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Detecting Cryptosporidium parvum and Giardia lamblia by coagulation concentration and real-time PCR quantification

Abstract

Rapid and sensitive pathogen detection methods are essential for the effective functioning of the water treatment industry, yet for many pathogens, effective detection and removal methods are still lacking. Cryptosporidium parvum oocysts and Giardia lamblia cysts are two of the most common waterborne pathogens currently infecting the water supply. In this study, a new method was developed for the detection of Cryptosporidium parvum oocysts and Giardia lamblia cysts. The method includes multi-steps as coagulation concentration of (oo)cysts in water, the dissolution of the resulting flocs into a small volume using acid, filtration of the (oo)cysts solution, and DNA extraction, purification, and examination using real-time PCR. The method was tested using spiked tap water and reservoir water as references, and the mean recovery ranged from 19.6% to 97.6% for oocysts, and from 51.4% to 98.7% for cysts. The method is economical and convenient, and is especially suitable for relatively high turbidity surface water.

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References

  1. Marshall M M, Naumovitz D, Ortega Y, Sterling C R. Waterborne protozoan pathogens. Clinical Microbiology Reviews, 1997, 10(1): 67–85

    CAS  Google Scholar 

  2. Hu J Y, Feng Y Y, Ong S L, Ng W J, Song L F, Tan X L, Chu X N. Improvement of recoveries for the determination of protozoa Cryptosporidium and Giardia in water using method 1623. Journal of Microbiological Methods, 2004, 58(3): 321–325

    Article  CAS  Google Scholar 

  3. DiGiorgio C L, Gonzalez D A, Huitt C C. Cryptosporidium and giardia recoveries in natural waters by using environmental protection agency method 1623. Applied and Environmental Microbiology, 2002, 68(12): 5952–5955

    Article  CAS  Google Scholar 

  4. Feng Y Y, Ong S L, Hu J Y, Song L F, Tan X L, Ng W J. Effect of particles on the recovery of Cryptosporidium oocysts from source water samples of various turbidities. Applied and Environmental Microbiology, 2003, 69(4): 1898–1903

    Article  CAS  Google Scholar 

  5. States S, Tomko R J. Enhanced coagulation and removal of Cryptosporidium. Journal- American Water Works Association, 2002, 94(11): 67–77

    CAS  Google Scholar 

  6. Walker N J. A technique whose time has come. Science, 2002, 296(5567): 557–559

    Article  CAS  Google Scholar 

  7. Verweij J J, Blangé R A, Templeton K, Schinkel J, Brienen E A T, van Rooyen M A A, van Lieshout L, Polderman A M. Simultaneous detection of Entamoeba histolytica, Giardia lamblia, and Cryptosporidium parvum in fecal samples by using multiplex real-time PCR. Journal of Clinical Microbiology, 2004, 42(3): 1220–1223

    Article  CAS  Google Scholar 

  8. Yu X, van Dyke M I, Portt A, Huck P M. Development of a direct DNA extraction protocol for real-time PCR detection of Giardia lamblia from surface water. Ecotoxicology, 2009, 18(6): 661–668

    Article  CAS  Google Scholar 

  9. Guy R A, Payment P, Krull U J, Horgen P A. Real-time PCR for quantification of Giardia and Cryptosporidium in environmental water samples and sewage. Applied and Environmental Microbiology, 2003, 69(9): 5178–5185

    Article  CAS  Google Scholar 

  10. Tanriverdi S, Tanyeli A, Başlamişli F, Köksal F, Kilinç Y, Feng X C, Batzer G, Tzipori S, Widmer G. Detection and genotyping of oocysts of Cryptosporidium parvum by real-time PCR and melting curve analysis. Journal of Clinical Microbiology, 2002, 40(9): 3237–3244

    Article  CAS  Google Scholar 

  11. Wang Z, Vora G J, Stenger D A. Detection and genotyping of Entamoeba histolytica, Entamoeba dispar, Giardia lamblia, and Cryptosporidium parvum by oligonucleotide microarray. Journal of Clinical Microbiology, 2004, 42(7): 3262–3271

    Article  CAS  Google Scholar 

  12. Adam R D. Biology of Giardia lamblia. Clinical Microbiology Reviews, 2001, 14(3): 447–475

    Article  CAS  Google Scholar 

  13. Harris J R, Petry F. Cryptosporidium parvum: structural components of the oocyst wall. The Journal of Parasitology, 1999, 85(5): 839–849

    Article  CAS  Google Scholar 

  14. Simmons O D, Sobsey M D, Heaney C D, Schaefer F W, Francy D S. Concentration and detection of Cryptosporidium oocysts in surface water samples by method 1622 using ultrafiltration and capsule filtration. Applied and Environmental Microbiology, 2001, 67(3): 1123–1127

    Article  Google Scholar 

  15. Brush C F, Walter M F, Anguish L J, Ghiorse W C. Influence of pretreatment and experimental conditions on electrophoretic mobility and hydrophobicity of Cryptosporidium parvum oocysts. Applied and Environmental Microbiology, 1998, 64(11): 4439–4445

    CAS  Google Scholar 

  16. Vesey G, Slade J S, Byrne M, Shepherd K, Fricker C R. A new method for the concentration of Cryptosporidium oocysts from water. Journal of Applied Bacteriology, 1993, 75(1): 82–86

    Article  CAS  Google Scholar 

  17. Zhang H N, Gu L, Yu X, Wei B. Removal of Cryptosporidium and Giardia microspherers from surface water by coagulation. Chinese Journal of Environmental Engineering, 2011, 5(1): 65–69 (in Chinese)

    CAS  Google Scholar 

  18. Cacciò SM, de Giacomo M, Aulicino F A, Pozio E. Giardia cysts in wastewater treatment plants in Italy. Applied and Environmental Microbiology, 2003, 69(6): 3393–3398

    Article  Google Scholar 

  19. Zong Z S, Hu H Y, Lu Y X, Zhang J S. Investigation on current water pollution by Giardia and Cryptosporidium in a city. China Water and Wastewater, 2005, 21(5): 44–46 (in Chinese)

    Google Scholar 

  20. Razzolini M T P, da Silva Santos T F, Bastos V K. Detection of Giardia and Cryptosporidium cysts/oocysts in watersheds and drinking water sources in Brazil urban areas. Journal of Water and Health, 2010, 8(2): 399–404

    Article  CAS  Google Scholar 

  21. Nam S, Lee G. A new duplex reverse transcription PCR for simultaneous detection of viable Cryptosporidium parvum oocysts and Giardia duodenalis cysts. Biomedical and Environmental Sciences, 2010, 23(2): 146–150

    Article  CAS  Google Scholar 

  22. Baque R H, Gilliam A O, Robles L D, Jakubowski W, Slifko T R. A real-time RT-PCR method to detect viable Giardia lamblia cysts in environmental waters. Water Research, 2011, 45(10): 3175–3184

    Article  CAS  Google Scholar 

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Correspondence to Qipei Jiang or Xin Yu.

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Zhang, H., Zhang, X., Zhang, S. et al. Detecting Cryptosporidium parvum and Giardia lamblia by coagulation concentration and real-time PCR quantification. Front. Environ. Sci. Eng. 7, 49–54 (2013). https://doi.org/10.1007/s11783-012-0455-2

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  • DOI: https://doi.org/10.1007/s11783-012-0455-2

Keywords

  • Cryptosporidium parvum
  • detection method
  • Giardia lamblia
  • surface water