Detection of Cryptosporidium parvum in secondary effluents using a most probable number-polymerase chain reaction assay.
Polymerase chain reaction (PCR) was used to detect Cryptosporidium parvum oocysts in secondary effluent samples collected from activated-sludge facilities. Serial dilutions of the purified nucleic acid extracts from the samples were made and PCR was conducted to estimate the C. parvum oocyst concentration via a Poisson distribution-based most probable number (MPN). The degree of oocysts associated with wastewater particles was also evaluated. The sensitivity of the MPN-PCR assay was 20 oocysts/PCR unit. The detection limit of the concentration, extraction, and purification protocols in phosphate buffer saline spiked with a known concentration of oocysts ranged from 1.1 to 4.6 oocysts/L; the detection limit for the wastewater samples ranged from 11 to 4200 oocysts/L depending on the extent of inhibition in each sample. The recovery efficiency of the oocysts ranged from 48 to 59% in most samples. Oocysts were found in two out of seven samples with concentrations of 203 and 308 oocysts/L, as estimated by the MPN-PCR method. The oocysts were found only in the filtrate of the grab samples; particle-associated oocysts were not detected. Association of spiked C. parvum oocysts with particles in secondary effluent drawn from wastewater plants with varying operating conditions indicated a weak correlation between the degree of association and the mean cell residence time of the system.
Tsuchihashi
Ryujiro
R
Department of Civil and Environmental Engineering, University of California, Davis, USA.
Loge
Frank J
FJ
Darby
Jeannie L
JL
D000818:Animals; D016785:Cryptosporidium parvum / Q000302:isolation & purification*; D004755:Enterobacteriaceae / Q000302:isolation & purification; D033661:Oocysts / Q000302:isolation & purification; D016012:Poisson Distribution; D016133:Polymerase Chain Reaction / Q000379:methods*; D012722:Sewage / Q000032:analysis* / Q000469:parasitology
D012722:Sewage