# Seaweed strandings as a nitrogen source for benthic algae: Evidence from in situ and in vitro experiments in the English Channel

Stéphanie Lemesle, Anne-Marie Rusig, Pascal Claquin, Isabelle Mussio

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Stéphanie Lemesle, Anne-Marie Rusig, Pascal Claquin, Isabelle Mussio. Seaweed strandings as a nitrogen source for benthic algae: Evidence from in situ and in vitro experiments in the English Channel. Aquatic Botany, 2026, 204, pp.103986. 10.1016/j.aquabot.2025.103986. hal-05447953

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# Seaweed strandings as a nitrogen source for benthic algae: Evidence from *in situ* and *in vitro* experiments in the English Channel

Stéphanie Lemesle<sup>a,b</sup>, Anne-Marie Rusig<sup>a,b</sup>, Pascal Claquin<sup>a,b</sup>, Isabelle Mussio<sup>a,b,*</sup>

## ARTICLE INFO

**Keywords:**

- Rocky-sandy beach ecosystem
- Seaweed strandings
- Regenerated nitrogen
- Intertidal zone
- Pore water
- Algal leachate
- δ¹⁵N

## ABSTRACT

Seaweed strandings are recurrent in the eastern Seine Bay (English Channel) and may strongly affect nutrient dynamics at the sediment-water interface. This study investigated whether nitrogen released from decaying stranded seaweeds contributes to nitrogen availability for benthic *Ulva* during summer, using a combined approach of 19 months of *in situ* monitoring and an *in vitro* experiment. Variation in δ¹⁵N in *Ulva* (δ¹⁵N*Ulva*) was observed, with lower values (<7 ‰) in spring and higher values (>9 ‰) in summer. In spring, low δ¹⁵N*Ulva* values and the positive correlation between concentrations of dissolved inorganic nitrogen (DIN) in intertidal seawater and the River Orne's flow indicate a terrigenous nitrogen origin, mainly from agricultural fertilizers. In summer, concentrations of dissolved organic nitrogen (DON) and ammonium increased in both intertidal and interstitial waters. Significant positive correlations were found between δ¹⁵N*Ulva*, the extent of algal wracks, and DON concentrations, the latter also increasing with the extent of wracks. These relationships suggest that, in summer, when riverine nitrogen inputs are minimal, decaying stranded seaweeds constitute an endogenous nitrogen source for benthic seaweeds. This hypothesis was supported by a controlled experiment showing an increase in δ¹⁵N in *Ulva* thalli after 9 days of culture in seawater enriched with algal leachate. Such endogenous nitrogen recycling supports the growth of opportunistic *Ulva*, which dominate summer strandings and negatively affect coastal water quality assessments under the European Water Framework Directive. These findings highlight the importance of accounting for seaweed strandings in strategies and management actions aimed at improving coastal water quality.

## 1. Introduction

Over the past few decades, coastal ecosystems have experienced increasing anthropogenic nitrogen inputs, primarily from sewage discharges and agricultural fertilizer use (Morand and Briand, 1996; Malone and Newton, 2020). These inputs have led to massive blooms of phytoplankton and/or opportunistic macroalgae, responsible for significant impacts on marine ecology and local economies (Wang et al., 2019; Malone and Newton, 2020; Joniver et al., 2021). Measuring nutrient concentrations in the water column is the conventional approach for monitoring nutrient enrichment in marine environments. However, this method does not allow discrimination between nitrogen sources and is less effective when nutrient loads in the water column are rapidly diluted or removed by microbial activity and plant uptake. Therefore, identifying nutrient sources is essential for developing

effective management strategies to reduce the negative effects of eutrophication (Bermejo et al., 2022). Stable nitrogen isotope ratios (δ¹⁵N) are useful for identifying sources of nitrogenous pollution in coastal systems, as different nitrogen sources have distinctive ¹⁵N signatures (Heaton, 1986) (for a review of δ¹⁵N values of different species of macroalgae exposed to various nitrogen sources see Piñón-Gimate et al. 2017). Synthetic N fertilizers which use atmospheric nitrogen (0 ‰) as their source, generally exhibit low or negative δ¹⁵N values, typically ranging from -4 ‰ to +4 ‰. Regenerated nitrogen from the decomposition of organic matter or from sewage effluent, livestock manure, or fish farm effluent is ¹⁵N-enriched due to the enzymatic preference in bacterial processes for ¹⁴N over ¹⁵N, resulting in elevated δ¹⁵N values. Mean δ¹⁵N values measured in algal tissue exposed to sewage effluents generally range between +7 ‰ and +50.1 ‰, with the highest δ¹⁵N values observed in algae growing directly in front of

<footnote><sup>a</sup> MARINE ECOSYSTEMS AND ORGANISMS RESEARCH LAB - Université de Caen Normandie, Normandie Univ, MERSEA UR 7482, F-14000 CAEN, France</footnote>

<footnote><sup>b</sup> Centre de Recherche en Environnement Côtier, Station marine, Université de Caen Normandie, F-14530 LUC-SUR-MER, France</footnote>

<footnote>* Corresponding author at: MARINE ECOSYSTEMS AND ORGANISMS RESEARCH LAB - Université de Caen Normandie, Normandie Univ, MERSEA UR 7482, F-14000 CAEN, France.</footnote>

<footnote>E-mail address: isabelle.mussio@unicaen.fr (I. Mussio).</footnote>
sewage outfall. This enrichment is due to denitrification processes occurring in wastewater treatment plants (Heaton, 1986; McClelland and Valiela, 1998; Gartner et al., 2002; Savage and Elmgren, 2004; Derse et al., 2004; Dailer et al., 2010; 2012; Piñón-Gimate et al., 2017; Gröcke et al., 2017; Bermejo et al., 2022; Alldred et al., 2023). Analysing nitrogen stable isotope ratios (δ¹⁵N) in macroalgae has become a widely used and particularly effective tool for tracing external sources of nutrient sources in coastal ecosystems across different spatial and temporal scales (McClelland and Valiela, 1998; Costanzo et al., 2001; 2003; Bruland and Mackenzie, 2010; Garcia-Sanz et al., 2010; Dailer et al., 2010; Raimonet et al., 2013; Ochoa-Izaguirre and Soto-Jiménez, 2014; Lemesle et al., 2015; Gröcke et al., 2017; Alldred et al., 2023, 2024a, 2024b). Benthic macroalgae are good indicators of nitrogen sources in aquatic ecosystems. They absorb and assimilate nutrients from the water column and accumulate them in their tissues, thus serving as reliable indicators of both continuous and intermittent nutrient loadings (Costanzo et al., 2001; Jones et al., 2001; Cole et al., 2005; Cohen and Fong, 2005; Lapointe and Bedford, 2010; Garcia-Sanz et al., 2010; Orlandi et al., 2014). Macroalgae are sessile organisms that integrate nitrogen exposure over a timespan of days to weeks (Gartner et al., 2002; Deutsch and Voss, 2006; Lemesle et al., 2016; Bailes and Gröcke, 2020; Parry and Black, 2023). Finally, macroalgal δ¹⁵N generally reflects N inputs from terrestrial sources accurately, with minimal nitrogen fractionation during uptake and assimilation (Naldi and Wheeler, 2002; Cohen and Fong, 2005; Viana et al., 2011), except in the presence of high nitrogen concentrations (Swart et al., 2014; Gröcke et al., 2017).

The Bay of Seine is located in the eastern French sector of the English Channel and is particularly sensitive to eutrophication (Passy et al., 2016). Nutrients levels in the Bay are influenced by freshwater inputs and strong tidal currents, which promotes water exchange and the efficient dispersion of nutrients in the water column (Dauvin, 2012). Despite relatively high nitrogen concentrations in the seawater, the intense mixing induced by tidal currents prevents vertical stratification and nutrient accumulation, thereby limiting large-scale eutrophication (Gentilhomme and Lizon, 1998; Menesguen and Gohin, 2006). The eastern coastline of the Bay of Seine is characterized by extensive intertidal and subtidal rocky plateaus colonized by dense populations of macroalgae. During summer, the sandy beaches in front of these rocky plateaus are regularly affected by significant strandings of various species - including Chlorophyta, Phaeophyceae and Rhodophyta - uprooted by wave action (Lemesle et al., 2015; 2016). Since 2008, strandings of macroalgae composed of over 30 % opportunistic green algae (Ulva spp.) occurring regularly in the Bay, resulted in the downgrading of coastal water bodies under the Water Framework Directive (WFD) (Lemesle et al., 2015; 2023). Luc-sur-Mer is located in the water body HC13 and has been classified as having poor water quality based on the WFD's indicator "opportunistic macroalgal blooms". The monitoring of the extent, biomass, and composition of stranded algal wrack over the same period as this study (March 2017 to October 2018) has been previously reported by Lemesle et al. (2023). Average monthly values of stranded algal biomass ranged from 0.3 ± 0.3 kg fresh wt.m⁻² in March 2018–3.4 ± 2.3 kg fresh wt.m⁻² in August 2017, with the highest peaks recorded in spring and summer, followed by a decline in the winter months. Analysis of wrack composition revealed a dominance of sheet-like, annual species - mainly Ulva - in spring and summer, and perennial brown seaweas in winter (Lemesle et al., 2023). In previous studies, a fine-scale survey of the nitrogen isotopic signal in benthic seaweeds was conducted at two sites of the Bay of Seine located to the west of Luc-sur-Mer - Courseulles-sur-Mer (latitude 49°19′49.79″N, longitude 0°27′21.585″W) and Grandcamp-Maisy (latitude 49°23′17.931″N, longitude 1°2′28.964″W) - which showed a seasonal variation in δ¹⁵N in macroalgae, characterized by low δ¹⁵N values in spring and elevated isotopic signals in summer, in contrast to the oceanic reference (Lemesle et al., 2015; 2016). In this latter site, which is characterized by the absence of terrigenous inputs and seaweed strandings, no seasonal variation in δ¹⁵N in macroalgae was observed

(Lemesle et al., 2015). At the two seaweed stranding sites, it was clearly demonstrated that the low δ¹⁵N values in spring were associated with inputs of inorganic fertilizers originating from the Seine River (Lemesle et al., 2015; 2016). In contrast, it was hypothesized that the elevated δ¹⁵N values measured in summer in macroalgae resulted from regenerated nitrogen inputs from decomposing stranded seaweeds. Thus, in summer, the decomposition of stranded seaweed could provide a new nitrogen source for benthic algae on rocky shores, explaining the absence of nitrogen limitation for their growth during this period, despite low terrestrial inputs (Lemesle et al., 2015; 2016).

Studies have highlighted the diverse roles of organic matter inputs in beach ecosystems, including nutrients recycling (Hyndes et al., 2022). Nitrogen produced by the decomposition of algal wrack on sandy beaches - primarily through bacterial activity - can follow several pathways: it may be incorporated into the food web via uptake by phytoplankton or microphytobenthos, released into the water column, lost through denitrification, absorbed by supralittoral vegetation, or buried in porous intertidal sand and accumulated in the underlying sediments (McLachlan and McGwynne, 1986; Rossi, 2007; Garcia-Robledo et al., 2008; Corzo et al., 2009; Dugan et al., 2011; Lastra et al., 2018; Gómez et al., 2018; Rodil et al., 2019; Luo et al., 2022; Hyndes et al., 2022). Nutrient release from benthic sediments can then be transported to nearshore waters, supplying nutrients to marine primary producers (Dugan et al., 2011; van Alstyne, 2016; Lowman et al., 2019). As a result, algal decomposition on sandy beaches could be a key process in enhancing or sustaining elevated primary productivity, particularly when nutrient availability in the water column is limited (Lavery et al., 1991; Hanisak, 1993). Thus, the growth of benthic seaweeds on rocky plateaus, especially Ulva spp., could be stimulated during summer by nitrogen released during the decomposition of stranded seaweeds. Green macroalgae of the genus Ulva (Ulvophyceae), due to its opportunistic nature, is widely recognized as a reliable bioindicator. It provides a time-integrated measure of nutrient availability in coastal waters, making it an effective tool for assessing water quality in the context of eutrophication (Areco et al., 2021).

In the context of significant summer macroalgal strandings, this study aimed to assess whether decomposing stranded algae could serve as a nitrogen source for intertidal macroalgae, thereby sustaining algal growth. To test this hypothesis, we combined a 19-month time series of biological and environmental data providing the ecological context with an *in vitro* approach to confirm the mechanistic pathways of nitrogen transfer. The field monitoring focused on (1) assessing the coupling between macroalgal wrack and δ¹⁵N variation in Ulva colonizing the rocky shore near the stranding area; (2) exploring the relationship between the extent of stranded algal coverage and the nitrogen content of intertidal pore water and seawater column above the intertidal zone; and (3) better understanding the dynamics of nitrogen compounds and their potential sources in relation to environmental parameters. In parallel, an *in vitro* experiment using leachate produced by the decomposition of stranded algae was conducted to verify whether Ulva could take up and assimilate nitrogen from this algal leachate by tracking the isotopic δ¹⁵N signal. Together, these complementary field and laboratory approaches will allow us to provide both ecological context and mechanistic evidence for the transfer of nitrogen from stranded macroalgae to intertidal primary producers.

## 2. Material and methods

### 2.1. *In situ* study

#### 2.1.1. *Study site*

The study was conducted at Luc-sur-Mer (Normandy, France), a seaside resort and tourist town (latitude 49°19′08″N; longitude 0°21′03″W) located on the west coast of the Seine Bay in the English Channel (Fig. 1). Sampling was conducted over approximately 2 kilometers along the Luc-sur-Mer coastline. The sampling area is
<figure><img src="image_2.png" /><figcaption>Fig. 1. Study area.</figcaption></figure>

characterized by a large sandy area, about 240 m wide, where detached algae wash ashore, followed by a large, slightly sloping rocky plateau (1.5–2 km in width) colonized by seaweeds (Fig. 1). The maximum depth on the foreshore does not exceed –15 m relative to hydrographic zero. The site is an open system oriented north-east, with a macrotidal regime that is semidiurnal, featuring a maximum tidal amplitude average of 6.5 m during spring tides and 2.5 m during neap tides. Tidal currents are generally moderate, ranging between 0.8 and 1 m s⁻¹ during maximum flood and ebb spring tides, and 90 % of waves are less than 1.25 m in height (Dauvin, 2012). The study site is located approximately 11 kilometers northeast of the estuary of the Orne River and 60 kilometers west of the Seine estuary (Fig. 1). It is located in the coastal water body HC13, which is directly influenced by the Orne River plume and indirectly by the Seine River (Table 1) (Aminot et al., 1998, 1997; Napoléon and Claquin, 2012; Garnier et al., 2021). Therefore, terrigenous inputs were assessed through the study of the discharge from the coastal Orne River.

### 2.1.2. Sampling of benthic Ulva

Approximately 500 g of Ulva were collected monthly from March 2017 to October 2018 (except December 2017) in the intertidal zone at low tide (tidal coefficient > 90) on the rocky plateau, within tide pools of the *Fucus serratus* belt, and near seaweed stranding areas, to assess the sources of nitrogen assimilated by seaweeds colonizing the rocky plateau. Algal samples were kept on ice during transport to the laboratory and were immediately processed for N isotopic analysis and for N and C content determination.

### 2.1.3. Abiotic sampling (water and sediment)

In order to monitor dissolved inorganic nitrogen (DIN: NO₃, NO₂, NH₄⁺) and dissolved organic nitrogen (DON) concentrations in interstitial seawater (DON<sub>interstitial</sub>) and in the water column above the intertidal zone, hereafter referred to as intertidal seawater (DON<sub>intertidal</sub>),

**Table 1**

Comparative characteristics of the Seine and Orne watersheds, including average discharge and nutrient fluxes at their outlets (2012–2014 average) (from Garnier et al., 2019).

<table><thead><tr><th></th><th>Seine</th><th>Orne</th></tr></thead><tbody><tr><td>Surface area</td><td>73 221 km²</td><td>2903 km²</td></tr><tr><td>Population</td><td>20 829 875</td><td>440 757</td></tr><tr><td>Cropland</td><td>50.5 %</td><td>35.7 %</td></tr><tr><td>Grassland</td><td>20.2 %</td><td>48.7 %</td></tr><tr><td>Average discharge</td><td>523. 1 m³/s</td><td>31.9 m³/s</td></tr><tr><td>Kton N/year</td><td>141.7</td><td>5.8</td></tr><tr><td>Kton P/year</td><td>4.8</td><td>0.1</td></tr><tr><td>Kton Si/year</td><td>69.6</td><td>4.5</td></tr></tbody></table>

samples were collected as follows. Sediment pore water samples were collected on 13 occasions from June 2018 to October 2018, corresponding to the period of significant seaweed strandings in the intertidal zone (Lemesle et al., 2023). At each sampling point, a pit was excavated with a spade to a depth where water began to fill the bottom of the excavation. The interstitial water was then collected using a 50 mL PE syringe and transferred into acid-washed polypropylene vials, which were filled to the brim to prevent the presence of air. In the lab, after decantation, the supernatant was removed with a syringe, immediately filtered through a Whatman GF/F filter into HCl-washed Nalgene bottles, and then frozen until DIN and DON analyses could be performed. Additionally, monthly sampling of intertidal water at a 1 m depth during high tide was carried out near the study site (49°19′7.921" N, 0°20′48.25" W) as part of the French Coastal Monitoring Network SOMLIT (https://doi.org/10.17882/96638), to investigate variations in nutrient concentrations. Sediment samples were collected bimonthly at low tide from April 2017 to September 2018 in triplicate from the mid-littoral soft substratum in the stranding area. Samples were extracted using 50 mL plastic syringes with a cut-off tip to collect the top 5 cm of surface sediment. The sediments were dried at 60°C until a constant weight was reached. The organic content was determined by measuring the loss on ignition in a 450°C muffle furnace (Heiri et al., 2001).

## 2.2. *In vitro* study

### 2.2.1. Production of a leachate from beached seaweeds

In April 2018, two kilograms of decaying beached algae were collected, briefly rinsed with seawater, and placed in a pre-washed container in a greenhouse. After 15 days of maceration and coarse filtration, the resulting filtrate was considered an algal-based organic fertilizer. The nutrient composition of the leachate was analysed (NO₃, NO₂, NH₄⁺ and PO₄³⁻). Three enrichment media were then prepared: a) offshore seawater alone (0 % - control), b) offshore seawater with 5 % leachate (vol/vol) (low enrichment) and c) offshore seawater with 10 % leachate (vol/vol) (high enrichment). These media were subsequently sterilized by filtration through a 0.22 µm filter.

### 2.2.2. Preparation of algal material

Ulva thalli were collected in April 2018 from the rocky plateau in front of the study site. The freshly collected thalli were acclimated by submerging them in bubbling seawater aquarium for 24 h under controlled conditions (16 ± 1°C, with a 12-hour photoperiod at 70 µmol photons m⁻² s⁻¹). Each thallus was then rinsed several times with filtrsterilized seawater and wiped with absorbent paper to remove all epiphytes.

### 2.2.3. Experimental design

Approximately 2 g (fresh weight) of previously acclimated thalli were placed in each Erlenmeyer flask containing 200 mL of the enrichment medium (0 %, 5 % or 10 %). The three conditions were prepared in triplicate, resulting in a total of 9 flasks per sampling day (day 2-D2, day 6-D6, day 9-D9). In this way, all samples remained independent of each other during time monitoring. For example, the 9 flasks sampled on Day 9 were not sampled on the previous days. Incubations were carried out over 9 days under controlled conditions at 16 ± 1°C with a 12-hour photoperiod at 70 µmol photons m⁻² s⁻¹. The δ¹⁵N was monitored at D0, D2, D6, and D9. To measure δ¹⁵N at D0, a small fragment of each Ulva thallus was sampled before being added to the flask.

## 2.3. Analysis

### 2.3.1. Isotopic analysis

Whole Ulva thalli were dried at 60°C for 48 h and ground into a homogeneous powder using a ball mill (MM 400). Triplicate subsamples
(2.0 ± 0.3 mg dry weight) were then placed in tin capsules. Isotopic abundance and nitrogen and carbon content were determined using a mass spectrometer (IsoPrime, GV Instrument, UK) coupled with an elemental analyser (EA 3000, Eurovector, Milan, Italy). The nitrogen isotope composition of the tissue samples was expressed as the relative per mil (‰) difference from the conventional standard atmospheric N₂, as follows: δ¹⁵N (‰) = [(R<sub>Sample</sub>/R<sub>standard</sub>) - 1] × 1000, where R is the ¹⁵N/¹⁴N ratio (Petersen and Fry, 1987). The analytical precision was 0.2 ‰.

### 2.3.2. Analysis of nitrogen concentrations

Concentrations of nitrate and nitrite in the interstitial and intertidal water samples were determined using a continuous-flow auto-analyzer (Technicon III) according to the method established by Aminot and Kérouel (2007). Ammonium was measured by fluorimetry following the method of Holmes et al. (1999). DON was analysed by a semi-automated method according to Shepherd and Davies (1981), in which DON was completely oxidized to nitrate by an alkaline persulfate digestion. The DON concentration was then calculated by difference (= Total Dissolved Nitrogen - Dissolved Inorganic Nitrogen).

### 2.3.3. Other abiotic and biotic parameters

Seawater temperature, salinity, dissolved oxygen concentrations, and Chl *a* concentration were obtained at high tide during the SOMLIT campaigns at the sampling station located in the intertidal zone (49°19′7.921″ N – 0°20′48.25″ W). Runoff data from the Orne River were measured at the Grimbosq station (49°03′56.29″N - 0°26′54.30″W) and provided by DREAL – Normandy (http://www.hydro.eaufrance.fr). The extent of seaweed strandings was assessed through field sampling conducted during low tide once or twice a week from March 2017 to October 2018. The extent of algal wrack (in hectares) was determined using GPS to map and record the position and the shape of the beaching zones along a 2-km stretch of shoreline at the study site, representing a total surface area of 30 Ha. The user positions around the stranding area were recorded at 5-second intervals with a Trimble TDC 100 GPS linked to a GPS enabled mapping software (Arpentgis mobil). Data on algal wrack extent used in the present study to assess the relationships between algal wrack extent and other biotic and abiotic parameters (δ¹⁵N<sub>Ulva</sub>, organic matter, dissolved oxygen, sea surface temperature, and DON<sub>intertidal</sub>) were previously published in Lemesle et al. (2023). The monitoring of algal wrack extent was conducted between March 2017 and October 2018 at the same study site (Luc-sur-Mer).

### 2.3.4. Statistical analysis

Statistical analyses were performed using Sigmaplot 11.0 software (SigmaStat, Systat Software Inc, San Jose, CA, USA). Shapiro and Wilk's tests were used to assess the normality and homogeneity of the data. For the *in situ* study, ANOVAs were used to examine how beach wrack area, δ¹⁵N<sub>Ulva</sub>, N content, and the C/N ratio varied between sampling months. A pairwise multiple comparison test (Holm-Sidak method) was performed when significant differences were observed. To assess relationships between stranded wrack, biological data (δ¹⁵N<sub>Ulva</sub>, N content and C/N ratio), hydrological parameters, environmental parameters, and organic matter enrichment of the sediment, non-parametric correlations (Spearman) were initially conducted. Additional analyses were performed using linear regression. For the *in vitro* study, a two-way ANOVA test was used to determine the effects of treatments and incubation duration on δ¹⁵N, N content, and C/N ratio values of *Ulva*. The normality of the data was tested using the Shapiro-Wilk test, and data homogeneity was verified. Pairwise multiple comparisons (Holm-Sidak method) were conducted when significant differences were identified. All statistical inferences were based on the adjusted p-value obtained from the Holm-Sidak procedure.

## 3. Results

### 3.1. In situ study

#### 3.1.1. Temporal distribution of δ¹⁵N, N content, and C/N ratio in *Ulva*

*Ulva* δ¹⁵N, N content, and C/N ratio all varied significantly across months (ANOVA, *P* < 0.001; see Fig. 2), with δ¹⁵N and C/N ratio showing the lowest values in spring and the highest values in summer and N content being the lowest in spring-summer and the highest in autumn-winter. δ¹⁵N*<sub>Ulva</sub>* values (6.7–11.4 ‰) formed three clusters: the lowest values in spring, intermediate values in winter (~8 ‰), and the highest values (>9 ‰) from summer to autumn, peaking in July–August. N content (2.4–4.9 ‰) decreased in spring-summer and increased in autumn-winter. The C/N ratio (8.08–15.07) showed an opposite seasonal pattern increasing during spring-summer, with a mid-summer peak (15.07 in July 2018), and decreasing in autumn-winter (<10). Although 2017 and 2018 exhibited similar overall seasonal trends, the timing of the lowest and the highest values differed, resulting in a one- to two-month temporal offset between the years.

#### 3.1.2. Characterization of abiotic and biotic parameters of intertidal sediment and seawater

Abiotic and biotic parameters measured in sediment (organic matter content) and seawater over the intertidal zone from March 2017 to October 2018 are summarized in Table 2. Sediment organic matter (OM) content showed little variation, ranging from 0.83 ± 0.28 % (September 2018) to 2.20 ± 0.84 % (September 2017). Seawater surface temperatures (SST) varied between 7.3°C (March 2018) and 19.8°C (July 2018) with the highest values in June 2017 and July 2018 (>18°C). Temperatures were notably colder in late winter and early spring 2018 compared to 2017. Chl *a* concentrations showed clear seasonal variation, peaking in spring and early summer (5.50 µg.L⁻¹ in April 2017; 6.72 µg.L⁻¹ in June 2018), and reaching minimum values in autumn and winter (0.90 µg.L⁻¹ in December 2017 and September 2018). Dissolved oxygen (DO) concentrations remained generally high (> 6 mL.L⁻¹) from March to May in 2017 and from December 2017 to May 2018, but declined during summer reaching the lowest values in September 2017 (4.98 mL.L⁻¹) and August 2018 (4.59 mL.L⁻¹). Freshwater runoff from the Orne River was low in summer and high in winter with minimal values in August 2017 (1.86 m³.s⁻¹) and in September 2018 (2.50 m³.s⁻¹) and the maximal value in January 2018 (100 m³.s⁻¹). Overall, river flow was higher in 2018 than in 2017 with a discharge of 34 m³.s⁻¹ in April 2018 compared 6.38 m³.s⁻¹ in April 2017.

#### 3.1.3. Temporal variation of nitrogen concentrations in intertidal seawater

Nitrate (NO₃⁻) concentrations (1.16–37.71 µM) in intertidal seawater showed clear seasonal variation, peaking in winter–early spring and decreasing in summer for both years (Fig. 3). Spring 2018 levels (~30 µM) were 2–3 times higher than in spring 2017 (14.89 µM in April and 8.69 µM in May). In contrast, ammonium (NH₄⁺) and nitrite (NO₂⁻) showed no clear seasonal pattern, with NH₄⁺ + NO₂⁻ ranging from 0.65 µM to 7.63 µM with NO₂⁻ always remaining below 0.70 µM. DON<sub>intertidal</sub> ranged from 2.37 µM (December 2017) to 11.93 µM (June 2017), peaking in spring–summer 2017 and late summer 2018 (~10 µM), and reaching minimum values (<5 µM) in December 2017, March 2018 and April 2018.

#### 3.1.4. Temporal variation of nitrogen concentrations in interstitial seawater

Table 3 shows DIN and DON concentrations in sediment pore water from June to October 2018, covering 13 samples during the period of algal growth and major seaweed strandings. DON and NH₄⁺ concentrations were generally higher (DON: 11.03–42.70 µM, NH₄⁺: 8.72–73.94 µM) than NO₂⁻ and NO₃⁻ (NO₂⁻: < 2.52 µM, NO₃⁻: mostly < 6.09 µM) with the exception of two NO₃⁻ peaks in August (83.11 µM) and October (24.94 µM). However, no clear overall trend was observed.
<figure><img src="image_3.png" /></figure>

<figure><img src="image_4.png" /></figure>

<figure><img src="image_5.png" /><figcaption>**Fig. 2.** Temporal variation of mean (A) δ¹⁵N, (B) nitrogen content, and (C) C/N ratio in *Ulva* tissue at Luc-sur-Mer (March 2017–October 2018; no sampling in December 2017). Values are mean ± SE (N = 3). Statistical differences between leachate concentrations (P < 0.05) are indicated by letters (a, b, c, d, e, f, g).</figcaption></figure>
<caption>**Table 2**</caption>

Organic matter content in sediments (OM), sea surface temperature (SST), chlorophyll a (Chl a), and dissolved oxygen concentrations (DO) in the water column above the intertidal zone at high tide, as well as Orne River flow, from March 2017 to October 2018. The minimum and maximum values are in bold.

<table><thead><tr><th>Date</th><th>OM (%)</th><th>SST<sup>a</sup> (°C)</th><th>Chl a (µg. L<sup>-1</sup>)</th><th>DO<sup>a</sup> (mL L<sup>-1</sup>)</th><th>Orne flow<sup>b</sup> (m<sup>3</sup> s<sup>-1</sup>)</th></tr></thead><tbody><tr><td>Mar-17</td><td>/</td><td>8.60</td><td>3.06</td><td>6.60</td><td>23.43</td></tr><tr><td>Apr-17</td><td>1.56</td><td>11.7</td><td>5.50</td><td><strong>6.99</strong></td><td>6.38</td></tr><tr><td>May-17</td><td>1.28</td><td>13.5</td><td>4.54</td><td>6.89</td><td>5.86</td></tr><tr><td>Jun-17</td><td>1.14</td><td>18.0</td><td>3.74</td><td>5.71</td><td>3.05</td></tr><tr><td>Jul-17</td><td>1.64</td><td>19.2</td><td>3.36</td><td>5.16</td><td>1.90</td></tr><tr><td>Aug-17</td><td>1.27</td><td>19.1</td><td>2.82</td><td>5.80</td><td><strong>1.86</strong></td></tr><tr><td>Sep-17</td><td><strong>2.20</strong></td><td>17.9</td><td>2.24</td><td>4.98</td><td>2.30</td></tr><tr><td>Oct-17</td><td>2.11</td><td>16.2</td><td>1.50</td><td>5.20</td><td>2.50</td></tr><tr><td>Nov-17</td><td>1.37</td><td>12.4</td><td>1.01</td><td>5.92</td><td>8.34</td></tr><tr><td>Dec-17</td><td>1.03</td><td>9.30</td><td><strong>0.90</strong></td><td>6.30</td><td>44.50</td></tr><tr><td>Jan-18</td><td>1.26</td><td>9.00</td><td>1.66</td><td>6.46</td><td><strong>100.0</strong></td></tr><tr><td>Feb-18</td><td>1.61</td><td>7.60</td><td>1.03</td><td>6.62</td><td>47.40</td></tr><tr><td>Mar-18</td><td>1.61</td><td><strong>7.30</strong></td><td>2.19</td><td>6.83</td><td>35.60</td></tr><tr><td>Apr-18</td><td>1.09</td><td>9.00</td><td>2.34</td><td>7.12</td><td>34.00</td></tr><tr><td>May-18</td><td>1.50</td><td>13.1</td><td>3.44</td><td>6.49</td><td>17.80</td></tr><tr><td>Jun-18</td><td>1.21</td><td>15.9</td><td><strong>6.72</strong></td><td>5.77</td><td>24.00</td></tr><tr><td>Jul-18</td><td>1.34</td><td><strong>19.8</strong></td><td>4.25</td><td>5.30</td><td>4.95</td></tr><tr><td>Aug-18</td><td>1.06</td><td>18.8</td><td>2.03</td><td><strong>4.59</strong></td><td>2.92</td></tr><tr><td>Sep-18</td><td><strong>0.83</strong></td><td>18.1</td><td><strong>0.90</strong></td><td>4.75</td><td>2.50</td></tr><tr><td>Oct-18</td><td>/</td><td>15.9</td><td>1.47</td><td>5.23</td><td>3.08</td></tr></tbody></table>

<figure><img src="image_6.png" /><figcaption>**Fig. 3.** Temporal variation of mean DIN (Ammonium + Nitrite, Nitrate) and DON<sub>intertidal</sub> in the water column above the intertidal zone at high tide from March 2017 to October 2018.</figcaption></figure>

### 3.1.5. Relationships between biotic and environmental variables

The extent of algal wrack accumulation was positively correlated with seawater temperature and negatively correlated with dissolved oxygen concentration in the intertidal water confirmed by linear regressions (p-values <0.001; r = 0.57 and 0.59 respectively) (Table 4). Spearman and linear correlations showed that δ<sup>15</sup>N in Ulva increased

<caption>**Table 3**</caption>

Average concentrations of DIN and DON in interstitial seawater of the intertidal zone at Luc-sur-Mer, measured from June 2018 to October 2018 (± standard error, N = 2). The minimum and maximum values are in bold.

<table><thead><tr><th>Date</th><th>DON (µM) ± se</th><th>NH<sub>4</sub><sup>+</sup> (µM) ± se</th><th>NO<sub>2</sub> (µM) ± se</th><th>NO<sub>3</sub><sup>-</sup> (µM) ± se</th></tr></thead><tbody><tr><td>06/22/2018</td><td>31.77 ± 1.78</td><td>50.54 ± 0.26</td><td><strong>0.18 ± 0.012</strong></td><td>2.19 ± 0.07</td></tr><tr><td>06/28/2018</td><td>12.12 ± 0.67</td><td>10.20 ± 0.87</td><td>0.31 ± 0.024</td><td>4.01 ± 0.52</td></tr><tr><td>07/17/2018</td><td>12.97 ± 0.74</td><td>40.83 ± 0.17</td><td>0.40 ± 0.070</td><td>3.53 ± 0.39</td></tr><tr><td>07/24/2018</td><td>24.13 ± 3.09</td><td>60.40 ± 8.47</td><td>0.39 ± 0.003</td><td><strong>0.73 ± 0.08</strong></td></tr><tr><td>08/03/2018</td><td>23.71 ± 3.22</td><td>25.12 ± 7.99</td><td>0.60 ± 0.002</td><td>6.09 ± 0.17</td></tr><tr><td>08/09/2018</td><td><strong>42.70 ± 9.96</strong></td><td>49.57 ± 0.66</td><td>0.28 ± 0.034</td><td>1.06 ± 0.06</td></tr><tr><td>08/16/2018</td><td>18.60 ± 4.23</td><td><strong>8.72 ± 1.01</strong></td><td><strong>2.52 ± 0.122</strong></td><td><strong>83.11 ± 3.11</strong></td></tr><tr><td>08/24/2018</td><td>11.28 ± 1.19</td><td>41.41 ± 3.20</td><td>0.24 ± 0.007</td><td>3.23 ± 0.78</td></tr><tr><td>08/28/2018</td><td>26.07 ± 1.80</td><td>56.36 ± 2.36</td><td>0.28 ± 0.068</td><td>3.23 ± 1.35</td></tr><tr><td>09/21/2018</td><td>29.79 ± 1.77</td><td>53.10 ± 2.98</td><td>0.45 ± 0.021</td><td>2.64 ± 0.52</td></tr><tr><td>10/05/2018</td><td>34.82 ± 0.32</td><td><strong>73.94 ± 2.87</strong></td><td><strong>0.18 ± 0.011</strong></td><td>1.41 ± 0.06</td></tr><tr><td>10/15/2018</td><td>16.25 ± 1.18</td><td>59.18 ± 12.98</td><td>0.50 ± 0.366</td><td>24.94 ± 20.62</td></tr><tr><td>10/23/2018</td><td><strong>11.03 ± 0.25</strong></td><td>16.61 ± 2.42</td><td><strong>0.18 ± 0.045</strong></td><td>1.12 ± 0.57</td></tr></tbody></table>

with the DON<sub>intertidal</sub>/DIN<sub>intertidal</sub> ratio, indicating <sup>15</sup>N enrichment in Ulva tissue as the ration rose. A Spearman correlation was also found between the N content of Ulva tissue and DIN<sub>intertidal</sub> concentrations (ρ = 0.47, P < 0.05), but no significant linear correlation was observed. No relationship was found between N content in Ulva and DON<sub>intertidal</sub> concentrations in intertidal seawater. DIN<sub>intertidal</sub> and DON<sub>intertidal</sub> concentrations were significantly correlated with the flow of the Orne River, showing a strong positive correlation with DIN<sub>intertidal</sub> concentrations (ρ = 0.85, p-value <0.0001) and a strong negative correlation with DON<sub>intertidal</sub> concentrations (ρ = -0.70, p-value <0.001) (Table 4).

### 3.1.6. Relationships between isotopic signal δ<sup>15</sup>N<sub>Ulva</sub> and extent of algal wrack

A significant positive correlation was found between δ<sup>15</sup>N in Ulva and the extent of algal wrack (R<sup>2</sup>= 0.57, P = 0.0002, F = 22.613) with δ<sup>15</sup>N in Ulva increasing with wrack area. Additionally, DON<sub>intertidal</sub> concentrations were significantly and positively correlated with the extent of algal wrack (R<sup>2</sup> = 0.35, P = 0.008, F = 9.12), indicating that DON<sub>intertidal</sub> levels increased with the extent of wrack. δ<sup>15</sup>N<sub>Ulva</sub> values decreased significantly with increasing DIN<sub>intertidal</sub> concentrations (R<sup>2</sup>=0.435, P = 0.003, F = 12.33), while a significant positive correlation was found between δ<sup>15</sup>N<sub>Ulva</sub> and DON<sub>intertidal</sub> concentrations (R=0.277, P = 0.025, F = 6.137) (Fig. 4).

### 3.2. In vitro study

Nutrient analysis showed that the leachate produced from beached seaweeds was rich in ammonium (364 µM NH<sub>4</sub><sup>+</sup>; 7.58 µM NO<sub>3</sub><sup>-</sup>; 0.652 µM NO<sub>2</sub>; 4.21 µM PO<sub>4</sub><sup>3-</sup>). The effect of leachate enrichment and incubation duration on δ<sup>15</sup>N<sub>Ulva</sub> is shown in Fig. 5. A significant interaction between the factors ‘duration’ and ‘enrichment level’ indicates that the effect of incubation time on δ<sup>15</sup>N<sub>Ulva</sub> varies across treatments (0 %, 5 % and 10 %) (two-way ANOVA, F(6)= 3.16, P = 0.02). Ulva cultured in un-enriched seawater showed no significant δ<sup>15</sup>N change over 9 days (7.13–7.86 ‰). In the enriched treatments, δ<sup>15</sup>N<sub>Ulva</sub> increased significantly after 2 days in the 5 % treatment and after 9 days in the 10 %
<caption>**Table 4**</caption>

Spearman and linear correlation analyses between biotic variables (WRACK: algal wrack extent; N<sub>Ulva</sub>: nitrogen content in *Ulva*; δ<sup>15</sup>N<sub>Ulva</sub>: δ<sup>15</sup>N in *Ulva* tissue) and environmental variables (DIN<sub>intertidal</sub> and DON<sub>intertidal</sub> concentrations in the water column above the intertidal zone at high tide; Orne flow; Chl *a*: chlorophyll a content; OM: organic matter concentration; SST: sea surface temperature; DO: dissolved oxygen concentration). Significant correlations are shown in bold.

<table><thead><tr><th rowspan="2">Variables</th><th colspan="2">Spearman correlations</th><th colspan="3">Linear correlations</th></tr><tr><th>Rho (ρ)</th><th>p-value</th><th>R<sup>2</sup></th><th>p-value</th><th>Equation</th></tr></thead><tbody><tr><td>WRACK<sup>a</sup> vs OM</td><td>-0.16</td><td>0.527</td><td></td><td></td><td></td></tr><tr><td>WRACK<sup>b</sup> vs DO</td><td><strong>-0.72</strong></td><td><strong>0.0001</strong></td><td><strong>0.59</strong></td><td><strong>&lt; 0.001</strong></td><td>DOC = 6.52-(0.42xWRACK)</td></tr><tr><td>WRACK<sup>b</sup> vs SST</td><td><strong>0.69</strong></td><td><strong>0.0006</strong></td><td><strong>0.57</strong></td><td><strong>&lt; 0.001</strong></td><td>WRACK = -2.12 + (0.25xSST)</td></tr><tr><td>DO vs SST</td><td><strong>-0.77</strong></td><td><strong>&lt; 0.0001</strong></td><td></td><td></td><td></td></tr><tr><td>Chl<sup>a</sup> vs DO</td><td>0.24</td><td>0.29</td><td></td><td></td><td></td></tr><tr><td>δ<sup>15</sup>N vs DON<sub>intertidal</sub>/DIN<sub>intertidal</sub></td><td><strong>0.62</strong></td><td><strong>0.005</strong></td><td><strong>0.32</strong></td><td><strong>0.018</strong></td><td>δ<sup>15</sup>N = 8.37 + (0.93x DON<sub>intertidal</sub>/DIN<sub>intertidal</sub>)</td></tr><tr><td>N<sub>Ulva</sub> vs DIN<sub>intertidal</sub></td><td>0.47</td><td>0.047</td><td>0.16</td><td>0.09</td><td></td></tr><tr><td>N<sub>Ulva</sub> vs DON<sub>intertidal</sub></td><td>-0.09</td><td>0.692</td><td></td><td></td><td></td></tr><tr><td>DIN<sub>intertidal</sub> vs Orne flow</td><td><strong>0.85</strong></td><td><strong>&lt; 0.0001</strong></td><td><strong>0.40</strong></td><td><strong>0.004</strong></td><td>DIN<sub>intertidal</sub>= 13.67 + (0.28xOrne flow)</td></tr><tr><td>DON<sub>intertidal</sub> vs Orne flow</td><td><strong>-0.70</strong></td><td><strong>&lt; 0.001</strong></td><td></td><td></td><td></td></tr></tbody></table>

<figure><img src="image_7.png" /><figcaption>**Fig. 4.** Relationships between A) δ<sup>15</sup>N in *Ulva* tissue and algal wrack extent, B) DON<sub>intertidal</sub> concentrations in high-tide seawater covering the intertidal zone and algal wrack extent, C) δ<sup>15</sup>N in *Ulva* tissue and DIN<sub>intertidal</sub> concentrations in high-tide seawater covering the intertidal zone, and D) δ<sup>15</sup>N in *Ulva* tissue and DON<sub>intertidal</sub> concentrations in high-tide seawater covering the intertidal zone with linear regression lines and 95 % confidence intervals.</figcaption></figure>

treatment (P < 0.05). In the 5 % treatment, δ<sup>15</sup>N increased from an initial value of 6.77 ± 0.19 ‰ to 8.49 ± 0.28 ‰, whereas in the 10 % treatment it increased from 7.53 ± 0.91 ‰ to 8.63 ± 0.75 ‰. No

significant changes were observed in tissue N content (~3 %) or C/N ratio (9.39–13.69 %) across treatments during the incubation period (data not shown).
<figure><img src="image_8.png" /><figcaption>**Fig. 5.** Variation in δ¹⁵N *Ulva* tissue incubated for 0, 2, 6 and 9 days in seawater enriched with low and high concentrations of an algal leachate and in unenriched seawater (control). Values are means ± SE, (n = 3). Statistical differences between leachate concentrations (P < 0.05) are indicated by letters (a, b).</figcaption></figure>

# 4. Discussion

Decaying algal wrack supports the growth of benthic *Ulva* on intertidal rocky shores during summer, as shown by an almost two-year δ¹⁵N time series and an experimental algal leachate assay. These demonstrate that nitrogen released from decaying seaweeds is effectively taken up by benthic algae and highlight the role of stranded macroalgae in supplying nitrogen to intertidal algal communities.

## 4.1. Identification of seasonal nitrogen sources for *Ulva* on rocky shores

In situ monitoring of δ¹⁵N in *Ulva* tissue (δ¹⁵N*Ulva*) confirmed the seasonal variation in nitrogen isotopic signatures previously observed at two other sites in the Bay of Seine, which are also affected by summer seaweed strandings (Lemesle et al., 2015; 2016). Specifically, δ¹⁵N*Ulva* values were the lowest in spring (mostly <7 ‰), the highest in summer (>9 ‰), and intermediate in autumn/winter (around 8 ‰). As in these earlier studies, the high-flow conditions of coastal rivers during winter and early spring in 2017 and 2018 - driven by precipitation and coinciding with the period of agricultural fertilization - led to increased nitrate concentrations in seawater. This likely explains the low δ¹⁵N*Ulva* values observed during this period, as evidenced by the significant relationship between δ¹⁵N*Ulva* and intertidal DIN concentrations (Costanzo et al., 2003; Raimonet et al., 2013). This interpretation is further supported by a significant correlation between DIN concentrations in the water column above the intertidal zone at high tide (DIN*intertidal*) - mainly composed of nitrate during spring, winter, and autumn - and the Orne River's discharge, confirming their terrigenous origin. Unlike in 2017, low isotopic signatures (δ¹⁵N*Ulva*) persisted until June 2018, which could be attributed to coastal freshwater discharge being 3-7 times higher in 2018 than during the same period in 2017. However, as also reported by Lemesle et al. (2015); (2016), δ¹⁵N*Ulva* values recorded were higher than those typically associated with synthetic N-fertilizers. At the watershed, N sources such as wastewater effluents or organic fertilizers exhibit overlapping δ¹⁵N signatures. The Seine and Orne River watersheds are dominated by croplands and grasslands, supporting the hypothesis of manure or organic fertilizer inputs and, consequently, higher δ¹⁵N*Ulva* values. Raimonet et al. (2013) also showed in the Charente estuary (France) that δ¹⁵N values of primary DIN sources from croplands were higher than expected (6.5 ± 0.6 ‰). Furthermore, the isotopic signature of nitrate also reflects soil processes that fractionate nitrogen isotopes (Böttcher et al., 1990;

Kendall, 1998; Briand et al., 2017). For example, Sebilo et al. (2003) demonstrated that in the Seine River, a large proportion of nitrate exported from agricultural soils was removed through riparian and benthic denitrification, resulting in the natural enrichment of residual nitrates in ¹⁵N. This may explain the high δ¹⁵N values observed, as also suggested by Derse et al. (2004). Unlike in spring, the input of terrigenous inorganic nitrogen into intertidal waters was the lowest in summer, while the N content of *Ulva* on the rocky shore remained high (mostly >3 %, except in July 2018). This contrasts with what is typically observed in temperate coastal waters, where nitrogen availability usually limits macroalgae growth in summer (Hanisak, 1993). The consistently high nitrogen content, combined with a C/N ratio generally ≤ 15, indicates that *Ulva* was not nitrogen-limited (Hurd et al., 2014). During summer, the high nitrogen content and elevated isotopic signature δ¹⁵N in *Ulva*, despite low DIN concentrations in seawater, suggest the presence of alternative nitrogen sources for *Ulva*.

## 4.2. Mechanistic evidence that stranded seaweeds serve as a source of nitrogen for benthic *Ulva* in summer

### 4.2.1. In situ evidence of stranded algae as a nitrogen source– increase in δ¹⁵N in benthic *Ulva*

During summer, the elevated isotopic signature in benthic *Ulva* indicates the uptake of regenerated nitrogen. Statistical analyses show that the extent of algal wrack accounts for 57 % of the variation in δ¹⁵N*Ulva*, suggesting that decomposing algal wracks may locally release bioavailable nitrogen. This process likely explains the higher δ¹⁵N values in benthic *Ulva* through recycling and isotopic enrichment of organic matter. Moreover, δ¹⁵N*Ulva* was positively correlated with DON concentrations in intertidal seawater at high tide (DON*intertidal*), which were themselves positively correlated with the extent of algal wrack. This pattern is also consistent with the significant positive relationship between δ¹⁵N *Ulva* and the DON/DIN ratio in intertidal seawater. These relationships highlight the role of organic nitrogen compounds from stranded algal biomass in driving the ¹⁵N enrichment of benthic seaweeds, particularly during spring and summer when the largest extents of algal wrack (>3 Ha) are recorded (Lemesle et al., 2023). A significant negative relationship was observed between DON concentrations in seawater over the intertidal zone at high tide (DON*intertidal*) and Orne River flow which excludes a terrestrial origin for DON*intertidal*. In contrast, DIN concentrations were strongly correlated with river flow. No correlation was found between DON*intertidal* concentrations and Chl *a* concentration in the water column above the intertidal zone at high tide, thereby rejecting the hypothesis that phytoplankton is a source of DON (data not shown

However, algal wracks alone cannot fully account for the observed variability in δ¹⁵N*Ulva*, as additional nitrogen sources and biogeochemical processes may also contribute. Small groundwater springs were present at the study site, but their contribution to nitrogen inputs - and thus to the δ¹⁵N variability in *Ulva* - is considered negligible due to the high dilution in this open system. Van Tussenbroek et al. (2017) reported that monthly nitrogen inputs from the decomposition of stranded *Sargassum* biomass in the Caribbean were approximately 30 times higher than those from groundwater discharge. In contrary, atmospheric nitrogen deposition represents a significant nitrogen source in some marine ecosystems. For example, Neumann et al. (2018) estimated that it accounts for 20-40 % of total nitrogen inputs to the North Sea and Baltic Sea, with contributions from human activities such as shipping and agriculture. However, no studies have yet quantified atmospheric nitrogen deposition in the Seine Bay. Finally, during the decomposition of stranded biomass, denitrification may occur, preferentially removing ¹⁴N and thereby enriching the remaining nitrogen pool in ¹⁵N. This process could partly explain the higher δ¹⁵N values observed in *Ulva* during summer. However, total nitrogen loss through denitrification appears to be relatively low in sandy-beach ecosystems (McLachlan and McGwynne, 1986).
### 4.2.2. Decomposition of stranded algae – release of DON and ammonium into intertidal and interstitial waters

The decomposition of stranded algal biomass remains a key mechanism driving the production of nitrogen available for benthic seaweeds. When stranded in the intertidal zone, algal biomass is broken down through physical fragmentation, grazing by intertidal fauna, and microbial activity. Bacteria from the seaweed biofilms proliferate and convert organic nitrogen into DON and DIN, which can be rapidly assimilated by benthic seaweeds. However, research on sandy-beach systems remains limited, particularly regarding microbial processes and nutrient cycling (Hyndes et al., 2022). The significant negative relationships observed between algal wrack deposits and dissolved oxygen concentrations in intertidal water suggest active bacterial decomposition, as also shown by Luo et al. (2022) during the decomposition of *Gracilaria* litter. During summer the decomposition of the seaweed strandings through mineralization increased DON and NH<sup>4</sup><sup>+</sup> concentrations in both the water column above the intertidal zone and in sediment pore water, as reported in previous studies (Dugan et al., 2011; van Tussenbroek, 2011; Hyndes et al., 2012; Rodriguez-Martinez et al., 2019; Luo et al., 2022). In sediment pore water, concentrations of DON and ammonium were higher than those of other inorganic forms (NO<sub>3</sub><sup>-</sup> and NO<sub>2</sub><sup>-</sup>). DON ranged from 11.03 µM to 42.70 µM, significantly exceeding the values reported by Dugan et al. (2011). Ammonium concentrations were also generally higher than nitrate concentrations, consistent with observations from other studies on beach pore water (Santos et al., 2009; Goodridge and Melack, 2014; Lowman et al., 2019). Similarly, concentrations of DON and DIN in beach pore water were generally higher than those measured in the water column above the intertidal zone during the same period, suggesting an accumulation of nitrogen in the intertidal pore water. In our study, no significant variation in organic matter content was observed, even in summer, with values consistently remaining below 2 %. This is consistent with intertidal sandy systems, in contrast to fine muddy sediments, which are rich in organic matter (Cowan et al., 1996; Berelson et al., 1998; Boyle et al., 2004; Boyer and Fong, 2005). This can be explained by the rapid aerobic decomposition of organic matter into DIN and DON due to regular oxygenation of pore water by tides and waves (Anschutz et al., 2009; Dugan et al., 2011). As organic matter was measured only monthly, these measurements may have missed short-term pulses, particularly following large algal stranding events. Similarly, monthly intertidal sampling may have missed brief but significant nutrient pulses following major strandings, underscoring the need for high-frequency sampling during peak wrack periods.

### 4.2.3. Evidence of nitrogen assimilation by benthic Ulva from decomposing algal wracks

The increase in δ¹⁵N observed in benthic *Ulva* during summer is most likely driven by the uptake of dissolved nitrogen forms, primarily NH<sub>4</sub><sup>+</sup> and DON, released during the decomposition of stranded algal biomass (Rossi, 2007; Dugan et al., 2011). Ammonium is generally the preferred nitrogen source for *Ulva* species (Tyler et al., 2005; Lemesle et al., 2016; Xiu et al., 2019). However, beyond inorganic forms, macroalgae can also assimilate dissolved organic nitrogen compounds, such as urea and free amino acids, as complementary nitrogen sources (Tyler et al., 2003; 2005; van Engeland et al., 2011; Shi et al., 2015). In New Zealand urea supplies up to 30 % of the nitrogen requirements of four intertidal seaweeds during summer (Phillips and Hurd, 2003), while in California it represents an important year-round nitrogen source for the brown seaweed *Macrocystis pyrifera* (Smith et al., 2018). Xiu et al. (2019) demonstrated that *Ulva prolifera*, assimilates ammonium, urea, nitrate, and amino acids in decreasing order of preference and that DON contributed significantly to support the green bloom underscoring the ecological relevance of DON for *Ulva* species.

Wrack-derived nitrogen contributes to δ¹⁵N enrichment in *Ulva* as demonstrated by an *in vitro* experiment in which *Ulva* incubated for 9 days in an algal leachate at low and high concentrations exhibited a

slight but significant increase in δ¹⁵N, indicating the uptake and assimilation of these nitrogen sources. The δ¹⁵N values measured *in vitro* were comparable to those measured *in situ* during summer (>9 ‰), consistent with observations by Orlandi et al. (2017) who found no significant differences in δ¹⁵N between *Ulva lactuca* cultured for 8 days in an organic fertilizer (cow manure) and in a non-enriched medium. The lower-than-expected δ¹⁵N values measured after 9 days may reflect isotopic fractionation. Macroalgal δ¹⁵N depends on both the δ¹⁵N of nitrogen sources and isotopic fractionation occurring under high nitrogen concentrations (Umezawa et al., 2007; Dudley et al., 2010; Swart et al., 2014; Orlandi et al., 2017; Pruell et al., 2020). Under such conditions, seaweeds preferentially take up the lighter isotope (¹⁴N), explaining the relatively lower δ¹⁵N values measured in macroalgae near wastewater discharges (Rogers, 1999; Viana and Bode, 2013; Carballeira et al., 2013; Parry and Black, 2023). In our study, *Ulva* samples were collected in April 2018, when DIN concentrations in seawater were high (~30 µM), algal nitrogen contents > 2 %, and algal C/N ratios were < 13, indicating non-limiting, eutrophic conditions (Lapointe and Bedford, 2010) consistent with isotopic fractionation. The observed timescale of isotopic change also aligns with Lemesle et al. (2016), who reported that *Ulva* cultured in a low-δ¹⁵N medium incorporated the isotopic signature of the medium within 20 days, with changes in isotopic signature detectable after only 5 days. The rapid δ¹⁵N change highlights the opportunistic nature of the genus *Ulva*, which can assimilate dissolved inorganic nitrogen within 48 h (Orlandi et al., 2017; Gröcke et al., 2017; Bailes and Gröcke, 2020), and completely turn over its nitrogen pool every 12–15 days (Aguiar et al., 2003). Nevertheless, laboratory conditions do not fully capture field variability. To better understand the contribution of wrack-derived nitrogen to benthic *Ulva*, in *situ* mesocosm experiments are needed to assess δ¹⁵N dynamics under more realistic environmental conditions. Furthermore, as only bulk DON concentrations were measured in this study, future work should focus on DON speciation using HPLC or LC-MS/MS to identify which specific organic nitrogen compounds—such as urea and amino acids—most strongly support *Ulva* nitrogen uptake. Han et al. (2023) showed that *Ulva pertusa* had the ability to directly assimilate urea and glycine.

### 4.3. Ecological implications

The genus *Ulva* is an appropriate bioindicator of eutrophication, since its rapid colonization ability is linked to its efficient nitrogen uptake from the surrounding environment. Unlike “green tides,” during summer on the foreshore of the Seine Bay, extensive populations of *Ulva* remain attached to the intertidal rocky substrate before detaching and accumulating with other macroalgal species, driven by both biological and physical processes (Lemesle et al., 2023). Our study highlights that DON and ammonium released from the decomposition of large accumulations of stranded macroalgae—into both the water column and interstitial water—can be directly assimilated by benthic *Ulva*. This creates a positive feedback mechanism, in which nutrients derived from decomposing algae promote further *Ulva* proliferation and subsequent stranding (Fig. 6). The genus *Ulva* quickly absorbs and metabolizes various forms of inorganic and organic nitrogen in contrast to the slower-growing rocky shore species (Pérez-Mayorga, et al., 2011; Xiu et al., 2019). A previous study conducted in the Seine Bay has shown that the annual species *Ulva* sp. and *Porphyra* sp. exhibit significantly higher NH<sub>4</sub><sup>+</sup> uptake rates than the perennial species *Fucus serratus* (Phaeophyceae) and *Chondrus crispus* (Rhodophyceae), with *Ulva* displaying a clear competitive advantage at high ammonium concentrations in seawater (Lemesle et al., 2015) consistent with Pedersen and Borum (1996). Lemesle et al. (2023) reported that at the study site, benthic *Ulva* represented over 30 % of algal cover, reaching a peak of 50 % in the summer 2018. Given that “blooms of green opportunistic macroalgae,” primarily the genus *Ulva*, are used as key indicators under the Water Framework Directive (WFD) for coastal ecological assessment
<figure><img src="image_9.png" /><figcaption>**Fig. 6.** Conceptual schematic of the hypothesized N-cycling pathway from *in situ* and *in vitro* findings. Dashed arrows represent nitrogen inputs (atmospheric deposition and resurgences) and processes (denitrification) whose impacts on the δ¹⁵N signature at the study site are negligible or have not been evaluated.</figcaption></figure>

(Scanlan et al., 2007), these findings underscore the importance of considering nitrogen inputs from decomposing stranded macroalgae in coastal management. Until today, in the Bay of Seine, beaches at seaside resorts have been cleaned during large algae accumulations by using tractors to push the algae back into the sea. However, this practice sustains the proliferation of *Ulva* and negatively impacts the beaches. Targeted removal of stranded algal biomass could help reduce nitrogen inputs during summer and limit *Ulva* proliferation, but such measures should be carefully evaluated for each site. While macroalgal wrack is harvested in some countries (Hyndes et al., 2022), removal of stranded algal biomass in France is only allowed when valorization pathways exist. Developing such pathways remains challenging, as stranded algae represent a highly variable resource – both in species composition and in biomass quantity and quality – which hinders the development of consistent and economically viable valorization strategies.

## 5. Conclusion

Our 19-month δ¹⁵N time series and leachate assays together demonstrate that algal wracks provide a source of nitrogen to benthic *Ulva*, highlighting the significant role of stranded macroalgal-derived organic nitrogen in coastal nitrogen cycling.

(1) Nineteen sampling campaigns conducted between March 2017 and October 2018 confirmed clear seasonal δ¹⁵N patterns in *Ulva* tissues, with the lowest values in spring and the highest in summer, consistent with previous observations in the Bay of Seine (Lemesle et al., 2023).

(2) The study highlights the close relationship between algal strandings and the nitrogen supply for local benthic seaweeds in summer, a period characterized by minimal nitrogen inputs from the coastal river.

(3) The δ¹⁵N increase, correlated with the extent of algal wracks, shows that DON and ammonium from decaying seaweeds released in intertidal seawater and accumulating in intertidal sediments represent an endogenous nitrogen source for benthic seaweeds, as confirmed by an *in vitro* experiment.

This study demonstrates that seaweed strandings are an important source of DON and ammonium for coastal primary producers during summer. Their decomposition promotes the growth of opportunistic algae such as *Ulva*, which can in turn lead to further accumulation of

biomass. These findings also suggest that stranded seaweeds should be considered in strategies aimed at improving coastal water quality.

## CRediT authorship contribution statement

**Isabelle Mussio:** Writing – review & editing, Validation, Supervision, Project administration, Funding acquisition, Conceptualization.

**Stéphanie Lemesle:** Writing – review & editing, Writing – original draft, Visualization, Validation, Resources, Methodology, Investigation, Formal analysis. **Pascal Claquin:** Validation, Conceptualization. **Anne-Marie Rusig:** Writing – review & editing, Validation, Supervision, Project administration, Funding acquisition, Conceptualization.

## Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

## Acknowledgments

This study is part of the project "Study of macroalgal heterospecific strandings along the Normandy coastline (Bay of Seine)" and was supported by the Seine-Normandy Water Agency (AESN) and the Normandy Region. We are very grateful to the PLATIN' (PLATeau d'Isotopie de Normandie) core facility for all the elemental and isotopic analysis used in this study. We also thank Guillaume Izabel from the Coastal Environment Research Centre - Marine Station of the University of Caen Normandie (Luc-sur-Mer) for creating the maps presented in Fig. 1. Additionally, we appreciate the support of all technical staff at the Coastal Environment Research Centre - Marine Station of the University of Caen Normandy (Luc-sur-Mer) for their invaluable assistance, notably with the SOMLIT data. The authors would also like to thank Céline Quint (Carré International, University of Caen Normandy) for her valuable assistance with the English language proofreading of this manuscript. We sincerely thank the anonymous reviewers for their constructive comments, which have greatly contributed to the improvement of this manuscript.

## Data availability

Data will be made available on request.
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