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Analysis of viability and infectivity of Cryptosporidium oocysts and Giardia cysts recovered from superficial and residual water by cell culture techniques integrated to Real Time Quantitative PCR (ICC-RTqPCR)

Grant number: 17/10175-3
Support type:Scholarships in Brazil - Post-Doctorate
Effective date (Start): January 01, 2018
Effective date (End): December 31, 2020
Field of knowledge:Engineering - Sanitary Engineering
Principal Investigator:Maria Ines Zanoli Sato
Grantee:Ronalda Silva de Araújo
Home Institution: Companhia Ambiental do Estado de São Paulo (CETESB). Secretaria do Meio Ambiente (São Paulo - Estado). São Paulo , SP, Brazil

Abstract

Access to potable water free of pathogenic parasites, especially Cryptosporidium and Giardia, continues to be a challenge for drinking water systems. Thus, the monitoring and identification of protozoa released in the environment with potential to trigger diseases, that is, viable, is essential for the control and decision-making of water management and agencies of control. The main goal of this project is to evaluate and validate an in vitro assay intended to quantify viable and infectious Cryptosporidium oocysts and Giardia cysts in superficial water and treated sewage intended for reuse, using reverse transcriptase/quantitative PCR techniques (RTqPCR) integrated to cell culture (ICC). Samples will be collected bimonthly for a year, and concentrated by 1623.1 method for superficial water and 1693 method for treated effluents, both recommended by the United States Environmental Protection Agency (USEPA). Nucleic acid amplification techniques and quantification of mRNA transcripts integrated to cell culture methods (ICC-RTqPCR) will be employed to assess the viability of Cryptosporidium hominis / parvum oocysts and Giardia intestinalis cysts isolated from the samples. The application of the ICC-RTqPCR technique will be the main scientific challenge of the study due to the lack of reliable and rapid quantitative testing methods to analyze the viability of oocysts and cysts in environmental samples. It is expected, however, to implement the protocols validated by this project, in order to overcome the difficulties faced by risk assessment studies associated with exposure to these protozoa and the control of cryptosporidiosis and giardiasis in our country,providing decision-makers subsides regarding concerns about these protozoa in catchment and reuse waters.