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Journal of Clinical Microbiology, September 2006, p. 3285-3291, Vol. 44, No. 9
0095-1137/06/$08.00+0     doi:10.1128/JCM.00541-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Rapid and Sensitive Detection of Single Cryptosporidium Oocysts from Archived Glass Slides

Centre for Molecular Biosciences, School of Biomedical Sciences, Faculty of Life and Health Sciences, University of Ulster, Cromore Road, Coleraine, Northern Ireland BT52 1SA,¹ Division of Parasitic Diseases, National Centers for Infectious Diseases, Centers for Disease Control and Prevention, 4770 Buford Highway, Chamblee, Georgia 30341,² Water Service Northern Ireland, Altnagelvin Laboratory, 1A Belt Road, Altnagelvin, Londonderry, Northern Ireland BT47 2LL,³ Northern Ireland Public Health Laboratory, Department of Bacteriology, Belfast City Hospital, Belfast, Northern Ireland BT9 7AD⁴

Received 13 March 2006/ Returned for modification 4 May 2006/ Accepted 5 July 2006

In this study we report on the development and application of a novel method for efficiently extracting and detecting single Cryptosporidium oocysts from archived glass slides. Laser capture microscopy was used to extract low numbers of oocysts from archived glass slides. Highly sensitive real-time PCR methods were then developed to enable the rapid detection and identification of Cryptosporidium oocysts from these samples. The method was applied to fecal smears stained with a variety of standard oocyst stains and water samples. This application, with samples derived from both public health and water service laboratories, highlighted the strong potential of this method to be used as a rapid high-throughput screening tool for the routine monitoring of Cryptosporidium and other medically important pathogens from clinical, veterinary, and environmental water samples. Importantly, the application of our protocol could be used to type Cryptosporidium and other pathogens from stored archived glass slides in public health and water service laboratories, providing vital epidemiological updates and helping to identify and trace pathogens and their routes of infection and ultimately improve their control.

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