Abundance and diversity of oil-degrading bacteria in produced water wetland treatment system, Rima-Oman
Daniel Blackburn, Fatema Khalifah Al-Mandhariyah , Mushtaque Ahmed
كلية العلوم الزراعية والبحرية-جامعة السلطان قابوس · عمان
Produced water is excess water from oil production, which contains hydrocarbon residuesand other pollutants. One method that can be used to treat wastewater is the constructionof artificial wetlands. The Rima Water Treatment Project (Oman) is a constructed wetlandthat was recently implemented to biologically treat produced water. In this study, theabundance and diversity of oil-degrading bacteria across the wetland system wereevaluated and compared to water quality, as it progressed through four subsequentterraces in three parallel tracks of the wetland. In addition, oil-degrading bacteria havebeen isolated and characterized based on their ability to withstand different environmentalconditions. The chemical properties of the sample were measured, including the oilcontent in water, pH, EC, total elemental composition, and hydrocarbon compounds,using gas chromatography-mass spectrometry (GC-MS). Culturable bacteria wereenumerated using nutrient agar, Bushnell Haas mineral salt (BHMS) agar, and mineralsalt medium (MSM) as minimal media. Morphologically different bacteria were isolatedand tested for their ability to degrade crude oil using a MicroResp™ system. The strainswith the highest oil degradation rates were further tested for their ability to withstand highionic strengths, a broad range of pH, and their response to NPK fertilizers using growthcurve analysis. The strains were then tested for their ability to form biofilms. Microbialdiversity was assessed by 16S rRNA V4 gene sequencing in the Rima wetland system.Our findings showed that the oil-in-water content decreased to less than 1 ppm at the endof the wetland. In contrast, EC, pH, and minerals such as Na progressively increased asthey flowed through the four stages of the wetland. Most hydrocarbon compoundsdecreased as they progressed through the system. Moreover, the bacterial enumerationprogressively decreased, being highest in the first terrace. Interestingly, there was also asignificant difference in bacterial enumeration between the three parallel tracks. Oildegrading bacteria were more abundant in the first two terraces of the wetland and in thesecond and third tracks. The isolated bacteria were all able to use crude oil as a carbonsource. Shewanella sp., Aeromonas sp., Aeromonas hydrophila, Ochrobactrum sp., andEnterobacter sp. had the highest respiration rates. However, Ochrobactrum sp. is a strainthat is tolerant to different pH ranges, high salinity, responsiveness to NPK, biofilmformation, and order of oil degradation. The 16S rRNA diversity showed higher bacterialdiversity in the last stages of the wetland system, with an overall very low number ofarchaea. Even though the differences in Shannon and Evenness diversity indices betweenthe terraces were small, the number of bacterial classes and community structure weresignificantly affected by both terraces and tracks. The most abundant class observed inthe initial stages of the wetland was Alphaproteobacteria, where most of the oildegradation took place. Bacterial community 2 (clustered by correlation) was responsiblefor the oil degradation and was dominated by Rhodobacter, Loktanella, and Thiofabagenera. Cyanobacteria negatively correlated with hydrocarbon degradation. Furtherstudies are needed to investigate the effects of autotrophic bacteria on the removal ofdifferent hydrocarbon compounds. To increase the efficiency of wetland systems, furthercross-inoculation and bioaugmentation trials can be conducted using the identified oil degrading bacterial isolates.