Marine Biofilms Research Articles

Bioprospecting of culturable marine biofilm bacteria for novel antimicrobial peptides

Bioprospecting of culturable marine biofilm bacteria for novel antimicrobial peptides

Marine biofilms: cyanobacteria factories for the global oceans.

Cyanobacteria, highly diverse microbial organisms, play a crucial role in Earth's oxygenation and biogeochemical cycling. However, their connection to these processes remains unclear, partly due to incomplete surveys of oceanic niches. Our study uncovered significant cyanobacterial diversity in marine biofilms, showing distinct niche differentiation compared to seawater counterparts. These patterns reflect three key stages of marine cyanobacterial diversification, coinciding with major geological events in the Earth's history.

Bioassays and virtual screening to identify potent natural antifouling compounds from the brown macroalga Dictyota dichotoma

Macroalgae are one of the potential natural sources for the isolation of novel eco-friendly antifouling compounds. In this study, the antifouling activity of an extract of the brown macroalga Dictyota dichotoma collected from the Red Sea was tested against bacteria isolated from the marine biofilm and larval forms of the barnacle. A maximum inhibition of barnacle larval settlement of 89.36% was observed in 25 µg ml-1 extract concentration at 24 h treatment. The secondary metabolite composition of the extract was analyzed by GC-MS and compounds were used as ligands for molecular docking with barnacle cement protein. The toxicity profile of secondary metabolites present in the extract was predicted through in silico analysis. The results indicate that the crude extract of the alga inhibited the biofilm formation by the bacteria and significantly reduced the settlement of the barnacle larvae. GC-MS analysis of the extract revealed the presence of five metabolites, including two fatty acids. All metabolites showed higher binding affinity with barnacle cement than the reference compound, copper. Among the secondary metabolites detected in the algal extract, the cholestane derivative exhibited maximum binding affinity (–14.2 kcal mol-1) with barnacle cement. The metabolites also showed positive for crustacean and fish toxicity in toxicity prediction using an in silico method.

Viral diversity within marine biofilms and interactions with corrosive microbes

In marine environments, a wide variety of microbes like bacteria, and archaea influence on the corrosion of materials. Viruses are widely distributed in biofilms among these microbes and may affect the corrosion process through interactions with key corrosive prokaryotes. However, understanding of the viral communities within biofilms and their interactions with corrosive microbes remains is limited. To improve this knowledge gap, 53 metagenomes were utilized to investigate the diversity of viruses within biofilms on 8 different materials and their interactions with corrosive microbes. Notably, the viruses within biofilms predominantly belonged to Caudoviricetes, and phylogenetic analysis of Caudoviricetes and protein-sharing networks with other environments revealed the presence of numerous novel viral clades in biofilms. The virus‒host linkages revealed a close association between viruses and corrosive microbes in biofilms. This means that viruses may modulate host corrosion-related metabolism through auxiliary metabolic genes. It was observed that the virus could enhance host resistance to metals and antibiotics via horizontal gene transfer. Interestingly, viruses could protect themselves from host antiviral systems through anti-defense systems. This study illustrates the diversity of viruses within biofilms formed on materials and the intricate interactions between viruses and corrosive microbes, showing the potential roles of viruses in corrosive biofilms.

Study of Amino Acid-Derived 3-Acyltetramic Acids as Herbicidal Agents.

The growing problem of herbicide resistance necessitates the development of novel herbicidal active ingredients, together with other integrated weed management approaches. Natural products are a major source of inspiration for novel actives. In previous research, we identified a 3-acyltetramic acid of microbial origin that inhibited algal growth in marine biofilms, at least in part through inhibition of photosystem II. In this work, we demonstrate the herbicidal effect of this lead compound and construct multiple libraries to test the impact of the different substituents of the central scaffold in order to study the structure-activity relationships. Among these analogues, the highest activities were found for medium- to long-chain acyl groups and apolar secondary amino acid residues. Finally, we provide first insights into the herbicidal mechanisms and present preliminary field-trial and ecotoxicological results for TA12-Pro, the most active analogue in our library. Together, this research shows the potential of 3-acyltetramic acids for herbicide development.

Integrating double-labeling HCR-FISH into a multidisciplinary pipeline for biofouling assessment on austenitic stainless steel in brackish seawater circuit

This study modified and integrated a bioimaging method of hybridization chain reaction fluorescence in situ hybridization (HCR-FISH) into a pipeline for assessing biofouling on stainless steel (SS). A modified protocol of double-labeling HCR-FISH was directly applied to two surface types of SS grade EN 1.4404 to detect localized bacteria and sulfate-reducing bacteria (SRB) by targeting bacterial 16 S rRNA genes and dissimilatory sulfite reductase (dsrB) genes, respectively. The protocol was first validated using microbial pure cultures and materials before being integrated into a biofouling assessment pipeline of SS in a laboratory-scale brackish water circuit, incorporating electrochemical, surface, and molecular biology characterization analyses. The double-labeling HCR-FISH improved bioimaging of surface biofilm morphology and microbial distribution, surpassing monochrome staining methods. This method was compatible and complemented other microscopy techniques and molecular biological analyses, providing additional insights into the biofilms and deposits on the alloy surfaces. The implemented assessment pipeline for biofouling determination frequently detected the ennoblement phenomenon in the evolution of marine biofilm on SS surfaces. However, within the experimental timeframe, microbial activities in the brackish seawater circuit did not flourish significantly, resulting in minimal impact on the steel material. Additionally, surface type and roughness may correlate with microbial adhesion, biofilm growth, and the deformation of passivation layers in SS. Despite abundant sessile bacteria, particularly opportunistic microorganisms, on the steel surfaces, no direct correlations with biodeterioration phenomena or influences of surface roughness of an alloy and the presence of biofilm were conclusively established.

Bacterial envelope polysaccharide cues settlement and metamorphosis in the biofouling tubeworm Hydroides elegans

Metamorphosis for many marine invertebrates is triggered by external cues, commonly produced by bacteria. For larvae of Hydroides elegans, lipopolysaccharide (LPS) from the biofilm-dwelling bacterium Cellulophaga lytica induces metamorphosis. To determine whether bacterial LPS is a common metamorphosis-inducing factor for this species, we compare larval responses to LPS from 3 additional inductive Gram-negative marine biofilm bacteria with commercially available LPS from 3 bacteria not known to induce metamorphosis. LPS from all the inductive bacteria trigger metamorphosis, while LPS from non-inductive isolated marine bacteria do not. We then ask, which part of the LPS is the inductive element, the lipid (Lipid-A) or the polysaccharide (O-antigen), and find it is the latter for all four inductive bacteria. Finally, we examine the LPS subunits from two strains of the same bacterial species, one inductive and the other not, and find the LPS and O-antigen to be inductive from only the inductive bacterial strain.

Genomic Analysis of Novel Sulfitobacter Bacterial Strains Isolated from Marine Biofilms.

Bacteria from the genus Sulfitobacter are distributed across various marine habitats and play a significant role in sulfur cycling. However, the metabolic features of Sulfitobacter inhabiting marine biofilms are still not well understood. Here, complete genomes and paired metatranscriptomes of eight Sulfitobacter strains, isolated from biofilms on subtidal stones, have been analyzed to explore their central energy metabolism and potential of secondary metabolite biosynthesis. Based on average nucleotide identity and phylogenetic analysis, the eight strains were classified into six novel species and two novel strains. The reconstruction of the metabolic pathways indicated that all strains had a complete Entner-Doudoroff pathway, pentose phosphate pathway, and diverse pathways for amino acid metabolism, suggesting the presence of an optimized central carbon metabolism. Pangenome analysis further revealed the differences between the gene cluster distribution patterns among the eight strains, suggesting significant functional variation. Moreover, a total of 47 biosynthetic gene clusters were discovered, which were further classified into 37 gene cluster families that showed low similarity with previously documented clusters. Furthermore, metatranscriptomic analysis revealed the expressions of key functional genes involved in the biosynthesis of ribosomal peptides in in situ marine biofilms. Overall, this study sheds new light on the metabolic features, adaptive strategies, and value of genome mining in this group of biofilm-associated Sulfitobacter bacteria.

Understanding the flow behavior around marine biofilms

In vitro platforms capable of mimicking the hydrodynamic conditions prevailing in natural aquatic environments have been previously validated and used to predict the fouling behavior on different surfaces. Computational Fluid Dynamics (CFD) has been used to predict the shear forces occurring in these platforms. In general, these predictions are made for the initial stages of biofilm formation, where the amount of biofilm does not affect the flow behavior, enabling the estimation of the shear forces that initial adhering organisms have to withstand. In this work, we go a step further in understanding the flow behavior when a mature biofilm is present in such platforms to better understand the shear rate distribution affecting marine biofilms. Using 3D images obtained by Optical Coherence Tomography, a mesh was produced and used in CFD simulations. Biofilms of two different marine cyanobacteria were developed in agitated microtiter plates incubated at two different shaking frequencies for 7 weeks. The biofilm-flow interactions were characterized in terms of the velocity field and shear rate distribution. Results show that global hydrodynamics imposed by the different shaking frequencies affect biofilm architecture and also that this architecture affects local hydrodynamics, causing a large heterogeneity in the shear rate field. Biofilm cells located in the streamers of the biofilm are subjected to much higher shear values than those located on the bottom of the streamers and this dispersion in shear rate values increases at lower bulk fluid velocities. This heterogeneity in the shear force field may be a contributing factor for the heterogeneous behavior in metabolic activity, growth status, gene expression pattern, and antibiotic resistance often associated with nutrient availability within the biofilm.

Isolation and Identification of Biofilm forming Bacteria from Glass Panels exposed in Sea Waters of South Andaman, India

Marine organisms attach to surfaces that are submerged in seawater in a process known as biofouling. These organisms can colonize on various surfaces such as ship hulls, pillars, jetties, undersea pipelines, oil rigs and buoys. Foulers can include bacteria, algae, barnacles, mussels and other invertebrates. The initial adhesion of fouling bacteria is the first stage of biofouling. The growth and attachment of marine fouling bacteria are influenced by various environmental factors including salinity, temperature and nutrient availability. The bacteria forming marine biofilm were isolated and identified from glass panels. Basic microbiological phenotypic identification methods and genotypic validation using 16s rRNA gene sequencing were used to identify the bacterial strains. The phylogenetic analysis of the 16s rRNA gene sequences confirmed 13 different species (Bacillus aquimaris (2), Vibrio hepatarius (1), Exiguobacterium profundum (3), Exiguobacterium aestuarii (2), Vibrio parahaemolyticus (2), Vibrio alginolyticus (3), Vibrio azureus (2), Vibrio fluvialis (2), Priestia flexa (1), Pseudomonas aeruginosa (2), Bacillus megaterium (2), Lysinibacillus macroides (2) and Bacillus subtilis (1)) of fouling bacteria belonging to six genera.

Anionic extracellular polymeric substances extracted from seawater-adapted aerobic granular sludge.

Anionic polymers, such as heparin, have been widely applied in the chemical and medical fields, particularly for binding proteins (e.g., fibroblast growth factor 2 (FGF-2) and histones). However, the current animal-based production of heparin brings great risks, including resource shortages and product contamination. Recently, anionic compounds, nonulosonic acids (NulOs), and sulfated glycoconjugates were discovered in the extracellular polymeric substances (EPS) of aerobic granular sludge (AGS). Given the prevalence of anionic polymers, in marine biofilms, it was hypothesized that the EPS from AGS grown under seawater condition could serve as a raw material for producing the alternatives to heparin. This study aimed to isolate and enrich the anionic fractions of EPS and evaluate their potential application in the chemical and medical fields. The AGS was grown in a lab-scale reactor fed with acetate, under the seawater condition (35 g/L sea salt). The EPS was extracted with an alkaline solution at 80 °C and fractionated by size exclusion chromatography. Its protein binding capacity was evaluated by native gel electrophoresis. It was found that the two highest molecular weight fractions (438- > 14,320 kDa) were enriched with NulO and sulfate-containing glycoconjugates. The enriched fractions can strongly bind the two histones involved in sepsis and a model protein used for purification by heparin-column. These findings demonstrated possibilities for the application of the extracted EPS and open up a novel strategy for resource recovery. KEY POINTS: • High MW EPS from seawater-adapted AGS are dominant with sulfated groups and NulOs • Fifty-eight percent of the EPS is high MW of 68-14,320 kDa • EPS and its fractions can bind histones and fibroblast growth factor 2.

Succession of microbial community composition and secondary metabolism during marine biofilm development.

In nature, secondary metabolites mediate interactions between microorganisms residing in complex microbial communities. However, the degree to which community dynamics can be linked to secondary metabolite potential remains largely unknown. In this study, we address the relationship between community succession and secondary metabolism variation. We used 16S and 18S rRNA gene and adenylation domain amplicon sequencing, genome-resolved metagenomics, and untargeted metabolomics to track the taxons, biosynthetic gene clusters, and metabolome dynamics in situ of microorganisms during marine biofilm succession over 113days. Two phases were identified during the community succession, with a clear shift around Day 29, where the alkaloid secondary metabolites, pseudanes, were also detected. The microbial secondary metabolite potential changed between the phases, and only a few community members, including Myxococotta spp., were responsible for the majority of the biosynthetic gene cluster potential in the early succession phase. In the late phase, bryozoans and benthic copepods were detected, and the microbial nonribosomal peptide potential drastically decreased in association with a reduction in the relative abundance of the prolific secondary metabolite producers. Conclusively, this study provides evidence that the early succession of the marine biofilm community favors prokaryotes with high nonribosomal peptide synthetase potential. In contrast, the late succession is dominated by multicellular eukaryotes and a reduction in bacterial nonribosomal peptide synthetase potential.

Synthesis and characterization of ZnO-SiO2 hybrid nanoparticles as an effective inhibitor for marine biofilm and biofouling

In the current study, we have created a highly efficient, economical, and environmentally friendly antifouling approach. ZnO nanoparticles were isolated from Avicennia Marina (a mangrove forest) and combined with SiO2 nanoparticles. ZnO-SiO2 hybrid nanoparticles were synthesized under reflux circumstances (with a 3:1 ratio, 45 mL of ethanol, 4 droplets of sulfuric acid, and 120 °C). ZnO-SiO2 hybrid nanoparticles were characterized through various techniques. Results showed the formation of Si-ZnO bond, indicated by a decrease in the strength of the peak at 470 cm−1 in the FTIR spectra. XRD analysis confirmed the presence of a crystal structure with a crystallite size of 30.72 nm. FE-SEM and TEM revealed spherical and hexagonal morphologies with sizes ranging from 17 to 49 nm and 8.94 nm, respectively. EDS analysis confirmed the presence of Zn, Si, and O elements in the nanoparticles. The minimum inhibitory concentration (MIC) of ZnO-SiO2 hybrid nanoparticles was assessed in the presence of Escherichia coli (50 μg/mL) and Staphylococcus aureus (25 μg/mL), and 10 mg/ml of the ZnO-SiO2 hybrid nanoparticles were capable of inhibiting 88 % of the Isochrysis galbana algae after 48 h, demonstrating their strong potential as an effective inhibitor. In addition, the ZnO-SiO2 hybrid nanoparticles were immersed in natural seawater at varying concentrations for a duration of 90 days to evaluate their effectiveness. The panel containing 3 wt% ZnO-SiO2 hybrid nanoparticles exhibited the most prosperous and efficient outcomes and had the least amount of biofouling (68.3 g). The findings of this research indicate that hybrid nanoparticles of ZnO-SiO2 possess promising and favorable characteristics for antifouling purposes.

Disturbance frequency directs microbial community succession in marine biofilms exposed to shear.

Disturbances are major drivers of community succession in many microbial systems; however, relatively little is known about marine biofilm community succession, especially under antifouling disturbance. Antifouling technologies exert strong local disturbances on marine biofilms, and resulting biomass losses can be accompanied by shifts in biofilm community composition and succession. We address this gap in knowledge by bridging microbial ecology with antifouling technology development. We show that disturbance by shear can strongly alter marine biofilm community succession, acting as a selective filter influenced by frequency of exposure. Examining marine biofilm succession patterns with and without shear revealed stable associations between key prokaryotic and eukaryotic taxa, highlighting the importance of cross-domain assessment in future marine biofilm research. Describing how compounded top-down and bottom-up disturbances shape the succession of marine biofilms is valuable for understanding the assembly and stability of these complex microbial communities and predicting species invasiveness.

Cuprous ions influence on the biocorrosion of a carbon steel in the presence of sulphate reducing bacteria marine biofilm

The influence of marine sulphate reducing bacteria (SRB) biofilm on naval carbon steel corrosion in seawater was studied in the presence or not of cuprous ions as polluting agent. The optical density measurement showed that a concentration of 15 mg L−1 in cuprous ions can cause a significant slowdown in the proliferation of planktonic bacteria. However, electrochemical impedance results indicated that there was still bacterial activity on the metal substrate. Indeed, biofilm was grown on the surface of material in seawater containing SRB. Initially, an increase in the charge transfer resistance values due to the iron sulphide layer formed by the bacteria precipitation reactions was recorded during growth phase. This film acts as a barrier, preventing further oxidation of the underlying metal; then, the localized breakdown of iron sulphide layer generates an acceleration of corrosion. Similar behaviour occurs in the presence of sulphate reducing bacteria flora with 15 mg L−1 cuprous ions. In order to protect themselves when the seawater media containing toxic metals, sulphate reducing bacteria embed into clusters and increase the production of extracellular polymeric substances on materials. Sulphur was present on specimens in both cases and EDS spectra revealed the formation of iron sulphide. After the removal of biofilm, carbon steel surfaces exhibited a plenty of micropits scattered on the entire surface compared to the surface immersed in the seawater without bacteria.

Genomic and transcriptomic insights into complex virus–prokaryote interactions in marine biofilms

Marine biofilms are complex communities of microorganisms that play a crucial ecological role in oceans. Although prokaryotes are the dominant members of these biofilms, little is known about their interactions with viruses. By analysing publicly available and newly sequenced metagenomic data, we identified 2446 virus–prokaryote connections in 84 marine biofilms. Most of these connections were between the bacteriophages in the Uroviricota phylum and the bacteria of Proteobacteria, Cyanobacteria and Bacteroidota. The network of virus–host pairs is complex; a single virus can infect multiple prokaryotic populations or a single prokaryote is susceptible to several viral populations. Analysis of genomes of paired prokaryotes and viruses revealed the presence of 425 putative auxiliary metabolic genes (AMGs), 239 viral genes related to restriction–modification (RM) systems and 38,538 prokaryotic anti-viral defence-related genes involved in 15 defence systems. Transcriptomic evidence from newly established biofilms revealed the expression of viral genes, including AMGs and RM, and prokaryotic defence systems, indicating the active interplay between viruses and prokaryotes. A comparison between biofilms and seawater showed that biofilm prokaryotes have more abundant defence genes than seawater prokaryotes, and the defence gene composition differs between biofilms and the surrounding seawater. Overall, our study unveiled active viruses in natural biofilms and their complex interplay with prokaryotes, which may result in the blooming of defence strategists in biofilms. The detachment of bloomed defence strategists may reduce the infectivity of viruses in seawater and result in the emergence of a novel role of marine biofilms.

Extreme weather events as an important factor for the evolution of plastisphere but not for the degradation process

Marine plastics, with their negative effects on marine life and the human health, have been recently recognized as a new niche for the colonization and development of marine biofilms. Members of the colonizing communities could possess the potential for plastic biodegradation. Thus, there is an urgent need to characterize these complex and geographically variable communities and elucidate the functionalities. In this work, we characterize the fungal and bacterial colonizers of 5 types of plastic films (High Density Polyethylene, Low Density Polyethylene, Polypropylene, Polystyrene and Polyethylene Terepthalate) over the course of a 242-day incubation in the south-eastern Mediterranean and relate them to the chemical changes observed on the surface of the samples via ATR-FTIR. The 16s rRNA and ITS2 ribosomal regions of the plastisphere communities were sequenced on four time points (35, 152, 202 and 242 days). The selection of the time points was dictated by the occurrence of a severe storm which removed biological fouling from the surface of the samples and initiated a second colonization period. The bacterial communities, dominated by Proteobacteria and Bacteroidetes, were the most variable and diverse. Fungal communities, characterized mainly by the presence of Ascomycota, were not significantly affected by the storm. Neither bacterial nor fungal community structure were related to the polymer type acting as substrate, while the surface of the plastic samples underwent weathering of oscillating degrees with time. This work examines the long-term development of Mediterranean epiplastic biofilms and is the first to examine how primary colonization influences the microbial community re-attachment and succession as a response to extreme weather events. Finally, it is one of the few studies to examine fungal communities, despite them containing putative plastic degraders.

Biofilm formation stabilizes metabolism in a Roseobacteraceae bacterium under temperature increase.

Ocean warming profoundly impacts microbes in marine environments; yet, how lifestyle (e.g., free living versus biofilm associated) affects the bacterial response to rising temperature is not clear. Here, we compared transcriptional, enzymatic, and physiological responses of free-living and biofilm-associated Leisingera aquaemixtae M597, a member of the Roseobacteraceae family isolated from marine biofilms, to the increase in temperature from 25℃ to 31℃. Complete genome sequencing and metagenomics revealed the prevalence of M597 in global ocean biofilms. Transcriptomics suggested a significant effect on the expression of genes related to carbohydrate metabolism, nitrogen and sulfur metabolism, and phosphorus utilization of free-living M597 cells due to temperature increase, but such drastic alterations were not observed in its biofilms. In the free-living state, the transcription of the key enzyme participating in the Embden-Meyerhof-Parnas pathway was significantly increased due to the increase in temperature, accompanied by a substantial decrease in the Entner-Doudoroff pathway, but transcripts of these glycolytic enzymes in biofilm-forming strains were independent of the temperature variation. The correlation between the growth condition and the shift in glycolytic pathways under temperature change was confirmed by enzymatic activity assays. Furthermore, the rising temperature affected the growth rate and the production of intracellular reactive oxygen species when M597 cells were free living rather than in biofilms. Thus, biofilm formation stabilizes metabolism in M597 when grown under high temperature and this homeostasis is probably related to the glycolytic pathways.IMPORTANCEBiofilm formation is one of the most successful strategies employed by microbes against environmental fluctuations. In this study, using a marine Roseobacteraceae bacterium, we studied how biofilm formation affects the response of marine bacteria to the increase in temperature. This study enhances our understanding of the function of bacterial biofilms and the microbe-environment interactions in the framework of global climate change.

Biodeterioration and Chemical Corrosion of Concrete in the Marine Environment: Too Complex for Prediction.

Concrete is the most utilized construction material worldwide. In the marine environment, it is subject to chemical degradation through reactions with chloride (the most important ion), and sulfate and magnesium ions in seawater, and to biodeterioration resulting from biological (initially microbiological) activities, principally acid production. These two types of corrosions are reviewed and the failure of attempts to predict the degree of deterioration resulting from each is noted. Chemical (abiotic) corrosion is greatest in the splash zone of coastal constructions, while phenomenological evidence suggests that biodeterioration is greatest in tidal zones. There have been no comparative experiments to determine the rates and types of microbial biofilm formation in these zones. Both chemical and microbiological concrete deteriorations are complex and have not been successfully modeled. The interaction between abiotic corrosion and biofilm formation is considered. EPS can maintain surface hydration, potentially reducing abiotic corrosion. The early marine biofilm contains relatively specific bacterial colonizers, including cyanobacteria and proteobacteria; these change over time, producing a generic concrete biofilm, but the adhesion of microorganisms to concrete in the oceans has been little investigated. The colonization of artificial reefs is briefly discussed. Concrete appears to be a relatively prescriptive substrate, with modifications necessary to increase colonization for the required goal of increasing biological diversity.

Substrate Specificity of Biofilms Proximate to Historic Shipwrecks.

The number of built structures on the seabed, such as shipwrecks, energy platforms, and pipelines, is increasing in coastal and offshore regions. These structures, typically composed of steel or wood, are substrates for microbial attachment and biofilm formation. The success of biofilm growth depends on substrate characteristics and local environmental conditions, though it is unclear which feature is dominant in shaping biofilm microbiomes. The goal of this study was to understand the substrate- and site-specific impacts of built structures on short-term biofilm composition and functional potential. Seafloor experiments were conducted wherein steel and wood surfaces were deployed for four months at distances extending up to 115 m away from three historic (>50 years old) shipwrecks in the Gulf of Mexico. DNA from biofilms on the steel and wood was extracted, and metagenomes were sequenced on an Illumina NextSeq. A bioinformatics analysis revealed that the taxonomic composition was significantly different between substrates and sites, with substrate being the primary determining factor. Regardless of site, the steel biofilms had a higher abundance of genes related to biofilm formation, and sulfur, iron, and nitrogen cycling, while the wood biofilms showed a higher abundance of manganese cycling and methanol oxidation genes. This study demonstrates how substrate composition shapes biofilm microbiomes and suggests that marine biofilms may contribute to nutrient cycling at depth. Analyzing the marine biofilm microbiome provides insight into the ecological impact of anthropogenic structures on the seabed.