. In fact, they can overcome the lack of nutri-
ents by diversifying their trophic modes (autotrophic,
mixotrophic, and heterotrophic; Jeong et al., 2010). About half
of dinoflagellate species in marine plankton lack chloroplasts
(Sherr and Sherr, 2007). Dinoflagellates comprise a large variety
of toxic species, which can produce many different toxic com-
pounds (Smayda, 1997) that can interfere with recruitment,
growth and viability of an important range of marine organisms
including their competitors (Plumley, 1997). In this study, poten-
tial toxic species such as Protoperidinium depressum (spring),
Protoperidinium steinii (autumn), Dinophysis caudata (autumn),
and Prorocentrum lima (autumn, spring, and summer) were
recorded (Hallegraeff, 1993). The dinoflagellates assemblages
also included high numbers of Gymnodinium, a genus that was
reported to occur under high phosphate loading (Daly-Yahia
Kéfi et al., 2005). However, in this study high Gymnodinium
abundance occurred under low phosphate and high nitrogen con-
centrations, suggesting that the reproduction of dinoflagellates
was mainly nitrogen-driven (Rekik et al., 2015a). High density
of diatoms in winter and spring may be due to their quick growth
Table 3. (Continued.)
Ciliates species (cells l
−1
) Autumn Winter Spring Summer
Tintinnopsis complex (Stein,
1867)
–RRC
Tintinnopsis fimbriata (Meunier,
1919)
R–––
Tintinnopsis nana (Lohmann,
1908)
RRRR
Tintinnopsis parva (Merkle, 1909) R –––
Tintinnopsis parvula (Jörgensen,
1912)
RR–R
Tintinnopsis sp. –R––
Undella claparedei (Daday, 1887) ––R–
Undella hemisphaerica
(Laackmann, 1910)
–R––
Undella hyalina (Daday, 1887) ––R–
Undella sp. –R––
Naked ciliates
Aspidisca lynceus (Ehrenberg,
1830)
R––R
Aspidisca sp. –R––
Balanion sp. –R––
Enchelyodon laevis
(Quennerstedt, 1869)
R–––
Euplotes charon (Müller, 1786) –RRR
Halteria sp. (Claparede and
Lachmann, 1853)
–RR–
Leegaardiella sol (Lynn and
Montagnes, 1988)
RRRR
Lohamaniella oviformis
(Leegaard, 1915)
RRR–
Mesodinium sp. R –––
Monodinium balbianii
(Fabre-Domergue, 1888)
––R–
Philasterine sp. –R––
Pleuronema crassum (Dujardin,
1841)
R–––
Strobilidium sp. R –RR
Strombidium acutum (Leegaard,
1915)
–R–R
Strombidium capitatum
(Leegaard, 1915)
–R––
Strombidium chlorophilum
(Montagnes et al., 1988)
–R––
Strombidium compressum
(Leegaard, 1915)
R–––
Strombidium conicum (Wulff,
1919)
RR––
Strombidium dalum (Lynn et al.,
1988)
–R––
Strombidium sp. R –RR
Tiarina fusus (Claparède and
Lachmann, 1857)
R–R–
Uronema marinum (Dujardin,
1841)
RR–R
(–) en dash means not detected.
(R) Rare means 0–100 cells l
−1
.
(C) Common means 100–300 cells l
−1
.
Fig. 7 - B/W online, B/W in print
Figure 7. Spatial and seasonal variations of the abundance of ciliate (cil), loricate
ciliate (loricate cil) and naked ciliates (naked cil).
10 Amira Rekik et al.
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capacity under turbulent and high nutrient conditions (Maranon
et al., 2012). Diatoms are known to be opportunistic organisms
(Fogg, 1991) having fast growth due to rapid nitrogen uptake
(Lomas and Glibert, 2000). These large species (Navicula (95
μm), Coscinodiscus (160 μm) and Leptocylindrus (150 μM)) are
characterized by a high tolerance to various environmental para-
meters and physical stress characteristic of shallow coastal ecosys-
tems, especially during spring blooms (Lomas and Glibert, 2000).
During our survey the south coast of Sfax, Euglenophyceae, repre-
sented by one species, Euglena acusformis, displayed their highest
abundance in summer (45% of the total microphytoplankton
abundance) but in previous surveys, high abundance of this spe-
cies was also recorded in winter in the same area (Ben Salem et al.,
2015). Euglena acusformis has been recognized as the most
opportunistic and saprobiontic species which assimilate lots of
organic matter and might be an indicator of pollution (Barrera
et al., 2008). Because of their high surface to volume ratio,
small cells like Euglena acusformis incorporate nutrients at low
energy cost (Agawin et al., 2000) and thus outperform large
cells (Sin and Wetzel, 2000). In our study Cyanobacteria were
not well represented in the microphytoplankton community but
reached their maximal density in Spring mainly due to the
nitrogen-fixing Anabaena sp., consistently with the lowest N/P
ratio, indicating N limitation at this period (Table 1). On the
north coast of Sfax, the main period for Cyanobacteriae growth
was in summer with the dominant opportunistic and nitrogen-
fixing species Trichodesmium erythraeum (Rekik et al., 2013b)
which can form important blooms in the gulf of Gabès during
Fig. 8 - Colour online, B/W in print
Figure 8. Non-metric dimensional scaling analyses (NBMDS) on mean values per sampling transects (T1–T5) and per season of abundance percentages of phyto-
plankton species (A) and ciliate species (B).
Journal of the Marine Biological Association of the United Kingdom 11
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warm periods (Hamza et al., 2016) and which is known to dom-
inate the microphytoplankton community in the oligotrophic Sea
(Nausch et al., 2008).
Ciliate community of the south coast of Sfax and its
environmental drivers
A total of 64 ciliates taxa representing 46 different genera were
identified during this study, few species of which could be char-
acterized as rare, found at only one or two stations. The species
number reported here is lower than the one reported by Rekik
et al.(2015c) in the northern coast of Sfax (Tunisia), over four
seasons (40 planktonic species at the surface and 53 at the bot-
tom). The divergence in terms of species number between the
north and the south coast of Sfax is probably partly due to differ-
ence in terms of sampling efforts applied to each study. Another
reason may be the mixing of planktonic and benthic ciliates (thus
increasing the number of sampled species) due to the shallow
water depth of the south coast of Sfax. Ciliates community
demonstrated a clear temporal pattern: high abundance values
in winter and summer with an obvious peak at station 3 in sum-
mer, low abundance values in spring and autumn. Ciliates are
dominant in southern coast of Sfax during summer as shown
by the maximal abundance recorded for a wide range of ciliates
(20 taxa) belonging to different size classes with different feeding
strategies (autotrophic, heterotrophic, and mixotrophic ciliates).
The ciliates community was dominated by loricate ciliates while
aloricate ciliates were relatively rare, as also reported in the
north coast of Sfax (Rekik et al., 2015c), the Gulf of Gabès
(Hannachi et al., 2009; Kchaou et al., 2009), the Adriatic Sea
(Bojanićet al., 2005), and the Yellow Sea (Jiang et al., 2011).
The high abundance of loricate species is probably due to the
eutrophic conditions in these marine areas, since these species
might possess a higher adaptability to eutrophic environments
than other ciliates (Bojanićet al., 2005). Some loricate ciliates spe-
cies, such as Favella ehrenbergii,Helicostomella subulata,
Tintinnopsis beroidea,Tintinnopsis campanula, and T. lobiancoi,
reach high abundances in highly anthropized marine coastal
areas, showing their tolerance to environmental stress (Rekik
et al., 2015a). However, there are some exceptions to this compos-
ition pattern in some marine regions where aloricate species dom-
inate the ciliate community; this is typical for many coastal and
oceanic waters, such as the Irish Sea (Edward and Burkill, 1995),
the Irminger Sea (Montagnes et al., 2010), the Baltic Sea
(Mironova et al., 2009), and the North Sea (Stelfox-Widdicombe
et al., 2004).
In this study, we found no significant correlations between
environmental factors and the abundance of the ciliates commu-
nity. The same results were reported by Gong et al.(2005). The
water temperature and various inorganic nutrients might not dir-
ectly control the structure and dynamics of the ciliates commu-
nity but indirectly influence it via food availability. For instance,
the dominance of the agglutinated species Tintinnopsis beroidea
was shown to be related to the availability of particles to construct
the lorica in addition to the presence of its preferred food
(Cyanobacteria) (Rakshit et al., 2015).
Relationships between ciliate and microphytoplankton
communities in the south coast of Sfax
Loricate ciliates are known to have significant relationships with
microphytoplankton groups suggesting a close ecological link to
Fig. 9 - Colour online, B/W in print
Figure 9. Principal component analysis (PCA) (axis I and II) of mean values per sampling transects (T1–T5) and per season abundance and selected environmental
variables.
12 Amira Rekik et al.
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this type of food and revealing further insights into the ecological
role of ciliates as grazers on microphytoplankton, especially in
autumn and summer when microphytoplankton is very abundant
(Yang et al., 2015). The seasonal distribution of microphytoplankton
and ciliates (dominated by the genus Tintinnopsis) suggests that cili-
ates community consume nanophytoplankton but also microphyto-
plankton, as shown in the north coast of Sfax (Rekik et al., 2012).
Significant relationships between loricate ciliates and microphyto-
plankton community abundance have been found in previous studies
in Tunisian coastal areas such as the Gulf of Gabes (Hannachi et al.,
2009). In our study, there were significant correlations between
loricate ciliates and Cyanobacteria (r= 0.85, n=20, P< 0.05) in
autumn and between loricate ciliates and dinoflagellates (r=0.56,
n=20, P< 0.05) in summer. On the other hand, dinoflagellates
may also be in direct feeding competition with ciliates for food.
Indeed, Protoperidinium is a heterotrophic genus known to feed
exclusivelyon diatoms (Sherrand Sherr, 2007)and several otherdino-
flagellates (including the genera Gymnodinium,Gyrodinium,
Gonyaulax,Tripos,andAlexandrium) are considered as grazers,
since most of them were previously shown to be mixotrophic
(Stoecker, 1999). This competition for foodbetween ciliates and dino-
flagellates may constitute another hypothesis explaining their simul-
taneous presence and the correlations recorded between them.
However, loricate ciliates rarely control the abundance or compos-
ition of their prey, as their aggregate feeding activity usually equates
to clearing a maximum of 1–2% per day of the surface layer waters
they occupy (Dolan et al., 2013).
Conclusion
The present study indicates that the environmental properties of
the southern coast of Sfax have typical characteristics of a stressed
area. The microphytoplankton community is highly tolerant and
dependant on environmental variables in particular pH and
nutrient availability. Diatoms are dominant in winter and spring
taking advantage of their high growth capacity. Dinoflagellates
dominate in autumn in low pH condition showing their high tol-
erance to environmental stress. Euglenophyceae are the most
numerous in summer in the lowest nutrient condition, may be
due to their high surface to volume ratio favouring nutrient
assimilation at low energy cost. In contrast with current observa-
tions in the open Mediterranean Sea the ciliate community of the
southern coast of Sfax is dominated by loricate ciliates (mostly the
genera Tintinnopsis,Codonellopsis, and Undella) which are more
abundant than naked ciliates. Ciliate abundance and community
structure is highly variable between seasons but this variability
seems not directly driven by environmental variables but indir-
ectly through dependence on prey availability, resulting in a
tight coupling with microphytoplankton community. Ciliates
should exert a top-down control on microphytoplankton but
the importance of mixotrophic and heterotrophic dinoflagellates
(known to feed on diatoms) also suggests a feeding competition
with this group.
At present, the phosphogypsum restoration had been acutely
necessary allowing microphytoplankton and ciliate species to
take optimal advantage of niche opportunities, which, in turn,
improve water quality along the southern coast.
Supplementary material. The supplementary material for this article can
be found at https://doi.org/10.1017/S0025315423000462.
Acknowledgements. This work was supported by the Taparura Project con-
ducted in the Laboratory LR/18ES30 Marine biodiversity and environment at
the University of Sfax. We have obtained permits for sampling and observation
field studies from the Taparura Project.
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