Adapting USEPA protocol for the detection and enumeration of Giardia and Cryptosporidium in an agricultural setting

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An ecological water treatment system that utilizes a series of plants and water tanks is being tested at the Ohio State University Waterman Dairy as an alternate method for agriculture waste management. Giardia and Cryptosporidium are two genuera of parasites that can be found in most surface water supplies due to runoff from agricultural facilities, sewage treatment plants, or wildlife areas such as forests. As a result, the level of these two parasites in the water supply is a concern in regards to public health. The purpose of this study, initially, was to evaluate viability of the United States Environmental Protection Agency (USEPA) Method 1623 commonly used for detection of Giardia and Cryptosporidium in surface water with an untraditional sample source—agricultural waste. An additional purpose was to evaluate the level of Giardia and Cryptosporidium cysts and oocysts entering the ecological treatment system and the level exiting the system, thus determining the efficiency of the system for reduction of parasitic load. Method limitations were established by enumerating stock organisms, running spiked distilled water controls, and finally using experimental samples from the agricultural waste entering the water treatment facility. The USEPA Method 1623 was used to evaluate the level of Giardia and Cryptosporidium in the samples. Eluted samples were concentrated, stained, and observed via immufluorescence. Modifications to the published protocol were made to account for the difference of this project’s sample type (agricultural waste verses surface water) and to account for not using immunomagnetic separation (IMS) for the concentration of the sample. The major finding from this project is the inability of a capsule filtration unit recommended in the the USEPA Method 1623 to evaluate the parasitic load in agricultural waste samples. The particle size and load, despite sieving of the sample, overtaxes the filter used in the sample processing system. This makes elution and recovery of the cysts and oocysts near impossible. An accidental finding occurred in regards to cysts and oocyst integrity and the ability to detect their presence efficiently with immunofluorescent staining. Alternate methods will need to be investigated in order to properly detect and quantify Giardia cyst and Cryptosporidium oocyst levels for the ecological water treatment system processing agricultural waste as this capsule filtration testing system is ineffective because of particle size and load in the waste. In addition, further research into cyst and oocyst integrity verses the ability to detect with immunofluorescent staining will need to be pursued.



waterborne parasites, ecological water treatment system