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Contrasting Effects of an Alien Worm on Benthic N Cycling in Muddy and Sandy Sediments

Department of Life Sciences and Biotechnology, University of Ferrara, 44121 Ferrara, Italy
Department of Chemistry, Life Sciences and Environmental Sustainability, University of Parma, 43124 Parma, Italy
Marine Science and Technology Center, Klaipeda University, 92294 Klaipeda, Lithuania
EA 4592 Géoressources et Environnement, ENSEGID, 33607 Pessac, France
UR EABX, Centre de Bordeaux, Irstea, 33612 Cestas, France
Author to whom correspondence should be addressed.
Water 2019, 11(3), 465;
Original submission received: 5 February 2019 / Revised: 24 February 2019 / Accepted: 28 February 2019 / Published: 5 March 2019
(This article belongs to the Special Issue The Role of Macrobiota in Aquatic Nutrient Cycling)


The North American oligochaete Sparganophilus tamesis is widespread in European freshwaters. Its ecological effects on benthic nitrogen (N) biogeochemistry were studied in two contrasting environments: the organic-rich muddy sediments of the eutrophic Mincio River (Italy) and the organic-poor sandy sediments of the oligotrophic Cazaux-Sanguinet Lake (France). Oxygen and inorganic N fluxes and denitrification rates (IPT) were measured by dark incubation of intact cores with different worm biomass. Sediment oxygen demand and denitrification were higher in muddy than in sandy sediments; however, at the two sites, bioturbation by the oligochaetes stimulated differing microbial O2 and NO3 respiration and NH4+ production. In particular, the relative effect of S. tamesis on sediment metabolism was greater in Cazaux-Sanguinet Lake than in the Mincio River. As a result, S. tamesis favored net N loss in the Mincio River, whereas it increased NH4+ recycling and lowered denitrification efficiency in the Cazaux-Sanguinet Lake. Our results suggest that the effects of S. tamesis on N biogeochemistry might differ depending on local trophic settings. These results have implications for the conservation of isoetids in the French Lake, whose persistence can be menaced by oligochaete-induced nutrient mobilization.

1. Introduction

In aquatic ecosystems invasive species have by default a negative connotation, with very limited exceptions in the literature [1]. Invasion by alien species may affect the interactions of species within communities and the cycling of energy and matter within ecosystems and produce a cascade of consequences and sequential shifts from pristine conditions [2]. This might be the case for bioturbating fauna, which may support both bacteria and primary producers by mobilizing refractory or scarcely bioavailable (e.g., deep and buried) organic matter, adding new nutrient input to the system [3]. If this can be true in well-preserved ecosystems, the invasion of heavily impacted ecosystems by alien species can paradoxically produce unexpected trajectories [1].
Invasive macrofauna, (able to colonize and spread, even under oligotrophic conditions), may determine a large nutrient mobilization within the sediments and from the sediments to the water column [4]. It is difficult to predict long-term net effects of alien species on biogeochemical functioning and on communities [5]. Generally, the highest impact takes place within the early invasion phase, when alien species spread and reach peak density as a result of a strong competition capacity or the abundance of resources [6]. Later on, pristine communities tend to reorganize and contrast the invasion, resulting in a decrease of density of the invader and a trajectory back to the original condition. This may require different time spans, depending on a large number of factors. In aquatic ecosystems, one of the most studied invasive species is Dreissena polymorpha, due to the impressive densities of this reef-forming filter feeder and its capacity to invade a wide range of environments, from plankton-rich eutrophic to oligotrophic environments [7,8,9,10]. D. polymorpha was studied due to its supposed capacity to control and reverse eutrophication, an apparent paradox for an alien species [11]. Specific hypotheses postulated the reduction in plankton biomass due to high filtration rates, increase of water transparency, increase of benthic production, and a regime shift that reverses a turbid status to a transparent one [12]. This trajectory was only partially verified, as multiple analyzed systems revealed that oligotrophication might be short-term or invalid in relatively deep systems where Dreissena has limited access to phytoplankton or where excreted nutrients stimulate new algal growth [13,14]. Another interesting example of the biogeochemical effects produced by an alien species in a marine environment is that of Marenzelleria spp. in the Baltic Sea. This invasion was studied due to the critical conditions (e.g., large suboxic areas) of the Baltic area and to the contrasting effects of this species’ production of nutrients. It was demonstrated that Marenzelleria spp. inhibited denitrification and stimulated the dissimilative nitrate reduction to ammonium (DNRA). The latter process is negative for a eutrophic ecosystem as it recycles ammonium. However, Marenzelleria spp. enhanced the oxidation of sediments and the long-term retention of phosphorus, which may favor the recolonization of pristine macrofauna. These effects may contribute to the recovery of impacted, anoxic, and poorly biodiverse sea bottom [4,15,16,17,18]. Macrofauna may produce contrasting effects on nutrient cycling, as it may stimulate permanent or temporal retention of nutrients, or loss or favor their mobilization [1]. In eutrophic environments, retention and losses are preferred over recycling (to contrast excess carbon fixation, infilling, and so on) [19]. In oligotrophic settings, macrofauna communities contribute to the slow cycling of elements and support the activity of primary producers [20].
In this study, we analyzed the effect of an alien worm in two freshwater environments: the eutrophic Mincio River (northern Italy) and the oligotrophic Cazaux-Sanguinet Lake (Atlantic coast of France). Sparganophilus tamesis is native in North America. It is now spreading all over Europe in a wide range of shallow environments generally colonized by macrophytes, but with variable trophic levels and sedimentary features [21]. We analyzed the effect of S. tamesis on rates of benthic respiration and nitrogen dynamics, hypothesizing a net stimulation of N2 production in the eutrophic site and N recycling in the oligotrophic site. The latter hypothesis is supported by the fact that this invasive worm may exploit refractory fragments of macrophytes as nutrient source, scattered within the upper sediment horizon. As such, it may mobilize N, otherwise associated and buried as particulate nitrogen (PN), and favor the displacement of N–PN (from the sediment to the water column) through burrow ventilation in the ammonium (NH4+) form. Furthermore, we hypothesized that there would be a limited stimulation of denitrification at the oligotrophic site due to low organic pools in sediment and low nitrate (NO3) in the water column, but with a high excretion and release of NH4+. Whereas ammonium mobilization by S. tamesis can produce negative consequences in nutrient-poor environments, the stimulation of processes such as denitrification might be positive in nitrate-rich environments. Therefore, the main aim was to compare the role of S. tamesis in two different freshwater environments in order to understand if a system could benefit from the invasion by an alien species.

2. Materials and Methods

2.1. Sampling Procedure and Sediment Characterization

Water, sediments, and oligochaetes were collected from two sites: a branch of the Mincio River in proximity of Goito (MN, northern Italy) and a littoral zone of the Cazaux-Sanguinet Lake (Atlantic coast of southern France). Both the sampling sites were shallow (~50 cm) with transparent water. The Italian site was characterized by a high nutrient concentration in the water column and muddy sediment, whereas Cazaux-Sanguinet Lake was a nutrient-poor and soft water environment with predominantly sandy sediments [22].
Intact sediment cores (n = 12 for Mincio and n = 16 for Cazaux-Sanguinet) were collected by means of plexiglass liners (inner diameter = 4 cm, height = 20 cm) vertically inserted into the sediment in order to have nearly 12 cm of sediment and nearly 8 cm of a water phase. Individuals of S. tamesis collected from the same sites were then added to the cores in order to have variable biomass: 0 gdw m−2 (n = 3), 35 ± 7 gdw m−2 (n = 3), 55 ± 11 gdw m−2 (n = 3), 65 ± 3 gdw m−2 (n = 3) for the Mincio River and 0 gdw m−2 (n = 4), 45 ± 13 gdw m−2 (n = 4), 65 ± 8 gdw m−2 (n = 4), 85 ± 4 gdw m−2 (n = 4) for the French site [21]. Nearly 50 L of in situ water was collected for preincubation and incubation procedures.
Four additional plexiglass liners (inner diameter = 4 cm, height = 20 cm) were collected from the sites for sediment characterization. The upper sediment layer (0–10 cm) was extruded with a piston and homogenized with a spatula. A sub-sample of 5 mL of fresh sediment was dried at 70 °C for 48 h for density, porosity, and organic matter analyses. The homogenized sediment was collected by means of a cut-off 5 mL syringe. Bulk density was measured as the weight of a volume of 5 mL fresh material, and porosity was calculated after drying at 70 °C until reaching a constant weight. Organic matter content (OM) was measured as a percentage of weight loss on ignition (450 °C, 2 h) from dried, powdered sediment.

2.2. Incubation Setup and Measurement of Benthic Fluxes

Once collected, water and intact sediment cores added with oligochaetes were transferred to the laboratory within two hours. Cores were submersed with the top open in a large incubation tank that contained in situ water that was well-mixed and aerated and maintained at ambient temperature (24 ± 0.5 °C). One week after the sampling, all cores were closed, and dissolved gas and nutrient fluxes were measured in the darkness. The incubation procedure was standard, with initial and final samplings from each core water phase as detailed in [23]. Incubations lasted 2 h for Mincio sediment cores and 3 h for the French sediment cores, and incubations started when a gas-tight lid with a sampling port and a compensation valve was positioned on the top of the liners. A Teflon-coated stirring bar gently mixed the water inside each liner to avoid stagnation and to guarantee homogeneous conditions within the cores. At the beginning and at the end of the incubations an aliquot of water was transferred to a 12 mL glass vial (Exetainer®, Labco Limited, High Wycombe, UK) and fixed with 100 µL of 7 M ZnCl2 for O2 analysis by means of Membrane Inlet Mass Spectrometer (MIMS, Bay Instrument, sensitivity 0.2 µM). In addition, an aliquot of 20 mL was filtered (Whatman GF/F glass fiber filters) and transferred to a plastic vial for NH4+, NO3, and nitrite (NO2) analyses performed with standard spectrophotometric techniques [24,25]. Fluxes were calculated according to the equation below:
F l u x   x = ( [ x ] f [ x ] i ) × V A × t ,
where xf and xi, expressed in µM or mM, are the concentrations of the solute x at the end and at the start of the incubation, respectively, V (L) is the volume of the core water phase, A (m2) is the area of the sediment, and t (h) is the incubation time.
The top lids were thereafter removed, and the water in the tank was replaced with fresh in situ water. In the afternoon, a sequential incubation was performed that aimed at measuring the denitrification rates with the isotope pairing technique (IPT) [26]. Briefly, 0.1 mL (Cazaux-Sanguinet) and 0.5 mL (Mincio) of a 20 mM 15NO3 stock solution was added to the water phase of each liner to reach 10 and 50 µM final concentrations of labelled nitrate at the oligotrophic and eutrophic sites, respectively. The top lids were then positioned and the cores were incubated in the dark for 3 h. At the end of the incubation the lids were removed, and the whole sediment and water phase was gently mixed to create a slurry, which was subsampled and transferred to the Exetainers, then poisoned with 200 µL of 7 M ZnCl2 for labelled N2 analysis by means of MIMS. At the end of the procedure the cores were sieved in order to check for the occurrence of other macrofauna and to retrieve the oligochaetes. The revised version of the IPT was not used at the sampling sites, as sediment slurries demonstrated the absence of anammox (Benelli unpublished). The rates of denitrification were calculated according to the equations and assumptions of [26]: D15 = p(15N14N) + 2p (15N15N) and D14 = p(15N14N) + 2p(14N14N), where D15 and D14 were equal to the rates of denitrification based on 15NO3 and 14NO3, respectively; and p(14N14N), p(15N14N), and p(15N15N) were equal to the rates of production of labelled and unlabeled N2 species. Because the p(14N14N) cannot be readily measured, estimation of D14 was obtained from: D14 = D15 × p(15N14N)/2p(15N15N). The proportion of D14 supported by unlabeled NO3 from the water column (DW) was calculated from: DW = D15 × f/(1 − f), where f is a mole fraction of 14NO3 in the water column. The coupled nitrification–denitrification (DN) was calculated as the difference: DN = D14 − DW.
Individuals of S. tamesis retrieved from the cores were analyzed for the wet (gww) and dry weights (gdw, after drying the soft tissue at 70 °C to a constant weight).
The sum of the fluxes of inorganic N forms was calculated in order to estimate the denitrification efficiency (DE) that is calculated as:
D E = D t o t D I N f l u x e s + D t o t × 100 ,
where Dtot is the sum of DW and DN, and DINfluxes is the sum of NH4+, NO3, and NO2 net fluxes (only values >0 are considered; when DINfluxes are negative DE is 100%). Denitrification efficiency represents the fraction of mineralized N that is released to the water column as N2. When DE is 100% it suggests tightly coupled ammonification, nitrification, and denitrification and no inorganic N efflux.

2.3. Statistical Analyses

Differences between sedimentary features and fluxes at the two sites were tested by one-way ANOVA. As process rates at the two sites were markedly different, in order to compare the site-specific effect produced by increasing worm biomass on benthic processes, an enhancement factor was calculated by dividing the rates measured in the bioturbated sediments by the rates measured in control sediments. The enhancement factor, representing the relative increase of different processes along with variable S. tamesis biomass, was analyzed via linear regression; the obtained slopes were compared with a t-test.

3. Results

3.1. Sedimentary Features

At the two sites, inorganic nitrogen concentration (DIN), sediment density, porosity, and organic matter content were significantly different (one-way ANOVA, p < 0.001). In the Mincio River, sediments were soft, muddy, and very organic, whereas the sandy sediment of Cazaux-Sanguinet Lake displayed a high density, low porosity, and low organic matter content (Table 1). The Mincio River sediments appeared dark and chemically reduced, with a sharp redox discontinuity a few mm below the interface, whereas those of the Cazaux-Sanguinet Lake were light brown and homogeneously oxidized along the upper 10–15 cm depth profile.

3.2. Benthic Fluxes Along Increasing Sparganophilus tamesis Biomass

Microbial respiration rates measured in sediments without macrofauna were significantly higher in the Mincio River, likely due to higher organic content (one-way ANOVA, p < 0.001). At both sites, the addition of increasing S. tamesis biomass resulted in increased sediment O2 demand (SOD) (Figure 1).
Denitrification rates measured with the IPT are reported in Figure 2. N2 production increased in the four treatments along with increasing S. tamesis biomass at both sites. Compared to the aerobic respiration, denitrification rates were high in the Mincio sediments (Figure 2a), whereas they were very low in the sandy sediments of Cazaux-Sanguinet Lake (Figure 2b), even with the highest biomass of oligochaetes. In the four treatments, denitrification supported by nitrification averaged 82 ± 22 and 0.38 ± 0.15 µmol m−2 h−1 in Mincio sediments and Cazaux-Sanguinet sediments, respectively, without significant differences between control and bioturbated sediments. In Cazaux-Sanguinet Lake, DN represented nearly 50% of total denitrification in control sediments, whereas this share was reduced to 6% in the highest biomass treatment.
Comparing the effect produced by the worm on the processes measured at the two sites, different enhancement factors were calculated for SOD and total denitrification (DW + DN) (Figure 3). At both sites, increasing worm biomass produced a higher relative effect on the nitrate-based than on the aerobic respiration. Furthermore, the stimulatory effect of S. tamesis on both processes was relatively higher in the Cazaux-Sanguinet Lake than in the Mincio River (t-test, SOD p < 0.001, Dtot p < 0.05).
Measured fluxes of the dissolved inorganic N forms had different trends, depending on the S. tamesis biomass (Figure 4). In the Mincio River, bare sediments acted as a source of NH4+ and NO3. Ammonium fluxes were augmented along increasing oligochaete biomass, whereas the presence of the worms reversed NO3 fluxes, which turned highly negative (Figure 4a). In Cazaux-Sanguinet Lake, in the absence and presence of the low oligochaete biomass, sediments acted as a sink for all N forms, whereas in sediments with the worm biomass equal or higher than 65 gdw m−2 the fluxes were reversed and NH4+ and NO3 were regenerated to the water column (NH4+ > NO3) (Figure 4b). Nitrite fluxes were low and negative in all treatments.
The mean value of DE in the four treatments increased along with the increase in the biomass of oligochaetes in Mincio sediments (Figure 5a), whereas the DE value decreased along with the increase in the biomass of oligochaetes in Cazaux-Sanguinet Lake, as a result of increasing and positive NH4+ fluxes (Figure 5b).

4. Discussion

We have investigated the biogeochemical effects of an alien, plastic species that is abundant in the contrasting environments of the eutrophic Mincio River and the oligotrophic Cazaux-Sanguinet Lake [21]. Our results suggest that S. tamesis produced opposite effects at the two sites on benthic N cycling. Such effects might be considered as positive (e.g., increased N removal from nutrient-enriched Mincio) or negative (e.g., increased N recycling in the oligotrophic Cazaux-Sanguinet) for the invaded ecosystem. Analogous results are reported after the invasion of the bivalve D. polymorpha, whose ecosystem consequences can be opposite and site-specific (e.g., spanning from the control of algal blooms to their enhancement) [14,27]. Also, for the invasive worm Marenzelleria spp., different authors report contrasting ecosystem level effects, depending upon the invaded site and the considered temporal scale [1,15,16].
S. tamesis was found at both sites in proximity to native macrophytes: Vallisneria spiralis in Italy [28] and Lobelia dortmanna in the Cazaux-Sanguinet Lake [21]. The close association with macrophytes might depend on the favorable chemical conditions promoted by roots via radial oxygen loss [29] that may increase the survival of worm cocoons. A large plasticity is suggested for this species [21] that allows successful invasion in dramatically different sedimentary environments compared to those from the study sites. Different food availabilities or qualities locally affect the worms’ growth, as adults retrieved from the two areas differ in size: they were larger in muddy sediments and smaller in sandy sediments. The two sites displayed a very different sediment oxygen demand, which was higher in Mincio, as muddy sediments were organic-rich and chemically reduced, and was lower in the organic-poor and oxidized sandy sediments at Cazaux-Sanguinet. The presence of oligochaetes increased O2 consumption at both sites because of the worms’ direct metabolic contribution and indirect stimulation of microbial or chemical processes [30,31]. However, the degree of stimulation was different and larger at the sandy site. We calculated that a worm biomass of 100 gdw m−2 might stimulate sediment oxygen demand by a factor of 1.8 in sandy sediments, compared to an enhancement factor of 0.7 calculated for muddy sediments. Such a difference could be due to different porosities and solute transports in sandy VS muddy sediments.
Similarly, rates of total denitrification were different and two orders of magnitude higher at the Mincio site. Such a difference is likely a combination of higher organic carbon and higher NO3 concentrations in the water column of the Mincio site, resulting in elevated N2 production mainly from DW [32]. The presence of S. tamesis increased the rates of denitrification at both sites, but, as for oxygen demand, the degree of stimulation was different and higher in Cazaux-Sanguinet Lake. With a worm biomass of 100 gdw m−2, the enhancement factor of N2 production in the sandy sediments was 5.5, whereas that calculated for the Mincio sediments was 2.9. Both denitrification enhancement factors were higher than those calculated for oxygen respiration. A higher relative stimulation of denitrification, as compared to O2 consumption, was also reported by other authors in similar bioturbation experiments [33]. In the Mincio River, denitrification rates support a substantial fraction of CO2 production (see next sections), whereas in the Cazaux-Sanguinet Lake, denitrification is quantitatively irrelevant as compared to the aerobic respiration.
In the Mincio sediments, the presence of the oligochaete reversed the role of the system from a source to a sink for NO3, whereas sediments acted always as an NH4+ source. The presence of S. tamesis augmented NH4+ production, suggesting an important contribution to N regeneration by the reworking activities and the excretion by the oligochaetes [30,31]. However, in the Mincio sediments S. tamesis stimulated NO3 consumption to an extent that always exceeded NH4+ production, resulting in negative DIN fluxes. In Cazaux-Sanguinet Lake, a worm biomass equal to or higher than 65 gdw m−2 reversed the role of sediment from N sink to N source, with NH4+ efflux as the main driver of the net DIN regeneration. The comparison between the two sites suggests that the effect of S. tamesis on net DIN fluxes depends on the environmental conditions, specifically the background nitrate level and the organic content.
Burrow ventilation and bioirrigation, with a few exceptions, always result in a higher consumption of O2 and NO3 [34,35]. This is due to increased sediment–water interfaces and to thinner oxic layers within burrows along a vertical sediment profile [36,37,38]. A few species of burrowers (e.g., Marenzelleria spp.) do not stimulate denitrification, but rather NO3 ammonification due to specific pore water movement [15]. Some studies on N cycling in faunated sediments support the major contribution of NO3 from the overlying water to the total N2 production [33,39,40,41]. This is likely true at the eutrophic sites with elevated NO3 concentrations in the water column [38]. Other studies demonstrated that nitrification and, hence, coupled nitrification–denitrification had the highest contribution to N2 production [42]. The latter result likely depends on factors such as the sediment redox or the NH4+ availability in the upper sediment layer. In sulfidic sediments, nitrification can be suppressed and the same can be true in oligotrophic sandy sites with elevated benthic microalgal activity and little exchangeable pools of NH4+. In the case of Cazaux-Sanguinet Lake, we believe that the low activity of the population of nitrifiers and the limited stimulation of nitrification by the oligochaete can be explained by NH4+ limitation, high competition with plants and benthic algae, and by general inhibition of these bacteria by primary producers [43]. On the other hand, the limited relevance of denitrification of water column NO3 in Cazaux-Sanguinet Lake strongly depended on NO3 concentration in the overlying water, which was very low (<6 µM). In the Mincio sediments, the high NH4+ availability enhanced the process of nitrification and, therefore, its share in the total N2 production.
From NO3 reduction, it was also estimated that the denitrification in bare sediments contributed differently in the two sites: 11.5% and 0.1% of the total mineralization rate in Mincio and Cazaux-Sanguinet sites, respectively. O2 fluxes were converted into CO2 production, assuming an O2:CO2 with a 1:1 stoichiometry, whereas denitrification was converted into CO2 production assuming a NO3:CO2 stoichiometry of 1.25 [44]. In the presence of the oligochaetes with the highest biomass, denitrification contributed 25.5% and 0.3% of the total CO2 production. Hence, in both sediments the highest biomass of S. tamesis enhanced the contribution of the anaerobic denitrification to total mineralization rates, but such contribution was completely different at the two sites. The contrasting effects of S. tamesis on benthic N cycling at the two sites were confirmed by the calculation of denitrification efficiency, which increased in the Mincio River along with an increasing worm biomass. S. tamesis stimulated the process of denitrification and reduced the recycling of inorganic N forms to the water column. The opposite result was found in the Cazaux-Sanguinet Lake. Here, the denitrification efficiency measured in bare sediment was 100%, which suggested a tightly coupled ammonification, nitrification and denitrification, and no inorganic N efflux. Along with an increasing oligochaete biomass, denitrification efficiency decreased in the Cazaux-Sanguinet Lake due to increased NH4+ recycling.
Oxygen and DIN fluxes were related to the dry biomass of the incubated organisms with a linear regression. Assuming a 1:1 ratio of O2 and CO2 fluxes, we calculated a C:N molar ratio of excreted nutrients (8 for the Mincio River and 19 for Cazaux-Sanguinet Lake) from the slopes of the regression. The C:N ratio calculated for Cazaux-Sanguinet was within the range reported by [45] for the detritus of L. dortmanna (C:N = 19–26). This result suggests that S. tamesis in Cazaux-Sanguinet Lake likely feeds on L. dortmanna detritus, which is characterized by a low N content. The C:N calculated for Mincio reflected the C:N molar ratio of more labile organic matter.
Overall results from the present study demonstrated that the alien worm S. tamesis could have both positive and negative effects on the invaded ecosystem, depending on the trophic conditions of the environment. At the eutrophic site the high density of burrowers that dig and rework the sediment, and flush oxic and nitrate-enriched water into a reduced sediment, stimulate the process of denitrification, which could alleviate the high nutrient load of the system. At the oligotrophic site, bare sediment and sediment with a low biomass of oligochaetes per m2 acted as buffers for inorganic N, whereas the increase of S. tamesis biomass enhanced the stimulation of benthic organic matter mineralization and nutrient cycling. Consequently, if the density of S. tamesis dramatically increased, the benthic–pelagic coupling of this shallow-water system would change, favoring the growth of pelagic primary producers and, hence, modify the overall function of the lake. The effects of alien species may vary with time, and a rapid expansion of S. tamesis combined with changes in environmental conditions (e.g., higher temperature regimes) may result in higher mobilization of nutrients buried within sediments. This may produce undesired effects such as the growth of epiphytic algae on the native isoetid L. dortmanna or of phytoplankton in the water column. In fact, previous studies demonstrated that >80% of the inorganic N requirements of L. dortmanna is absorbed by the roots, in the forms of NO3 rather than NH4+ [46]. It is, therefore, likely that higher ammonium availability would favor microalgal primary production. Higher phytoplanktonic or epiphytic productivity may in turn enrich sediments with labile organic matter and create other positive feedbacks on benthic respiration and nutrient recycling [45]. All these cascade effects may hamper protected isoetid vegetation and ultimately result in a regime shift of the lake.

5. Conclusions

Results from this study support the hypothesis that the biogeochemical effects of an invasive worm on sedimentary processes are not univocal, but site-specific. In particular, they demonstrated that bioturbation of muddy sediments in contact with nitrate-rich water resulted in net removal of inorganic N and elevated denitrification efficiency. These biogeochemical effects can be considered positive, as they counteract eutrophication. On the contrary, bioturbation of sandy sediments in contact with a nutrient-poor water column produced a net effect of increased ammonium recycling and decreased denitrification efficiency. These biogeochemical effects have a negative connotation, as they might stimulate algal growth and produce positive feedback. The ecosystemic effects of invasions by alien species needs careful and comprehensive evaluations along large temporal scales.

Author Contributions

S.B. carried out the experiments in both sites, interpreted the data, and wrote the first draft of the manuscript. M.B. designed and conducted the experiments in both sites, interpreted the data, and worked on the draft of the article. C.R. carried out the experiment in the Cazaux-Sanguinet Lake, the chemical analyses, and edited the advanced draft of the manuscript. E.A.F. provided feedback on the research approach and edited the advanced draft of the manuscript. All authors reviewed the manuscript.


This research received no external funding.


The authors want to thank Vincent Bertrin and Gwilherm Jan for the field and laboratory support. Sara Benelli received a financial support from the French National Research Agency (ANR) in the framework of the Investments for the Future Programme, within the COTE Cluster of Excellence (ANR-10-LABX-45). Marco Bartoli was supported by the INBALANCE (Invertebrate-BacteriAL Associations as hotspots of benthic Nitrogen Cycling in Estuarine ecosystems) project, funded by the European Social Fund according to the activity ‘Improvement of researchers’ qualification by implementing world-class R&D projects’ of Measure No. 09.3.3-LMT-K-712-01-0069.

Conflicts of Interest

The authors declare no conflict of interest.


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Figure 1. Sediment oxygen demand in the four treatments in the Mincio River (a) and in the Cazaux-Sanguinet Lake (b). Averages ± standard errors are reported. Note different scales on y-axis.
Figure 1. Sediment oxygen demand in the four treatments in the Mincio River (a) and in the Cazaux-Sanguinet Lake (b). Averages ± standard errors are reported. Note different scales on y-axis.
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Figure 2. Denitrification rates measured via isotope pairing technique (IPT) in the four treatments in the Mincio River (a) and in the Cazaux-Sanguinet Lake (b). DW represents the denitrification of NO3 diffusing to anoxic sediments from the water column, whereas DN is the denitrification of NO3 produced by nitrification in the sediment. Averages ± standard errors are reported. Note different scales on y-axis.
Figure 2. Denitrification rates measured via isotope pairing technique (IPT) in the four treatments in the Mincio River (a) and in the Cazaux-Sanguinet Lake (b). DW represents the denitrification of NO3 diffusing to anoxic sediments from the water column, whereas DN is the denitrification of NO3 produced by nitrification in the sediment. Averages ± standard errors are reported. Note different scales on y-axis.
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Figure 3. Enhancement factors of Sparganophilus tamesis biomass on sediment O2 demand (SOD) and denitrification rates. Results of linear regressions are reported in the graphs. The relative stimulations were calculated by dividing the rates measured in the bioturbated sediments by the rates measured in control sediments, in the Mincio River (a) and in the Cazaux-Sanguinet Lake (b). Averages ± standard errors are reported.
Figure 3. Enhancement factors of Sparganophilus tamesis biomass on sediment O2 demand (SOD) and denitrification rates. Results of linear regressions are reported in the graphs. The relative stimulations were calculated by dividing the rates measured in the bioturbated sediments by the rates measured in control sediments, in the Mincio River (a) and in the Cazaux-Sanguinet Lake (b). Averages ± standard errors are reported.
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Figure 4. Benthic fluxes of N forms (NH4+, NO3, and NO2) expressed in µmol m−2 h−1 measured during dark incubation in the sediment from the Mincio River (a) and from the Cazaux-Sanguinet Lake (b). Averages ± standard errors are reported. Note different scales on y-axes.
Figure 4. Benthic fluxes of N forms (NH4+, NO3, and NO2) expressed in µmol m−2 h−1 measured during dark incubation in the sediment from the Mincio River (a) and from the Cazaux-Sanguinet Lake (b). Averages ± standard errors are reported. Note different scales on y-axes.
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Figure 5. Denitrification efficiency (%) calculated in the 4 treatments of (a) the Mincio River and of (b) the Cazaux-Sanguinet Lake. Averages ± standard errors are reported.
Figure 5. Denitrification efficiency (%) calculated in the 4 treatments of (a) the Mincio River and of (b) the Cazaux-Sanguinet Lake. Averages ± standard errors are reported.
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Table 1. Inorganic nitrogen concentrations measured in replicated water column samples (n = 3) collected at the two investigated sites and sediment features (n = 4) obtained by pooling the upper 10 cm sediment horizon. The reported values correspond to the average ± standard error.
Table 1. Inorganic nitrogen concentrations measured in replicated water column samples (n = 3) collected at the two investigated sites and sediment features (n = 4) obtained by pooling the upper 10 cm sediment horizon. The reported values correspond to the average ± standard error.
NH4+ (µM)2.0 ± 0.23.6 ± 0.1
NO3 (µM)160.7 ± 5.61.5 ± 0.1
NO2 (µM)3.7 ± 0.30.4 ± 0.1
Sediment typologyMuddySandy
Density (g cm−3)1.34 ± 0.081.83 ± 0.09
Porosity0.77 ± 0.040.36 ± 0.05
Organic matter content (%)9.20 ± 0.700.11 ± 0.03

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Benelli, S.; Bartoli, M.; Ribaudo, C.; Fano, E.A. Contrasting Effects of an Alien Worm on Benthic N Cycling in Muddy and Sandy Sediments. Water 2019, 11, 465.

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Benelli S, Bartoli M, Ribaudo C, Fano EA. Contrasting Effects of an Alien Worm on Benthic N Cycling in Muddy and Sandy Sediments. Water. 2019; 11(3):465.

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Benelli, Sara, Marco Bartoli, Cristina Ribaudo, and Elisa Anna Fano. 2019. "Contrasting Effects of an Alien Worm on Benthic N Cycling in Muddy and Sandy Sediments" Water 11, no. 3: 465.

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