Biocorrosion and Biofilm Formation in a Nutrient Limited Heating System
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CORE Metadata, citation and similar papers at core.ac.uk Provided by Universidade do Minho: RepositoriUM Biofouling Vol. 25, No. 8, November 2009, 727–737 Biocorrosion and biofilm formation in a nutrient limited heating system subjected to alternating microaerophilic conditions B.V. Kjellerupa,b*, K.U. Kjeldsenc, F. Lopesd, L. Abildgaardc, K. Ingvorsenc, B. Frølunde, K.R. Sowersb and P.H. Nielsena aDepartment of Biotechnology, Chemistry and Environmental Engineering, Aalborg University, Sohngaardsholmsvej 49, DK-9000 Aalborg, Denmark; bCenter of Marine Biotechnology, University of Maryland Biotechnology Institute, 701 E. Pratt St, Baltimore, MD 21202, USA; cDepartment of Biological Sciences, Section for Microbiology, Aarhus University, Ny Munkegade, Building 1540, DK-8000 A˚rhus C., Denmark; dDepartment of Biological Engineering, University of Minho, Campus de Gualtar, 4710–057 Braga, Portugal; eDanish Technological Institute, Centre for Chemical and Water Technology, Teknologiparken, DK-8000 A˚rhus C., Denmark (Received 17 March 2009; final version received 10 June 2009) Severe biofilm formation and biocorrosion have been observed in heating systems even when the water quality complied with existing standards. The coupling between water chemistry, biofilm formation, species composition, and biocorrosion in a heating system was investigated by adding low concentrations of nutrients and oxygen under continuous and alternating dosing regimes. Molecular analysis of 16S rRNA gene fragments demonstrated that the amendments did not cause changes in the overall bacterial community composition. The combined alternating dosing of nutrients and oxygen caused increased rates of pitting (bio-) corrosion. Detection of bacteria involved in sulfide production and oxidation by retrieval of the functional dsrAB and apsA genes revealed the presence of Gram-positive sulfate- and sulfite-reducers and an unknown sulfur-oxidizer. Therefore, to control biocorrosion, sources of oxygen and nutrients must be limited, since the effect of the alternating operational conditions apparently is more important than the presence of potentially corrosive biofilm bacteria. Keywords: biocorrosion; biofilm; heating systems; sulfate reducing bacteria (SRB); functional genes dsrAB and apsA Introduction respiration resulting in increased biofilm formation Biofilm formation in heating systems and related and promotion of diverse microbial populations. problems such as biofouling and biocorrosion have Biocorrosion in heating systems can occur due to the for many years been considered insignificant because presence of many potentially corrosive groups of of good water quality with high pH, anaerobic microorganisms such as sulfate-reducing bacteria conditions, low conductivity, and limited nutrient (SRB), methanogenic Archaea, organic acid-producing availability. However, several studies have found that bacteria, sulfur-oxidizing bacteria, iron- and manga- this is not always the case. While general corrosion nese-oxidizing and reducing bacteria (Domingo et al. rates were low, biocorrosion rates, as inferred from 1998; Little et al. 2000; Hamilton 2003; Rozanova et al. pitting corrosion, were generally an order of magni- 2003; Zhang et al. 2003). SRB, in particular, have been tude higher (Kjellerup et al. 2003; Olesen et al. 2003). related to corrosion in anoxic systems (Dinh et al. Biocorrosion has been estimated to cost millions of 2004; Videla and Herrera 2005) and they have been dollars annually because of increased costs of opera- detected in the majority of heating systems investigated tion and maintenance in addition to the reduced (Goeres et al. 1998). However, heating systems harbor lifetime caused by perforation/corrosion of pipes and diverse microbial communities and so far it has not other installations (Flemming 1996). been possible to link the occurrence of specific The pretreatment of the heating water normally microbial associations and biocorrosion (Kjellerup implies ion exchange, pH adjustment (9.2–10.0), and et al. 2003, 2004, 2005; Kjeldsen et al. 2007). Although deaeration to reduce the occurrence of chemically poorly understood, biocorrosion is likely the result of induced corrosion. Measurements of oxygen are interplay between the activity of corrosive microorgan- regularly performed showing low concentrations be- isms and environmental physico-chemical conditions. cause the steel surfaces serve as oxygen scavengers In particular, oxygen intrusion events may play an (Heitz 1996). Oxygen may also be removed by bacterial important role because concomitant or sequential *Corresponding author. Email: [email protected] ISSN 0892-7014 print/ISSN 1029-2454 online Ó 2009 Taylor & Francis DOI: 10.1080/08927010903114611 http://www.informaworld.com 728 B.V. Kjellerup et al. presence of reduced sulfur compounds derived from Materials and methods the activity of SRB and oxygen can create a corrosive environment (Lee and de Beer 1995). Operational conditions Aerobic bacteria are often observed in heating Corrosion monitoring units (CMUs) based on the systems without oxygen being detected, indicating the rotortorque principle (Griebe and Flemming 2000) and presence of oxygen in concentrations below the equipped with mild steel coupons were installed in a detection limit or temporarily at elevated concentra- side stream of the investigated heating system in tions (Kjellerup et al. 2003, 2006). Additionally, SRB Aalborg, Denmark to ensure in situ conditions have been found in aerobic environments such as throughout the experiment. This heating system was activated sludge aeration tanks (Itoh et al. 2002; selected because of its high water quality and Kjeldsen et al. 2004) and the top layers of microbial uninterrupted operation resulting in low in situ mats (Minz et al. 1999), where they can survive and corrosion rates compared to other heating systems consume oxygen even though respiration with sulfate (Kjellerup et al. 2004, 2005). Therefore, this heating as electron acceptor does not occur (Dilling and system was suitable for evaluating the corrosion effects Cypionka 1990; Krekeler et al. 1998; Teske et al. caused by induced changes in the water quality. 1998). Based on this, the role SRB plays in biocorro- Eight CMUs were run during the experiment sion of heating systems in the presence of potentially to monitor biocorrosion under different conditions microaerophilic conditions is important. (Table 1). The flow through the CMUs was *1l Nevertheless, studies of the effect of water chem- h71 (hydraulic resistance time of *1 h) with istry on biofilm formation, community composition, turbulent conditions (surface velocity of *2m s71) and biocorrosion in heating systems are lacking, imitating the conditions of a heating pipe. The possibly because of the technical challenges in experi- temperature in the return circuit ranged from 39 to mentally manipulating heating systems under in situ 418C (summer/winter). The water quality of the like conditions. Such studies are important to under- heating system was: pH 9.8+0.2; conductivity stand and prevent biocorrosion. Until recently, little 550 mScm71; oxygen 510 mgl71; chloride 53 has been known about corrosive bacteria because they mg l71; sulfate 52mgl71; total iron 550 mg l71. are difficult to grow; thus the application of culture- Part 1 of the experiment contained five CMUs dependent methods has been shown to be difficult to supplied with nutrients (sodium acetate and yeast apply and has led to incorrect results and conclusions extract, 5 mg l71 of each), oxygen (0.3 mg l71), or (Wagner et al. 1993; Eschenhagen et al. 2003). There- glutaraldehyde (50 mg l71) once per day by fore, the application of culture-independent techniques anaerobically adding (with syringes flushed with for detection and identification of microorganisms nitrogen) as concentrated solutions as possible to involved in biocorrosion is necessary. A selection of minimize the added volumes, while oxygen was techniques has been applied to study the microbial added via saturated heating water at 258C. Oxygen populations in environmental samples, such as fluor- and nutrients were added at 12 h intervals to avoid escence in situ hybridization (FISH), the polymerase growth of heterotrophic aerobic bacteria. Part 2 chain reaction (PCR), denaturing gradient gel electro- contained three CMUs, where nutrients were con- phoresis (DGGE), and terminal restriction fragment tinuously added to two CMUs with different pH length polymorphism (TRFLP) (Muyzer et al. 1993; values and one CMU was run as a control without Amann et al. 1995; Wilderer et al. 2002; Wagner et al. any additions. The start concentration of organic 2003). matter corresponded to five times the recommended This study examined whether corrosion of mild maximum limit for Danish heating systems (Metro- steel could be related to variations in physical– politan Copenhagen Heating Transmission Company chemical conditions such as alternating concentrations 1994), while the oxygen concentration was 15 times of oxygen and nutrient availability in the heating water above the recommendation (Danish Council of in the presence of potentially corrosive biofilm District Heating 1999). These conditions simulate bacteria. The effect of microaerophilic conditions on conditions where the heating system would be biocorrosion is of special interest as large numbers of continuously contaminated from inflowing sources aerobic bacteria previously have been detected in having a higher pressure than the heating pipe or heating systems