The effect of substratum type, orientation and depth on the development of bacterial deep-sea biofilm communities grown on artificial substrata deployed in the Eastern Mediterranean

An increasing number of deep-sea studies have highlighted the importance of deep-sea biofouling, especially in relation to the protection of deep-sea instruments. In this study, the microbial communities developed on different substrata (titanium, aluminum, limestone, shale and neutrino telescope glass) exposed for 155 days at different depths (1500?m, 2500?m, 3500?m and 4500 m) and positions (vertical and horizontal) in the Eastern Mediterranean Deep Sea were compared. Replicated biofilm samples were analyzed using a Terminal Restriction Fragment Length Polymorphisms (T-RFLP) method. The restriction enzymes CfoI and RsaI produced similar total numbers (94, 93) of different T-RFLP peaks (T-RFs) along the vertical transect. In contrast, the mean total T-RF number between each sample according to substratum type and depth was higher in more samples when CfoI was used. The total species richness (S) of the bacterial communities differed significantly between the substrata, and depended on the orientation of each substratum within one depth and throughout the water column (ANOVA). T-RFLP analyses using the Jaccard similarity index showed that within one depth layer, the composition of microbial communities on different substrata was different and highly altered among communities developed on the same substratum but exposed to fouling at different depths. Based on Multidimensional Scaling Analyses (MDS), the study suggests that depth plays an important role in the composition of deep-sea biofouling communities, while substratum type and orientation of substrata throughout the water column are less important. To the authors' knowledge, this is the first study of biofilm development in deep waters, in relation to the effects of substratum type, orientation and depth.     

The effect of surface colour on the formation of marine micro and macrofouling communities

The effect of substratum colour on the formation of micro- and macro fouling communities was investigated. Acrylic tiles, painted either black or white were covered with transparent sheets in order to ensure similar surface properties. All substrata were exposed to biofouling at 1?m depth for 40?d in the Marina Bandar al Rowdha (Muscat, Sea of Oman). Studies were conducted in 2010 over a time course of 5, 10 and 20?d, and in 2012 samples were collected at 7, 14 and 21?d. The densities of bacteria on the black and white substrata were similar with the exception of day 10, when the black substrata had a higher abundance than white ones. Pyrosequencing via 454 of 16S rRNA genes of bacteria from white and black substrata revealed that Alphaproteobacteria and Firmicutes were the dominant groups. SIMPER analysis demonstrated that bacterial phylotypes (uncultured Gammaproteobacteria, Actibacter, Gaetbulicola, Thalassobius and Silicibacter) and the diatoms (Navicula directa, Navicula sp. and Nitzschia sp.) contributed to the dissimilarities between communities developed on white and black substrata. At day 20, the highest amount of chlorophyll a was recorded in biofilms developed on black substrata. SIMPER analysis showed that Folliculina sp., Ulva sp. and Balanus amphitrite were the major macro fouling species that contributed to the dissimilarities between the communities formed on white and black substrata. Higher densities of these species were observed on black tiles. The results emphasise the effect of substratum colour on the formation of micro and macro fouling communities; substratum colour should to be taken into account in future studies.     

Effects of environment on electric field detection by small spotted catshark Scyliorhinus canicula (L.)

This study evaluated the influence of environment (substratum type and depth) on the electroreception capabilities of small spotted catsharks Scyliorhinus canicula in response to prey-simulating electric fields. In experiments where electric fields (applied current 15 μA) were presented beneath different substrata (sand, pebbles, rocks and control) it was found that search effort was not different between substrata or S. canicula sexes, however, both rates of turning and biting towards active electrodes were decreased over pebbles and rocks compared with sand and the control (no substratum). There was no significant effect of sex on turn and bite rates over any substrata. Electric fields were then presented beneath different depths of sand to examine the depth-limits of fish electroreception. Turn and bite rates were significantly lower at depths below 10 mm, with no bites towards electrodes made when they were >30 mm depth. Search effort was not found to be different between different burial depth treatments or between sexes. These results indicate substratum type and depth influences the ability of S. canicula to detect prey-simulating electric fields. This variation in electroreceptive performance may influence space use of sharks.     

Bacterial communities in the initial stage of marine biofilm formation on artificial surfaces

Succession of bacterial communities during the first 36 h of biofilm formation in coastal water was investigated at 3 approximately 15 h intervals. Three kinds of surfaces (i.e., acryl, glass, and steel substratum) were submerged in situ at Sacheon harbor, Korea. Biofilms were harvested by scraping the surfaces, and the compositions of bacterial communities were analyzed by terminal restriction fragment length polymorphism (T-RFLP), and cloning and sequencing of 16S rRNA genes. While community structure based on T-RFLP analysis showed slight differences by substratum, dramatic changes were commonly observed for all substrata between 9 and 24 h. Identification of major populations by 16S rRNA gene sequences indicated that gamma-Proteobacteria (Pseudomonas, Acinetobacter, Alteromonas, and uncultured gamma-Proteobacteria) were predominant in the community during 0 approximately 9 h, while the ratio of alpha-Proteobacteria (Loktanella, Methylobacterium, Pelagibacter, and uncultured alpha-Proteobacteria) increased 2.6 approximately 4.8 folds during 24 approximately 36 h of the biofilm formation, emerging as the most predominant group. Previously, alpha-Proteobacteria were recognized as the pioneering organisms in marine biofilm formation. However, results of this study, which revealed the bacterial succession with finer temporal resolution, indicated some species of gamma-Proteobacteria were more important as the pioneering population. Measures to control pioneering activities of these species can be useful in prevention of marine biofilm formation.     

Mineralogy Influences Structure and Diversity of Bacterial Communities Associated with Geological Substrata in a Pristine Aquifer

Our understanding of mineralogical influences on subsurface microbial community structure and diversity has been difficult to assess due to difficulties in isolating this variable from others in the subsurface environment. In this study, biofilm coupons were used to isolate specific geological substrata from the surrounding geological matrix during colonization by microorganisms suspended in the surrounding groundwater for an 8-week period. Upon retrieval, the structure and diversity of the microbial community associated with each type of substratum was evaluated using 16S rDNA-based terminal-restriction fragment length polymorphism (T-RFLP). Phylogenetic affiliations of the populations associated with each type of substratum were established based on sequence analysis of near full-length 16S rDNA obtained through construction of a clone library. Hematite, quartz, and saprolite each harbored a community dominated by members of the division Proteobacteria (>67% of community). However, the different substrata selected for different subdivisions of bacteria within the Proteobacteria. After accounting for the influence exerted by substratum type on recovery of DNA from the attached populations, both phylogenetic data and Jaccard and Bray–Curtis similarity indices derived from terminal-restriction fragment (T-RF) profiles suggested a strong mineralogical influence on the structure and composition of the solid phase-associated community. The results suggest that mineralogical heterogeneity influences microbial community structure and diversity in pristine aquifers. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized authors.     

Biofilm diatom community structure: Influence of temporal and substratum variability

Abstract

Diatoms, which are early autotrophic colonisers, are an important constituent of the biofouling community in the marine environment. The effects of substratum and temporal variations on the fouling diatom community structure in a monsoon-influenced tropical estuary were studied. Fibreglass and glass coupons were exposed every month for a period of 4 days and the diatom population sampled at 24 h intervals, over a period of 14 months. The planktonic diatom community structure differed from the biofilm community. Pennate diatoms dominated the biofilms whilst centric diatoms were dominant in the water column. Among the biofilm diatoms, species belonging to the genera Navicula, Amphora, Nitzschia, Pleurosigma and Thalassionema were dominant. On certain occasions, the influence of planktonic blooms was also seen on the biofilm community. A comparative study of biofilms formed on the two substrata revealed significant differences in density and diversity. However species composition was almost constant. In addition to substratum variations, the biofilm diatom community structure also showed significant seasonal variations, which were attributed to physico-chemical and biological changes in both the water and substratum. Temporal variations in the tychopelagic diatoms of the water were also observed to exert an influence on the biofilm diatom community. Variations in diatom communities may determine the functional ecosystem of the benthic environment.     

Archaeal Diversity in Deep-Sea Sediments Estimated by Means of Different Terminal-Restriction Fragment Length Polymorphisms (T-RFLP) Protocols

Despite the increasing recognition of the quantitative importance of Archaea in all marine systems, the protocols for a rapid estimate of Archaeal diversity patterns in deep-sea sediments have been only poorly tested yet. Sediment samples from 11 deep-sea sites (from 79°N to 36°N, at depths comprised from 469 to 5,571 m) were used to compare the performance of two different primer sets (ARCH21f/ARCH958r and ARCH109f/ARCH 915r) and three restriction enzymes (AluI, Rsa I, and HaeIII) for the fingerprinting analysis of Archaeal diversity using terminal-restriction fragment length polymorphisms (T-RFLP). In silico and experimental analyses indicated that different combinations of primer sets and restriction enzymes provided different values of benthic Archaeal ribotype richness and different Archaeal assemblage compositions. The use of the ARCH109f/ARCH 915r primer set in combination with AluI provided the best results (a number of ribotypes up to four folds higher than other combinations), suggesting that this primer set should be used in future studies dealing with the analysis of the patterns of Archaeal diversity in deep-sea sediments. Multivariate multiple regression analysis revealed that, whatever the T-RFLP protocol utilized, latitude and temperature explained most of the variance in benthic Archaeal ribotype richness, while water depth had a negligible role.     

Depth distributions of coral reef fishes: the influence of microhabitat structure,settlement, and post-settlement processes

Many coral reef fishes have restricted depth ranges that are established at settlement or soon after, but the factors limiting these distributions are largely unknown. This study examines whether the availability of microhabitats (reef substrata) explains depth limits, and evaluates whether juvenile growth and survival are lower beyond these limits. Depth-stratified surveys of reef fishes at Kimbe Bay (Papua New Guinea) showed that the abundance of new settlers and the cover of coral substrata differed significantly among depths. A field experiment investigated whether settling coral reef fishes preferred particular depths, and whether these depth preferences were dependent on microhabitat. Small patch reefs composed of identical coral substrata were set up at five depths (3, 6, 10, 15 and 20 m), and settlement patterns were compared to those on unmanipulated reef habitat at the same five depths. For all species, settlement on patch reefs differed significantly among depths despite uniform substratum composition. For four of the six species tested, depth-related settlement patterns on unmanipulated habitat and on patch reefs did not differ, while for the other two, depth ranges were greater on the patch reefs than on unmanipulated habitat. A second experiment examined whether depth preferences reflected variation in growth and survival when microhabitat was similar. Newly settled individuals of Chrysiptera parasema and Dascyllus melanurus were placed, separately, on patch reefs at five depths (as above) and their survival and growth monitored. D. melanurus, which is restricted to shallow depths, had highest survival and growth at the shallowest depth. Depth did not affect either survival or growth of C. parasema, which has a broader depth range than D. melanurus (between 6 and 15 m). This suggests that the fitness costs potentially incurred by settling outside a preferred depth range may depend on the strength of the depth preference.     

Using Species-Area Relationships to Inform Baseline Conservation Targets for the Deep North East Atlantic

Demands on the resources of the deep-sea have increased in recent years. Consequently, the need to create and implement a comprehensive network of Marine Protected Areas (MPAs) to help manage and protect these resources has become a global political priority. Efforts are currently underway to implement MPA networks in the deep North East Atlantic. To ensure these networks are effective, it is essential that baseline information be available to inform the conservation planning process. Using empirical data, we calculated conservation targets for sessile benthic invertebrates in the deep North East Atlantic for consideration during the planning process. We assessed Species-Area Relationships across two depth bands (200–1100 m and 1100–1800 m) and nine substrata. Conservation targets were predicted for each substratum within each depth band using z-values obtained from fitting a power model to the Species-Area Relationships of observed and estimated species richness (Chao1). Results suggest an MPA network incorporating 10% of the North East Atlantic’s deep-sea area would protect approximately 58% and 49% of sessile benthic species for the depth bands 200–1100 m and 1100–1800 m, respectively. Species richness was shown to vary with substratum type indicating that, along with depth, substratum information needs to be incorporated into the conservation planning process to ensure the most effective MPA network is implemented in the deep North East Atlantic.     

A picture on the wall: innovative mapping reveals cold-water coral refuge in submarine canyon

Cold-water corals are azooxanthellate species found throughout the ocean at water depths down to 5000 m. They occur in patches, reefs or large mound structures up to 380 m high, and as ecosystem engineers create important habitats for a diverse fauna. However, the majority of these habitats are now within reach of deep-sea bottom trawling. Many have been severely damaged or are under threat, despite recent protection initiatives. Here we present a cold-water coral habitat type that so far has been overlooked--quite literally--and that has received minimal impact from human activities. Vertical and overhanging cliffs in deep-sea canyons, revealed using an innovative approach to marine habitat mapping, are shown to provide the perfect substratum for extensive cold-water coral-based communities. Typical canyon-related processes, including locally enhanced internal tides and focussed downslope organic carbon transport, provide favourable environmental conditions (current regime, food input) to sustain the communities, even outside the optimal depth and density envelopes reported elsewhere in the NE Atlantic. Our findings show that deep-sea canyons can form natural refuges for faunal communities sensitive to anthropogenic disturbance, and have the potential to fulfil the crucial role of larval sources for the recolonisation of damaged sites elsewhere on the margin.     

Response of Estuarine Biofilm Microbial Community Development to Changes in Dissolved Oxygen and Nutrient Concentrations

The information content and responsiveness of microbial biofilm community structure, as an integrative indicator of water quality, was assessed against short-term changes in oxygen and nutrient loading in an open-water estuarine setting. Biofilms were grown for 7-day periods on artificial substrates in the Pensacola Bay estuary, Florida, in the vicinity of a wastewater treatment plant (WWTP) outfall and a nearby reference site. Substrates were deployed floating at the surface and near the benthos in 5.4 m of water. Three sampling events covered a 1-month period coincident with declining seasonal WWTP flow and increasing dissolved oxygen (DO) levels in the bottom waters. Biomass accumulation in benthic biofilms appeared to be controlled by oxygen rather than nutrients. The overriding effect of DO was also seen in DNA fingerprints of community structure by terminal restriction fragment length polymorphism (T-RFLP) of amplified 16S rRNA genes. Ribotype diversity in benthic biofilms at both sites dramatically increased during the transition from hypoxic to normoxic. Terminal restriction fragment length polymorphism patterns showed pronounced differences between benthic and surface biofilm communities from the same site in terms of signal type, strength, and diversity, but minor differences between sites. Sequencing of 16S rRNA gene clone libraries from benthic biofilms at the WWTP site suggested that low DO levels favored sulfate-reducing prokaryotes (SRP), which decreased with rising oxygen levels and increasing overall diversity. A 91-bp ribotype in the CfoI-restricted 16S rRNA gene T-RFLP profiles, indicative of SRP, tracked the decrease in relative SRP abundance over time.     

Profile of Microbial Community Structure and Presence of Endolithic Microorganisms Inside a Deep-Sea Rock

Microbial community structure in the depth profile of a deep-sea sedimentary rock collected from the Sanriku Escarpment in the Japan Trench at a depth of 6337 m were analyzed using enrichment culture methods and culture-independent molecular phylo-genetic techniques. The rock was subsampled at four depths (S1 to S4; from the surface to the inside), and carbon concentrations and colony-forming units (CFU) were determined under several culture conditions. Terminal-restriction fragment length polymorphism (T-RFLP) analysis of PCR-amplified 16S rRNA gene (rDNA) sequences indicated that a shift in bacterial and archaeal ribotype structures occurred in the sections at different depths from the surface. rDNA clone analysis revealed a significant change in microbial rDNA community structure. Bacterial community rDNA in sections S1 to S3 consisted of typical marine bacteria mainly members of the f and n -subclass of Proteobacteria, while the inner most section, S4, contained rDNA signatures for the g -subclass of Proteobacteria and the High G + C Gram-Positive Group. Major archaeal rDNA clones shifted from Marine Group I (S1) to Thermococcales (S2-S4). The changes in bacterial and archaeal rDNA community structure indicated the possible infiltration of seawater and microorganisms into the rock and strongly suggested the isolation of endolithic microbial communities over the geological history of the rock.     

Patterns in the benthic communities on the shelf of the sub-Antarctic Prince Edward Islands

Summary Benthic communities at the sub-Antarctic Prince Edward Islands were sampled qualitatively with an epi-benthic sled at 57 stations over a five year period. Additional information on these communities was obtained from replicate underwater photographs. In total, 546 macrofaunal species were found. Despite the extreme isolation and geological youth of the islands, there seems to have been a diverse colonisation. A cluster-analysis based on species abundance aggregated most stations into 8 groups. Each of these groups had a unique depth and substratum combination. Thourella variabilis (Octocoralea), and Cabarea darwinii (Bryozoa) characterised deep rocky assemblages and Serpula vermicularis, Lanice conchilega (Polychaeta) and Magellania kerguelenensis (Brachiopoda) characterised soft substrata. Indicator species separating each of the 8 clusters are given. Trends in percentage cover and densities of major taxa with respect to depth and substratum, as shown by photographs, are described. Deep, rocky substrata were dominated by Porifera, Bryozoa and Cnidaria, while soft-substrata were dominated by Polychaeta, Bivalvia and Brachiopoda. Errant forms consisted predominantly of Echinodermata, with Crustacea being important in shallower habitats. Filter-feeders dominated the benthos of the islands in all habitats.     

Antifouling diketopiperazines produced by a deep-sea bacterium,Streptomyces fungicidicus

Abstract

Modern antifouling coatings use heavy metals and toxic organic molecules to prevent biofouling, the undesirable growth of marine organisms on man-made substrata. In an ongoing survey of deep-sea microorganisms aimed at finding low toxic antifouling metabolites, an actinomycete bacterium was isolated from the Pacific sediment at the depth of about 5000 m. The bacterium was closely related to Streptomyces fungicidicus (99% similarity) according to 16S ribosomal RNA sequence information. The spent culture medium of this bacterium inhibited barnacle larval attachment. Bioassay-guided fractionation was employed to isolate antifouling compounds. The ethyl acetate extract was fractionated by using an open silica gel column. Active fractions were further purified on a HPLC C₁₈ column. Five diketopiperazines, cyclo-(L-Leu-L-Pro), cyclo-(L-Phe-L-Pro), cyclo-(L-Val-L-Pro), cyclo-(L-Trp-L-Pro), and cyclo-(L-Leu-L-Val) were isolated for the first time from a deep sea bacterium, and the structures of the compounds were elucidated by nuclear magnetic resonance spectroscopy and mass spectrometry. The pure diketopiperazines were tested for antilarval activity using the barnacle Balanus amphitrite. Effective concentrations that inhibited 50% larval attachment (EC₅₀) after 24 h ranged from 0.10 – 0.27 mM. The data suggest that diketopiperazines and other compounds from deep-sea bacteria may be used as novel antifoulants.     

Biofilm diatom community structure: influence of temporal and substratum variability

Diatoms, which are early autotrophic colonisers, are an important constituent of the biofouling community in the marine environment. The effects of substratum and temporal variations on the fouling diatom community structure in a monsoon-influenced tropical estuary were studied. Fibreglass and glass coupons were exposed every month for a period of 4 days and the diatom population sampled at 24 h intervals, over a period of 14 months. The planktonic diatom community structure differed from the biofilm community. Pennate diatoms dominated the biofilms whilst centric diatoms were dominant in the water column. Among the biofilm diatoms, species belonging to the genera Navicula, Amphora, Nitzschia, Pleurosigma and Thalassionema were dominant. On certain occasions, the influence of planktonic blooms was also seen on the biofilm community. A comparative study of biofilms formed on the two substrata revealed significant differences in density and diversity. However species composition was almost constant. In addition to substratum variations, the biofilm diatom community structure also showed significant seasonal variations, which were attributed to physico-chemical and biological changes in both the water and substratum. Temporal variations in the tychopelagic diatoms of the water were also observed to exert an influence on the biofilm diatom community. Variations in diatom communities may determine the functional ecosystem of the benthic environment.     

Baseline characterization of aerobic hydrocarbon degrading microbial communities in deep-sea sediments of the Great Australian Bight,Australia

Exploratory drilling for deep-sea oil and gas resources is planned for the Great Australian Bight (GAB). There is scant knowledge of the region's benthic ecosystems and no baseline information of the region's indigenous oil degrading bacteria. To address this knowledge gap, we used next generation sequencing (NGS) of three marker genes (alkB, c23o and pmoA) to detect and characterize the microbial communities capable of aerobic hydrocarbon degradation. Unique, highly novel microbial communities capable of degrading hydrocarbons occur in surface sediments at depths between 200 and 2800 m. Clustering at 97% demonstrated differences in community structure with depth, changing most markedly between 400 and 1000 m depth on the continental slope, and identified putative functional ‘ecotypes’ related to depth. Observed differences in community structure showed strong correlations with temperature, other physicochemical properties of the overlying water column and are further modulated by differences in sediment grain size. This study provides important baseline data on hydrocarbon degrading microbial communities prior to the start of petroleum resource extraction. Our data will inform future ecological monitoring of the GAB deep-sea ecosystem.     

Influence of substratum on the variability of benthic biofilm stable isotope signatures: implications for energy flow to a primary consumer

Benthic biofilms have been identified using stable isotope analysis (SIA) as an important resource supporting many freshwater food webs. However, biofilm δ¹³C signatures are highly variable in freshwaters, which may hamper our understanding of energy flow through food webs in these systems. There has been little consideration of the influence that substratum may have on biofilm δ¹³C signature variability and energy flows to primary consumers. We investigated the effect of organic and inorganic substrata on biofilm dynamics by examining: (1) temporal variability of biofilm stable isotope (δ¹³C, δ¹⁵N) signatures on allochthonous leaf-litter (Eucalyptus camaldulensis) and cobble substrata over 12 months in a lowland river in south-eastern Australia; and (2) the effect of substrata on biofilm energy flows to a grazer snail, Physa acuta (Gastropoda: Physidae), using SIA and ecological stoichiometry in a laboratory experiment. The temporal study indicated that cobble biofilm varied significantly in δ¹³C signature during the 12 months (up to 11‰), whereas the δ¹³C signature of leaf biofilm was less variable (less than 2‰). In contrast, biofilm δ¹⁵N signatures varied temporally on both cobble (2.6‰) and leaf (1‰) substrata. This suggests that leaf biofilm was more reliant on leaf tissue for carbon and therefore less limited by carbon supply than cobble biofilm whereas for nitrogen biofilm on both substrata was reliant on external sources. In the laboratory experiment, snails fed leaf biofilm reflected more of an allochthonous δ¹³C signature than cobble biofilm fed snails, suggesting assimilation of leaf carbon via the heterotrophic microbial community within the biofilm. Snails grew largest on cobble biofilm, which had lower C:N ratios than leaf biofilm. Our results demonstrate that the type of substratum can influence the temporal variability of biofilm δ¹³C signatures and energy flow to primary consumers.     

Dynamic surface deformation of silicone elastomers for management of marine biofouling: laboratory and field studies using pneumatic actuation

Many strategies have been developed to improve the fouling release (FR) performance of silicone coatings. However, biofilms inevitably build on these surfaces over time. Previous studies have shown that intentional deformation of silicone elastomers can be employed to detach biofouling species. In this study, inspired by the methods used in soft-robotic systems, controlled deformation of silicone elastomers via pneumatic actuation was employed to detach adherent biofilms. Using programmed surface deformation, it was possible to release > 90% of biofilm from surfaces in both laboratory and field environments. A higher substratum strain was required to remove biofilms accumulated in the field environment as compared with laboratory-grown biofilms. Further, the study indicated that substratum modulus influences the strain needed to de-bond biofilms. Surface deformation-based approaches have potential for use in the management of biofouling in a number of technological areas, including in niche applications where pneumatic actuation of surface deformation is feasible.     

Optimal colonization and growth of marine benthic diatoms on artificial substrata: protocol for a routine use in bioindication

Benthic diatoms growing on hard substrata are used for their bioindication ability in freshwater quality monitoring. Artificial substrata are needed in cases where any natural substrate is present or to achieve similar sampling conditions between sites. Prior to use marine benthic diatoms for monitoring, a standardized protocol for sampling on artificial substrata must be set up. Two major types of information are required: (1) the time needed for a diatom community to be well developed and mature (climax stage); (2) the optimal growth conditions, given that the substrataum nature and texture are important parameters for the initial phase of biofilm development and can influence the future diatom assemblage. Three substrataum types were tested: frosted Plexiglass®, frosted glass, and rough enameled tiles. They were submerged for 8 weeks and sampled weekly. The experiment was conducted at five sites of distinct morphology and water chemistry, along the coastal area of Martinique Island, French West Indies. Development of diatom community was studied through biofilm dry weight, valve density, species richness, and species relative abundances. Globally, substratum type had no significant effect on any parameter. Frosted Plexiglass® was found to be the most interesting substratum because of higher valve densities and practical use. The asymptotic phase of biofilm development was encountered between 5 and 8 weeks depending on site and parameter. A compromise between community development and vandalism or loss through time was fixed to 5 weeks. This period is longer than for stream environments and is valid for tropical oligotrophic marine environments.     

Effects of localised,low-voltage pulsed electric fields on the development and inhibition of Pseudomonas aeruginosa biofilms

Abstract

This work describes the use of low-voltage (0.5 – 5 V) pulsed electric fields to prevent Pseudomonas aeruginosa biofilm development. Interdigitated electrodes (IDEs) with 29-μm spacing between 22-μm-wide electrodes, were used as a platform where the effect of localised, high-strength electric fields could be tested. Alternating current, square-wave pulses were applied to the IDEs in 1 sec intervals. A two-level, three-variable factorial design experiment was used to detect the effects of applied voltage, frequency, and pulse duty ratio (i.e. percentage of pulsing time over one cycle) on the inhibition of biofilm formation. The observations indicated that a pulse configuration of 1% duty ratio, 5 V, and 200 Hz frequency reduced the area of the electrodes covered by biofilm by 50%. In general, the application of low-duty ratio pulses had a positive effect on preventing biofouling. Comparatively, frequency and applied voltage were observed to have less influence on biofouling.