AWWA ACE58303

Waterborne transmission of the protozoan parasite Cryptosporidium parvum remains a significant source of disease with severe consequences for immunocompromised people. Over the past decade much progress has been made in addressing the issue of Cryptosporidium in water. Both the Interim Enhanced Surface Water Treatment Rule (IESWTR) and the Long Term 1 Enhanced Surface Water Treatment Rule (LT1ESWTR) were promulgated by the US Environmental Protection Agency (USEPA) to reduce the risk of cryptosporidiosis. The pending Long Term 2 Enhanced Surface Water Treatment Rule (LT2ESWTR) will require additional treatment for utilities with poor source water quality. In addition, many utilities, including all of the utility subsidiaries of American Water, have joined a voluntary Partnership for Safe Drinking Water to further tighten operating standards to improve the treatment of Cryptosporidium oocysts. The immunofluorescent antibody (IFA) test has been used to detect Cryptosporidium oocysts in filtered drinking water but the assay has a number of limitations. The IFA assay cannot determine the public health significance of oocysts detected in drinking water. As a result, American researchers developed a patented test for detecting live, infectious Cryptosporidium oocysts in water. The method called the "cell culture, polymerase chain reaction" (CC-PCR) test is accurate and precise because it measures the DNA of Cryptosporidium that are able to grow in human intestinal cells after the isolation of infectious oocysts from water samples. Application of the CC-PCR assay to filtered drinking water samples can help determine the central question facing water utilities: do current water treatment techniques adequately control the risks of Cryptosporidium in water? The current study was initiated to answer this question and provide a strategy for control of Cryptosporidium in water. Monthly finished plant effluent samples were collected from 82 conventional surface water treatment plants, located in 14 states, for a total of 1690 100-L finished water samples analyzed by the CC-PCR technique. Finished water samples were neutralized with a solution of 2% sodium thiosulfate (Sigma, St. Louis, MO) using an inline DEMA injector (DEMA, St. Louis, MO) at a flow rate of 10.0 ml/min. Cryptosporidium samples were collected according to Method 1622 (USEPA, 1998) using the Envirochek(TM) sampling capsule (Pall Life Sciences, Ann Harbor, MI) and filtration of 100 L of finished water at a flow rate of 2.0 L/minute. Infectious Cryptosporidium oocysts were detected using the cell culture PCR (CC-PCR) assay (Di Giovanni et al. 1999). For positive controls, live Cryptosporidium parvum oocysts from the Harley-Moon strain (NADC-USDA, Ames, IA), bovine genotype; no older than four weeks were purchased from the Sterling Parasitology Laboratory, University of Arizona, I(Tucson, AZ). To determine the genotype of the Cryptosporidium isolates, the hsp70 amplicon was cloned using the TOPO TA Cloning Kit (Invitrogen; Carlsbad, CA) in accordance with the manufacturer's instructions. To rule out the possibility that laboratory positive control samples (Harley-Moon strain oocysts) could have contaminated finished drinking water samples during processing, original hsp70 PCR reactions, which had been kept frozen (at -80C or -20C) for two to twenty-four months were reexamined for the gp60 marker as described by Strong et al. (2000). Includes 27 references, tables, figures.