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Microorganism can be divided into bacteria, fungi, algae and viruses. Among all these types of microorganism, bacteria and fungi play important role in corrosion related problems. All the bacteria that involve in corrosion activities are grouped based on the oxygen tolerance (Normal Shaw,1974). Below are the groups:
Strict (obligate) - usually this microbial unable to function with the presence of oxygen
Aerobes - able to function with the presence of oxygen
Facultative anaerobes-works in both environment that is with and without the presence of oxygen
Microaerophiles - need oxygen for metabolism but at very low levels
In this project focus will be on bacteria.
Sulfate reducing bacteria under electron microscope (Aramco ExPats , 2007)
2.2 Introduction to Sulfate Reducing Bacteria
High sulfate concentration in water system can create problem such as corrosion of water transport system and of concrete structure. SRB is a non-pathogenic (will not cause disease) and anaerobic bacteria and they can produce enzyme to accelerate the reduction of sulfate compounds to hydrogen sulphide, that corrodes copper, iron and carbon steel ( Zhang Yan,Lin Jing, Yu Guiwei,2009) In order for this reduction to occur, four components must be present which are SRB (Sulfate Reducing Bacteria), sulfate, external energy source in the form of free electrons and the temperature of the water must be less than 65Â°C (G.F Yuzwa,1991)
A water system naturally consists of sulfate based components. When sulfate exist in a closed water system, the sodium sulphite is oxidized to sodium sulphate as indicated in the following reaction:
2Na+1 + SO3-1+ 0.5O2-2 = 2Na+1 + SO4-2 (1)
SRB can accelerate the corrosion mechanism of iron. The resultant accelerated corrosion mechanism of iron by the sulfate reducing bacteria is shown in below equations
SO 4-2 + 8H = S-2 + 4H2O (2)
8H2O = 8 OH-1 + 8H+ 1 (3)
2H+1 + S-2 = H2 S (4)
Fe+2 + S-2 = FeS (5)
3Fe+2 + 6(OH)-1 = 3Fe (OH)2 (6)
Since hydrogen sulphide is a by-product of SRB metabolism, this hydrogen sulphide can be used as an indicator that SRB may present in the water.
2.2.1 Classification of Sulfate Reducing Bacteria
SRB falls under the category of anaerobic obligate, which means it will be active without the presence of oxygen. Meanwhile if oxygen exists, SRB will go into 'resting mode' and they tend to survive but will not grow actively
SRB can be classified into five groups (Larry L. Barton, W. Allan Hamilton et all, 2007) :
Mesophilic (incubation temperature from 25Â°C to 30Â°C) Î´-proteobacteria with genera Desulfovibrio, Desulfobacterum, Desulfobacteria and Desulfobulbus
Thermophilic (incubation temperature above 45Â°C) - gram negative bacteria with genera Thermodesulfovibrio
Gram positive with genera Desulfotomaculum
Euryarchaeota with genera Archaeoglabus (Castro et al, 2000)
Thermodesulfobiaceae which is the most recent one (Mori, K., Kim, H., Kakegawa, T. & Hanada, S et al,2003 )
2.2.2 Impact of Sulfate Reducing Bacteria
SRB create advantages and disadvantage to mother nature. The advantages of SRB are:
i) Capability to degrade lignin compounds in municipal solid waste materials
in landfills (Kim JongHo, Kim Monll,Baewooken , 2009)
ii) Used to treat soil that is contaminated with mercury. H2S released by SRB
will be able to remove metals from water.
(T.Hakansson, P.Suer, B.Mattiasson, B.Allard, 2008)
The disadvantages of SRB are :
Water piping system that is exposed to SRB will easily go through
corrosion . Furthemore combination of hydrogen sulphide and sulphate makes piping system corrode at faster rate compated to the presence of hydrogen sulphide only (V.Assaad, 2006). Corrosion and embrittlement of steel in marine environment especially
seawater (M.J Robinson and P.J Kilgallon, 1998)(Yan Linna, 2008)
Fouling of pipe system -In most cases SRB will clogg piping system and effect the quality of system ( Rao TS, Kora AJ,Chandramohan P, Panigrahi
BS, Narasimhan SV,2009)
Souring of reservoirs- In most fuel piping system SRB will contaminate water or fuel and affect the quality of water and fuel. This will cost a lot from maintenance aspect. (Mei Liu, Neil S. Summer, Jason J. Gill, Ry
Young, and Chris Janes,2011)
Microbial fouling and corrosion inside water injection system (Aramco ExPats , 2007)
2.2.3 Cultivation of Sulfate Reducing Bacteria
Sulfate reducing bacteria were successfully enriched from sludge taken from pond in USM. It went through the process of isolation and inoculation. This process took nearly 2 months in the Biotechnology Industry Faculty in USM. The sample of SRB was then cultured and incubated for this project.
Before culturing SRB the proper medium has to be prepared so that SRB growth will be in better condition. For this purpose 2.8g of NaCl was added into 300ml distilled water. This mixture was boiled for around 6 minutes. This mixture was poured into 10 universal bottles and the remaining was poured into 250ml medium bottle. These bottles were autoclaved at 121â°C as shown in figure 1.Any microorganism will die at this temperature. Hence a medium which is sterilized is produced. Then the medium which is in liquid form was poured into 10 plates ( this is only done once the liquid is warm as the vapor produced will spoil the medium) . This is done in a Laminar Flow as shown in figure 2 which will give a sterilized surrounding.
Figure 1: Autoclave Figure 2 : Laminar Flow
SRB is transformed through the streaking method. A loop rod is used by heating it up first at the Bunsen burner. This is to ensure no other bacteria exist on the loop before touching SRB. A small sample from universal bottle which consist SRB is touched using the loop and then it is streaked into the petri dish. This process is done nearer to Bunsen burner. Purity of the cultures was checked by streaking on solidified sulfate reducing agar under anaerobic conditions (Jeffry George, 2008). After this process the plate is sealed with parafilm. This plate is kept inside mesophilic bacteria incubator at 30â°C, as shown in figure 3. Less than 10 hours later can see traces of SRB colony as shown in figure 4. Most common strains of SRB grow best at temperatures from 25Â°C to 35Â°C.( CLI Houstan)
Figure 3 : Incubator Figure 4 : Sample of SRB colony in plate
Petri dish has wider space for the colony to spread but it also has higher risk of being contaminated. SRB can live 1 to 2 weeks in petri dish. Since SRB is needed for longer duration, the sample is also streaked into universal bottles. The same method that was done for petri dish is applied for universal bottle. After incubation could see traces of SRB inside the bottle as shown in figure 5. The laboratory process to prepare the medium for SRB and incubating SRB takes less than 2 days
Figure 5 : Universal bottle with sample of SRB
For this experiment purpose, tube with medium is used. SRB is transferred from universal bottle using a capillary tube by filling the capillary tube completely with sample of SRB. The capillary tube is pushed deep inside the tube and then closed with the cap which is attached to a sensor as shown in Figure 6. This tube is incubated at 34â°C in field incubator as shown in figure 7.
2.2.4 Current Detection Methods
In literature there are several studies relating to methods of detecting SRB. To detect bio-corrosion a person should have a great idea of which types of microbes contaminate water and the effects of microorganism on water piping system. All these are done based on observation and theory. By the time SRB is detected it could be late and uncontrollable.
At present there are conventional methods and also lab testing method to detect SRB. Since the very nature of SRB is the odor that it emanate due to hydrogen sulphide, SRB can be detected through the smell. Hydrogen sulphide has pungent smell which can be connected to 'rotten egg' smell (John H. Schnieder,2005) Other conventional method is to use color forming unit and Most Probable Number (MPN) method (H.A. Videla,1996)
There is biological method that involves either laboratory analysis or field analysis (G.F.Yuzwa, 1991). The laboratory process takes longer time because sample of SRB needs to be kept in sterilized bottle and must be sent to laboratory within 24 hours of collection to avoid SRB from going into 'resting mode'. The procedures that are used in laboratory also take longer period.
Meanwhile the field analysis is easier because one need not to be expert to detect SRB. Usually there will be transparent dipslides with medium for SRB growth. These dipslides will be kept in incubator and incubated at 30â°C to 35â°C and after one to two days can see traces of SRB. The level of growth can be seen from the colony traces of SRB whether it is 'Nil', 'Slight', 'Moderate' or 'Severe'.( Kiitiwake Developments LTD ,2011).
Books and Journals:
G.F Yuzwa, P.Eng, Corrosion by Sulfate Reducing Bacteria, ALBERTA Public Works, Supply & Services Property Management Operations Divisions Water (1991)
Larry L. Barton, W.Allan Hamilton. (2007). Sulfate Reducing Bacteria, Environmental and Engineered System, Cambridge University Press
Jeffy George, C.S Purushothaman, Yogesh S Shouche.(2008). Isolation and
characterization of sulphate reducing bacteria Desulfovibrio vulgaris from Vajreshwari thermal springs in Maharashtra, India, World J Microbiol Biotechnol 24: 681-685, DOI 10.1007/s11274-007-9524-2
V.Assaad Abdelmseeh,J.C.Jofriet,S.C.Negi and G.Hayward (2006).Sulphide,
Sulphate and Sulphuric Acide Corrosion of Concrete in Laboratory Tests, Springer
Castro, M.C., Stute, M., and Schlosser, P. (2000), Comparison of 4He ages and
14C ages in simple aquifer systems: implications for groundwater flow and chronologies. Applied Geochemistry, v. 15, p. 1137-1167.
Mori, K., Kim, H., Kakegawa, T. & Hanada, S. (2003). A novel lineage of
sulfate-reducing microorganisms: Thermodesulfobiaceae fam. nov., Thermodesulfobium narugense, gen. nov., sp. nov., a new thermophilic isolate from a hot spring. Extremophiles 7, 283-290.
Kim JongHo, Kim Monll, Baewooken.(2009). Effect of oxidized leachate
on degradation of lignin by sulfate reducing bacteria, Journal waste management and Research ,Vol.27 No.5 pp.520-526
T.Hakansson,P.Suer,B.Mattiasson,B.Allard.(2008).Sulfate reducing bacteria
to precipitate mercury after electrokinetic soil remediation. Int.J.Environ.Sci.Tech,5(2),267-274,Spring
M.J. Robinson and P.J Kilgallon .(1998). A review of the effects of sulfate
reducing bacteria in the marine environment on the corrosion fatigue and hydrogen embrittlement of high strength steels. Health & safety Executive-Offshore Technology Report, ISBN 0-7176-1642-8
Yan.Linna.(2008).Corrosion behavior of 30X stainless steel in cultured
microbial medium,Journal of Chinese Society for Corrosion and Protection, Vol.28,pp.24-37
Zhang Yan, Lin Jing, Yu Guiwei.(2009). 30X stainless steel microbiological
influenced corrosion characteristics research, Surface Technology Vol 38,pp.44-45
T.Liu,H.Liu,Y.Hui,L.Zhou,B.Zheng.(2009).Growth characteristics of
thermophile sulfate reducing bacteria and its effect on carbon steel, Journal of Chinese Society for Corrosion and Protection, Vol.29,pp.93-97
13. Rao TS, Kora AJ,Chandramohan P, Panigrahi BS, Narasimhan SV.(2009). Biofouling and microbial corrosion problem in the thermo-fluid heat exchanger and cooling water system of a nuclear test reactor. Biofouling,Oct;25(7):581-91.
Mei Liu, Neil S. Summer, Jason J. Gill, Ry Young, and Chris Janes.(2011). Phage of Sulfate Reducing Bacteria Isolated from High Saline Environment. NACE International Corrosion 2011 Conference & EXPO,Houstan Texas USA
15. John H. Schnieder. (2005). Sulfate reducing bacteria, Water Well Journal,pg 68-
H.A. Videla, Manual of Biocorrosion.(1996). Lewis Publishers, Boca Raton.
16. Norman Shaw.( 1974).Structure and classification of bacteria. Advances in Applied Microbiology, Volume 17,1974,pg:63-108
Web Sites :
Aramco ExPats.(2007).Workshop gets on microbes good site.[Online], [Acessed 2011]. Available from World Wide Web:
Kiitiwake Developments LTD (2011).Filed Incubator for Dipslides, Kittiwake Measuring, Monitoring and Managing.[Online],[Acessed 2010]. Available from World Wide Web: