Submitted to: Emerging Infectious Diseases
Publication Type: Abstract Only
Publication Acceptance Date: 4/30/2000
Publication Date: 7/1/2001
Technical Abstract: By 2001, systems supplying water to US municipalities with over 10,000 people, will be required to meet new USEPA standards for levels of microbial contaminants, such as Cryptosporidium parvum . Detection methodologies for these and other pathogens will be critical in determining whether purification regimens are adequate. We are investigating the use of fnovel detection technologies, developed for use in biowarfare scenarios, i detecting C. parvum and other waterborne pathogens. We have designed a real time, PCR-based assay, for C. parvum using the 18S rRNA gene sequence as a target. The assay is genus-specific and can detect as few as 5 oocysts in a 50 £l reaction volume. The assay is capable of detecting 1000 oocysts in filtrate from a 10 liter volume of water. While developed for use on the ABI 7700 Prism sequence detection platform, the assay can also be used on new, portable thermal cycler instruments. We have demonstrated that the assay provides satisfactory results on a handheld, automated nucleic acid analyzer (HANAA). The HANAA offers real time monitoring of fluorogenic probe-based PCR reactions with discrimmination between two dyes. We believe such instruments, as well as anticipated improvements in sample processing and detection technologies, can offer water resource laboratories convenient, cost-effective protocols for pathogen surveilance and quality assurance.