AWWA WQTC55137

Since the placement of Encephalitozoon intestinalis on the USEPA's drinking water Emerging Pathogens Contaminant Candidate List (CCL), development of detection assays and disinfection criteria have become a priority for assessing the true extent of the risk to humans for this microsporidian parasite. This study reports on two methods used to address the issue of the sensitivity of low pressure UV light on the microsporidial pathogens Encephalitozoon intestinalis, E. hellem and E. cuniculi for determining UV log reduction. The first method is the in vitro Microwell Cell Culture coverslip assay reported by Wolk. The assay is sensitive and specific. Dose response curves have been generated for all three Encephalitozoon species in the RK-13 cell line and the untreated controls can be easily compared to these curves for ID50 determination. However, this assay is labor and time intensive since the whole stained 15mm coverslip has to be scanned at 400x by a skilled technician and data can not be generated until 6 days post infection. With the various molecular techniques reported for detection of infectivity in cell culture, it was decided to investigate an in vitro microwell RT-PCR cell culture assay method. These methods have been reported for Cryptosporidium parvum infectivity studies. Various Encephalitozoon published primer pairs and newly designed primers were investigated for the method development as was a standard PCR assay with beta-tublin primers. Includes 5 references, table, figure.