Author of Response: Chip Simmons, Ph.D., Assistant Research Professor, University of North Carolina-Chapel Hill School of Public Health, osimmons@email.unc.edu
There currently is a reliable and robust EPA-approved method for detection and quantification of both Cryptosporidium and Giardia in water (including both environmental surface and groundwaters, as well as finished drinking waters). This is method 1623 and involves initial filtration, elution of (oo)cysts from filters, concentration by centrifugation, separation by immuno-magnetic beads (IMS), staining with a fluorescent antibody stain, counterstaining with DAPI (to observe internal structures within the organisms), and final detection and quantification by microscopic evaluation.
This method uses a combination of monoclonal antibodies that attach to surface epitopes of the (oo)cysts for both IMS and fluorescent staining prior to microscopic evaluation. The method has been tested in a variety of waters in round-robin studies by the EPA, as well as in independent research. The method examines 10-L volumes of water and has a theoretical detection limit of 1 (oo)cyst. Subsequent research has demonstrated the utility of a commercially available internal positive control for measuring diminished recovery efficiency of the method by difficult-to-characterize matrix effects associated with some environmental waters.
Additional reading:
Francy, D.S., O.D. Simmons, III, M.W. Ware, E.J. Grainger, M.D. Sobsey, and F.W. Schaefer, III. 2004. Effects of spiking procedures and water quality on recovery of Cryptosporidium in stream water using USEPA Method 1623. Applied and Environmental Microbiology 70:4118-4128.
Simmons, III, O.D., C.D. Heaney, D.S. Francy, R.A. Nally, F.W. Schaefer, III, and M.D. Sobsey. 2001. Detection of Cryptosporidium Oocysts in Stream Water Samples Using U.S. Environmental Protection Agency Method 1622. Journal of the American Water Works Association 93:78-87.
Simmons, III, O.D., C.D. Heaney, D.S. Francy, R.A. Nally, F.W. Schaefer, III, and M.D. Sobsey. 2001. Concentration and Detection of Cryptosporidium Oocysts in Surface Water Samples by Method 1622 Using Ultrafiltration and Capsule Filtration. Applied and Environmental Microbiology 67:1123-1127.
USEPA. 2005. Method 1623: Cryptosporidium and Giardia in Water by Filtration/IMS/FA. Publication EPA-815-R-05-002. Office of Water, Washington, D.C.
