Molecular characterization of intestinal protozoa in an oil wastewater treatment facility; risk for contamination of downstream farmlands

BACKGROUND: Oil refinery (OR) facilities consume vast volumes of water during their processes, generating a large amount of wastewater (WW). The oil refinery wastewater (ORWW) contains toxic materials, and few microorganisms are believed to remain active in this environment. METHODS: Water samples were collected from an ORWW facility for nine months. After recording physical-chemical features, filtration and sedimentation were employed. DNA extraction was performed for all samples and the presence of targeted parasites was evaluated using real-time PCR. Genotype/subtype of positive samples was characterized using sequencing. RESULTS: Real-time PCR showed that from 76 ORWW samples, targeted protozoa were identified in 26 (34.2%) samples. Accordingly, Blastocystis sp., Enterocytozoon bieneusi, and Giardia duodenalis were detected among 9.21% (7/76), 25.0% (19/76), and 2.63% (2/76) of samples, respectively. Enterocytozoon bieneusi was the most common detected species in samples followed by Blastocystis sp. and G. duodenalis. There was a statistically association between the presence of Blastocystis sp. and the type of wastewater treatment plants (WWTPs). Enterocytozoon bieneusi genotypes BEB6 (3/5; 60%), C (1/5; 20%), and an unknown genotype were detected among samples. Also, Blastocystis sp., ST1 (2/4; 50%), ST3 (1/4; 25%), uncultured uncharacterized stramenopile (1/4; 25%) and ST2 (1; 100%), were characterized in samples. Giardia duodenalis was the assemblage AII (1/2; 50%). Phylogenetic analysis showed that E. bieneusi genotypes were grouped within the groups 1 and 2, human-infecting genotypes. In addition, Blastocystis sp., subtypes were clustered within subtypes isolated from humans. CONCLUSION: This is the first study investigating intestinal protozoa in a super harsh condition. Our findings signify the concern due to the distribution of potentially pathogenic parasites to downstream farmlands through industrial treated wastewaters which are used for irrigation.