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Diatoms are abundant unicellular microalgae, responsible for ≈20 % of global photosynthetic CO2 fixation. Nevertheless, we know little about fundamental aspects of their biology, such as their sexual reproduction. Pheromone‐mediated chemical communication is crucial for successful mating. An attraction pheromone was identified in the diatom Seminavis robusta, but metabolites priming cells for sex and synchronizing search and mating behavior remained elusive. These sex‐inducing pheromones (SIP) induce cell cycle arrest and trigger the production of the attraction pheromone. Here we describe the challenging structure elucidation of an S. robusta SIP. Guided by metabolomics, a candidate metabolite was identified and elucidated by labeling experiments, NMR, ESI MSn analyses, and chemical transformations. The use of negative ion mode MS was essential to decipher the unprecedented hydroxyproline and β‐sulfated aspartate‐containing cyclic heptapeptide that acts in femtomolar concentrations. [ABSTRACT FROM AUTHOR]

The development of metagenomics has opened up a new era in the study of marine microbiota, which play important roles in biogeochemical cycles. In recent years, the global ocean sampling expeditions have spurred this research field toward a deeper understanding of the microbial diversities and functions spanning various lifestyles, planktonic (free-living) or sessile (biofilm-associated). In this review, we deliver a comprehensive summary of marine microbiome datasets generated in global ocean expeditions conducted over the last 20 years, including the Sorcerer II GOS Expedition, the Tara Oceans project, the bioGEOTRACES project, the Micro B3 project, the Bio-GO-SHIP project, and the Marine Biofilms. These datasets have revealed unprecedented insights into the microscopic life in our oceans and led to the publication of world-leading research. We also note the progress of metatranscriptomics and metaproteomics, which are confined to local marine microbiota. Furthermore, approaches to transforming the global ocean microbiome datasets are highlighted, and the state-of-the-art techniques that can be combined with data analyses, which can present fresh perspectives on marine molecular ecology and microbiology, are proposed. [ABSTRACT FROM AUTHOR]

3D printing has become widely used to rapidly prototype and manufacture novel or bespoke objects or replacement components in a wide range of marine industries, engineering, and research. 3D-printed objects are subject to marine biofouling, impacting their operation and longevity. Application of antifouling paints or coatings adds costly and time-consuming steps and may interfere with the function of fine surface features, counteracting some of the benefits of 3D-printing technology. We measured the antifouling performance of two 3D-printing thermoplastics embedded with antifouling biocides to create 3D-printed materials with inherent antifouling properties: 1) polycaprolactone (PCL) mixed with the organic biocide dichlorooctylisothiazolinone (DCOIT) and extruded as 3D-printing filament, and 2) a commercial polylactic acid (PLA) 3D-printing filament with embedded copper powder. Settlement plates printed from these thermoplastics (“PCL-DCOIT” and “PLA-Cu”, respectively) and deployed in temperate, coastal marine water for 17 weeks during summer remained free of macrofouling. A biofilm developed, and 16S and 18S rRNA metabarcoding analyses revealed that early stage biofilms (at 5 and 12 weeks) had dramatically altered assemblage structures of both prokaryotes and eukaryotes compared to natural biofilms. The assemblage on PCL-DCOIT had reduced microbial diversity, strong dominance of Proteobacteria and chlorophytes, and almost complete absence of Flavobacteriia, Cyanobacteria, and diatoms. In contrast, the biofilm on PLA-Cu had a dominance of Flavobacteriia over Proteobacteria, and resistance to chlorophytes, yet similar to PCL-DCOIT it resisted Cyanobacteria and diatoms. Such alterations to biofilm microbial assemblages could influence microbial dynamics, biofilm growth, and settlement cues to which biofouler propagules respond. At 17 weeks, the two biocide-embedded thermoplastics completely resisted macrofouling, equally well as three commercial antifouling coatings (Intercept 8500, Hempaguard X7, Hempasil X3); however, PCL-DCOIT was more extensively covered by a microalgal film (79%, evidently chlorophytes) than were the commercial coatings, and PLA-Cu had the most settled detritus (100% cover). Biofilm assemblages on the commercial coatings were investigated for comparison, with PCL-DCOIT standing out due to its almost complete resistance to Flavobacteriia. Thermoplastic 3D-printing filaments with embedded biocides show promise for producing 3D-printed objects with inherent antifouling properties, avoiding or lessening the need to apply antifouling coatings, and possibly extending their service lifetime.

Marine biofilms were newly revealed as a giant microbial diversity pool for global oceans. However, the cyanobacterial diversity in marine biofilms within the upper seawater column and its ecological and evolutionary implications remains undetermined. Here, we reconstructed a full picture of modern marine cyanobacteria habitats by re-analyzing 9.3 terabyte metagenomic data sets and 2,648 metagenome-assembled genomes (MAGs). The abundances of cyanobacteria lineages exclusively detected in marine biofilms were up to ninefold higher than those in seawater at similar sample size. Analyses revealed that cyanobacteria in marine biofilms are specialists with strong geographical and environmental constraints on their genome and functional adaption, which is in stark contrast to the generalistic features of seawater-derived cyanobacteria. Molecular dating suggests that the important diversifications in biofilm-forming cyanobacteria appear to coincide with the Great Oxidation Event (GOE), "boring billion" middle Proterozoic, and the Neoproterozoic Oxidation Event (NOE). These new insights suggest that marine biofilms are large and important cyanobacterial factories for the global oceans., Importance: 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.

Most members of the Pseudoalteromonas genus have been isolated from living surfaces as members of epiphytic and epizooic microbiomes on marine macroorganisms. Commonly Pseudoalteromonas isolates are reported as a source of bioactive exoproducts, i.e., secondary metabolites, such as exopolymeric substances and extracellular enzymes. The experimental conditions for the production of these agents are commonly associated with sessile metabolic states such as biofilms or liquid cultures in the stationary growth phase. Despite this, the molecular mechanisms that connect biofilm formation and the biosynthesis of exoproducts in Pseudoalteromonas isolates have rarely been mentioned in the literature. This review compiles empirical evidence about exoproduct biosynthesis conditions and molecular mechanisms that regulate sessile metabolic states in Pseudoalteromonas species, to provide a comprehensive perspective on the regulatory convergences that generate the recurrent coexistence of both phenomena in this bacterial genus. This synthesis aims to provide perspectives on the extent of this phenomenon for the optimization of bioprospection studies and biotechnology processes based on these bacteria. [ABSTRACT FROM AUTHOR]

The development of metagenomics has opened up a new era in the study of marine microbiota, which play important roles in biogeochemical cycles. In recent years, the global ocean sampling expeditions have spurred this research field toward a deeper understanding of the microbial diversities and functions spanning various lifestyles, planktonic (free-living) or sessile (biofilm-associated). In this review, we deliver a comprehensive summary of marine microbiome datasets generated in global ocean expeditions conducted over the last 20 years, including the Sorcerer II GOS Expedition, the Tara Oceans project, the bioGEOTRACES project, the Micro B3 project, the Bio-GO-SHIP project, and the Marine Biofilms. These datasets have revealed unprecedented insights into the microscopic life in our oceans and led to the publication of world-leading research. We also note the progress of metatranscriptomics and metaproteomics, which are confined to local marine microbiota. Furthermore, approaches to transforming the global ocean microbiome datasets are highlighted, and the state-of-the-art techniques that can be combined with data analyses, which can present fresh perspectives on marine molecular ecology and microbiology, are proposed.

Introduction: Marine biofilms are the most widely distributed mode of life on Earth and drive biogeochemical cycling processes of most elements. Phosphorus (P) is essential for many biological processes such as energy transfer mechanisms, biological information storage and membrane integrity. Objectives: Our aim was to analyze the effect of a gradient of ecologically relevant phosphate concentrations on the biofilm-forming capacity and the metabolome of the marine bacterium Pseudoalteromonas lipolytica TC8. Methods: In addition to the evaluation of the effect of different phosphate concentration on the biomass, structure and gross biochemical composition of biofilms of P. lipolytica TC8, untargeted metabolomics based on liquid chromatography-mass spectrometry (LC-MS) analysis was used to determine the main metabolites impacted by P-limiting conditions. Annotation of the most discriminating and statistically robust metabolites was performed through the concomitant use of molecular networking and MS/MS fragmentation pattern interpretation. Results: At the lowest phosphate concentration, biomass, carbohydrate content and three-dimensional structures of biofilms tended to decrease. Furthermore, untargeted metabolomics allowed for the discrimination of the biofilm samples obtained at the five phosphate concentrations and the highlighting of a panel of metabolites mainly implied in such a discrimination. A large part of the metabolites of the resulting dataset were then putatively annotated. Ornithine lipids were found in increasing quantity when the phosphate concentration decreased, while the opposite trend was observed for oxidized phosphatidylethanolamines (PEs). Conclusion: This study demonstrated the suitability of LC-MS-based untargeted metabolomics for evaluating the effect of culture conditions on marine bacterial biofilms. More precisely, these results supported the high plasticity of the membrane of P. lipolytica TC8, while the role of the oxidized PEs remains to be clarified. [ABSTRACT FROM AUTHOR]

Marine biofilms are diverse microbial communities and important ecological habitats forming on surfaces submerged in the ocean. Biofilm communities resist environmental disturbance, making them a nuisance to some human activities ("biofouling"). Antifouling solutions rarely address the underlying stability or compositional responses of these biofilms. Using bulk measurements and molecular analyses, we examined temporal and UV-C antifouling-based shifts in marine biofilms in the coastal western North Atlantic Ocean during early fall. Over a 24-day period, bacterial communities shifted from early dominance of Gammaproteobacteria to increased proportions of Alphaproteobacteria, Bacteroidia, and Acidimicrobiia. In a network analysis based on temporal covariance, Rhodobacteraceae (Alphaproteobacteria) nodes were abundant and densely connected with generally positive correlations. In the eukaryotic community, persistent algal, protistan, and invertebrate groups were observed, although consistent temporal succession was not detected. Biofilm UV-C treatment at 13 and 20 days resulted in losses of chlorophyll a and transparent exopolymer particles, indicating biomass disruption. Bacterial community shifts suggested that UV-C treatment decreased the biofilm maturation rate and was associated with proportional shifts among diverse bacterial taxa. UV-C treatment was also associated with increased proportions of protists potentially involved in detritivory and parasitism. Older biofilm communities had increased resistance to UV-C, suggesting that early biofilms are more susceptible to UVC-based antifouling. The results suggest that UV-C irradiation is potentially an effective antifouling method in marine environments in terms of biomass removal and in slowing maturation. However, as they mature, biofilm communities may accumulate microbial members that are tolerant or resilient under UV treatment. [ABSTRACT FROM AUTHOR]

Microbes use signal transduction systems in the processes of swarming motility, antibiotic resistance, virulence, conjugal plasmid transfer, and biofilm formation. However, the signal transduction systems in natural marine biofilms have hardly been profiled. Here we analyzed signal transduction genes in 101 marine biofilm and 91 seawater microbial metagenomes. The abundance of almost all signal transduction-related genes in biofilm microbial communities was significantly higher than that in seawater microbial communities, regardless of substrate types, locations, and durations for biofilm development. In addition, the dominant source microbes of signal transduction genes in marine biofilms were different from those in seawater samples. Co-occurrence network analysis on signal communication between microbes in marine biofilms and seawater microbial communities revealed potential inter-phyla interactions between microorganisms from marine biofilms and seawater. Moreover, phylogenetic tree construction and protein identity comparison displayed that proteins related to signal transductions from Red Sea biofilms were highly similar to those from Red Sea seawater microbial communities, revealing a possible biological basis of interspecies interactions between surface-associated and free-living microbial communities in a local marine environment. Our study revealed the special profile and enrichment of signal transduction systems in marine biofilms and suggested that marine biofilms participate in intercellular interactions of the local ecosystem where they were seeded. [ABSTRACT FROM AUTHOR]

Microbes use signal transduction systems in the processes of swarming motility, antibiotic resistance, virulence, conjugal plasmid transfer, and biofilm formation. However, the signal transduction systems in natural marine biofilms have hardly been profiled. Here we analyzed signal transduction genes in 101 marine biofilm and 91 seawater microbial metagenomes. The abundance of almost all signal transduction-related genes in biofilm microbial communities was significantly higher than that in seawater microbial communities, regardless of substrate types, locations, and durations for biofilm development. In addition, the dominant source microbes of signal transduction genes in marine biofilms were different from those in seawater samples. Co-occurrence network analysis on signal communication between microbes in marine biofilms and seawater microbial communities revealed potential inter-phyla interactions between microorganisms from marine biofilms and seawater. Moreover, phylogenetic tree construction and protein identity comparison displayed that proteins related to signal transductions from Red Sea biofilms were highly similar to those from Red Sea seawater microbial communities, revealing a possible biological basis of interspecies interactions between surface-associated and free-living microbial communities in a local marine environment. Our study revealed the special profile and enrichment of signal transduction systems in marine biofilms and suggested that marine biofilms participate in intercellular interactions of the local ecosystem where they were seeded.

Free‐living (FL) marine microbial communities differ from those attached to particles (PA). Likewise, biofilms (B) colonizing artificial surfaces, including plastics or ship hulls, hardly resemble their planktonic surroundings. However, few studies have examined the effect of the environment on these lifestyles and on the source of organisms colonizing marine surfaces. Using 16S rRNA gene metabarcoding, we identified specificities of marine prokaryotic community lifestyles (FL, PA or B) sampled in three coastal polluted locations with dissimilar environmental conditions: the North‐Western Mediterranean Sea and the Atlantic and Indian Oceans. Biofilms developed over polyvinyl chloride (PVC) were found to be significantly different from FL or PA collected during the immersions. Alpha‐diversity increased from FL to PA and to B, illustrating the integrative aspect of the latter, with little proportion of operational taxonomic units shared with the first two. Beta‐diversity clustered first the lifestyles and then the sites. FL and PA were more affected by water quality, especially by trace metal contamination, whereas B were as sensitive to trace metals as to nutrients. Although biofilms should be supplied by the planktonic (ultra) rare biosphere, source tracking could only detect small contributions of FL or PA taxa to B communities. [ABSTRACT FROM AUTHOR]

Planctomycetes occur in almost all aquatic ecosystems on earth. They have a remarkable cell biology, and members of the orders Planctomycetales and Pirellulales feature cell division by polar budding, perform a lifestyle switch from sessile to motile cells and have an enlarged periplasmic space. Here, we characterise a novel planctomycetal strain, Pla110T, isolated from the surface of polystyrene particles incubated in the Baltic Sea. After phylogenetic analysis, the strain could be placed in the family Planctomycetaceae. Strain Pla110T performs cell division by budding, has crateriform structures and grows in aggregates or rosettes. The strain is a chemoheterotroph, grows under mesophilic and neutrophilic conditions, and exhibited a doubling time of 21 h. Based on our phylogenetic and morphological characterisation, strain Pla110T (DSM 103387T = LMG 29693T) is concluded to represent a novel species belonging to a novel genus, for which we propose the name Polystyrenella longa gen. nov., sp. nov. [ABSTRACT FROM AUTHOR]

Microbes respond to environmental stimuli through complicated signal transduction systems. In microbial biofilms, because of complex multiple species interactions, signals transduction systems are of an even higher complexity. Here, we performed a signal-molecule-treatment experiment to study the role of different signal molecules, including N-hexanoyl-L-homoserine lactone (C6-HSL), N-dodecanoyl-L-homoserine lactone (C12-HSL), Pseudomonas quinolone signal (PQS), and cyclic di-GMP (c-di-GMP), in the development of marine biofilms. Comparative metagenomics suggested a distinctive influence of these molecules on the microbial structure and function of multi-species biofilm communities in its developing stage. The PQS-treated biofilms shared the least similarity with the control and initial biofilms. The role of PQS in biofilm development was further explored experimentally with the strain Erythrobacter sp. HKB8 isolated from marine biofilms. Comparative transcriptomic analysis showed that 314 genes, such as those related to signal transduction and biofilm formation, were differentially expressed in the untreated and PQS-treated Erythrobacter sp. HKB8 biofilms. Our study demonstrated the different roles of signal molecules in marine biofilm development. In particular, the PQS-based signal transduction system, which is frequently detected in marine biofilms, may play an important role in regulating microbe-microbe interactions and the assemblage of biofilm communities.

Marine biofilms are known to influence the corrosion of metal surfaces in the marine environment. Despite some recent research, the succession of bacterial communities colonizing artificial surfaces remains uncharacterized in some temporal settings. More specifically, it is not fully known if bacterial colonizers of artificial surfaces are similar or distinct in the different seasons of the year. In particular the study of early biofilms, in which the bacterial cells communities first adhere to artificial surfaces, are crucial for the development of the subsequent biofilm communities. In this work, we used amplicon-based NGS (next-generation sequencing) and universal 16S rRNA bacterial primers to characterize the early biofilm bacterial communities growing on 316 L stainless steel surfaces in a Northern Portugal port. Sampling spanned 30-day periods in two distinct seasons (spring and winter). Biofilm communities growing in steel surfaces covered with an anti-corrosion paint and planktonic communities from the same location were also characterized. Our results demonstrated that distinct temporal patterns were observed in the sampled seasons. Specifically, a significantly higher abundance of Gammaproteobacteria and Mollicutes was found on the first days of biofilm growth in spring (day 1 to day 4) and a higher abundance of Alphaproteobacteria during the same days of biofilm growth in winter. In the last sampled day (day 30), the spring biofilms significantly shifted toward a dominance of photoautotrophic groups (mostly diatoms) and were also colonized by some macrofouling communities, something not observed during the winter sampling. Our results revealed that bacterial composition in the biofilms was particularly affected by the sampled day of the specific season, more so than the overall effect of the season or overall sampling day of both seasons. Additionally, the application of a non-fouling-release anti-corrosion paint in the steel plates resulted in a significantly lower diversity compared with plates without paint, but this was only observed during spring. We suggest that temporal succession of marine biofilm communities should be taken in consideration for future antifouling/anti-biofilm applications.

Microbes respond to environmental stimuli through complicated signal transduction systems. In microbial biofilms, because of complex multiple species interactions, signals transduction systems are of an even higher complexity. Here, we performed a signal-molecule-treatment experiment to study the role of different signal molecules, including N-hexanoyl-L-homoserine lactone (C6-HSL), N-dodecanoyl-L-homoserine lactone (C12-HSL), Pseudomonas quinolone signal (PQS), and cyclic di-GMP (c-di-GMP), in the development of marine biofilms. Comparative metagenomics suggested a distinctive influence of these molecules on the microbial structure and function of multi-species biofilm communities in its developing stage. The PQS-treated biofilms shared the least similarity with the control and initial biofilms. The role of PQS in biofilm development was further explored experimentally with the strain Erythrobacter sp. HKB8 isolated from marine biofilms. Comparative transcriptomic analysis showed that 314 genes, such as those related to signal transduction and biofilm formation, were differentially expressed in the untreated and PQS-treated Erythrobacter sp. HKB8 biofilms. Our study demonstrated the different roles of signal molecules in marine biofilm development. In particular, the PQS-based signal transduction system, which is frequently detected in marine biofilms, may play an important role in regulating microbe-microbe interactions and the assemblage of biofilm communities. [ABSTRACT FROM AUTHOR]

Marine biofilms are known to influence the corrosion of metal surfaces in the marine environment. Despite some recent research, the succession of bacterial communities colonizing artificial surfaces remains uncharacterized in some temporal settings. More specifically, it is not fully known if bacterial colonizers of artificial surfaces are similar or distinct in the different seasons of the year. In particular the study of early biofilms, in which the bacterial cells communities first adhere to artificial surfaces, are crucial for the development of the subsequent biofilm communities. In this work, we used amplicon-based NGS (next-generation sequencing) and universal 16S rRNA bacterial primers to characterize the early biofilm bacterial communities growing on 316 L stainless steel surfaces in a Northern Portugal port. Sampling spanned 30-day periods in two distinct seasons (spring and winter). Biofilm communities growing in steel surfaces covered with an anti-corrosion paint and planktonic communities from the same location were also characterized. Our results demonstrated that distinct temporal patterns were observed in the sampled seasons. Specifically, a significantly higher abundance of Gammaproteobacteria and Mollicutes was found on the first days of biofilm growth in spring (day 1 to day 4) and a higher abundance of Alphaproteobacteria during the same days of biofilm growth in winter. In the last sampled day (day 30), the spring biofilms significantly shifted toward a dominance of photoautotrophic groups (mostly diatoms) and were also colonized by some macrofouling communities, something not observed during the winter sampling. Our results revealed that bacterial composition in the biofilms was particularly affected by the sampled day of the specific season, more so than the overall effect of the season or overall sampling day of both seasons. Additionally, the application of a non-fouling-release anti-corrosion paint in the steel plates resulted in a significantly lower diversity compared with plates without paint, but this was only observed during spring. We suggest that temporal succession of marine biofilm communities should be taken in consideration for future antifouling/anti-biofilm applications. [ABSTRACT FROM AUTHOR]

Planctomycetes are bacteria that were long thought to be unculturable, of low abundance, and therefore neglectable in the environment. This view changed in recent years, after it was shown that members of the phylum Planctomycetes can be abundant in many aquatic environments, e.g., in the epiphytic communities on macroalgae surfaces. Here, we analyzed three different macroalgae from the North Sea and show that Planctomycetes is the most abundant bacterial phylum on the alga Fucus sp., while it represents a minor fraction of the surface-associated bacterial community of Ulva sp. and Laminaria sp. Especially dominant within the phylum Planctomycetes were Blastopirellula sp., followed by Rhodopirellula sp., Rubripirellula sp., as well as other Pirellulaceae and Lacipirellulaceae, but also members of the OM190 lineage. Motivated by the observed abundance, we isolated four novel planctomycetal strains to expand the collection of species available as axenic cultures since access to different strains is a prerequisite to investigate the success of planctomycetes in marine environments. The isolated strains constitute four novel species belonging to one novel and three previously described genera in the order Pirellulales, class Planctomycetia, phylum Planctomycetes.

Abstract It is not straightforward to link biofilm parameters to frictional drag, because of the heterogeneous distribution and viscoelasticity of the produced matrix. Here we present the design and calibration of a flow cell in which marine biofilms can be cultured under flow and then assessed for drag, structural and mechanical properties. The flow cell test section comprised a rectangular channel constructed by sandwiching together rigid PVC panels and side panels of clear acrylic, which allow natural light to enter. The Fanning friction factor (C f) of the flow cell was found by measuring the pressure drop (ΔP) at various flow velocities (u). Flow cell calibration was carried out using a clean inert marine coating, and various roughness grades of sandpaper sheets to find C f for each rigid roughness. ΔP was proportional to u 2, indicating flow was turbulent (R2 = 0.99). The top panel of the flow cell can be substituted with a clear acrylic lid to allow simultaneous measurement of C f and biofilm physico-mechanical properties by optical coherence tomography (OCT). Here, we demonstrated that the flow cell can be used to image microbial fouling using OCT. Future experiments will assess physico-mechanical and drag properties of marine fouling biofilms under flow. [ABSTRACT FROM AUTHOR]

Abstract A novel fouling marine flow cell was designed and fitted with a clear plastic lid to allow real-time imaging of biofilms using optical coherence tomography (OCT). Marine biofilms were grown under controlled shear flow on coupons coated with 6 different biocidal antifouling coatings (SPC1, SPC2, SPC3, SPC4, CDP1 and CDP2, AkzoNobel) and one inert coating which contained no biocidal actives (NB-D) for 8 weeks. One set of coupons coated with NB was statically immersed in sea water during the same time period (NB-S). Biofilm removal was assessed by increasing the flow velocity while OCT simultaneously measured the biofilm cross-sectional area (CSA). The highest initial removal rates were observed for NB-S, NB-D, and SPC2. Percent biofilm cross-sectional area reduction (%CSAred) was higher on SPCs (>60%) compared to CDPs (<50%). SPCs had the highest percent reduction in biofilm surface area coverage (%SACred >60%) compared to the CDPs (<20%). The marine biofilm flow cell combined with OCT can be used to screen for coating-specific differences in biofilm growth and removal in real time rather than traditional before and after surface area coverage measurements. Future testing will focus on how the biofilm-coatings interactions interact with biofilm mechanical and structural properties to produce drag. [ABSTRACT FROM AUTHOR]

Metabarcoding and metabolomics were used to explore the taxonomic composition and functional diversity of eukaryotic biofouling communities on plates with antifouling paints at two French coastal sites: Lorient (North Eastern Atlantic Ocean; temperate and eutrophic) and Toulon (North-Western Mediterranean Sea; mesotrophic but highly contaminated). Four distinct coatings were tested at each site and season for one month. Metabarcoding showed biocidal coatings had less impact on eukaryotic assemblages compared to spatial and temporal effects. Ciliophora, Chlorophyceae or Cnidaria (mainly hydrozoans) were abundant at Lorient, whereas Arthropoda (especially crustaceans), Nematoda, and Ochrophyta dominated less diversified assemblages at Toulon. Seasonal shifts were observed at Lorient, but not Toulon. Metabolomics also showed clear site discrimination, but these were associated with a coating and not season dependent clustering. The meta-omics analysis enabled identifications of some associative patterns between metabolomic profiles and specific taxa, in particular those colonizing the plates with biocidal coatings at Lorient. [ABSTRACT FROM AUTHOR]

The cyclic peptides portoamides produced by the cyanobacterium Phormidium sp. LEGE 05292 were previously isolated and their ability to condition microcommunities by allelopathic effect was described. These interesting bioactive properties are, however, still underexplored as their biotechnological applications may be vast. This study aims to investigate the antifouling potential of portoamides, given that a challenge in the search for new environmentally friendly antifouling products is to find non-toxic natural alternatives with the ability to prevent colonization of different biofouling species, from bacteria to macroinvertebrates. A multi-bioassay approach was applied to assess portoamides antifouling properties, marine ecotoxicity and molecular mode of action. Results showed high effectiveness in the prevention of mussel larvae settlement (EC50 = 3.16 µM), and also bioactivity towards growth and biofilm disruption of marine biofouling bacterial strains, while not showing toxicity towards both target and non-target species. Antifouling molecular targets in mussel larvae include energy metabolism modifications (failure in proton-transporting ATPases activity), structural alterations of the gills and protein and gene regulatory mechanisms. Overall, portoamides reveal a broad-spectrum bioactivity towards diverse biofouling species, including a non-toxic and reversible effect towards mussel larvae, showing potential to be incorporated as an active ingredient in antifouling coatings.

Temperature variability is a major driver of ecological pattern, with recent changes in average and extreme temperatures having significant impacts on populations, communities and ecosystems. In the marine realm, very few experiments have manipulated temperature in situ, and current understanding of temperature effects on community dynamics is limited. We developed new technology for precise seawater temperature control to examine warming effects on communities of bacteria, microbial eukaryotes (protists) and metazoans. Despite highly contrasting phylogenies, size spectra and diversity levels, the three community types responded similarly to seawater warming treatments of +3°C and +5°C, highlighting the critical and overarching importance of temperature in structuring communities. Temperature effects were detectable at coarse taxonomic resolutions and many taxa responded positively to warming, leading to increased abundances at the community-level. Novel field-based experimental approaches are essential to improve mechanistic understanding of how ocean warming will alter the structure and functioning of diverse marine communities. [ABSTRACT FROM AUTHOR]

At low tide, intertidal mudflat biofilms cover large surfaces and are mainly responsible for the high productivity of these marine areas. In the European Atlantic coast, such biofilms are mainly composed of diatoms, especially Navicula phyllepta, bacteria, and microbial extracellular polymeric substances (EPS). To better understand interactions occurring between microorganisms, we first axenized a N. phyllepta culture with a new and simple protocol. Colloidal and bound EPS secreted by diatom cells during the exponential growth and the stationary phase were then harvested, and we tested their effects on the in vitro formation of biofilms by three marine bacteria. The latter had been isolated from a French Atlantic intertidal mudflat and were previously selected for their strong in vitro biofilm-forming ability. They belong to the Flavobacterium, Roseobacter, and Shewanella genera. Navicula phyllepta-bound EPS synthesized during the stationary phase specifically inhibited the biofilm formation by the Flavobacterium sp. strain, whereas they stimulated biofilm development by the two other strains. The EPS acted in all cases during the first stages of the biofilm establishment. Saccharidic molecules were found to be responsible for these activities. This is the first report on marine bacterial antibiofilm saccharides of microalgal origin. This work points out the complexity of the benthic natural biofilms with specific microalgae/bacteria interactions and underlines the possibility to use axenic diatoms as a source of bioactive compounds. [ABSTRACT FROM AUTHOR]

Marine biofilms are multispecies microbial communities on surfaces that are crucial to the marine environment. They cause marine corrosion, biofouling, and transmission of marine pathogens and thus pose a great threat to public health and the maritime industry. To control marine biofilms, effective and environmentally friendly antibiofilm compounds are highly needed. Elasnin is a potent antibiofilm compound that exhibits high efficiency in inhibiting marine biofilms and biofouling, but its mode of action remains unclear. In the present study, multiomic analysis combined with quorum-sensing assays and in silico study revealed that elasnin acted as a signaling molecule in the microbial community. Elasnin promoted the growth of dominant species in the biofilm but deprived their ability of sensing and responding to environmental changes by disturbing their regulations of the two-component system, i.e. , the ATP-binding cassette transport system and the bacterial secretion system. Consequently, biofilm maturation and subsequent biofouler settlement were inhibited. Elasnin also exhibited higher antibiofilm efficiency than dichlorooctylisothiazolinone and had low toxicity potential on the embryos and adults of marine medaka fish. Overall, this study provided molecular and ecological insights into elasnin's mode of action, highlighting its application potential in controlling marine biofilms and the feasibility and advantages of using signal molecules to develop eco-friendly technologies.

16S rRNA gene profiling using a pipeline involving the Greengenes database revealed that bacterial populations in innermost (proximal to the steel surface) and outer regions of biofilms on carbon steel exposed 3 m below the surface at an offshore site in the Gulf of Guinea differed from one another and from seawater. There was a preponderance of gammaproteobacterial sequences, representing organisms known for hydrocarbon degradation. Total DNA from the innermost layer was 1500 times that recovered from the outermost. Stramenopiles (diatom) sequences were prevalent in the former. Rhodobacteriaceae, key biofilm formers, comprised 14.9% and 4.22% OTUs of inner and outer layers, respectively. Photosynthetic anaerobic sulfur oxidizer sequences were also prominent in the biofilms. Analysis of data using a different pipeline with Silva111 allowed detection of 0.3–0.4% SRB in the biofilms. The high abundance of aerobic micro-algal sequences in inner biofilm suggests they are initial colonizers of carbon steel surfaces in a marine environment. This is the first time that the microbial population of the strongly attached inner layer of the biofilm on steel has been differentiated from the outer, readily removed layer. The accepted scraping removal method is obviously inadequate and the resulting microbial analysis does not offer complete information on the biofilm community structure. [ABSTRACT FROM AUTHOR]

In this experimental study the patterns in early marine biofouling communities and possible implications for surveillance and environmental management were explored using metabarcoding,viz.18S ribosomal RNA gene barcoding in combination with high-throughput sequencing. The community structure of eukaryotic assemblages and the patterns of initial succession were assessed from settlement plates deployed in a busy port for one, five and 15 days. The metabarcoding results were verified with traditional morphological identification of taxa from selected experimental plates. Metabarcoding analysis identified > 400 taxa at a comparatively low taxonomic level and morphological analysis resulted in the detection of 25 taxa at varying levels of resolution. Despite the differences in resolution, data from both methods were consistent at high taxonomic levels and similar patterns in community shifts were observed. A high percentage of sequences belonging to genera known to contain non-indigenous species (NIS) were detected after exposure for only one day. [ABSTRACT FROM PUBLISHER]

One of the greatest challenges in the development of Microbial Electrochemical Technologies (METs) is the achievement of efficient bioanodes, which not only can operate for long term but which can also be effectively and promptly regenerated for the sustained and successive production of electricity out of waste organics contained in aqueous streams. Simple strategies that facilitate the engineering of these systems are then pursued. Sustainable electricity generation was here achieved using electrochemically-active marine biofilms, which reached up to 6.8 A m −2 in the best case. These biofilms showed deteriorated current generation after successive transfers in fresh natural media. The electricity-generation functionality of these marine biofilms was recuperated after their relocation into synthetic minimal media (i.e. up to about 3.8 A m −2 after a decay down to about 1–2 A m −2 ). Upon this relocation, the overall electrochemical mechanisms were preserved. Fluctuating nutrient stress intensified the effect of minimal media on current generation. The change from natural to minimal media showed an important impact on the selection and adaptation of microbial communities, characterized by CE-SSCP profiles; yet, robust bioanodes in which some microbial species were preserved were obtained in synthetic minimal media, these being sufficient for a reproducible electrochemical functionality. The systematic cycling between natural media and minimal media, regarded as periodic stress conditioning, is therefore proposed as a convenient strategy to boost current generation in robust electrochemically-active marine bioanodes. [ABSTRACT FROM AUTHOR]

Diatoms are tenacious fouling organisms and have been observed to remain on ship hull coatings at speeds in excess of 30 knots. This study was designed to investigate the adhesion of diatoms to three fouling release ship hull coatings, Intersleek 700 (IS 700), Intersleek 900 (IS 900), and Hempasil X3 (HX3) and one reference coating Dow Corning 3140 (DC 3140), that were exposed at a static immersion test site. The coatings were subjected to water jet testing to determine biofilm adhesion and the subsequent pressures needed to remove specific diatom species. Differences in community composition were observed among coatings in the pre and post water jet evaluation. All diatoms were removed from HX3 after 0.08 MPa; however, the pressure required to completely remove biofilm and diatoms on DC 3140, IS 700, and IS 900 varied. Biofilm removal pressure ranged from 0.08 to 0.40 MPa on DC 3140, 0.16 to 0.40 MPa on IS 700, and 0.08 to 0.40 MPa on IS 900. Significant differences were observed among coatings for removal pressure, as well as initial diatom abundance; however, differences were dependent on the sampling date. The diatom genera which required the highest pressure for removal from the fouling release coatings were Achnanthes, Amphora, Cocconeis, Navicula, and Synedra. The results show differences in the adhesion strength of diatoms to different fouling release coatings and highlight those species, which have the potential to contribute to biofilms remaining on ship hulls once a vessel is underway. [ABSTRACT FROM AUTHOR]

Biofilms are integral to many marine processes but their formation and function may be affected by anthropogenic inputs that alter environmental conditions, including fertilisers that increase nutrients. Density composition and connectivity of biofilms developedin situ(under ambient and elevated nutrients) were compared using 454-pyrosequencing of the 16S gene. Elevated nutrients shifted community composition from bacteria involved in higher processes (egPseudoalteromonasspp.invertebrate recruitment) towards more nutrient-tolerant bacterial species (egTerendinibactersp.). This may enable the persistence of biofilm communities by increasing resistance to nutrient inputs. A core biofilm microbiome was identified (predominantly Alteromonadales and Oceanospirillales) and revealed shifts in abundances of core microbes that could indicate enrichment by fertilisers. Fertiliser decreased density and connectivity within biofilms indicating that associations were disrupted perhapsviachanges to energetic allocations within the core microbiome. Density composition and connectivity changes suggest nutrients can affect the stability and function of these important marine communities. [ABSTRACT FROM AUTHOR]

Periphyton communities are complex phototrophic, multispecies biofilms that develop on surfaces in aquatic environments. These communities harbor a large diversity of organisms comprising viruses, bacteria, algae, fungi, protozoans, and metazoans. However, thus far the total biodiversity of periphyton has not been described. In this study, we use metagenomics to characterize periphyton communities from the marine environment of the Swedish west coast. Although we found approximately ten times more eukaryotic rRNA marker gene sequences compared to prokaryotic, the whole metagenome-based similarity searches showed that bacteria constitute the most abundant phyla in these biofilms. We show that marine periphyton encompass a range of heterotrophic and phototrophic organisms. Heterotrophic bacteria, including the majority of proteobacterial clades and Bacteroidetes, and eukaryotic macroinvertebrates were found to dominate periphyton. The phototrophic groups comprise Cyanobacteria and the alpha-proteobacterial genus Roseobacter, followed by different micro- and macro-algae. We also assess the metabolic pathways that predispose these communities to an attached lifestyle. Functional indicators of the biofilm form of life in periphyton involve genes coding for enzymes that catalyze the production and degradation of extracellular polymeric substances, mainly in the form of complex sugars such as starch and glycogen-like meshes together with chitin. Genes for 278 different transporter proteins were detected in the metagenome, constituting the most abundant protein complexes. Finally, genes encoding enzymes that participate in anaerobic pathways, such as denitrification and methanogenesis, were detected suggesting the presence of anaerobic or low-oxygen micro-zones within the biofilms. [ABSTRACT FROM AUTHOR]

Biofilm formation is the initial step of the complex marine biofouling process, limiting the performance of submerged surfaces in numerous applications. No technology is known up to now capable to stop the marine biofilm development even on biocidal paints coated surfaces. Several non-toxic approaches for control over marine biofilms based on physical, chemical or/and biological methods, most of them inspired by the nature, are currently on search. Surface patterning, enzyme alternatives of toxic biocides and nature derived anti-biofilm agents (natural biocides, bio-surfactants/dispersants and quorum sensing inhibitors) are in the scope of this review. [ABSTRACT FROM AUTHOR]

Abstract: Settlement-inducing cues for larvae of the green-lipped mussel, Perna canaliculus, were characterized from biofilm exudates of mono-species bacteria. Biofilm exudates of Macrococcus sp. AMGM1 and Bacillus sp. AMGB1, previously known to induce settlement of P. canaliculus larvae, were chosen for this study. An innovative step-wise multi-technique approach, including physical methods (molecular weight fractionation and heat treatment) and chemical methods (crude fractionation, sub-sample fractionation, sequential fractionation, lipid fractionation and protease treatment), was used to characterize the chemical cues from these bacterial biofilm exudates. The resulting fractions obtained from these methods were individually tested for their ability to induce larval settlement. Results from the settlement assays indicate that settlement cues from Macrococcus sp. AMGM1 were between 1–3, 3–10 and 10–30kDa. These molecules were thermo-labile to over 70°C, and contained polar proteins and non-polar lipoprotein molecules. Protease-treated biofilm exudates did not elicit a settlement response, indicating the involvement of proteins from Macrococcus sp. AMGM1 in the larval settlement process. In addition, settlement cues from Bacillus sp. AMGB1 were found to be between 10–30 and 30–50kDa, were stable to up to 100°C, and contained glycolipid molecules. These results provide evidence for the presence of multiple settlement cues in the biofilm exudates from both Macrococcus sp. AMGM1 and Bacillus sp. AMGB1. This study provides a comprehensive and inexpensive set of biochemical techniques, which can be used in a step-wise manner, to deduce partial chemical identity of both inductive and non-inductive settlement cues produced by biofilm exudates. The results can be used to decipher complex bacterially mediated larval settlement processes for P. canaliculus and other marine invertebrates. [Copyright &y& Elsevier]

Abstract: Although the importance of picophytoplankton (<3μm) as the major primary producers is well established in the oceanic and coastal marine environment, occurrence of this community in the marine biofilms is not explored. Considering their small size and high abundance which is next to bacteria, the initial heterotrophic colonizers, it was hypothesized that picophytoplankton are the first autotrophic colonizers in marine biofilms. In this study biofilms were developed by immersing glass slides in the tropical waters of the Dona Paula Bay up to a period of 192h. The presence of picophytoplankton and the temporal variations in community structure was investigated flow cytometrically after every 24h. Biofilms were also developed in the laboratory by immersing the glass slides in fresh natural seawater and the sequence of appearance of the microorganisms was assessed with high frequency samplings. Field observations showed the presence of three groups of picophytoplankton, two prokaryotes, Prochlorococcus-like organisms (PRO), Synechococcus (SYN) and the picoeukaryotes (PEUK). Nanoeukaryotes, which are mostly represented by diatoms were also monitored. In the total biofilm community, prokaryotes were dominant throughout the study period wherein contribution of SYN was highest (50%) in the earlier stages which were later overtaken by PRO-like cells. The contribution of PEUK and nanoeukaryotes was always below 20%. Picophytoplankton contribution to the total photosynthetic biomass was >60% in the initial period of biofilm formation, both in terms of numbers and biomass with PEUK as the major contributors. However, after 2days of incubation, their contribution to total chlorophyll declined thus revealing that although their numbers were increasing, picophytoplankton were succeeded by nanoeukaryotes in terms of biomass. Laboratory experiments revealed that heterotrophic bacteria and picophytoplankton appeared within 5min in the biofilms which were followed by the nanoeukaryotes after 5h. This study shows that picophytoplankton are the pioneer autotrophic colonizers in the tropical marine biofilms, suggesting an essential role in the biofilm food web dynamics, especially in the initial stages. [Copyright &y& Elsevier]

Abstract: Investigations were undertaken to elucidate causes of accelerated low water corrosion (ALWC) of steel piling in a harbour in Southern England. Visual inspection revealed features characteristic of ALWC such as the presence of poorly adherent, thick corrosion products of varying morphology, often seen as large blisters randomly located on sections of the structure at the low water mark. Upon the removal of blisters, a bright surface covered with shallow pits was exposed. Representative samples of the corrosion products were collected from the structure and water and sediment specimens were retrieved from selected areas in the harbour for microbiological, chemical and microscopy testing. In the laboratory, field samples were enriched to detect and enumerate communities of sulphur-oxidising bacteria (SOB) and sulphate-reducing bacteria (SRB). Biofilms, comprising SRB and SOB populations isolated from a sediment sample were grown under static conditions on surfaces of electrodes manufactured from steel piling material. Linear polarisation resistance (LPR) measurements revealed that the corrosion rate of steel with biofilms (0.518mmy−1) was higher than that recorded in sterile seawater alone (0.054mmy−1) and in sterile seawater to which nutrient was added (0.218mmy−1). Scanning electron microscopy (SEM) imaging demonstrated enhanced pitting under biofilms. The results of our investigation revealed for the first time that the attack on steel piling in the presence of sediment SRB and SOB populations was characteristic of ALWC. [Copyright &y& Elsevier]

In the marine environment, biofilms on submerged surfaces can promote or discourage the settlement of invertebrate larvae and macroalgal spores. The settlement-mediating effects of biofilms are believed to involve a variety of biofilm attributes including surface chemistry, micro-topography, and a wide range of microbial products from small-molecule metabolites to high-molecular weight extracellular polymers. The settled organisms in turn can modify microbial species composition of biofilms and thus change the biofilm properties and dynamics. A better understanding of biofilm dynamics and chemical signals released and/or stored by biofilms will facilitate the development of antifouling and mariculture technologies. This review provides a brief account of 1) existing knowledge of marine biofilms that are relevant to settlement mediation, 2) biotechnological application of biofilms with respect to developing non-toxic antifouling technologies and improving the operation of aquaculture facilities, and 3) challenges and future directions for advancing our understanding of settlement-mediating functions of biofilms and for applying this knowledge to real-life situations. [ABSTRACT FROM AUTHOR]

Abstract: The antifouling (AF) paint model of Kiil et al. [S. Kiil, C.E. Weinell, M.S. Pedersen, K. Dam-Johansen, Analysis of self-polishing antifouling paints using rotary experiments and mathematical modelling, Ind. Eng. Chem. Res. 40 (2001) 3906–3920] and the simplified biofilm growth model of Gujer and Wanner [W. Gujer, O. Wanner, Modeling mixed population biofilms, in: W.G. Characklis, K.C. Marshall (Eds.), Biofilms, Wiley–Interscience, New York, 1990] are used to provide a reaction engineering-based insight to the effects of marine microbial slimes on biocide leaching and, to a minor extent, polishing behaviour of AF paints. It is concluded that the perturbation of the local sea water conditions (e.g. pH), as a consequence of the metabolic activity of the biofilm should not affect the net biocide leaching and binder reaction rates significantly. This results from the thin and poorly active biofilms which presumably grow onto the highly effective modern AF paints. According to simulations, the experimental decrease in the biocide leaching rate caused by biofilm growth must be mainly attributed to adsorption of the biocide by the exopolymeric substances secreted by the microorganisms. The effects of biofilms on the leaching of any generic active compound (e.g. natural antifoulants) are discussed in relation to their potential release mechanisms. The largest influence of biofilms is predicted for those active compounds that are released by a diffusion-controlled mechanism (typically tin-free algaecides). [Copyright &y& Elsevier]

Marine microorganisms are capable of successfully colonizing toxic surfaces through the formation of biofilm structures. In this article, most of the literature reporting the presence of marine biofilms on chemically-active antifouling paints is briefly reviewed. Of special concern is the influence of the dense extracellular polymeric substances (EPS) matrix on the release rate of the compounds involved in antifouling paint performance (i.e. active compounds and controlled-release binder molecules). A deeper understanding of these phenomena is of interest for both environmental legislators and paint formulators. [ABSTRACT FROM AUTHOR]

A protocol is described for rapid DNA isolation from marine biofilm microorganisms embedded in large amounts of exopolysaccharides. The method is a modification of the hot phenol protocol used for plants tissues, where nonexpensive and easily available enzymes were used. The method is based on the incubation of biofilm biomass samples in an extraction buffer mixed with phenol preheated at 65°C. The procedure can be completed in 2 h and up to 20 samples can be processed simultaneously with ease and DNA of excellent quality, as shown by successfully amplification of polymerase chain reaction (PCR) products. DNA was recovered from a range of intertidal marine biofilms with varying amounts of exopolysaccharides. [ABSTRACT FROM AUTHOR]

• Nutrient amendment of natural waters for evaluating of multispecies biofilms control. • Biofilm growth was more pronounced in seawater than freshwater. • [C 12 MIM][Cl] and [C 16 MIM][Cl] retarded growth of multispecies biofilms in nutrient amended natural waters. • [C 16 MIM][Cl] showed biofilm dispersal action on pre-formed multispecies biofilms. • [C 16 MIM][Cl] prevented biofilm formation in a re-circulating flow-cell reactor. Biofilm associated problems and inherent resistance of biofilm-bacteria to antimicrobial agents drives the need for alternate agents with potent antibiofilm activities. Alkylimidazolium ionic liquids were reported as novel antimicrobial and antibiofilm agents for preventing single species biofilms. However, naturally occurring biofilms are multispecies in nature constituting diverse organisms and have high prevalence, significance in medical, industrial and ecological perspective. In this study, the effect of three alkylimidazolium ionic liquids ([CnMIM]+[X]−, n = 4, 12 and 16) were evaluated against formation of natural and multispecies biofilms in freshwater and seawater. In 96-well biofilm assays, [C 4 MIM][Cl] with –butyl alkyl group did not exert antibiofilm activity even at 500 µM. [C 12 MIM][I] with –dodecyl alkyl group showed antibiofilm activity at 32 and 250 µM, in natural freshwater and seawater, respectively. Among the tested ionic liquids, [C 16 MIM][Cl] with –hexadecyl group showed superior antibiofilm activity. Presence of [C 16 MIM][Cl] in freshwater and seawater, respectively, at 8 and 125 µM caused complete inhibition in biofilm formation. Among the tested compounds, only [C 16 MIM][Cl] showed biofilm dispersal activity on pre-formed biofilms. Biofilm dispersal was near complete at 125 and 250 µM for freshwater and seawater biofilms, respectively. Potent antimicrobial ionic liquid ([C 16 MIM][Cl]) was further evaluated for biofilm prevention under dynamic conditions in a laboratory scale recirculation system. Over a period of 12 d of operation, biofilm formation was retarded in the presence of 100 μM and 250 μM of [C 16 MIM][Cl], in freshwater and seawater, respectively. The results suggest that antimicrobial [C 16 MIM][Cl] may find potential application for preventing multispecies biofilms in cooling water systems. [ABSTRACT FROM AUTHOR]

Reliable and accurate in situ sensors capable of detecting and quantifying troublesome marine biofilms on metallic surfaces are increasingly necessary. A 0.2 mm diameter gold electrochemical sensor was fully characterized using cyclic voltammetry in abiotic and biotic artificial seawater media within a continuous culture flow cell to detect the growth and development of an aerobic Pseudoalteromonas sp. biofilm. Deconvolution of the abiotic and biotic responses enable the constituent extracellular electron transfer and biofilm responses to be resolved. Differentiation of enhanced oxygen reduction kinetics within the aerobic bacterial biofilm is linked to enzyme and redox mediator activities.