Marine biofilm bacterial community response and carbon steel loss following Deepwater Horizon spill contaminant exposure.

Steel marine structures provide foci of biodiversity when they develop into artificial reefs. Development begins with deposition of a biofilm. The effects of contaminants from oil spills on biofilm microbiomes, microbially-induced corrosion (MIC) and metal loss may impact preservation of marine metal structures. A microcosm experiment exposed biofilms on carbon steel disks (CSDs) to crude oil, dispersant, and dispersed oil to address their impacts on bacterial composition and metal loss and pitting. Biofilm diversity increased over time in all exposures. Community composition in dispersant and dispersed oil treatments deviated from the controls for the duration of a 12-week experiment. As biofilms matured, Pseudomonadaceae increased while Rhodobacteraceae decreased in abundance in dispersed oil treatments compared to the controls and dispersant treatments. Greatest mass loss and deepest pitting on CSDs were observed in dispersed oil treatments, suggesting impacts manifest as a consequence of increased MIC potential on carbon steel.

The corrosion process caused by the activity of the anaerobic sporulated bacterium Clostridium celerecrescens on API XL 52 steel.

The microbial corrosion of oil and gas pipes is one of the problems occurring in the oil industry. Various mechanisms explaining microbial corrosion have been demonstrated. Commonly, biocorrosion is attributed to sulfate-reducing bacteria. Also, it has recently been reported that microbial species can connect their electron transport system to metal electrodes. In this research, two spore-forming bacteria isolated in different years from a gas pipeline were identified by biochemical techniques and by 16S rDNA amplification, sequencing, and comparison with the NCBI database. Isolates were also compared between them using molecular techniques as the restriction patterns, unique for 16S rDNA (ARDRA), and the profile of the amplified bit from the genomic DNA, using an unspecific primer (RAPD). The results obtained showed that both isolates corresponded to Clostridium celerecrescens with a 99% similarity according to the sequence reported on the NCBI database. Also, the ARDRA and RAPD electrophoretic profiles of both strains were identical, and no plasmids were found in the strains. Thus, it can be settled that this bacterium is persistent in the environment prevailing in gas pipelines. Also, it was demonstrated that the bacterial secretion of organic acids contributes to the pitting and general biocorrosion of API XL 52 steel. The rates of corrosion obtained, approximately after 40 days, were correlated with the presence and metabolic activity of C. celerecrescens on the metallic surfaces.

Estimating Bacterial Concentrations in Fibrous Substrates Through a Combination of Scanning Electron Microscopy and ImageJ.

Conventional signal-based microanalytical techniques for estimating bacterial concentrations are often susceptible to false signals. A visual quantification, therefore, may compliment such techniques by providing additional information and support better management decisions in the event of outbreaks. Herein, we explore a method that combines electron microscopy (EM) and image-analysis techniques and allows both visualization and quantification of pathogenic bacteria adherent even to complex nonuniform substrates. Both the estimation and imaging parameters were optimized to reduce the estimation error ( E, %) to close to ±5%. The method was validated against conventional microbiological techniques such as the use of optical density, flow cytometry, and quantitative real-time PCR (qPCR). It could easily be tailored to estimate different species of pathogens, such as Escherichia coli O157, Listeria innocua, Staphylococcus aureus, Enterococcus faecalis, and Bacillus anthracis, on samples similar to those in real-time contamination scenarios. The present method is sensitive enough to detect ∼100 bacterial CFU/mL but has the potential to estimate even lower concentrations with increased imaging and computation times. Overall, this imaging-based method may greatly complement any signal-based pathogen-detection technique, especially in negating false signals, and therefore may significantly contribute to the field of analytical microbiology and biochemistry.

Microbial fouling and corrosion of carbon steel in deep anoxic alkaline groundwater.

Understanding the corrosion of carbon steel materials of low and intermediate level radioactive waste under repository conditions is crucial to ensure the safe storage of radioactive contaminated materials. The waste will be in contact with the concrete of repository silos and storage containers, and eventually with groundwater. In this study, the corrosion of carbon steel under repository conditions as well as the microbial community forming biofilm on the carbon steel samples, consisting of bacteria, archaea, and fungi, was studied over a period of three years in a groundwater environment with and without inserted concrete. The number of biofilm forming bacteria and archaea was 1,000-fold lower, with corrosion rates 620-times lower in the presence of concrete compared to the natural groundwater environment. However, localized corrosion was detected in the concrete-groundwater environment indicating the presence of local microenvironments where the conditions for pitting corrosion were favorable.

Corrosion behaviour and biocorrosion of galvanized steel water distribution systems.

Galvanized steel tubes are a popular mean for water distribution systems but suffer from corrosion despite their zinc or zinc alloy coatings. First, the quality of hot-dip galvanized (HDG) coatings was studied. Their microstructure, defects, and common types of corrosion were observed. It was shown that many manufactured tubes do not reach European standard (NBN EN 10240), which is the cause of several corrosion problems. The average thickness of zinc layer was found at 41µm against 55µm prescribed by the European standard. However, lack of quality, together with the usual corrosion types known for HDG steel tubes was not sufficient to explain the high corrosion rate (reaching 20µm per year versus 10µm/y for common corrosion types). Electrochemical tests were also performed to understand the corrosion behaviours occurring in galvanized steel tubes. Results have shown that the limiting step was oxygen diffusion, favouring the growth of anaerobic bacteria in steel tubes. EDS analysis was carried out on corroded coatings and has shown the presence of sulphur inside deposits, suggesting the likely bacterial activity. Therefore biocorrosion effects have been investigated. Actually sulphate reducing bacteria (SRB) can reduce sulphate contained in water to hydrogen sulphide (H2S), causing the formation of metal sulphides. Although microbial corrosion is well-known in sea water, it is less investigated in supply water. Thus, an experimental water main was kept in operation for 6months. SRB were detected by BART tests in the test water main.

Influence of respiratory substrate in carbon steel corrosion by a Sulphate Reducing Prokaryote model organism.

Sulphate Reducing Prokaryotes (SRP) are an important group of microorganisms involved in biocorrosion processes. Sulphide production is recognized as a fundamental cause of corrosion and nitrate is often used as treatment. The present work analyses the influence of respiratory substrates in the metal, from off-shore installations, SRP influenced corrosion, using Desulfovibrio desulfuricans ATTC 27774 as model organism, since this can switch from sulphate to nitrate. Open Circuit Potential over 6days in different conditions was measured, showing an increase around 200 and 90mV for the different media. Tafel plots were constructed allowing Ecorr and jcorr calculations. For SRP in sulphate and nitrate media Ecorr values of -824 and -728mV, and jcorr values of 2.5 and 3.7µAcm(-2), respectively, were attained indicating that in nitrate, the resultant corrosion rate is larger than in sulphate. Also, it is shown that the equilibrium of sulphide in the solution/gas phases is a key factor to the evolution of corrosion Nitrate prevents pitting but promotes general corrosion and increases the corrosion potential and iron dissolution 40 times when compared to sulphate. Our results demonstrate that nitrate injection strategy in oil fields has to be considered carefully as option to reduce souring and localized corrosion.

Impact of Desulfovibrio alaskensis biofilms on corrosion behaviour of carbon steel in marine environment.

Sulfate reducing prokaryotes are associated with the steel deterioration. They build heterogeneous biofilms, capable of accelerating corrosion processes. In this study metabolic activity and the biofilm development of Desulfovibrio alaskensis were correlated to electrochemical response of carbon steel surface. In the exponential growth phase sulfide concentration reached its maximum of about 10mM. This phenomenon was responsible for the parallel increase in the corrosion potential (Ecorr) up to -720mV (vs. SCE). Subsequently, during the intensive biofilm formation and development another Ecorr peak (-710mV vs. SCE) occurred. Decrease in Ecorr was registered during the biofilm maturation and kept stable, being 20mV lower than in the control. While carbon steel was protected from the microbial attachment and exposed to metabolic products, only one potential maximum (-730mV vs. SCE) was recorded. Here Ecorr variations coincided with sulfide concentration changes and kept at 120mV lower vs. the control. Weight loss examinations revealed corrosion rates, which did not exceed 0.05mm/y. Confocal microscopy suggested the importance of extracellular proteins in the biofilm formation. Above 150 proteins were detected in the EPS matrix. Surface effects of biofilm and metabolic products were visualised, revealing the role of attached microorganisms in the localised corrosion.

Combined geochemical and electrochemical methodology to quantify corrosion of carbon steel by bacterial activity.

The availability of respiratory substrates, such as H2 and Fe(II,III) solid corrosion products within nuclear waste repository, will sustain the activities of hydrogen-oxidizing bacteria (HOB) and iron-reducing bacteria (IRB). This may have a direct effect on the rate of carbon steel corrosion. This study investigates the effects of Shewanella oneidensis (an HOB and IRB model organism) on the corrosion rate by looking at carbon steel dissolution in the presence of H2 as the sole electron donor. Bacterial effect is evaluated by means of geochemical and electrochemical techniques. Both showed that the corrosion rate is enhanced by a factor of 2-3 in the presence of bacteria. The geochemical experiments indicated that the composition and crystallinity of the solid corrosion products (magnetite and vivianite) are modified by bacteria. Moreover, the electrochemical experiments evidenced that the bacterial activity can be stimulated when H2 is generated in a small confinement volume. In this case, a higher corrosion rate and mineralization (vivianite) on the carbon steel surface were observed. The results suggest that the mechanism likely to influence the corrosion rate is the bioreduction of Fe(III) from magnetite coupled to the H2 oxidation.

Influence of hydrogen-oxidizing bacteria on the corrosion of low carbon steel: Local electrochemical investigations.

Low carbon steel has been considered a suitable material for component of the multi-barrier system employed on the geological disposal of high-level radioactive waste (HLW). A non negligible amount of dihydrogen (H2) is expected to be produced over the years within the geological repository due to the anoxic corrosion of metallic materials and also to the water radiolysis. The influence of the activity of hydrogen-oxidizing bacteria (HOB) and iron-reducing bacteria (IRB) on carbon steel corrosion is considered in this study because of the high availability of energetic nutriments (H2, iron oxides and hydroxides) produced in anoxic disposal conditions. Local electrochemical techniques were used for investigating the activity of IRB as a promoter of local corrosion in the presence of H2 as electron donor. A local consumption of H2 by the bacteria has been evidenced and impedance measurements indicate the formation of a thick layer of corrosion products.

Gamma irradiation of cultural artifacts for disinfection using Monte Carlo simulations.

In this study, it has been investigated the disinfection of Korean cultural artifacts by gamma irradiation, simulating the absorbed dose distribution on the object with the Monte Carlo methodology. Fungal contamination was identified on two traditional Korean agricultural tools, Hongdukkae and Holtae, which had been stored in a museum. Nine primary species were identified from these items: Bjerkandera adusta, Dothideomycetes sp., Penicillium sp., Cladosporium tenuissimum, Aspergillus versicolor, Penicillium sp., Entrophospora sp., Aspergillus sydowii, and Corynascus sepedonium. However, these fungi were completely inactivated by gamma irradiation at an absorbed dose of 20kGy on the front side. Monte Carlo N Particle Transport Code was used to simulate the doses applied to these cultural artifacts, and the measured dose distributions were well predicted by the simulations. These results show that irradiation is effective for the disinfection of cultural artifacts and that dose distribution can be predicted with Monte Carlo simulations, allowing the optimization of the radiation treatment.

Factors influencing the microbial composition of metalworking fluids and potential implications for machine operator's lung.

Hypersensitivity pneumonitis, also known as "machine operator's lung" (MOL), has been related to microorganisms growing in metalworking fluids (MWFs), especially Mycobacterium immunogenum. We aimed to (i) describe the microbiological contamination of MWFs and (ii) look for chemical, physical, and environmental parameters associated with variations in microbiological profiles. We microbiologically analyzed 180 MWF samples from nonautomotive plants (e.g., screw-machining or metal-cutting plants) in the Franche-Comté region in eastern France and 165 samples from three French automotive plants in which cases of MOL had been proven. Our results revealed two types of microbial biomes: the first was from the nonautomotive industry, showed predominantly Gram-negative rods (GNR), and was associated with a low risk of MOL, and the second came from the automotive industry that was affected by cases of MOL and showed predominantly Gram-positive rods (GPR). Traces of M. immunogenum were sporadically detected in the first type, while it was highly prevalent in the automotive sector, with up to 38% of samples testing positive. The use of chromium, nickel, or iron was associated with growth of Gram-negative rods; conversely, growth of Gram-positive rods was associated with the absence of these metals. Synthetic MWFs were more frequently sterile than emulsions. Vegetable oil-based emulsions were associated with GNR, while mineral ones were associated with GPR. Our results suggest that metal types and the nature of MWF play a part in MWF contamination, and this work shall be followed by further in vitro simulation experiments on the kinetics of microbial populations, focusing on the phenomena of inhibition and synergy.

Prevention of biofilm growth on man-made surfaces: evaluation of antialgal activity of two biocides and photocatalytic nanoparticles.

As algal growth on man-made surfaces impacts their appearance, biocides and surfaces with self-cleaning properties are widely used in the building and paint industries. The objective of this study was to evaluate the antialgal activity of two biocides (triazine and isothiazoline) and photocatalytic nanoparticles of zinc oxide (20-60 nm). An aeroterrestrial green, microalgal strain of the genus Stichococcus was chosen as the test organism. By comparing a set of different structural and physiological performance parameters, lethal and also sublethal (chronic) effects were determined. Even though the herbicide triazine effectively inhibited growth (EC50 = 1.6 micromol l(-1)) and photosynthetic performance, structural properties (eg membrane integrity) were unaffected at the EC100 (250 micromol l(-1)), hence this herbicide did not kill the algal cells. In contrast, and due to their multiple modes of action, isothiazoline and the photocatalytic nanoparticles (the latter activated with low UV radiation) severely impacted all performance and structural parameters.

A practical approach for the validation of sterility, endotoxin and potency testing of bone marrow mononucleated cells used in cardiac regeneration in compliance with good manufacturing practice.

BACKGROUND: Main scope of the EU and FDA regulations is to establish a classification criterion for advanced therapy medicinal products (ATMP). Regulations require that ATMPs must be prepared under good manufacturing practice (GMP). We have validated a commercial system for the determination of bacterial endotoxins in compliance with EU Pharmacopoeia 2.6.14, the sterility testing in compliance with EU Pharmacopoeia 2.6.1 and a potency assay in an ATMP constituted of mononucleated cells used in cardiac regeneration. METHODS: For the potency assay, cells were placed in the upper part of a modified Boyden chamber containing Endocult Basal Medium with supplements and transmigrated cells were scored. The invasion index was expressed as the ratio between the numbers of invading cells relative to cell migration through a control insert membrane. For endotoxins, we used a commercially available system based on the kinetic chromogenic LAL-test. Validation of sterility was performed by direct inoculation of TSB and FTM media with the cell product following Eu Ph 2.6.1 guideline. RESULTS AND DISCUSSION: The calculated MVD and endotoxin limit were 780x and 39 EU/ml respectively. The 1:10 and 1:100 dilutions were selected for the validation. For sterility, all the FTM cultures were positive after 3 days. For TSB cultures, Mycetes and B. subtilis were positive after 5 and 3 days respectively. The detection limit was 1-10 colonies. A total of four invasion assay were performed: the calculated invasion index was 28.89 +/- 16.82% (mean +/- SD). CONCLUSION: We have validated a strategy for endotoxin, sterility and potency testing in an ATMP used in cardiac regeneration. Unlike pharmaceutical products, many stem-cell-based products may originate in hospitals where personnel are unfamiliar with the applicable regulations. As new ATMPs are developed, the regulatory framework is likely to evolve. Meanwhile, existing regulations provide an appropriate structure for ensuring the safety and efficacy of the next generation of ATMPs. Personnel must be adequately trained on relevant methods and their application to stem-cell-based products.