Molecular characterization of putative biocorroding microbiota with a novel niche detection of Epsilon- and Zetaproteobacteria in Pacific Ocean coastal seawaters

Submerged metal surfaces in marine waters undergo rapid microbial colonization and biocorrosion, causing huge damage to marine engineering facilities and significant financial losses. In coastal areas, an accelerated and particularly severe form of biocorrosion termed accelerated low water corrosion (ALWC) is widespread globally. While identification of biocorroding microorganisms and the dynamics of their community structures is the key for understanding the processes and mechanisms leading to ALWC, neither one is presently understood. In this study, analysis of constructed clone libraries and qPCR assays targeting group-specific 16S rRNA or functional marker genes were used to determine the identity and abundance of putative early carbon steel surface-colonizing and biocorroding microbes in coastal seawater. Diverse microbial groups including 10 bacterial phyla, archaea and algae were found to putatively participate in the surface-colonizing process. Analysis of the community structure of carbon steel surface microbiota revealed a temporal succession leading to ALWC. By extending the current state of knowledge, our work demonstrates the global importance of Alphaproteobacteria (mainly Rhodobacterales), Gammaproteobacteria (mainly Alteromonadales and Oceanospirillales), Bacteroidetes (mainly Flavobacteriales) and microalgae as the pioneer and sustaining surface colonizers that contribute to initial formation and development of surface biofilms. We also discovered Epsilonproteobacteria and the recently described Zetaproteobacteria as putative corrosion-causing microorganisms during early steps of the ALWC process. Hence, our study reports that Zetaproteobacteria may be ubiquitous also in non-hydrothermal coastal seawaters and that ALWC of submerged carbon steel surfaces in coastal waters may involve a highly diverse, complex and dynamic microbial consortium. Our finding that Epsilon- and Zetaproteobacteria may play pivotal roles in ALWC provides a new starting point for future investigation of the ALWC process and mechanism in marine environments. Further studies of Epsilon- and Zetaproteobacteria in particular may thus help with the design of effective corrosion prevention and control strategies.     

Study of parameters implicated in the biodeterioration of mild steel in the presence of different species of sulphate-reducing bacteria

Two different species of sulphate-reducing bacteria, strain classified by NCIMB as Desulfovibrio desulfuricans subspecies desulfuricans New Jersey (8313) isolated from the corroding heat exchanger, and SRB species recovered from a corroding ship hull anchored off the Indonesian coast (Indo isolate) were grown as laboratory batch cultures. Several factors such as the surface finish of substratum, metabolic activity of planktonic and sessile bacterial populations, initial attachment of cells to surfaces and subsequent formation of biofilms on the process of biodeterioration of mild steel in the presence of these two different species of SRB were investigated. The corrosion rates of mild steel were estimated by weight loss measurements and correlated with the density of sessile SRB population. The yield and composition of extracellular polymers released into the bulk phase of culture media were determined and the amount of dissolved hydrogen sulphide was monitored. The results revealed differences between SRB species in their aggressiveness towards mild steel under identical growth conditions, emphasising the importance of biochemistry and physiology of SRB for the biocorrosion process. Biochemical and genetic characterisation of SRB isolates chosen for this study are currently in progress.     

Characterization of bacterial community associated to biofilms of corroded oil pipelines from the southeast of Mexico

Microbial communities associated to biofilms promote corrosion of oil pipelines. The community structure of bacteria in the biofilm formed in oil pipelines is the basic knowledge to understand the complexity and mechanisms of metal corrosion. To assess bacterial diversity, biofilm samples were obtained from X52 steel coupons corroded after 40 days of exposure to normal operation and flow conditions. The biofilm samples were directly used to extract metagenomic DNA, which was used as template to amplify 16S ribosomal gene by PCR. The PCR products of 16S ribosomal gene were also employed as template for sulfate-reducing bacteria (SRB) specific nested-PCR and both PCR products were utilized for the construction of gene libraries. The V3 region of the 16S rRNA gene was also amplified to analyse the bacterial diversity by analysis of denaturing gradient gel electrophoresis (DGGE). Ribosomal library and DGGE profiles exhibited limited bacterial diversity, basically including Citrobacter spp., Enterobacter spp. and Halanaerobium spp. while Desulfovibrio alaskensis and a novel clade within the genus Desulfonatronovibrio were detected from the nested PCR library. The biofilm samples were also taken for the isolation of SRB. Desulfovibrio alaskensis and Desulfovibrio capillatus, as well as some strains related to Citrobacter were isolated. SRB consists in a very small proportion of the community and Desulfovibrio spp. were the relatively abundant groups among the SRB. This is the first study directly exploring bacterial diversity in corrosive biofilms associated to steel pipelines subjected to normal operation conditions.     

Inhibiting mild steel corrosion from sulfate-reducing bacteria using antimicrobial-producing biofilms in Three-Mile-Island process water

Biofilms were used to produce gramicidin S (a cyclic decapeptide) to inhibit corrosion-causing, sulfate-reducing bacteria (SRB). In laboratory studies these biofilms protected mild steel 1010 continuously from corrosion in the aggressive, cooling service water of the AmerGen Three-Mile-Island (TMI) nuclear plant, which was augmented with reference SRB. The growth of both reference SRB (Gram-positive Desulfosporosinus orientis and Gram-negative Desulfovibrio vulgaris) was shown to be inhibited by supernatants of the gramicidin-S-producing bacteria as well as by purified gramicidin S. Electrochemical impedance spectroscopy and mass loss measurements showed that the protective biofilms decreased the corrosion rate of mild steel by 2- to 10-fold when challenged with the natural SRB of the TMI process water supplemented with D. orientis or D. vulgaris. The relative corrosion inhibition efficiency was 50–90% in continuous reactors, compared to a biofilm control which did not produce the antimicrobial gramicidin S. Scanning electron microscope and reactor images also revealed that SRB attack was thwarted by protective biofilms that secrete gramicidin S. A consortium of beneficial bacteria (GGPST consortium, producing gramicidin S and other antimicrobials) also protected the mild steel.     

Influence of cathodic protection of mild steel on the growth of sulphate‐reducing bacteria at 35°C in marine sediments

In order to protect metallic structures from marine corrosion, cathodic protection using sacrificial anodes or impressed current is widely used. In aerated seawater steel is considered to be protected when a cathodic potential of — 800 mV/SCE (Saturated Calomel Electrode) is applied. However, in many cases, this potential must be lowered due to the presence and activity of microorganisms such as acid‐producing bacteria or sulphate‐reducing bacteria (SRB). SRB are obligate anaerobes using sulphate as an electron acceptor with resultant production of sulfides. Some SRB are able to use hydrogen as an electron donor causing thereby depolarization of steel surfaces.

An experiment was performed in marine sediments to determine the relationship between cathodically produced hydrogen and growth of SRB in marine sediments both at ambiant temperature (Therene, 1988) and at 35°C. Results concerning the latter experiments are reported here.

Analytical techniques included microbiological analyses, lipid biomarker studies and electrochemical measurements including AC impedance spectroscopy. Results indicated a change in the bacterial community structure both on the steel and sediment as a function of time and potential. The results also showed that cathodically‐produced hydrogen promoted the growth of SRB with the Desulfovibrio genus predominating.     

Diverse bacterial groups are associated with corrosive lesions at a Granite Mountain Record Vault (GMRV)

Aims: This study applied culture‐dependent and molecular approaches to examine the bacterial communities at corrosion sites at Granite Mountain Record Vault (GMRV) in Utah, USA, with the goal of understanding the role of microbes in these unexpected corrosion events. Methods and Results: Samples from corroded steel chunks, rock particles and waters around the corrosion pits were collected for bacterial isolation and molecular analyses. Bacteria cultivated from these sites were identified as members of Alphaproteobacteria, Gammaproteobacteria, Firmicutes and Actinobacteria. In addition, molecular genetic characterization of the communities via nested‐polymerase chain reaction‐denaturing gradient gel electrophoresis (DGGE) indicated the presence of a broad spectrum of bacterial groups, including Alphaproteobacteria, Betaproteobacteria, Deltaproteobacteria, Actinobacteria, Firmicutes and Bacteroidetes. However, neither cultivation nor molecular approaches identified sulfate‐reducing bacteria (SRB), the bacteria commonly implicated as causative organisms were found associated with corrosive lesions in a process referred to as microbially influenced corrosion (MIC). The high diversity of bacterial groups at the corrosion sites in comparison with that seen in the source waters suggested to us a role for the microbes in corrosion, perhaps being an expression of a redox‐active group of microbes transferring electrons, harvesting energy and producing biomass. Conclusions: The corrosion sites contained highly diverse microbial communities, consistent with the involvement of microbial activities along the redox gradient at corrosion interface. We hypothesize an electron transport model for MIC, involving diverse bacterial groups such as acid‐producing bacteria (APB), SRB, sulfur‐oxidizing bacteria (SOB), metal‐reducing bacteria (MRB) and metal‐oxidizing bacteria (MOB). Significance and Impact of the Study: The characterization of micro‐organisms that influence metal‐concrete corrosion at GMRV has significant implications for corrosion control in high‐altitude freshwater environments. MIC provides a potential opportunity to further our understandings of extracellular electron transfer and interspecies communications.     

Inhibiting sulfate-reducing bacteria in biofilms on steel with antimicrobial peptides generated in situ

In batch and continuous fermentations, the reduction in corrosion of SAE 1018 mild steel and 304 stainless steel caused by inhibition of the reference sulfate-reducing bacterium (SRB) Desulfovibrio vulgaris by a protective, antimicrobial-producing Bacillus brevis biofilm was investigated. The presence of D. vulgaris produced a thick black precipitate on mild steel and a higher corrosion rate in batch cultures than that seen in a mono-culture of non-antimicrobial-producing Pseudomonas fragi K upon the addition of SRB to the aerobic P. fragi K biofilm. In continuous reactors, the polarization resistance R _p decreased for stainless steel and increased for mild steel upon the addition of SRB to a P. fragi K biofilm. Addition of either 200 μg/ml ampicillin, chloramphenicol, or ammonium molybdate to batch and continuous reactors after SRB had colonized the metal was ineffective in killing SRB, as inferred from the lack of change in both R _p and the impedance spectra. However, when ampicillin was added prior to SRB colonization, the growth of SRB was completely inhibited on stainless steel in continuous reactors. Prior addition of ampicillin was only able to delay the growth of SRB on mild steel in continuous reactors. External addition of the purified peptide antimicrobial agent gramicidin S prior to the addition of SRB also inhibited the growth of SRB on stainless steel in continuous reactors, and the SRB were also inhibited on stainless steel in both batch and continuous reactors by producing gramicidin S in situ in a protective biofilm when the gramicidin-S-overproducing strain Bacillus brevis 18 was used. Received: 29 October 1998 / Received revision: 18 February 1999 / Accepted: 26 February 1999     

Phylogenetic characterization of a corrosive consortium isolated from a sour gas pipeline

Biocorrosion is a common problem in oil and gas industry facilities. Characterization of the microbial populations responsible for biocorrosion and the interactions between different microorganisms with metallic surfaces is required in order to implement efficient monitoring and control strategies. Denaturing gradient gel electrophoresis (DGGE) analysis was used to separate PCR products and sequence analysis revealed the bacterial composition of a consortium obtained from a sour gas pipeline in the Gulf of Mexico. Only one species of sulfate-reducing bacteria (SRB) was detected in this consortium. The rest of the population consisted of enteric bacteria with different characteristics and metabolic capabilities potentially related to biocorrosion. Therefore, several types of bacteria may be involved in biocorrosion arising from natural biofilms that develop in industrial facilities. The low abundance of the detected SRB was evidenced by environmental scanning electron microscopy (ESEM). In addition, the localized corrosion of pipeline steel in the presence of the consortium was clearly observed by ESEM after removing the adhered bacteria.     

The use of biocides to control sulphate‐reducing bacteria in biofilms on mild steel surfaces

Three different types of biocides, viz. formaldehyde (FM), glutaraldehyde (GA) and isothiozolone (ITZ) were used to control planktonic and sessile populations of two marine isolates of sulphate‐reducing bacteria (SRB). The influence of these biocides on the initial attachment of cells to mild steel surfaces, on subsequent biofilm formation and on the activity of hydrogenase enzymes within developed biofilms was evaluated. In the presence of biocides the rate and degree of colonization of mild steel by SRB depended on incubation time, bacterial isolate and the type of biocide used. Although SRB differed in their susceptibility to biocides, for all isolates the biofilm population was more resistant to the treatment than the planktonic population. GA showed highest efficiency in controlling planktonic and sessile SRB compared with the other two biocides. The activity of the enzyme hydrogenase measured in SRB biofilms varied between isolates and with the biocide treatment. No correlation was found between the number of sessile cells and hydrogenase activity.     

The corrosion behaviour of galvanized steel in cooling tower water containing a biocide and a corrosion inhibitor

The corrosion behaviour of galvanized steel in cooling tower water containing a biocide and a corrosion inhibitor was investigated over a 10-month period in a hotel. Planktonic and sessile numbers of sulphate reducing bacteria (SRB) and heterotrophic bacteria were monitored. The corrosion rate was determined by the weight loss method. The corrosion products were analyzed by energy dispersive X-ray spectroscopy and X-ray diffraction. A mineralized, heterogeneous biofilm was observed on the coupons. Although a biocide and a corrosion inhibitor were regularly added to the cooling water, the results showed that microorganisms, such as SRB in the mixed species biofilm, caused corrosion of galvanized steel. It was observed that Zn layers on the test coupons were completely depleted after 3?months. The Fe concentrations in the biofilm showed significant correlations with the weight loss and carbohydrate concentration (respectively, p?<?0.01 and p?<?0.01).     

Comparative studies of bacterial biofilms on steel surfaces using atomic force microscopy and environmental scanning electron microscopy

Environmental scanning electron microscopy (ESEM) and atomic force microscopy (AFM) were compared as tools for the observation of bacterial biofilms developed on carbon steel and AISI 316 stainless steel surfaces under stagnant conditions. Biofilms were generated in batch cultures of two different isolates of marine sulphate reducing bacteria (SRB) and in cultures consisting of mixed populations of acidophilic bacteria, known as "acid streamers";. Imaging of single SRB cells on mica was also carried out to reveal the surface topography of individual bacterial cells at nanometre resolution. Following the removal of biofilms, the stainless steel surfaces were profiled using AFM to determine the degree of steel deterioration. ESEM and AFM studies of bacterial biofilms in-situ, gave both qualitative and quantitative information on biofilm structure at high resolution. The use of AFM image analysis software allowed estimation of the width and height of bacterial cells, the thickness and width of exopolymeric (EPS) capsule and bacterial flagella, as well as characterisation of the surface roughness of the steel, including measurements of depth and diameter of individual pits. Exposure of stainless steel specimens to acid streamers resulted in a significant increase in the surface roughness of the steel, compared to specimens placed in sterile medium.     

Sulphate-reducing bacteria which do not induce accelerated corrosion

The corrosive activity of the recently isolated SRB genera has not previously been reported in the literature. In this investigation, three genera of sulphate-reducing bacteria (SRB). Desulfovibrio vulgaris, Desulfobacter postgatei and Desulfobulbus propionicus were tested for their ability to induce accelerated corrosion of mild steel in laboratory growth media. Desulfovibrio vulgaris, well-recognised for its corrosive activity, caused a 78·2% increase in weight loss compared to the control, uninoculated medium (95% confidence limits +37·0% to +130·6%). Desulfobacter postgatei and Desulfobulbus propionicus had no significant effect on corrosion. For Desulfobacter postgatei the mean rate of corrosion was 10·6% more than the control (95% confidence limits −12·0% to +39·0%). For Desulfobulbus propionicus the mean corrosion rate was increased by 5·8% over control (95% confidence limits −9·8% to +24·2%).     

Extracellular Electron Transfer Is a Bottleneck in the Microbiologically Influenced Corrosion of C1018 Carbon Steel by the Biofilm of Sulfate-Reducing Bacterium Desulfovibrio vulgaris

Carbon steels are widely used in the oil and gas industry from downhole tubing to transport trunk lines. Microbes form biofilms, some of which cause the so-called microbiologically influenced corrosion (MIC) of carbon steels. MIC by sulfate reducing bacteria (SRB) is often a leading cause in MIC failures. Electrogenic SRB sessile cells harvest extracellular electrons from elemental iron oxidation for energy production in their metabolism. A previous study suggested that electron mediators riboflavin and flavin adenine dinucleotide (FAD) both accelerated the MIC of 304 stainless steel by the Desulfovibrio vulgaris biofilm that is a corrosive SRB biofilm. Compared with stainless steels, carbon steels are usually far more prone to SRB attacks because SRB biofilms form much denser biofilms on carbon steel surfaces with a sessile cell density that is two orders of magnitude higher. In this work, C1018 carbon steel coupons were used in tests of MIC by D. vulgaris with and without an electron mediator. Experimental weight loss and pit depth data conclusively confirmed that both riboflavin and FAD were able to accelerate D. vulgaris attack against the carbon steel considerably. It has important implications in MIC failure analysis and MIC mitigation in the oil and gas industry.     

Biodiversity analysis by polyphasic study of marine bacteria associated with biocorrosion phenomena

Aims: A polyphasic approach was used to study the biodiversity bacteria associated with biocorrosion processes, in particular sulfate‐reducing bacteria (SRB) and thiosulfate‐reducing bacteria (TRB) which are described to be particularly aggressive towards metallic materials, notably via hydrogen sulfide release. Methods and Results: To study this particular flora, an infrared spectra library of 22 SRB and TRB collection strains were created using a Common Minimum Medium (CMM) developed during this study and standardized culture conditions. The CMM proved its ability to allow for growth of both SRB and TRB strains. These sulfurogen collection strains were clearly discriminated and differentiated at the genus level by fourier transform infrared (FT‐IR) spectroscopy. In a second step, infrared spectra of isolates, recovered from biofilms formed on carbon steel coupons immersed for 1 year in three different French harbour areas, were compared to the infrared reference spectra library. In parallel, molecular methods (M13‐PCR and 16S rRNA gene sequencing) were used to qualitatively evaluate the intra‐ and inter‐species genetic diversity of biofilm isolates. The biodiversity study indicated that strains belonging to the Vibrio genus were the dominant population; strains belonging to the Desulfovibrio genus (SRB) and Peptostreptococcaceae were also identified. Conclusion: Overall, the combination of the FT‐IR spectroscopy and molecular approaches allowed for the taxonomic and ecological study of a bacterial flora, cultivated on CMM, associated with microbiology‐induced corrosion (MIC) processes. Significance and Impact of the Study: Via the use of the CMM medium, the culture of marine bacteria (including both SRB and TRB bacteria) was allowed, and the implication of nonsulforogen bacteria in MIC was observed. Their involvement in the biocorrosion phenomena will have to be studied and taken into account in the future.     

Nitrate treatment effects on bacterial community biofilm formed on carbon steel in produced water stirred tank bioreactor

To better understand the impact of nitrate in Brazilian oil reservoirs under souring processes and corrosion, the goal of this study was to analyse the effect of nitrate on bacterial biofilms formed on carbon steel coupons using reactors containing produced water from a Brazilian oil platform. Three independent experiments were carried out (E1, E2 and E3) using the same experimental conditions and different incubation times (5, 45 and 80 days, respectively). In every experiment, two biofilm-reactors were operated: one was treated with continuous nitrate flow (N reactor), and the other was a control reactor without nitrate (C reactor). A Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis approach using the 16S rRNA gene was performed to compare the bacterial groups involved in biofilm formation in the N and C reactors. DGGE profiles showed remarkable changes in community structure only in experiments E2 and E3. Five bands extracted from the gel that represented the predominant bacterial groups were identified as Bacillus aquimaris, B. licheniformis, Marinobacter sp., Stenotrophomonas maltophilia and Thioclava sp. A reduction in the sulfate-reducing bacteria (SRB) most probable number counts was observed only during the longer nitrate treatment (E3). Carbon steel coupons used for biofilm formation had a slightly higher weight loss in N reactors in all experiments. When the coupon surfaces were analysed by scanning electron microscopy, an increase in corrosion was observed in the N reactors compared with the C reactors. In conclusion, nitrate reduced the viable SRB counts. Nevertheless, the nitrate dosing increased the pitting of coupons.     

In situ detection of bacteria involved in cathodic depolarization and stainless steel surface corrosion using microautoradiography

Aims: To examine the activity of bacteria involved in cathodic depolarization and surface corrosion on stainless steel in an in situ model system. Methods and Results: The microautoradiographic technique (MAR) was used to evaluate the activity of bacterial populations on stainless steel surfaces with a single cell resolution. Anaerobic uptake and fixation of ¹⁴C‐labelled bicarbonate occurred within corrosion sites in the absence of atmospheric hydrogen or other external electron donors, whereas it was taken up and fixed by bacteria at all other stainless steel surfaces in the presence of atmospheric hydrogen. This indicates that the bacteria utilized electrons originating from the corrosion sites due to the ongoing corrosion (cathodic depolarization). Conclusion: Under in situ conditions, bacteria were fixating ¹⁴C‐labelled bicarbonate at corrosion sites in the absence of atmospheric hydrogen. This indicates that electrons transferred to the bacteria provided energy for bicarbonate fixation due to cathodic depolarization. Significance and Impact of the Study: Application of the MAR method showed ongoing biocorrosion in the applied in situ model system and allowed in situ examination of bacterial activity on a single cell level directly on a metal surface providing information about potential corrosion mechanisms. Furthermore, application of fluorescence in situ hybridization in combination with MAR allows for identification of the active bacteria.     

Sulfato/thiosulfato reducing bacteria characterization by FT-IR spectroscopy: a new approach to biocorrosion control

Sulfato and Thiosulfato Reducing Bacteria (SRB, TRB) can induce corrosion process on steel immersed in seawater. This phenomenon, called biocorrosion, costs approximatively 5 billion euros in France each year. We provide the first evidence that Fourier Transformed InfraRed (FTIR) spectroscopy is a competitive technique to evaluate the sulfurogen flora involved in biocorrosion in comparison with time consuming classical identification methods or PCR analyses. A great discrimination was obtained between SRB, TRB and some contamination bacteria known to be present in seawater and seem to be able to reduce sulfate under particular conditions. Moreover, this preliminary study demonstrates that FTIR spectroscopic and genotypic results present a good correlation (these results are confirmed by other data obtained before or later, data not shown here). The advantages gained by FTIR spectroscopy are to give information on strain phenotype and bacterial metabolism which are of great importance in corrosion processes.     

Inhibiting sulfate-reducing bacteria in biofilms by expressing the antimicrobial peptides indolicidin and bactenecin

To identify novel, less-toxic compounds capable of inhibiting sulfate-reducing bacteria (SRB), Desulfovibrio vulgaris and Desulfovibrio gigas in suspension cultures were exposed to several antimicrobial peptides. The bacterial peptide antimicrobials gramicidin S, gramicidin D, and polymyxin B as well as the cationic peptides indolicidin and bactenecin from bovine neutrophils decreased the viability of both SRB by 90% after a 1-h exposure at concentrations of 25–100 μg ml⁻¹. To reduce corrosion by inhibiting SRB in biofilms, the genes for indolicidin and bactenecin were expressed in Bacillus subtilisBE1500 and B. subtilis WB600 under the control of the constitutive alkaline protease (apr) promoter, and the antimicrobials were secreted into the culture medium using the apr signal sequence. Bactenecin was also synthesized and expressed as a fusion to the pro-region of barnase from Bacillus amyloliquefaciens. Concentrated culture supernatants of B. subtilis BE1500 expressing bactenecin at 3 μg ml⁻¹ decreased the viability of Escherichia coli BK6 by 90% and the reference SRB D. vulgaris by 83% in suspension cultures. B. subtilis BE1500 and B. subtilis WB600 expressing bactenecin in biofilms also inhibited the SRB-induced corrosion of 304 stainless steel six to 12-fold in continuous reactors as evidenced by the lack of change in the impedance spectra (resistance polarization) upon addition of SRB and by the reduction in hydrogen sulfide and iron sulfide in batch fermentations with mild steel. A 36-fold decrease in the population of D. vulgaris in a B. subtilis BE1500 biofilm expressing bactenecin was also observed. This is the first report of an antimicrobial produced in a biofilm for in vivo applications and represents the first application of a beneficial, genetically-engineered biofilm for combating corrosion. Received 27 October 1998/ Accepted in revised form 21 February 1999     

Influence of carbon steel grade on the initial attachment of bacteria and microbiologically influenced corrosion

The influence of the composition and microstructure of different carbon steel grades on the initial attachment (≤ 60 min) of Escherichia coli and subsequent longer term (28 days) corrosion was investigated. The initial bacterial attachment increased with time on all grades of carbon steel. However, the rate and magnitude of bacterial attachment varied on the different steel grades and was significantly less on the steels with a higher pearlite phase content. The observed variations in the number of bacterial cells attached across different steel grades were significantly reduced by applying a fixed potential to the steel samples. Longer term immersion studies showed similar levels of biofilm formation on the surface of the different grades of carbon steel. The measured corrosion rates were significantly higher in biotic conditions compared to abiotic conditions and were found to be positively correlated with the pearlite phase content of the different grades of carbon steel coupons.