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.     

Impact of nitrate-mediated microbial control of souring in oil reservoirs on the extent of corrosion

The effect of microbial control of souring on the extent of corrosion was studied in a model system consisting of pure cultures of the nitrate-reducing, sulfide-oxidizing bacterium (NR-SOB) Thiomicrospira sp. strain CVO and the sulfate-reducing bacterium (SRB) Desulfovibrio sp. strain Lac6, as well as in an SRB consortium enriched from produced water from a Canadian oil reservoir. The average corrosion rate induced by the SRB consortium (1.4 g x m(-2) x day(-1)) was faster than that observed in the presence of strain Lac6 (0.2 g x m(-2) x day(-1)). Examination of the metallic coupons at the end of the tests indicated a uniform corrosion in both cases. Addition of CVO and 10 mM nitrate to a fully grown culture of Lac6 or the SRB consortium led to complete removal of sulfide from the system and a significant increase in the population of CVO, as determined by reverse sample genome probing. In the case of the SRB consortium addition of just nitrate (10 mM) had a similar effect. When grown in the absence of nitrate, the consortium was dominated by Desulfovibrio sp. strains Lac15 and Lac29, while growth in the presence of nitrate led to dominance of Desulfovibrio sp. strain Lac3. The addition of CVO and nitrate to the Lac6 culture or nitrate to the SRB consortium accelerated the average corrosion rate to 1.5 and 2.9 g x m(-2) x day(-1), respectively. Localized corrosion and the occurrence of pitting were apparent in both cases. Although the sulfide concentration (0.5-7 mM) had little effect on corrosion rates, a clear increase of the corrosion rate with increasing nitrate concentration was observed in experiments conducted with consortia enriched from produced water.     

Metal corrosion by aerobic bacteria isolated from stimulated corrosion systems: Effects of additional nitrate sources

Many microorganisms are reported to influence the corrosive behaviour of mild steel and stainless steel in different habitats. In this study, 40 bacterial strains were isolated from corroded mild steel and stainless steel coupons in the nitrate supplemented environments. The corrosion abilities of the isolates against the mild steel and stainless steel coupons were tested with or without additional nitrate sources. The presence of bacterial isolates alone stimulated the corrosion of mild steel coupons. Most of the bio-corrosion processes of mild steel coupons were mitigated by adding nitrate supplement with bacterial isolates. The effects of bacterial isolates and additional nitrogen sources on corrosion of stainless steels were varied. Not all bacterial isolates stimulated the corrosion on stainless steel during the study period. Unlike the effects on mild steel coupons, additional NaNO₃ might stimulate, retard the corrosion rate by the bacterial isolates or have limited effects. Similar results were obtained when NH₄NO₃ was used. Phylogenetic analysis demonstrated that all isolates were closely related. The majority of the bacterial isolates from corroded metal coupons were identified as Bacillus species. Others were identified as Pseudomonas sp., Marinobacter sp., and Halomonas species. The results prove that the isolated aerobic microorganisms do play a role in the corrosion process of stainless and mild steel. Adding additional nitrate sources might be a tool to mitigate corrosion of mild steel which was stimulated by the presence of bacteria. However, to prevent the corrosion of stainless steels, it might need a trial and errors approach in each case.     

Effects of diverse water pipe materials on bacterial communities and water quality in the annular reactor

To investigate the effects of pipe materials on biofilm accumulation and water quality, an annular reactor with the sample coupons of four pipe materials (steel, copper, stainless steel, and polyvinyl chloride) was operated under hydraulic conditions similar to a real plumbing system for 15 months. The bacterial concentrations were substantially increased in the steel and copper reactors with progression of corrosion, whereas those in stainless steel (STS) and polyvinyl chloride (PVC) reactors were affected mainly by water temperature. The heterotrophic plate count (HPC) of biofilms was about 100 times higher on steel pipe than other pipes throughout the experiment, with the STS pipe showing the lowest bacterial number at the end of the operation. Analysis of the 16S rDNA sequences of 176 cultivated isolates revealed that 66.5% was Proteobacteria and the others included unclassified bacteria, Actinobacteria, and Bacilli. Regardless of the pipe materials, Sphingomonas was the predominant species in all biofilms. PCR-DGGE analysis showed that steel pipe exhibited the highest bacterial diversity among the metallic pipes, and the DGGE profile of biofilm on PVC showed three additional bands not detected from the profiles of the metallic materials. Environmental scanning electron microscopy showed that corrosion level and biofilm accumulation were the least in the STS coupon. These results suggest that the STS pipe is the best material for plumbing systems in terms of the microbiological aspects of water quality.     

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).     

Microbial fouling community analysis of the cooling water system of a nuclear test reactor with emphasis on sulphate reducing bacteria

Culture and molecular-based techniques were used to characterize bacterial diversity in the cooling water system of a fast breeder test reactor (FBTR). Techniques were selected for special emphasis on sulphate-reducing bacteria (SRB). Water samples from different locations of the FBTR cooling water system, in addition to biofilm scrapings from carbon steel coupons and a control SRB sample were characterized. Whole genome extraction of the water samples and SRB diversity by group specific primers were analysed using nested PCR and denaturing gradient gel electrophoresis (DGGE). The results of the bacterial assay in the cooling water showed that the total culturable bacteria (TCB) ranged from 10(3) to 10(5)?cfu?ml(-1); iron-reducing bacteria, 10(3) to 10(5)?cfu?ml(-1); iron oxidizing bacteria, 10(2) to 10(3)?cfu?ml(-1) and SRB, 2-29?cfu?ml(-1). However, the counts of the various bacterial types in the biofilm sample were 2-3 orders of magnitude higher. SRB diversity by the nested PCR-DGGE approach showed the presence of groups 1, 5 and 6 in the FBTR cooling water system; however, groups 2, 3 and 4 were not detected. The study demonstrated that the PCR protocol influenced the results of the diversity analysis. The paper further discusses the microbiota of the cooling water system and its relevance in biofouling.     

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     

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.     

Microbial fouling community analysis of the cooling water system of a nuclear test reactor with emphasis on sulphate reducing bacteria

Culture and molecular-based techniques were used to characterize bacterial diversity in the cooling water system of a fast breeder test reactor (FBTR). Techniques were selected for special emphasis on sulphate-reducing bacteria (SRB). Water samples from different locations of the FBTR cooling water system, in addition to biofilm scrapings from carbon steel coupons and a control SRB sample were characterized. Whole genome extraction of the water samples and SRB diversity by group specific primers were analysed using nested PCR and denaturing gradient gel electrophoresis (DGGE). The results of the bacterial assay in the cooling water showed that the total culturable bacteria (TCB) ranged from 10³ to 10⁵ cfu ml^?1; iron-reducing bacteria, 10³ to 10⁵ cfu ml^?1; iron oxidizing bacteria, 10² to 10³ cfu ml^?1 and SRB, 2–29 cfu ml^?1. However, the counts of the various bacterial types in the biofilm sample were 2–3 orders of magnitude higher. SRB diversity by the nested PCR-DGGE approach showed the presence of groups 1, 5 and 6 in the FBTR cooling water system; however, groups 2, 3 and 4 were not detected. The study demonstrated that the PCR protocol influenced the results of the diversity analysis. The paper further discusses the microbiota of the cooling water system and its relevance in biofouling.     

Monitoring of a Vibrio natriegens and Desulfovibrio vulgaris marine aerobic biofilm on a stainless steel surface in a laboratory tubular flow system

N.D. BENBOUZID-ROLLET, M. CONTE, J. GUEZENNEC AND D. PRIEUR. 1991. In an aerobic bulk environment sulphate-reducing bacteria (SRB) can find suitable growth conditions on surfaces where other micro-organisms have developed an extensive biofilm. On metal surfaces they may induce or enhance corrosion. A laboratory tubular flow system was designed to study this phenomenon by creating a biofilm on stainless steel under dynamic conditions with Vibrio natriegens and Desulfovibrio vulgaris. The sulphate reducer colonized the surface, constituting approximately 5% of the total population. Its in situ growth rate, calculated by a simplified mathematical model, showed that the attached SRB multiplied at their settling locations. No significant difference with respect to corrosion enhancement was found in the electrochemical reactions of the metal betwen the control and the reactor, where D. vulgaris was present in the biofilm.     

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.     

Comparison of microbial communities involved in souring and corrosion in offshore and onshore oil production facilities in Nigeria

Samples were obtained from the Obigbo field, located onshore in the Niger delta, Nigeria, from which oil is produced by injection of low-sulfate groundwater, as well as from the offshore Bonga field from which oil is produced by injection of high-sulfate (2,200 ppm) seawater, amended with 45 ppm of calcium nitrate to limit reservoir souring. Despite low concentrations of sulfate (0–7 ppm) and nitrate (0 ppm), sulfate-reducing bacteria (SRB) and heterotrophic nitrate-reducing bacteria (NRB) were present in samples from the Obigbo field. Biologically active deposits (BADs), scraped from corrosion-failed sections of a water- and of an oil-transporting pipeline (both Obigbo), had high counts of SRB and high sulfate and ferrous iron concentrations. Analysis of microbial community composition by pyrosequencing indicated anaerobic, methanogenic hydrocarbon degradation to be a dominant process in all samples from the Obigbo field, including the BADs. Samples from the Bonga field also had significant activity of SRB, as well as of heterotrophic and of sulfide-oxidizing NRB. Microbial community analysis indicated high proportions of potentially thermophilic NRB and near-absence of microbes active in methanogenic hydrocarbon degradation. Anaerobic incubation of Bonga samples with steel coupons gave moderate general corrosion rates of 0.045–0.049 mm/year, whereas near-zero general corrosion rates (0.001–0.002 mm/year) were observed with Obigbo water samples. Hence, methanogens may contribute to corrosion at Obigbo, but the low general corrosion rates cannot explain the reasons for pipeline failures in the Niger delta. A focus of future work should be on understanding the role of BADs in enhancing under-deposit pitting corrosion.     

Changes in physico-chemical characteristics and viable bacterial communities during fermentation of alfalfa silages inoculated with Lactobacillus plantarum

Sulfate-reducing bacteria (SRB) are culprits for microbiologically influenced corrosion, and biofilms are believed to play essential roles in the corrosion induced by SRB. However, little is known about the regulation of SRB biofilms. Quorum sensing signal molecules acyl-homoserine lactones (AHLs) and autoinducer-2 (AI-2) regulate biofilm formation of many bacteria. In this study, the production of AHLs and AI-2 by one SRB strain, Desulfovibrio sp. Huiquan2017, was detected, and the effect of exogenous AI-2 on bacterial biofilm formation was discussed. It was found that the cell-free supernatants of Desulfovibrio sp. Huiquan2017 induced luminescence in a ?luxS mutant strain Vibrio harveyi BB170, indicating the production of functional AI-2 by the bacterium. In the presence of exogenous AI-2, the growth of Desulfovibrio sp. Huiquan2017 and early biofilm formation were not affected, but the later stage of biofilm development was inhibited significantly. The biofilms became looser, smaller, and thinner, and contained less bacteria and extracellular polymeric substances (EPS). The inhibition effect of AI-2 on the biofilm development of Desulfovibrio sp. Huiquan2017 was mainly achieved through reducing the amount of EPS in biofilms. These findings shed light on the biofilm regulation of SRB.

     

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.     

A synergistic d-tyrosine and tetrakis hydroxymethyl phosphonium sulfate biocide combination for the mitigation of an SRB biofilm

Microbiologically influenced corrosion (MIC) is a major problem in various industries such as oil and gas, and water utilities. Billions of dollars are lost to microbiologically influenced corrosion (MIC) each year in the US. The key to MIC control is biofilm mitigation. Sulfate-reducing bacteria (SRB) are often the culprits. They are also involved in souring and biofouling. SRB biofilms are notoriously difficult to eradicate. Due to environmental concerns and increasing costs, better biocide treatment strategies are desired. Recent studies suggested that D: -tyrosine and some other D: -amino acids may signal biofilm dispersal. Experimental results in this work indicated that D: -tyrosine is an effective biocide enhancer for tetrakis hydroxymethyl phosphonium sulfate (THPS) that is a green biocide. Desulfovibrio vulgaris (ATCC 7757) was used in biofilm prevention and biofilm removal tests. It was found that 100?ppm D: -tyrosine alone and 50?ppm THPS alone were both ineffective against the SRB biofilm. However, when 1?ppm D: -tyrosine was combined with 50?ppm THPS, the synergy between the two chemicals successfully prevented the establishment of the SRB biofilm on C1018 mild steel coupon surfaces in batch treatment tests. It also eradicated established SRB biofilms from coupon surfaces in both 1 and 3-h shock treatment tests.     

A green biocide enhancer for the treatment of sulfate-reducing bacteria (SRB) biofilms on carbon steel surfaces using glutaraldehyde

Generally speaking, a much higher concentration of biocide is needed to treat biofilms compared to the dosage used to for planktonic bacteria. With increasing restrictions of environmental regulations and safety concerns on large-scale biocide uses such as oil field applications, it is highly desirable to make more effective use of biocides. In this paper a green biocide enhancer ethylenediaminedisuccinate (EDDS) that is a biodegradable chelator, was found to enhance the efficacy of glutaraldehyde in its treatment of sulfate-reducing bacteria (SRB) biofilms. Experiments were carried out in 100 ml anaerobic vials with carbon steel coupons. The ATCC 14563 strain of Desulfovibrio desulfuricans was used. Biofilms on coupon surfaces were visualized using scanning electron microscopy (SEM). Experimental results showed that EDDS reduced the glutaraldehyde dosages considerably in the inhibition of SRB biofilm establishment and the treatment of established biofilms on carbon steel coupon surfaces.     

Influence of sulfate reducing bacterial biofilm on corrosion behavior of low-alloy,high-strength steel (API-5L X80)

The utilization of high strength carbon steels in oil and gas transportation systems has recently increased. This work investigates microbiologically influenced corrosion (MIC) of API 5L X80 linepipe steel by sulfate reducing bacteria (SRB). The biofilm and pit morphology that developed with time were characterized with field emission scanning electron microscopy (FESEM). In addition, electrochemical impedance spectroscopy (EIS), polarization resistance (R_p) and open circuit potential (OCP) were used to analyze the corrosion behavior. Through circuit modeling, EIS results were used to interpret the physicoelectric interactions between the electrode, biofilm and solution interfaces. The results confirmed that the extensive localized corrosion activity of SRB is due to a formed biofilm and a porous iron sulfide layer on the metal surface. Energy Dispersive Spectroscopy (EDS) revealed the presence of different sulfide and oxide constituents in the corrosion products for the system exposed to SRB.     

The effect of long-term nitrate treatment on SRB activity,corrosion rate and bacterial community composition in offshore water injection systems

Biogenic production of hydrogen sulphide (H₂S) is a problem for the oil industry as it leads to corrosion and reservoir souring. Continuous injection of a low nitrate concentration (0.25–0.33 mM) replaced glutaraldehyde as corrosion and souring control at the Veslefrikk and Gullfaks oil field (North Sea) in 1999. The response to nitrate treatment was a rapid reduction in number and activity of sulphate-reducing bacteria (SRB) in the water injection system biofilm at both fields. The present long-term study shows that SRB activity has remained low at ≤0.3 and ≤0.9 μg H₂S/cm²/day at Veslefrikk and Gullfaks respectively, during the 7–8 years with continuous nitrate injection. At Veslefrikk, 16S rRNA gene based community analysis by PCR–DGGE showed that bacteria affiliated to nitrate-reducing sulphide-oxidizing Sulfurimonas (NR–SOB) formed major populations at the injection well head throughout the treatment period. Downstream of deaerator the presence of Sulfurimonas like bacteria was less pronounced, and were no longer observed 40 months into the treatment period. The biofilm community during nitrate treatment was highly diverse and relative stable for long periods of time. At the Gullfaks field, a reduction in corrosion of up to 40% was observed after switch to nitrate treatment. The present study show that nitrate injection may provide a stable long-term inhibition of SRB in sea water injection systems, and that corrosion may be significantly reduced when compared to traditional biocide treatment.     

Fouling and corrosion in an open recirculating cooling system

The paper discusses the problem of fouling and corrosion in the Service Water System of a nuclear reactor employing an open recirculating cooling system. Field and laboratory experiments were carried out using carbon steel as test material. The studies included corrosion rate measurements, corrosion product characterisation (using XRD, EDAX and IRAS), and the effects of addition of biocide (chlorine) and corrosion inhibitor (polyphosphates) on the bacterial population of the cooling water. The fouling films which developed on metal and glass coupons as well as debris collected from within the cooling system were chemically characterised. The results indicated the prevalence of microbiologically induced corrosion (MIC) within the system. The findings are discussed in the light of the information available on MIC of cooling water systems.     

Importance of biofilm formation for corrosion inhibition of SAE 1018 steel by axenic aerobic biofilms

To investigate if corrosion inhibition by aerobic biofilms is a general phenomenon, carbon steel (SAE 1018) coupons were exposed to a complex liquid medium (Luria–Bertani) and seawater-mimicking medium (VNSS) containing fifteen different pure-culture bacterial suspensions representing seven genera. Compared to sterile controls, the mass loss in the presence of these bacteria (which are capable of developing a biofilm to various degrees) decreased by 2- to 15-fold. The extent of corrosion inhibition in LB medium depended on the nature of the biofilm: an increased proportion of live cells, observed with confocal scanning laser microscopy (CSLM) and image analysis, decreased corrosion. Corrosion inhibition in LB medium was greatest with Pseudomonas putida (good biofilm formation), while metal coupons exposed to Streptomyces lividans in LB medium (poor biofilm formation) corroded in a manner similar to the sterile controls. Pseudomonas mendocina KR1 reduced corrosion the most in VNSS. It appears that only a small layer of active, respiring cells is required to inhibit corrosion, and the corrosion inhibition observed is due to the attached biofilm. Received 09 December 1996/ Accepted in revised form 19 March 1997