Cathodic behaviour of stainless steel in coastal Indian seawater: calcareous deposits overwhelm biofilms

Type-316 stainless steel (SS) was investigated as the cathode in galvanic couples in full-strength seawater from the Gulf of Mannar on the southeast coast of India. Tests were devised to examine the impact of SS cathodes on anode materials with or without the accrual of marine biofilms. Biofilmed SS cathodes significantly enhanced the rate of corrosion of nickel, causing noble shifts in the couple potentials. With mild steel and zinc as the anodes, calcareous deposits developed quite rapidly on the SS cathodes and led to a significant reduction of bacterial numbers. The calcareous deposits also caused substantial reduction of galvanic corrosion rates for mild steel, whereas there was no difference for zinc. The deposits were identified by XRD as essentially carbonates, oxides and hydroxides of calcium and magnesium. Potentiodynamic polarization performed on the actual couples after disconnection and equilibration provided reasonable interpretations of the galvanic corrosion trends. Data from this work suggest that a potential of about ?0.70 V vs. saturated calomel electrode (SCE) should provide optimum protection of SS in warmer, full-strength seawater that supports the precipitation of calcareous deposits. The criterion commonly recommended for temperate conditions of lower water temperature and estuarine waters of lower alkalinity is ?1.0 V (SCE).     

Antifouling properties of 10beta-formamidokalihinol-A and kalihinol A isolated from the marine sponge Acanthella cavernosa

Many soft-bodied sessile marine invertebrates such as sponges and soft corals defend themselves against fouling directly through the production of antifouling compounds, or indirectly through regulating the epibiotic microbes that affect larval settlement. In this study, 10beta-formamidokalihinol-A and kalihinol A were isolated and purified from the marine sponge Acanthella cavernosa (Dendy). The results indicated that both compounds inhibited the growth of bacteria isolated from the natural environment whereas kalihinol A suppressed larval settlement of a major fouling polychaete, Hydroides elegans with an EC50 of 0.5 microg ml(-1). Kalihinol A was incorporated in Phytagel that was exposed to the bacterial consortia in natural seawater for biofilm formation. Biofilms that developed on the Phytagel surfaces were analysed for bacterial abundance and bacterial species composition using a DNA fingerprinting technique, terminal restriction fragment length polymorphism (T-RFLP). The results showed that kalihinol A only slightly reduced bacterial abundance (t-test, p = 0.0497), but modified the bacterial species composition of the biofilms. Inhibition of H. elegans larval settlement was observed when biofilms developed under the influence of kalihinol A were exposed to larvae, suggesting that compounds like kalihinol A from the sponge A. cavernosa may change bacterial community composition on the sponge surface, which in turn, modulates larval settlement of fouling organisms.     

Biomineralisation of manganese on titanium surfaces exposed to seawater

A 2-year long study was carried out to isolate and characterise various bacterial species present in the biofilm formed on titanium surfaces exposed to seawater and to assess the manganese oxidizing potential of the marine isolates. The amount of manganese present in the biofilm was also measured using atomic absorption spectrometry (AAS). The results showed that titanium was susceptible to biofouling. More than 50% of the culturable marine bacterial isolates were capable of bringing about oxidation of Mn(II). All these manganese oxidizing bacteria were heterotrophic. Autotrophic manganese oxidizing bacteria such as Leptothrix was not isolated in the present study. The AAS results confirmed that the manganese content in the biofilms increased with increasing exposure time. Hence, the study indicates that the titanium surfaces when exposed to seawater were colonised by a large number of heterotrophic bacteria, which have the ability of bringing about biomineralisation of manganese.     

Interactions between cathodic protection and bacterial settlement on steel in seawater

Cathodic protection (CP) is a widely utilised method of corrosion prevention in aqueous systems. There is evidence that the electrochemical changes it produces on a metal surface influence the chemistry of the water and the settlement of fouling organisms. The interaction is in both directions, and fouling organisms and their products have an influence on the CP and on calcareous deposits formed on the metal in marine environments. The studies described show that on both stainless and nonstainless steels, CP retards the development of aerobic bacterial fouling. Increasing the CP decreases the number of bacteria. Additionally, the presence of organic material affects both the current required to maintain CP and the nature of the calcareous deposits formed. The results are discussed in relation to the electrochemistry of the system.     

Chemical defenses of seaweeds against microbial colonization

Any living or non-living surface immersed in seawaterrapidly acquires a bacterial biofilm. For living marineorganisms, biofilm formation can result in the death ofthe host, and thus there is strong evolutionary pressure formarine eukaryotes to evolve mechanisms which inhibit orcontrol the development of biofilms on their surfaces.Some marine eukaryotes are indeed successful incontrolling biofilms on their surfaces, and in manyinstances this control is achieved by the production ofinhibitory chemicals which act at or near the surface ofthe organism. In some cases these natural inhibitors aresimply toxic to bacteria. However, increasingly it appearsthat at least some of these compounds act by interferingspecifically with bacterial characteristics which effect theability of bacteria to colonize their hosts, such asattachment, surface spreading, or the production ofextracellular macromolecules. As an example, theAustralian seaweed Delisea pulchra appears tocontrol bacterial colonization by interfering with abacterial regulatory system (the acylated homoserinelactone system) that regulates several colonizationrelevant bacterial traits. Understanding how marineorganisms control specific bacterial colonization traitsshould provide us with insights into new technologies forthe control of biofilms on artificial surfaces.     

Bacterial biofilms as a natural form of existence of bacteria in the environment and host organism

Advances in microscopic analysis and molecular genetics research methods promoted the acquisition of evidence that natural bacteria populations exist predominately as substrate attached biofilms. Bacteria in biofilms are able to exchange signals and display coordinated activity that is inherent to multicellular organisms. Formation of biofilm communities turned out to be one of the main survival strategies of bacteria in their ecological niche. Bacteria in attached condition in biofilm are protected from the environmental damaging factors and effects of antibacterial substances in the environment and host organism during infection. According to contemporary conception, biofilm is a continuous layer of bacterial cells that are attached to a surface and each other, and contained in a biopolymer matrix. Such bacterial communities may be composed of bacteria of one or several species, and composed of actively functioning cells as well as latent and uncultured forms. Particular attention has recently been paid to the role of biofilms in the environment and host organism. Microorganisms form biofilm on any biotic and abiotic surfaces which creates serious problems in medicine and various areas of economic activity. Currently, it is established that biofilms are one of the pathogenetic factors of chronic inflection process formation. The review presents data on ubiquity of bacteria existence as biofilms, contemporary methods of microbial community analysis, structural-functional features of bacterial biofilms. Particular attention is paid to the role of biofilm in chronic infection process formation, heightened resistance to antibiotics of bacteria in biofilms and possible mechanisms of resistance. Screening approaches for agents against biofilms in chronic infections are discussed.     

Identification of Bacterial Strains Isolated from the Mediterranean Sea Exhibiting Different Abilities of Biofilm Formation

The Mediterranean Sea has rarely been investigated for the characterization of marine bacteria as compared to other marine environments such as the Atlantic or Pacific Ocean. Bacteria recovered from inert surfaces are poorly studied in these environments, when it has been shown that the community structure of attached bacteria can be dissimilar from that of planktonic bacteria present in the water column. The objectives of this study were to identify and characterize marine bacteria isolated from biofilms developed on inert surfaces immersed in the Mediterranean Sea and to evaluate their capacity to form a biofilm in vitro. Here, 13 marine bacterial strains have been isolated from different supports immersed in seawater in the Bay of Toulon (France). Phylogenetic analysis and different biological and physico-chemical properties have been investigated. Among the 13 strains recovered, 8 different genera and 12 different species were identified including 2 isolates of a novel bacterial species that we named Persicivirga mediterranea and whose genus had never been isolated from the Mediterranean Sea. Shewanella sp. and Pseudoalteromonas sp. were the most preponderant genera recovered in our conditions. The phenotypical characterization revealed that one isolate belonging to the Polaribacter genus differed from all the other ones by its hydrophobic properties and poor ability to form biofilms in vitro. Identifying and characterizing species isolated from seawater including from Mediterranean ecosystems could be helpful for example, to understand some aspects of bacterial biodiversity and to further study the mechanisms of biofilm (and biofouling) development in conditions approaching those of the marine environment.     

Interaction of Desulfovibrio desulfuricans biofilms with stainless steel surface and its impact on bacterial metabolism

AIMS: To study the influence of some metallic elements of stainless steel 304 (SS 304) on the development and activity of a sulfate-reducing bacterial biofilm, using as comparison a reference nonmetallic material polymethylmethacrylate (PMMA). METHODS AND RESULTS: Desulfovibrio desulfuricans biofilms were developed on SS 304 and on a reference nonmetallic material, PMMA, in a flow cell system. Steady-state biofilms were metabolically more active on SS 304 than on PMMA. Activity tests with bacteria from both biofilms at steady state also showed that the doubling time was lower for bacteria from SS 304 biofilms. The influence of chromium and nickel, elements of SS 304 composition, was also tested on a cellular suspension of Des. desulfuricans. Nickel decreased the bacterial doubling time, while chromium had no significant effect. CONCLUSIONS: The following mechanism is hypothesized: a Des. desulfuricans biofilm grown on a SS 304 surface in anaerobic conditions leads to the weakening of the metal passive layer and to the dissolution in the bulk phase of nickel ions that have a positive influence on the sulfate-reducing bacteria metabolism. This phenomenon may enhance the biocorrosion process. SIGNIFICANCE AND IMPACT OF THE STUDY: A better understanding of the interactions between metallic surfaces such as stainless steel and bacteria commonly implied in the corrosion phenomena which is primordial to fight biocorrosion.     

Bacterial Community Structure of Biofilms on Artificial Surfaces in an Estuary

This study examined bacterial community structure of biofilms on stainless steel and polycarbonate in seawater from the Delaware Bay. Free-living bacteria in the surrounding seawater were compared to the attached bacteria during the first few weeks of biofilm growth. Surfaces exposed to seawater were analyzed by using 16S rDNA libraries, fluorescence in situ hybridization (FISH), and denaturing gradient gel electrophoresis (DGGE). Community structure of the free-living bacterial community was different from that of the attached bacteria according to FISH and DGGE. In particular, alpha-proteobacteria dominated the attached communities. Libraries of 16S rRNA genes revealed that representatives of the Rhodobacterales clade were the most abundant members of biofilm communities. Changes in community structure during biofilm growth were also examined by DGGE analysis. We hypothesized that bacterial communities on dissimilar surfaces would initially differ and become more similar over time. In contrast, the compositions of stainless steel and polycarbonate biofilms were initially the same, but differed after about 1 week of biofilm growth. These data suggest that the relationship between surface properties and biofilm community structure changes as biofilms grow on surfaces such as stainless steel and polycarbonate in estuarine water.     

A review: microbiologically influenced corrosion and the effect of cathodic polarization on typical bacteria

Microbiologically influenced corrosion is a serious type of corrosion as approximately 20% of the total economic losses. Sulfate reducing bacteria and Iron oxidizing bacteria are one of the typical representatives of the anaerobic and aerobic bacteria, which are ubiquitous in natural environments and corrode steel structures. Cathodic polarization has been recognized as an effective method for preventing steels from microbial corrosion. Although cathodic polarization method has been widely studied, the specific properties of cathodic current that influences the bacterial removal and inactivation remained largely unclear. This review is to show the main effects of Sulfate reducing bacteria and Iron oxidizing bacteria on metal decay as well as the inhibition mechanism of cathodic polarization in the study of bio-corrosion.     

Effect of antimicrobial residues on early adhesion and biofilm formation by wild-type and benzalkonium chloride-adapted Pseudomonas aeruginosa

Antimicrobial residue deposition can change the physico-chemical properties of bacteria and surfaces and thus promote or impair bacterial adhesion. This study focuses on benzalkonium chloride (BC) deposition on polystyrene (PS) surfaces and the influence of this conditioning film on the physico-chemical properties of PS and on early adhesion and biofilm formation by Pseudomonas aeruginosa wild-type and its laboratory BC-adapted strain. The latter readily acquired the ability to grow in BC, and also exhibited physico-chemical surface changes. The existence of residues on PS surfaces altered their hydrophobicity and favoured adhesion as determined by the free energy and early adhesion characterization. Adapted bacteria revealed a higher ability to adhere to surfaces and to develop biofilms, especially on BC-conditioned surfaces, which thereby could enhance resistance to sanitation attempts. These findings highlight the importance of investigations concerning the antimicrobial deposition effect after cleaning procedures, which may encourage bacterial adhesion, especially of bacteria that have been previously exposed to chemical stresses.     

Effect of antimicrobial residues on early adhesion and biofilm formation by wild-type and benzalkonium chloride-adapted Pseudomonas aeruginosa

Antimicrobial residue deposition can change the physico-chemical properties of bacteria and surfaces and thus promote or impair bacterial adhesion. This study focuses on benzalkonium chloride (BC) deposition on polystyrene (PS) surfaces and the influence of this conditioning film on the physico-chemical properties of PS and on early adhesion and biofilm formation by Pseudomonas aeruginosa wild-type and its laboratory BC-adapted strain. The latter readily acquired the ability to grow in BC, and also exhibited physico-chemical surface changes. The existence of residues on PS surfaces altered their hydrophobicity and favoured adhesion as determined by the free energy and early adhesion characterization. Adapted bacteria revealed a higher ability to adhere to surfaces and to develop biofilms, especially on BC-conditioned surfaces, which thereby could enhance resistance to sanitation attempts. These findings highlight the importance of investigations concerning the antimicrobial deposition effect after cleaning procedures, which may encourage bacterial adhesion, especially of bacteria that have been previously exposed to chemical stresses.     

Environmental fate of sacrificial zinc anodes and influence of a biofilm

Laboratory experiments were performed to determine the environmental fate of zinc compounds formed during sacrificial dissolution of zinc anodes in seawater and to further determine the role of biofilms in mobilizing/immobilizing zinc corrosion products. Experimental conditions simulated cathodic protection and overprotection (40x normal) of ship ballast tanks with an anode to cathode ratio of 0.091. Anodic dissolution and percentage of soluble zinc increased with the level of protection. For normal cathodic protection in abiotic seawater, approximately 3% of the sacrificed zinc was dissolved in solution. Additionally, weight loss and percent soluble products increased dramatically in the presence of an agar film, but were unchanged due to the presence of natural marine biofilms. Under overprotection conditions, weight loss did not vary in the presence of surface films. Solid zinc corrosion products were precipitated, suspended in solution, and attached to electrode surfaces. Experimental data were compared with zinc concentrations measured in ballast tanks for two United States Navy surface ships.     

Induction of larval metamorphosis of the coral Acropora millepora by tetrabromopyrrole isolated from a Pseudoalteromonas bacterium

The induction of larval attachment and metamorphosis of benthic marine invertebrates is widely considered to rely on habitat specific cues. While microbial biofilms on marine hard substrates have received considerable attention as specific signals for a wide and phylogenetically diverse array of marine invertebrates, the presumed chemical settlement signals produced by the bacteria have to date not been characterized. Here we isolated and fully characterized the first chemical signal from bacteria that induced larval metamorphosis of acroporid coral larvae (Acropora millepora). The metamorphic cue was identified as tetrabromopyrrole (TBP) in four bacterial Pseudoalteromonas strains among a culture library of 225 isolates obtained from the crustose coralline algae Neogoniolithon fosliei and Hydrolithon onkodes. Coral planulae transformed into fully developed polyps within 6 h, but only a small proportion of these polyps attached to the substratum. The biofilm cell density of the four bacterial strains had no influence on the ratio of attached vs. non-attached polyps. Larval bioassays with ethanolic extracts of the bacterial isolates, as well as synthetic TBP resulted in consistent responses of coral planulae to various doses of TBP. The lowest bacterial density of one of the Pseudoalteromonas strains which induced metamorphosis was 7,000 cells mm(-2) in laboratory assays, which is on the order of 0.1-1% of the total numbers of bacteria typically found on such surfaces. These results, in which an actual cue from bacteria has been characterized for the first time, contribute significantly towards understanding the complex process of acroporid coral larval settlement mediated through epibiotic microbial biofilms on crustose coralline algae.     

Both sulfate-reducing bacteria and Enterobacteriaceae take part in marine biocorrosion of carbon steel

AIMS: In order to evaluate the part played in biocorrosion by microbial groups other than sulfate-reducing bacteria (SRB), we characterized the phylogenetic diversity of a corrosive marine biofilm attached to a harbour pile structure as well as to carbon steel surfaces (coupons) immersed in seawater for increasing time periods (1 and 8 months). We thus experimentally checked corroding abilities of defined species mixtures. METHODS AND RESULTS: Microbial community analysis was performed using both traditional cultivation techniques and polymerase chain reaction cloning-sequencing of 16S rRNA genes. Community structure of biofilms developing with time on immersed coupons tended to reach after 8 months, a steady state similar to the one observed on a harbour pile structure. Phylogenetic affiliations of isolates and cloned 16S rRNA genes (rrs) indicated that native biofilms (developing after 1-month immersion) were mainly colonized by gamma-proteobacteria. Among these, Vibrio species were detected in majority with molecular methods while cultivation techniques revealed dominance of Enterobacteriaceae such as Citrobacter, Klebsiella and Proteus species. Conversely, in mature biofilms (8-month immersion and pile structure), SRB, and to a lesser extent, spirochaetes were dominant. CONCLUSIONS: Corroding activity detection assays confirmed that Enterobacteriaceae (members of the gamma-proteobacteria) were involved in biocorrosion of metallic material in marine conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: In marine biofilms, metal corrosion may be initiated by Enterobacteriaceae.     

Bacterial corrosion of iron in seawater in situ, and in aerobic and anaerobic model systems

Abstract A screening of twenty-two marine isolates was made to examine their effects on corrosion of carbon steel ASTM A619. In batch cultures, sixteen of the isolates gave a lower corrosion than the control. Aerobic and anaerobic biofilm populations were formed by immersing iron coupons in natural seawater under aerobic and anaerobic conditions. The effects of the biofilms depended on a balance between the presence of oxygen and the type of population. An anaerobic population attached to the surface increased the corrosion rate if immersed in a suspension of Vibrio sp. DW1. The vibrio population probably 'protected' the anaerobic population from oxygen and may have provided nutrients, thereby creating conditions that allowed production of corrosive metabolites close to the metal. In contrast, coupons without a biofilm showed a decrease in the corrosion when immersed in the same vibrio suspension. The protective effect of a dense suspension of bacteria found earlier [5,6] was tested in situ in seawater. Iron coupons were immersed in dialysis bags with a suspension of Vibrio sp. DW1. Coupons immersed in dialysis bags with DW1 showed a lower degree of corrosion than coupons immersed in bags with seawater.     

Biofilms: strategies for metal corrosion inhibition employing microorganisms

Corrosion causes dramatic economic loss. Currently widely used corrosion control strategies have disadvantages of being expensive, subject to environmental restrictions, and sometimes inefficient. Studies show that microbial corrosion inhibition is actually a common phenomenon. The present review summarizes recent progress in this novel strategy: corrosion control using beneficial bacteria biofilms. The possible mechanisms may involve: (1) removal of corrosive agents (such as oxygen) by bacterial physiological activities (e.g., aerobic respiration), (2) growth inhibition of corrosion-causing bacteria by antimicrobials generated within biofilms [e.g., sulfate-reducing bacteria (SRB) corrosion inhibition by gramicidin S-producing Bacillus brevis biofilm], (3) generation of protective layer by biofilms (e.g., Bacillus licheniformis biofilm produces on aluminum surface a sticky protective layer of γ-polyglutamate). Successful utilization of this novel strategy relies on advances in study at the interface of corrosion engineering and biofilm biology.     

Cross-Ocean Distribution of Rhodobacterales Bacteria as Primary Surface Colonizers in Temperate Coastal Marine Waters

Bacterial surface colonization is a universal adaptation strategy in aquatic environments. However, neither the identities of early colonizers nor the temporal changes in surface assemblages are well understood. To determine the identities of the most common bacterial primary colonizers and to assess the succession process, if any, of the bacterial assemblages during early stages of surface colonization in coastal water of the West Pacific Ocean, nonnutritive inert materials (glass, Plexiglas, and polyvinyl chloride) were employed as test surfaces and incubated in seawater off the Qingdao coast in the spring of 2005 for 24 and 72 h. Phylogenetic analysis of the 16S rRNA gene sequences amplified from the recovered surface-colonizing microbiota indicated that diverse bacteria colonized the submerged surfaces. Multivariate statistical cluster analyses indicated that the succession of early surface-colonizing bacterial assemblages followed sequential steps on all types of test surfaces. The Rhodobacterales, especially the marine Roseobacter clade members, formed the most common and dominant primary surface-colonizing bacterial group. Our current data, along with previous studies of the Atlantic coast, indicate that the Rhodobacterales bacteria are the dominant and ubiquitous primary surface colonizers in temperate coastal waters of the world and that microbial surface colonization follows a succession sequence. A conceptual model is proposed based on these findings, which may have important implications for understanding the structure, dynamics, and function of marine biofilms and for developing strategies to harness or control surface-associated microbial communities.     

Effect of bacterial biofilms formed on fouling-release coatings from natural seawater and Cobetia marina,on the adhesion of two marine algae

Previous studies have shown that bacterial biofilms formed from natural seawater (NSW) enhance the settlement of spores of the green alga Ulva linza, while single-species biofilms may enhance or reduce settlement, or have no effect at all. However, the effect of biofilms on the adhesion strength of algae, and how that may be influenced by coating/surface properties, is not known. In this study, the effect of biofilms formed from natural seawater and the marine bacterium Cobetia marina, on the settlement and the adhesion strength of spores and sporelings of the macroalga U. linza and the diatom Navicula incerta, was evaluated on Intersleek® 700, Intersleek® 900, poly(dimethylsiloxane) and glass. The settlement and adhesion strength of these algae were strongly influenced by biofilms and their nature. Biofilms formed from NSW enhanced the settlement (attachment) of both algae on all the surfaces while the effect of biofilms formed from C. marina varied with the coating type. The adhesion strength of spores and sporelings of U. linza and diatoms was reduced on all the surfaces biofilmed with C. marina, while adhesion strength on biofilms formed from NSW was dependent on the alga (and on its stage of development in the case of U. linza), and coating type. The results illustrate the complexity of the relationships between fouling algae and bacterial biofilms and suggest the need for caution to avoid over-generalisation.     

Aggregation of ammonia-oxidizing bacteria in microbial biofilm on oyster shell surface

Summary Formation and activity of bacterial nitrifying biofilms play an important role in the closed seawater systems for shrimp cultivation. The structure of microbial biofilm on empty oyster shells, used as a biofilm carrier in biofiltration of aquacultural water, was studied using fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy. FISH was performed with specific oligonucleotide probes for Bacteria and ammonia-oxidizing Nitrosomonas spp. The bacterial cells were arranged within the biofilm as a layer of vertically elongated aggregates. Aggregates of ammonia-oxidizing bacteria were embedded within the matrix formed by other bacteria. Vertically elongated cell aggregates may be ecologically important in bacterial biofilms because they have a higher surface-to-volume ratio than that of laminated biofilms.