title: Polymerase chain reaction and nested-PCR approaches for detecting Cryptosporidium in water catchments of water treatment plants in Curitiba, State of Paraná, Brazil.
abstract: BACKGROUND
  Cryptosporidium is an important protozoan cause of waterborne disease worldwide of concern to public health authorities. To prevent outbreaks of cryptosporidiosis, the monitoring of this parasite in drinking water is necessary. In the present work, the polymerase chain reaction (PCR) and nested-PCR techniques were used to detect Cryptosporidium in raw water from catchment points of four water treatment plants (WTP) in Curitiba, Paraná, Brazil.
  
  
  METHODS
  First, DNA extraction techniques were tested in samples containing decreasing amount of oocysts in reagent water, and PCR and nested-PCR with specific primers for 18SSU rDNA of Cryptosporidium were conducted to determine their sensitivity. In reagent water, a commercial extraction kit provided the best analytical sensitivity, and PCR and nested-PCR allowed the detection of five and two oocysts, respectively, with the primers XIAOR/XIAOF and XIAO1F/XIAO2R.
  
  
  RESULTS
  In the spiking experiments, only the PCR with the primers AWA995F/AWA1206R was successful at detecting concentrations of 0.1 oocysts/mL. Two catchments samples of raw water and/or water sludge from four WTPs were contaminated with Cryptosporidium.
  
  
  CONCLUSIONS
  The application of the techniques to monitor Cryptosporidium in water and detect contamination in water catchments of WTPs in Curitiba are discussed in the present work.
authors.affiliation: Programa de Pós-Graduação em Processos Biotecnológicos e Biotecnologia, Centro Politécnico, Universidade Federal do Paraná, Curitiba, PR, Brazil. sil_osaki@ufpr.br
authors.forename: Silvia Cristina
authors.initials: SC
authors.lastname: Osaki
authors.forename: Vanete Thomaz
authors.initials: VT
authors.lastname: Soccol
authors.forename: Adriana Oliveira
authors.initials: AO
authors.lastname: Costa
authors.forename: Márcia Benedita
authors.initials: MB
authors.lastname: Oliveira-Silva
authors.forename: Juliana Tracz
authors.initials: JT
authors.lastname: Pereira
authors.forename: Antônio Eduardo
authors.initials: AE
authors.lastname: Procópio
mesh_terms: D001938:Brazil; D003458:Cryptosporidium / Q000235:genetics / Q000302:isolation & purification*; D016054:DNA, Protozoan / Q000032:analysis; D004275:DNA, Ribosomal / Q000032:analysis*; D005618:Fresh Water / Q000469:parasitology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012722:Sewage / Q000469:parasitology; D018508:Water Purification; D014881:Water Supply / Q000032:analysis
chemical_list: D016054:DNA, Protozoan; D004275:DNA, Ribosomal; D012722:Sewage