Occurrence and genetic diversity of Cryptosporidium and Giardia in urban wastewater treatment plants in north-eastern Spain.

This study was designed to investigate the presence and removal efficiency of Cryptosporidium and Giardia in wastewater treatment plants at the 20 most populated towns in Aragón (north-eastern Spain). Samples of influent and effluent wastewater and dewatered sewage sludge were collected seasonally from 23 plants and processed according to USEPA Method 1623. All samples from raw and treated wastewater tested positive for Giardia, at an average concentration of 3247±2039cysts/l and 50±28cysts/l, respectively. Cryptosporidium was identified in most samples from both raw (85/92) and treated (78/92) wastewaters in a concentration significantly lower than Giardia, at both influent (96±105oocysts/l) and effluent samples (31±70oocysts/l) (P<0.001). The (oo)cyst counts peaked in summer in most plants. The removal efficiency was higher for Giardia (1.06-log to 2.34-log) than Cryptosporidium (0.35-log to 1.8-log). Overall, high removal efficiency values were found for Giardia after secondary treatment based on activated sludge, while tertiary treatment (microfiltration, chlorination and/or ultraviolet irradiation) was needed to achieve the greatest removal or inactivation of Cryptosporidium. Most samples of treated sludge were positive for Giardia (92/92) and Cryptosporidium (45/92), at an average concentration of 20-593cysts/g and 2-44oocyst/g, respectively. The molecular characterization of Cryptosporidium oocysts and Giardia cysts were attempted at the SSU rRNA/GP60 and bg/tpi loci, respectively. G. duodenalis sub-assemblage AII was identified in all plants, with a large proportion of samples (15/47) harboring mixed assemblages (AII+B). Nine Cryptosporidium species and six subtypes were identified, with C. parvum IIaA15G2R1 being the most prevalent. The presence of significant numbers of (oo)cysts in samples of final effluents and treated sludge reveals the limited efficacy of conventional treatments in removing (oo)cysts and highlights the potential environmental impact and public health risks associated with disposal and reclamation of wastewater.