Phenotypic and genotypic characterization of Salmonella species isolated from treated wastewater effluents and receiving rivers in Durban.

Abstract

Salmonella and Shigella spp. are major pathogens of humans and they cause diseases ranging from mild food poisoning to chronic diarrhoea, especially in children under the age of 5. They are commonly found in the gastrointestinal tract of animals and humans and contaminate water surfaces through faecal pollution. Discharge of inadequately treated wastewater has been known to be conduits of these pathogens to surface waters. Emergence of antibiotic resistant bacteria is a public health concern worldwide especially in developing countries where disease burden is high. This study investigated the efficiency of two Wastewater Treatment Plants (WWTPs) in Durban for wastewater treatment, and assessed the impact of treated effluent discharge on the receiving surface water. The genotypic characteristics and antibiogram profile of Salmonella spp. recovered from the treated effluent samples of the WWTPs and the receiving river was also determined. Water samples were collected from the WWTPs over a 12 month period and analyzed for physico-chemical parameters including temperature, pH, turbidity, BOD and COD using standard methods; while presumptive Salmonella and Shigella spp. were enumerated on Salmonella-Shigella and xylose-lysine-desoxycholate agar, respectively, via membrane filtration technique. Isolation of Salmonella spp. was done by enrichment of samples in Rappaort Vassiliadis soy broth followed by spread plating on Salmonella chromogenic agar and aerobic incubation at 37°C for 18 to 24 h. Presumptive isolates were biochemically characterized and confirmed via PCR amplification of the invA gene. Isolates were tested against 20 selected antibiotics to determine their antibiotic resistance profile. Presence of virulence markers; spiC, misL, orfL and pipD genes were also determined using PCR. Unacceptably high levels of turbidity (5.52-37.58 NTU), BOD (2.19-9.1 mg/l) and COD (67.67-294 mg/l) were observed in the water samples, while temperature (14°C-25°C) and pH (6.72-7.3) fell within the recommended maximum of 25°C and 10 7.5, respectively, for treated wastewater effluent. Significant positive correlation (p < 0.05) was observed between pH and BOD, temperature and COD, and between turbidity and presumptive Salmonella count. Presumptive Salmonella and Shigella spp. were prevalent at all sampling points, with population ranging from 8.5×102 to 1.59×105 CFU/ml and 0.1×102 to 7.5×103 CFU/ml, respectively. The isolates were highly susceptible to β-lactams, Chloramphenicol, Tetracycline, Quinolones and Trimethoprim-Sulfamethoxazole (99% to 100%). Complete antibiotics resistance was observed against Sulfamethoxazole (100%), Nalidixic acid (27%) and Streptomycin (14%). Intermediate resistance was observed against Streptomycin (74%), Nalidixic acid (44%) and Fosfomycin (8.5%). Of the 200 isolates tested, 93% harboured the spiC gene, 84% harbored the misL gene, while 87.5% and 87 % of the isolates harboured the orfL and pipD gene, respectively. Results from this study indicate the inefficiency of the WWTPs investigated to totally eradicate Salmonella spp. from the final effluent and discharge of such effluent. Discharge of these effluent to surface water resources could pose health threat to the end-users of the surface water for daily domestic and recreational activities . Thus, appropriate intervention by the regulatory agencies is required to ensure compliance of WWTPs to the stipulated guidelines for safe disposal of treated effluent.