Severe Pitting Corrosion of Carbon Steel Caused by Sulfate Reducing Bacteria in Aqueous Medium

Sanaullah Sattar1, Ali Hussain2*, Javed Iqbal Qazi2, Arshad Javid1 and

Shahid Mehmood1

1Department of Wildlife and Ecology, University of Veterinary and Animal Sciences, Outfall Road, Lahore, Pakistan

2Institute of Zoology, University of the Punjab, Quaid-i-Azam Campus, Lahore, Pakistan

3Department of Poultry Production, University of Veterinary and Animal Sciences, Outfall Road, Lahore, Pakistan

ABSTRACT

The current study was conducted to assess the corrosive action of SRB on carbon steel coupons in aqueous media with and without the addition of nutrients of Postgate B medium. For the purpose, SRB strain was isolated from a highly polluted wastewater stream flowing in Lahore, Punjab, Pakistan. The bacterial isolate was motile, Gram-negative, non-spore former and identified as Desulfovibrio vulgaris after 18 S rRNA gene sequencing. The corrosive action of the bacteria was assessed in a 60-day trial of anaerobic incubation. After an incubation period of 60 days, it was found that CR was significantly higher for the coupons that were dipped in inoculated media. For these coupons, the average CR appeared as 210 ± 17 µg dm−2 d−1. However, CR was significantly lower (210 ± 17 µg dm−2 d−1) for the coupons dipped in inoculated water only (Fig. 2). In un-inoculated vials having nutrients, CR appeared higher (97 ± 16 µg dm−2 d−1) as compared to those without nutrients (48 ± 21 µg dm−2 d−1). Bacterial growth appeared higher in nutrient added vials. At termination of the experiments, total viable counts for SRB were 2.7 ± 0.3 × 109 CFU mL−1 in vials supplemented with nutrients. However, no bacterial growth was seen in nutrient-deprived vials. No change in pH was observed in all vials except those which were inoculated under optimum provision of nutrients. In these vials, pH changed from neutral to slightly basic in range (7.0 ± 0.5 to 6.0 ± 0.5). SEM analysis of the coupon which was recovered from a vial supplemented with nutrients revealed severe pitting corrosion of the steel coupon. Our findings will be helpful in understanding and protecting bacterial corrosion of water-dipped metallic bodies.

Article Information

Received 05 February 2023

Revised 18 March 2023

Accepted 10 April 2023

Available online 01 June 2023

(early access)

Published 31 October 2024

Authors’ Contribution

SS methodology. AH supervision. JIQ co-supervision. AJ data analysis. SM data compilation.

Key words

Bacterial corrosion, Biocorrosion, Biodeterioration and biodegradation, Biosulfidogens, Metal corrosion

DOI: https://dx.doi.org/10.17582/journal.pjz/20230205110237

* Corresponding author: ali.zool@pu.edu.pk

0030-9923/2024/0006-2837 $ 9.00/00

Copyright 2024 by the authors. Licensee Zoological Society of Pakistan.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

INTRODUCTION

Microbiologically influenced corrosion (MIC) is a type of corrosion that is caused by the presence of microorganisms, such as bacteria, fungi, and algae. These microorganisms can produce various by-products, such as acids, gases, and enzymes, that can corrode metal surfaces (Usher et al., 2014; Kiani et al., 2019; Kokilaramani et al., 2021). One common type of MIC is sulfate-reducing bacterial (SRB) corrosion, which occurs when SRB use sulfates as an electron acceptor in their metabolism and produce hydrogen sulfide (H2S) gas. The H2S gas can then react with water and oxygen to form sulfuric acid, which can corrode metal surfaces (Dou et al., 2018; Gu et al., 2019; Luo et al., 2023).

It is difficult to quantify the exact annual cost of MIC globally, as it depends on a variety of factors such as the type of industry, the type of microorganisms involved, and the location of the corrosion. However, it is estimated that MIC can account for a significant portion of the total cost of corrosion, which is estimated to be around 3% of the global gross domestic product (GDP) (Yazdi et al., 2023; Zhu et al., 2023). In the United States, the total annual cost of corrosion is estimated to be around $276 billion, and it is believed that a significant portion of this cost is due to MIC (Bhaskaran et al., 2005; Thompson et al., 2007; Chevallier et al., 2023). In the oil and gas industry, MIC is a major concern as it can cause corrosion of pipelines and other infrastructure, leading to costly repairs and disruptions in service (Popoola et al., 2013; Askari et al., 2019). MIC can also cause corrosion in a variety of other industries, including the marine, water treatment, and power generation industries (Jia et al., 2019; Etim et al., 2022).

SRB can cause corrosion of a variety of metals, including iron, steel, copper, aluminum, and zinc (Zarasvand and Rai, 2014; Sun et al., 2017; Sattar et al., 2023). The type of corrosion that occurs can depend on the specific characteristics of the metal, the environment in which the corrosion takes place, and the type and concentration of SRB present (Beech and Sunner, 2007; Javaherdashti, 2011; Jia et al., 2019; Zhao et al., 2022). Some of the different types of corrosion that can be caused by SRB include pitting corrosion, crevice corrosion and galvanic corrosion (Lane, 2005; Alamri, 2020).

MIC can occur in a variety of environments, including marine, freshwater, soil, and industrial settings (Coetser and Cloete, 2005; Ibrahim et al., 2018). It can affect a wide range of metal structures, including pipelines, tanks, ships, and offshore platforms (Loto, 2017; Kannan et al., 2018). MIC can be difficult to detect and diagnose, as it often occurs in hidden or hard-to-reach areas and may not be visible until significant damage has already been done (Little et al., 2006; Xu et al., 2020). Keeping in view the metal-damaging potential of SRB, the current study was designed to isolate and characterize an indigenous SRB strain from wastewater and to check its corrosive action on a comparatively corrosion-resistant form of steel (carbon steel).

MATERIALS AND METHODS

Bacterial isolation and identification

The SRB species was isolated from a wastewater stream (Hudiara Drain) passing through Mohlanwal, Lahore, Pakistan. In order to isolate SRB, water (approximately 100 mL) was collected for the purpose, placed in a sterile glass container, and brought to the lab. Using the Postgate B medium (concentration in g L−1: CaSO4, 1.0; FeSO4.7H2O, 0.5; KH2PO4, 0.5; MgSO4.7H2SO4, 2.0; NH4Cl, 1.0; Ascorbic acid, 0.1; sodium lactate, 3.5; Thioglycolic acid, 0.1; yeast extract, 1.0) and the dilution-to-extinction technique described by Postgate (1984), the bacteria were isolated in deep agar.

Motility detection, Gram’s staining, and endospore staining were used to phenotypically describe the bacterial pure culture. Following that, 16S rRNA gene sequencing was used to identify the bacterial strain at the molecular level. SRB growth was revived for this purpose in Postgate B medium. Following Hussain et al. (2014), total genomic DNA from freshly grown bacterial cells was isolated and amplified using universal primers [27f (5′-AGAGTTTGATCMTGGCTCAG-3′) and 1492r (5′-GGTTACCTTGTTACGACTT-3′)]. After that, the amplified gene was sequenced by Macrogen, Korea. Together with reference sequences retrieved from the NCBI GenBank database, a phylogenetic tree using neighbor-joining methods was constructed for the bacteria isolated for the current investigation.

After molecular identification, the bacterial growth was optimized for temperature, pH and inoculum size. Optimum growth was found at 30 oC in the medium having pH 7 and inoculated with 1.7 × 106 C.F.U. mL−1 of the medium.

Preparation of carbon steel coupons

Carbon steel coupons (CSCs) were the small pieces of CS that were used for studying corrosion. CSCs (1 × 1 × 0.1 cm) for the current study were obtained from the Department of Metallurgy and Materials Engineering, University of the Punjab, Lahore, Pakistan. First of all, the coupons were thoroughly cleaned with analytical grade acetone (Cat. No. 10014, Merck, Germany) following Bano (2008) to remove any dirt, oil, or other contaminants that could affect the corrosion rate (CR). The surface of the coupons was then prepared to ensure that the CR is uniform. This was done by polishing the surface of the coupons with polishing paper (Cat. No. WD240, Caswell, Taiwan). After polishing, the coupons were cleaned with a lint-free paper towel, washed with ethanol (Cat. No. 100983, Merck, Germany) and dried completely at 80 oC for 10 min in an electric oven. The weight of the coupons was then measured before and after the corrosion test to determine the amount of metal that has been lost due to corrosion following Zuo and Wood (2004).

Experimental design

The corrosive impact of SRB was studied in water with and without the provision of nutrients of Postgate B medium. The experiments were conducted in serum bottles of 20-mL capacity. For the purpose, half of the total serum bottles were filled upto the brim with Postgate B medium, while remaining half of the bottles were filled with distilled water. At start of the experimental trial, pH was adjusted between 7.0 and 7.5. After pH adjustment, one cleaned coupon was inserted in each bottle, all the bottles containing distilled water and postgate B medium were properly capped with butyl rubber stoppers, sealed with aluminum crimp seals, sterilized by autoclaving and cooled at room temperature. After this, half of these vials were inoculated with freshly cultivated SRB culture @ 5 % (v/v) harbouring around 1.7×106 C.F.U. mL−1, while remaining vials were kept uninoculated as control group. All the vials were then incubated at 30 oC for 60 days. At end of the testing period, coupons were extracted aseptically from the bottles. Following Angeles-Chavez et al. (2001), Clark’s solution (Sb2O3, 20 g; SnCl2, 50 g; HCl, 774 mL) was used to clean the coupons from corrosion products and other impurities. Clean coupons were thoroughly dried after rinsing in distilled water and weighed to determine weight reduction. After that, the rate of corrosion (µg dm−2 d−1) was calculated using the following formula as reported by Majumdar et al. (1999).

Where, C represents CR (µg dm−2 d−1), while W1 and W2 are initial and final weights of coupons, respectively. Surface area of the coupons and the experimental period (days) are represented by A and T, respectively.

After determining CR, SRB growth was also estimated in Postgate B medium following Postgate (1984).

Scanning electron microscopic analysis of the corroded coupon

In order to study surface morphology and type of corrosion, the corroded coupon was analyzed by SEM. To prepare a metal coupon for SEM analysis, the coupon was cleaned for the removal of any dirt or other contaminants. The coupon was then coated with a thin layer of palladium to make it electron-conductive. Once the coupon was prepared, it was placed in the SEM and scanned to generate an image of the surface. The SEM facility at Institute of Chemical Engineering and Technology, University of the Punjab, Lahore, was availed for the said purpose.

RESULTS AND DISCUSSION

The current study was conducted to assess the corrosive action of SRB on CSCs in aqueous media with and without the addition of nutrients of Postgate B medium. For the purpose, SRB strain was isolated from a highly polluted wastewater stream flowing in Lahore, Punjab, Pakistan. The bacterial isolate was motile, Gram-negative, non-spore former and identified as Desulfovibrio vulgaris after 18 S rRNA gene sequencing. A phylogenetic tree was constructed to show its neighbour-joining microbial species as shown in Figure 1. It is well known that SRB are commonly found in aquatic environments, including wastewaters and can contribute to the production of hydrogen sulfide gas, which has a strong, unpleasant odour (Karunakaran et al., 2016; Hamoda and Alshalahi, 2021; Jantharadej et al., 2021; Aalto et al., 2022). In polluted wastewaters, the presence of SRB can be an indicator of the presence of organic matter and can be used as a measure of the effectiveness of wastewater treatment processes. However, the presence of SRB in wastewaters can also have negative impacts. For example, the production of hydrogen sulfide can lead to corrosion of infrastructure, such as pipes and treatment facilities (Wiener and Salas, 2005; Foorginezhad et al., 2021; Wang et al., 2023).

 

After an incubation period of 60 days, it was found that CR was significantly higher for the coupons that were dipped in inoculated media. For these coupons, the average CR appeared as 210±17 µg dm−2 d−1. However, CR was significantly lower (210±17 µg dm−2 d−1) for the coupons dipped in inoculated water only (Fig. 2A). In un-inoculated vials having nutrients, CR appeared higher (97 ± 16 µg dm−2 d−1) as compared to those without nutrients (48 ± 21 µg dm−2 d−1) as shown in Figure 2B. In a previously reported study, CR was higher (249 ± 2 µg dm−2 d−1) than that of current study (Sattar, 2023). In that study, incubation period, pH and temperature were similar to those of the current study. However, coupon composition and bacterial species were different. In that study, mild steel coupons were used instead of CSCs. In mild steel coupons, the lower percentage of carbon makes microbial action more stronger (Mehanna et al., 2009; Chaerun et al., 2023; Golafshani et al., 2023). In addition, the rate of corrosion is expected to vary both with the type and ecology of SRB (Beech and Sunner, 2007; Madirisha et al., 2022). There are several factors that can contribute to the corrosion of metal structures by SRB. These include the presence

 

of organic matter that can serve as a food source for the bacteria, the pH and temperature of the environment, and the availability of nutrients such as sulfur and hydrogen (AlAbbas et al., 2013; Enning and Garrelfs, 2014; Lv et al., 2021). CR can also be influenced by the type of metal, the presence of other microorganisms, and the presence of other corrosive agents (Javaherdashti, 2011; Jia et al., 2019; Zhao et al., 2022).

SRB growth appeared higher in nutrient added vials. At termination of the experiments, total viable counts for SRB were 2.7 ± 0.3 × 109 CFU mL−1 in vials supplemented with nutrients. However, no bacterial growth was observed in nutrient-deprived vials. No change in pH was observed in all vials except those which were inoculated under optimum provision of nutrients. In these vials, pH changed from neutral to slightly basic in range (7.0 ± 0.5 to 6.0 ± 0.5). It has been reported that during SRB growth, medium pH changes towards alkalinity (Hussain and Qazi, 2016; Hussain et al., 2016).

At termination of the experiment, of all the coupons, a highly corroded coupon was analyzed through SEM. SEM analysis of the coupon which was recovered from a vial supplemented with nutrients revealed severe pitting corrosion of CS (Fig. 3). Pitting corrosion of CS by SRB

 

can be a serious problem in certain environments, particularly in the oil and gas industry (Wang and Melchers, 2017; Kannan et al., 2018; Qi et al., 2023; Wan et al., 2023; Zulkafli et al., 2023). SRB consume sulfates as a source of energy and produce hydrogen sulfide as a byproduct. This hydrogen sulfide can react with the iron in CS to form iron sulfide, which can lead to pitting corrosion (Sherar et al., 2011; AlAbbas et al., 2013; Giorgi-Pérez et al., 2021). Pitting corrosion is a type of localized corrosion that occurs when small, deep pits form on the surface of the metal. These pits can weaken the metal and make it more susceptible to failure (Frankel and Sridhar, 2008; Alamri, 2020; Messinese et al., 2022).

Conclusion

In the current study, severe pitting corrosion of carbon steel was caused by SRB and observed in nutrient-supplemented vials. The pits on the coupon surface were studied by SEM. In order to prevent pitting corrosion caused by SRB, it is important to control the presence of these bacteria in the environment. This can be achieved through the use of biocides, which are chemicals that are toxic to bacteria. It is also important to maintain the pH of the environment at a level that is not conducive to the growth of SRB. In addition, using corrosion-resistant materials or coatings can help to protect CS from pitting corrosion caused by SRB.

Acknowledgement

We acknowledge the support of Department of Metallurgy and Materials Engineering, University of the Punjab, Lahore, Pakistan, for providing mild steel coupons.

Funding

The study received no funding.

IRB approval

The current research work was approved by Board of Studies of Department of Wildlife and Ecology, University of Veterinary and Animal Sciences, Lahore.

Ethical statement

The current research work didn’t involve any animal model and thus ethical statement was not needed.

Statement of conflict of interest

The authors have declared no conflict of interest.

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