# Functional profiles and organochlorine degradation potential in microbial communities of tropical epilithic biofilms: a case study in Guadeloupe (Lesser Antilles)

Anthony Gouyer, Dominique Monti, Sonia Mion, Philippe Oger, Olivier Gros

► To cite this version:

Anthony Gouyer, Dominique Monti, Sonia Mion, Philippe Oger, Olivier Gros. Functional profiles and organochlorine degradation potential in microbial communities of tropical epilithic biofilms: a case study in Guadeloupe (Lesser Antilles). Canadian Journal of Microbiology, 2025, 71, pp.1-13. 10.1139/cjm-2025-0074. hal-05278443

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Submitted on 27 Nov 2025

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# Bacterial communities of epilithic biofilms in rivers contaminated by pesticides: a case study of Guadeloupe (Lesser Antilles).

¹Gouyer A., ¹Monti D., ²Mion S., ²Oger P., and ¹Gros O.*.

1- Institut de Systématique, Evolution, Biodiversité (ISYEB), Muséum National d'Histoire Naturelle, CNRS, Sorbonne Université, EPHE, Université des Antilles, Pointe-à-Pitre, France.

2-

*corresponding author: olivier.gros@univ-antilles.fr

## Abstract

This study examines the composition, the structure and the predictive associated functions of epilithic bacteria living in a freshwater (RPE) or a mixed-saline (RGA) tropical river biofilm. The bacterial assemblages were explored using high-throughput sequencing and showed 69% of common richness between the two sites, but predominance of cyanobacteria in freshwaters, instead of heterotrophic bacterial classes with Alphaproteobacteria, Betaproteobacteria, Saprospiria and Chitinophagia found in mixed-saline biofilm. This latter showed a more mature and, through predictive functional analysis (FAPROTAX), a more diverse community favoring organic matter degradation and nutrient cycling. Strong relationships were observed with co-occurrence network modeling between nitrogen-fixing and denitrifying bacteria emphasizing their functional interdependencies in the nitrogen cycle. Key microbial interactions between phototrophic cyanobacteria and heterotrophs such as *Fulvivirga* (Cytophagia) were revealed, suggesting a pivotal role of this genera in the carbon cycle. Bacteria species considered able to remove chlorine from pollutants or used worldwide for organochlorine pollutant remediation were found in the epilithic biofilms of these Caribbean rivers.

## 1. Introduction

Since the early 2000s, the French Caribbean territories have been grappling with a major environmental and health crisis. Contamination by organochlorine chemicals was first detected in numerous freshwater resources both in Guadeloupe and Martinique. Chlordecone (CLD) (Kepone®, Curlone®) is currently the most concerning pollutant (see Le Déaut and Procaccia 2009 for extensive review). Once used to control the banana weevil, this chemical is still widespread across aquatic ecosystems, contaminating fish and crustaceans consumed by local populations, with serious health implications (Multigner et al., 2010). Managing, conserving, and preserving both environments and people from organochlorine contamination rely on tools for monitoring and assessing ecosystem quality. The unique functional characteristics of these tropical island aquatic ecosystems (such as the number of migratory species and the high turbulence of waterways, which simplify aquatic trophic networks thereby reducing the candidates for bioindication Coat et al. (2011), have required the development of targeted research. This research was to focus on creating reliable bioindication tools of chlordecone contamination and to be applicable on a large scale. Among the ecological candidates the epilithic biofilm, a spatially stable and long-lasting component, was chosen for its demonstrated effectiveness as a strong bioaccumulator of pesticides (Fernandes et al. 2020) and a good usual
indicator of pesticide pollution on both prokaryotic and eukaryotic communities (Tlili *et al.*, 2008, Dorigo *et al.*, 2010).

Caribbean river's epilithic biofilm has been first chemically and physically analyzed by Hubas *et al.* (2022) and the results highlighted biofilm's bacterial modifications with CLD levels (consistent with earlier studies on the relation between Kepone® and bacteria (Mahaffey *et al.*, 1982)) and mesoscale chemical and physical reactivity correlated with CLD contamination (Monti *et al.*, 2020). The next step in research came with the MetHalo program (2022-2025), funded by the French National Research Agency, which aims to search for bacterial strains capable of metabolizing chlordecone with the hope of bioremediation. Our study takes place in this context, i.e. acquiring basic knowledge about the natural and poorly known bacterial compartments of aquatic biofilms naturally present in contaminated waters, and investigated both in freshwater and in mixed-saline waters to compare and find environments of interest. In this objective, our study characterizes bacterial communities within epilithic biofilm collected from freshwater *sensu stricto* and marine-influenced environment, investigating their composition and potential differences in bacterial structure, organization, and functional roles. Taxonomic composition, predicted metabolic functions using FAPROTAX, and modeled bacterial co-occurrence networks were investigated. We hypothesize that salted transition environments could favor larger and varied communities specialized in heterotrophy and biogeochemical cycles, and bacteria of interest in bioremediation.

## 2. Materials and methods

### 2.1. Sampling

The two study sites are part of a long-term monitoring network managed by the Guadeloupe Water Office, focusing on the physicochemical properties of the waters in Guadeloupe Island (four analyses per year over the past 12 years). The selection of the two stations chosen for comparison, located in two proximate (a few kilometers apart) but different rivers, was the result of a compromise aimed at minimizing possible differences caused by environmental factors—other than the intrusion of marine-origin waters—known to deeply influence the composition and the structure of the epilithic biofilms : the upstream freshwater section of the Rivière Grande Anse being characterized by extreme turbulence, while the Rivière du Pérou lacks a mixing zone in its downstream section. Furthermore, the two sites are part of the same hydro-ecoregions (Wasson *et al.*, 2004) and are the only two with a same landscape of contamination with an average CLD concentration ranging between 1 to 2 µg/L (locations FRIR17 and FRIR22, Office de l'Eau Guadeloupe databases, 2020). They are located on the island of Basse-Terre (Guadeloupe archipelago) and were designed as Rivière Grande Anse (RGA) (15°57'44.5"N 61°39'33.9"W) and Rivière du Pérou (RPE) (16°03'15.0"N 61°34'01.8"W) (Figure 1). The context is a freshwater river *sensu stricto* for the RPE site, while in RGA, it is a mixed water situation, considered to be a transition zone with marine intrusion. The facies in which the samples were immersed were selected in a similar manner, in calm situations with a water velocity of less than 10 cm/s to avoid the influence of turbulence known to be the main factor causing changes in the composition of bacterial communities in epilithic biofilms (Romero *et al.*, 2020). Two baskets made with a 5mm-plastic mesh containing 18 glass microscope slides were immersed for two weeks. In these two stations, for confirmation, the salinity was checked twice during the campaign (WTW Multiline P4). At the end of the two-week immersion, each slide colonized by biofilm was stored in a freezer at -20°C until DNA extraction.
<figure><img src="image_2.png" /><figcaption>Figure 1: Mapping of sampling sites. The RPE sampling site (Rivière Pérou) represents a strictly freshwater environment. The RGA sampling site (Rivière Grande Anse) represents a mixed water environment, considered to be a transition zone with marine intrusions.</figcaption></figure>

## 2.2. Sequencing of 16S rRNA genes

Bulk DNA was extracted from each slide using the FastDNA™ SPIN Kit for Soil (MPBio). The quantity and quality of DNA were measured using NanoDrop (Thermo Scientific). The extracted genomic DNA was stored at -20 °C until PCR amplification. The conditions for PCR amplifications were defined as follows: initial denaturation at 95 °C for 3 min; 29 cycles at 95 °C for 45 s; annealed at 50 °C for 45 s; extension at 72 °C for 1 min 30 s; and a final extension of 5 min at 72 °C. The V3 and V4 regions of the microbial 16s rRNA were amplified using two universal PCR primers (Bac357F (α-CCTACGGGAGGCAGCAG) and Bac926R (β-CCGTCAATTCMTTTRAGT)). The PCR products were performed in triplicates and sent to the company Microsynth (9436 Balgach, Switzerland) for high-throughput sequencing steps and the construction of the OTU abundance tables.

## 2.3. Statistical analyses, functional group prediction and network modeling

Shannon and Piélou's alpha diversity indices were calculated on R using the *Vegan* package. A non-metric multidimensional scaling (NMDS) analysis was performed based on the Bray-Curtis formula, to identify the community structures of all samples. A similarity analysis (ANOSIM) was conducted to examine whether bacterial communities were significantly different. NMDS and ANOSIM analyses were applied to bacterial genera with a relative abundance ≥ 0.2% at least one of the sampled sites.

The prediction of the functional bacterial groups was carried out by cross-referencing OTU data from 16S r DNA sequences with the FAPROTAX database (Louca et al., 2016). The OTUs were annotated with functional information, attributing them to groups involved in metabolic functions related to different biogeochemical cycles and the degradation of organic matter (Yang et al., 2022). The number of OTUs involved in these functions was calculated. Based on this data, a table including the samples and the number of OTUs associated with the different functions was constructed. This table enabled the comparison of relative abundances of various functions between the two sites.
A bacterial taxa co-occurrence model was developed for each sampling site in the form of a network. The networks were created by selecting genera with significant Pearson correlations (ρ ≥ 0.6 and P < 0.05). This analysis of correlations reflects the relationships between the different genera (Yu *et al.*, 2019). The exploration and the visualization of bacterial networks were conducted using the open source software Gephi (Bastian *et al.*, 2009). The central nodes of the networks, which may correspond to the key bacterial genera in the community, were determined on the basis of a degree (number of connections to other nodes) higher than the average degree of the network and a value of the eigenvector centrality values > 0.8 (Ruhnau, 2000). Gephi can also calculate the “modularity” of the network that corresponds to the ability of a network to divide itself into subunits (or modules) containing nodes that are strongly connected to each other. Networks with a high modularity index have dense connections between nodes within modules but sparse connections between nodes in different modules.

# 3. Results

## 3.1. Biodiversity within epilithic biofilm bacterial communities

Salinity checks revealed 0 g/L (conf. int. 0 g/L, N=4) at the RPE station and 15 g/L (conf. int. 6.7 g/L, N=4) at the RGA station, confirming the strictly freshwater status of the former and the presence of saline intrusions at the latter. A total of 1,944,711 high-quality bacterial sequences were obtained from the two sites analyzed. The total OTU richness of all samples was 1,103. The taxonomic analysis revealed 22 bacterial phyla, grouping 39 classes, 88 orders, 163 families, 346 genera for a total of 569 species. A table provided in the supplementary material shows the number of OTUs for each bacterial genus at the two sites. To summarize, in the RPE samples, the main bacterial classes were Cyanophyceae (41.28%), Alphaproteobacteria (26.17%), Betaproteobacteria (6.92%), Cytophagia (5.58%), and Flavobacteriia (4.98%) (Figure 2). For the RGA samples, the main bacterial classes were Alphaproteobacteria (28.18%), Cyanophyceae (18.83%), Betaproteobacteria (17.4%), Saprospiria (13.9%) and Chitinophagia (9.87%) (Figure 2). Significant differences in alpha diversity between the two sites (Wilcoxon test and Student's test, p < 0.05) were observed by measuring species richness and Shannon and Piélou indices (Figure 3). The alpha diversity of bacterial communities was significantly higher and more homogeneously distributed in RPE. This observation is consistent with the analysis of the rank-abundance curves which shows a less pronounced curvature, meaning a lesser dominance by a small number of bacteria genera, in freshwater and a more homogeneous distribution of abundances (Figure 3). Concerning RGA, a slightly steeper curve reduced equitability compared to RPE, with 55% of biodiversity explained by the 7 most abundant species. To reach this same percentage, it would be necessary to take into account the 22 most abundant species in RPE. NMDS analysis showed a strong distinction between the bacterial biodiversity of the two sites (Figure 4). The ANOSIM statistical test (R = 0.9587, p < 0.05) confirms this significant difference in beta diversity between sites.
<figure><img src="image_3.png" /><figcaption>Figure 2: Mean relative abundances (%) of bacterial communities at the class level for the RGA site (mixed water, with marine intrusions) and the RPE site (fresh water).</figcaption></figure>
<figure><img src="image_4.png" /><figcaption>Figure 3: Comparison of the indices associated with biodiversity between the two sampling sites (RGA : mixed water, with marine intrusions and RPE : freshwater) : A - Species richness; B - Shannon index; C - Pielou's index; D – Venn diagram showing shared and unique bacterial species between sites; E – Rank-Abundance Curve (RAC) of bacterial species for each site. For Figures A, B and C, the significance of the results was assessed by a Student's or Wilcoxon's test.</figcaption></figure>
<figure><img src="image_5.png" /><figcaption>Figure 4: Non-metric multidimensional scaling analysis of bacterial community structure in RGA site (mixed water, with marine intrusions) and RPE site (freshwater). made to visualize the proximity of bacterial genera to samples</figcaption></figure>

## 3.2. Metabolism and functional groups

FAPROTAX analysis was used to predict the focal groups present in the biofilm samples and to assess the difference between sites. A total of 768,934 OTUs are involved in the functional groups in RGA while 743,134 OTUs were detected in RPE. In RGA, chemoheterotrophy (47.40%) is the dominant function (Figure 5). Other functions, such as phototrophy (13.95%), chitinolysis (7.27%), nitrate reduction (6.91%), and ureolysis (5.44%) can be noted. For the RPE, phototrophy is dominant (40.49%), followed by chemoheterotrophy (32.92%), nitrate reduction (5.19%), ureolysis (4.22%) and chitinolysis (3.13%) (Figure 5).
<figure><img src="image_6.png" /><figcaption>Figure 5: Mean relative abundances (%) of functional groups predicted by the FAPROTAX database for the RGA site (mixed water, with marine intrusions) and the RPE site (fresh water).</figcaption></figure>

Significant variations in processes associated with the C, N and S cycles could be observed between the two sampling sites. The FAPROTAX analysis revealed that the processes involved in carbon transformation were distributed differently between the two sampling sites. Biofilms collected in RGA had higher numbers of bacterial OTUs involved in chiomioheterotrophy, chitinolysis and hydrogen oxidation under dark conditions (Wilcoxon, p > 0.001) (Figure 6). In RPE, a higher abundance of bacterial OTUs involved in phototrophy, methanotrophy, lignolysis and degradation of aromatic compounds, hydrocarbons and plastics was found (Wilcoxon, p > 0.001) (Figure 6).
<figure><img src="image_7.png" /><figcaption>Figure 6: A functional annotation analysis of prokaryotic taxa (FAPROTAX) was performed to assess the abundance of OTUs associated with the biogeochemical carbon cycle and degradation at the two sites (RGA: mixed water, with marine intrusions and RPE: freshwater). The Y-axis shows the number of OTUs involved in these functions. The significance of the results was assessed by a Wicoxon test.</figcaption></figure>

Concerning biogeochemical cycle of nitrogen, bacteria contributing to nitrogen fixation, nitrate reduction, denitrification and ureolysis were significantly more abundant in RGA compared to RPE (Wilcoxon, p > 0.001) (Figure 7). The bacteria involved in the biogeochemical cycle of sulfur, although being a mechanism used in a minor way within the biofilms studied (Figure 5), were also better represented in RGA, which have significantly higher abundance values for sulfate respiration and sulfur compound respiration (Wilcoxon, p > 0.001) (Figure 7). When looking at OTUs, genera such as *Desulfurivibrio*, *Desulfonatronum* and *Desulfonatronospira*, related to sulfate reduction are, for example, present only in RGA samples. It should be also noted that the abundance of OTUs associated with dark iron oxidation was significantly higher in RGA (Wilcoxon, p > 0.001) (Figure 7).
<figure><img src="image_8.png" /><figcaption>Figure 7: A functional annotation analysis of prokaryotic taxa (FAPROTAX) was performed to assess the abundance of OTUs associated with the biogeochemical cycling of nitrogen, sulfur and iron oxidation at the two sites (RGA: pink; RPE: blue). The RPE site represents a strictly freshwater environment, while the RGA site represents a mixed water environment, with marine intrusions. The Y-axis shows the number of OTUs involved in these functions. The significance of the results was assessed by a Wicoxon test.</figcaption></figure>

### 3.3. Co-occurrence patterns within bacterial communities

Co-occurrence networks were modelled using strong correlations ($p \ge 0.6$, $P < 0.05$). In RGA, the network was composed of 52 nodes (genera belonging to 10 bacterial classes) and 105 edges (significantly important correlations between two nodes). The dominant classes being Alphaproteobacteria, Betaproteobacteria, and Cyanophyceae (32.69%, 19.23%, and 19.23%, respectively) (Figure 8A). The modularity index has a value of 0.634 for this network. On the basis of this index, 8 modules could be isolated (Figure 8B). Concerning RPE, the bacterial network consists of 65 nodes (genera belonging to 11 bacterial classes) and 236 edges. The dominant classes being Alphaproteobacteria and Cyanophyceae (33.95% and 26.15% respectively) (Figure 8C). The modularity index had a value of 0.548 and 6 modules were isolated (Figure 8D).
<figure><img src="image_9.png" /><figcaption>Figure 8: Networks of coexisting bacterial classes based on Pearson correlation analysis at the two sites (RGA: mixed water with marine intrusions; RPE: freshwater). A connection corresponds to a strong (ρ ≥ 0.6) and significant (P < 0.05) correlation. The size of each node is proportional to the number of connections (i.e., degree). RGA (A) and RPE (C) networks colored according to bacterial classes. RGA (B) and RPE (D) networks are colored according to the modules (a module corresponds to a group of nodes that are strongly connected to each other relative to the rest of the network) identified within the network.</figcaption></figure>

# 4. Discussion

## 4.1. Diversity of bacterial communities

The Venn diagram (Figure 3D) reveals that nearly 69% of the species richness was common between the two sites suggesting that abundant bacteria are often associated with a wider distribution
(Nemergut *et al.* 2011). Despite this shared richness, significant differences were observed in the relative abundance of OTUs with a higher Piélou index at RPE showing a more equitable distribution of species in the freshwater environment. This result is confirmed by the rank-abundance curve, where RPE displayed reduced dominance by specific taxa compared to RGA, which exhibited a steeper curve and a lower equitability. Furthermore, NMDS ordination and ANOSIM statistical testing (R = 0.9587, p < 0.05) show clear distinctions in community composition between the sites and significant differences are noted: the predominance of cyanobacteria in RPE suggesting a pioneering colonization stage during which these microorganisms contribute to the primary production and the establishment of other microbial groups (Rossi *et al.*, 2015). The hypothesis of a more pioneering biofilm in RPE is consistent with an abundance of Gammaproteobacteria and species from *Flavobacterium* and *Sphingobacterium*, which are bacteria with more favorable growth in the early stages of biofilm formation (Besemer *et al.*, 2012). In mixed-saline waters (RGA), the lower alpha diversity, with a few dominant species and a larger proportion of rare species, suggests a biofilm formed by more competitive species eclipsing species with poorer fitness under changing conditions (Besemer *et al.*, 2007). In this site, a greater diversity and homogeneity of heterotrophic bacterial classes, with Alphaproteobacteria, Betaproteobacteria, Chitinophagia, and Saprospiria suggest a bacterial community more advanced in the colonization process and the establishment of various functional roles related to degradation activities, decomposition of organic matter and nutrient recycling (Dang *et al.*, 2016; Wagner *et al.*, 2002; McIlroy *et al.*, 2014; Kämpfer *et al.*, 2012). These groups would provide functional complementarity that can accelerate biofilm maturation by introducing more specialization in nutrient recycling.

## 4.2. Functional profiles and co-occurrence networks of biofilms

The FAPROTAX analysis highlighted the predominance of chemoheterotrophic activities in the mixed-saline water samples (RGA). These activities, combined with the degradation of organic matter and the recycling of nutrients, are evidence in ecosystems rich in organic matter as well as environments with fine sediments (Romani and Sabater 2001, Romani *et al.*, 2004) and in which respiration (heterotrophy) dominates over primary production (Middelburg *et al.*, 2007). In RGA, phototrophy could be limited by turbidity, which reduces light input, while the load of various nutrients promotes chemoheterotrophy. In these environments, intense respiration could be generated within the biofilms, depending on oxygen gradients, promoting the establishment of metabolisms such as denitrification and alternative breathing pathways to oxygen, such as those using nitrates, nitrites, sulfates and hydrogen (Stief, 2013) or even provide growth niches for organohalide consortia of respiring bacteria (OHRB) employed in microcosms using polluted river sludge for bioremediation (Vandermeeren *et al.*, 2014, Qiu *et al.*, 2020). At the two sampling sites, five bacteria known for their ability to make a consortium able to remove Cl from pollutants such as trichloroethylene, trans-dichloroethene and chloroform (Cui *et al.*, 2020) were found : *Acidovorax*, *Acinetobacter*, *Comamonas*, *Curvibacter* and *Sediminibacterium*, with a greater dominance of *Acinetobacter* and *Sediminibacterium* genera in freshwater. *Acinetobacter* species have been mainly isolated from environments polluted by humans or by hydrocarbons (see Al Atrouni 2016 for review). Knowledge about the natural distribution of this genus in tropical environments is limited (Xin *et al.*, 2014), but in our study the non-baumanii species *A. brisouii*, *A. lactucae*, *A. nosocomialis*, and *A. septicus* the last three of which have proven pathogenicity, dominate largely the occurrences in freshwater, this appeared being strongly decreased with salinity. Often correlated with *E. coli* levels in freshwaters and frequently resistant to antibiotics, bacterial species of the genus *Acinetobacter* could present potential hazard to public health (Serwecińska *et al.*, 2021), and due to their resistance to desiccation and many disinfectants, *Acinetobacter* strains have been proposed as model microorganisms in environmental studies and water monitoring (Jung and Park, 2015). The *Sediminibacterium* genus, as
for it, only represented in RPE by *S. salmoneum*, first described in a Chinese reservoir by Qu and
Wuang (2008), has been widely encountered in natural or even engineered environments as
activated sludge (Ayardza *et al.*, 2014). This dominance completely shifts in mixed-saline waters
(RGA) towards *S. soli*, recently isolated from tropical soils (Wu *et al.*, 2021), which confirms the
functional importance of sediment in the constitution of the biofilm's bacterial assemblages in the
RGA environment.

The results reveal significantly greater phototrophy and methanotrophy in freshwater biofilm
compared to mixed-saline water. For phototrophy, its functional predominance is consistent with the
omnipresence of cyanobacteria. The freshwater assemblages of cyanobacteria largely dominated by
*Anagnostidinema visiae*, freshly described in non-tropical, oxygen-poor, and sulfur-rich groundwater
mats (Mc Govern 2023) and *Arthronema gygaxiana*, widely distributed in both freshwaters and soils
(Casamatta *et al.*, 2005) confirm the great ubiquity of this photosynthetic part of the biofilm.
Concerning methanotrophy, our results reveal the presence of a unique bacterium in RPE:
*Methylocapsa palsarum* which possesses a methane monooxygenase enzyme and uses methane and
methanol as carbon and energy sources (Dedysh *et al.*, 2015). Freshwaters, particularly in tropical
regions, are known to be a significant reservoir of methane, and the cyanobacteria releasing oxygen
in the biofilm could promote activities such as aerobic methanotrophy (Vigderovich *et al.*, 2023).

Interestingly, a significant part of the heterotrophic bacterial consortia observed in the biofilms is
associated with ureolysis. The large number of OTUs linked to this metabolic activity, detected in
both saline and non-saline environments, suggests an influence of external nutrient inputs. One
plausible explanation is the proximity of the sampling sites to agricultural lands, where urea is
commonly used as a nitrogen-rich fertilizer in the cultivation of crops such as sugarcane and bananas,
which are economically important for the region (Archimède et al., 2011).

To complete the FAPROTAX analysis, the co-occurrence networks are based on correlations between
bacterial genera that could facilitate the understanding of the complex bacterial assemblages in the
biofilms. Modular structures could be identified, inferring the complexity of the network and the
functional relationships between bacteria, based on their coexistence. A higher modularity index, as
well as a greater number of modules was observed in mixed-saline biofilm, indicating a more
complex structure. In freshwaters (RPE), according to the large number of nodes belonging to the
cyanobacteria class, a module III (Figure 8D) plays an important role in phototrophy within the
network. Moreover, a strong co-occurrence between these cyanobacteria and the genus *Fulvivirga*
which belongs to the class Cytophagia was observed. According to the recent study of Nguyen *et al.*
(2023), this genus has the particularity of carrying a large number of genes encoding carbohydrate-
active enzymes participating in the biodegradation of polysaccharides, these latter being an
important part of the biofilm matrix biomass. Within the mixed-saline network (RGA), a similar
situation could be observed with a module I, co-occurrence between cyanobacteria and heterotrophs
belonging in particular to the classes of Cytophagia and Betaproteobacteria. These strong
relationships between the photosynthetic guild and these heterotrophic bacteria, brought to light by
our network analysis, reveal the essential role of heterotrophic bacteria in the carbon cycle in these
tropical aquatic environments. Through this biodegradation capacity of a wide variety of organic
compounds, genera such as *Fulvivirga* participate in making available a diversity of oligosaccharides,
potential substrates for other organisms, playing a pivotal role in biofilm functioning.

For the other modules of the two bacterial networks (I, II and IV in RPE; II, III and IV in RGA), a large
part of the nodes are involved in the carbon cycle because they are essentially made up of members
of Alphaproteobacteria, Betaproteobacteria and Chitinophagia. The central nodes are the genera
*Arcicella*, *Rhizobacter*, *Rhodobacter*, *Sphingomonas* and *Sphingorhabdus*, the species related to the
latter being particularly known to use simple organic substrates (mono- or disaccharides) and amino
acids (Nikitin et al., 2004; Stackebrandt et al., 2009; Ko et al., 2017; Jung et al., 2019). For module III
of the mixed-saline network, the genera Haoranjiania, Acidibacter, Niastella and Tsuneonella are
known to be attached to varied heterotrophic metabolisms, using monosaccharides, disaccharides,
and even polysaccharides (Weon et al., 2006; Fan et al., 2011; Falagán et al., 2014; Zhang et al.,
2016). In addition, some central nodes are involved in the nitrogen cycle such as Aquirufa and
Novosphingobium, which are known to reduce nitrates (Tiirola et al., 2005; Pitt et al., 2019). While
the FAPROTAX analysis predicted a relative importance of nitrogen fixation and denitrification in
mixed-saline biofilms, the modelling of co-occurrences reveal interactions between these two
metabolisms. with significant positive correlation between diazotrophic bacteria (I: Rheinheimera; II:
Nordella and Hyphomicrobium; VI: Azohydromonas) and bacteria capable of denitrification (I:
Aquirufa; II: Defluviimonas; VI: Albitalea) (Fesefeldt et al., 1998; Xie et al., 2005; Foesel et al., 2011;
Thajudeen et al., 2017; Qi et al., 2022; Pitt et al., 2019; Khan et al., 2020). Denitrification and
nitrogen fixation are regulatory mechanisms for nitrogen bioavailability in aquatic ecosystems; while
diazotrophs make nitrogen bioavailable by converting nitrogen gas into ammonium, denitrifying
bacteria limit the amount of nitrogen that can be used by most organisms by convert it to nitrogen
(Fulweiler et al., 2013). The combination of these two metabolisms associated with the nitrogen
cycle has already been demonstrated in marine environment (Fulweiler et al., 2013).

4.3. Bioremediation potential of epilithic biofilms in a context of organochlorine pollution

In a context of persistent pollution of the soils and waters of the French West Indies by the remnant
chlordecone, the study of the microbial diversity of epilithic biofilms offers an opportunity to identify
bacteria potentially capable of achieving dechlorination. This reaction, catalyzed by reducing
dehalogenases, is a key step in the biodegradation of organochlorine compounds. In the case of
chlordecone, it would open up the perchlorinated bishomocubane structure, which is essential for its
degradation (Chaussonnerie et al., 2016). The genera Flavobacterium, Nitratireductor, and
Pseudomonas observed mainly in the freshwater biofilms in this study are known to possess reducing
dehalogenases (Xun et al., 1992; Halliwell et al., 2022; Chen et al., 2024). In addition, bacteria of the
Sphingomonadaceae family, such as Novosphingobium barchaimii, Novosphingobium
lindaniclasticum, and Sphingomonas paucimobilis have been implicated in the dechlorination of
hexachlorocyclohexane (HCH) (Saxena et al., 2013; Laquitaine et al., 2016; Wu et al., 2022). Other
bacterial genera, including Azoarcus and Brevundimonas can also degrade lindane (Anfei et al., 2021;
Egorova et al., 2021) and the consortium capable to remove chlorine atoms from pollutants involving
Acidovorax, Acinetobacter, Comamonas, Curvibacter, and Sediminibacterium (Qiu et al. 2020) is also
promising

Acknowledgements

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