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Applied and Environmental Microbiology, July 2005, p. 3848-3857, Vol. 71, No. 7
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.7.3848-3857.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Environmental Temperature Controls Cryptosporidium Oocyst Metabolic Rate and Associated Retention of Infectivity

Co-operative Research Centre for Water Quality and Treatment, Australian Water Quality Centre, SA Water Corporation, Salisbury, South Australia 5108, Australia

Received 5 November 2004/ Accepted 8 February 2005

Cryptosporidium is a significant cause of water-borne enteric disease throughout the world and represents a challenge to the water industry and a threat to public health. In this study we report the use of a cell culture-TaqMan PCR assay to measure oocyst inactivation rates in reagent-grade and environmental waters over a range of temperatures. While oocysts incubated at 4°C and 15°C remained infective over the 12-week holding period, we observed a 4 log₁₀ reduction in infectivity for both 20 and 25°C incubation treatments at 12 and 8 weeks, respectively, for all water types examined, a faster rate of inactivation for oocysts than previously reported. This temperature-dependent inactivation was further investigated using a simple and rapid ATP assay described herein. Time course experiments performed in reagent-grade water at incubation temperatures of 4, 15, 20, 25, 30, and 37°C identified a close relationship between oocyst infectivity and oocyst ATP content, demonstrating that temperature inactivation at higher temperatures is a function of increased oocyst metabolic activity. While water quality did not affect oocyst inactivation, biological antagonism appears to be a key factor affecting oocyst removal from environmental waters. Both the cell culture-TaqMan PCR assay and the ATP assay provide a sensitive and quantitative method for the determination of environmental oocyst inactivation, providing an alternative to the more costly and time-consuming mouse infection assay. The findings presented here relating temperature to oocyst inactivation provide valuable information for determining the relative risks associated with Cryptosporidium oocysts in water.

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