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Project title

Pulsed-UV light inactivation of Cryptosporidium spp. oocysts in drinking water supplies in Ireland

Primary Funding Agency

Environmental Protection Agency

Co-Funding Organisation(s)

n/a

Lead Organisation

Athlone Institute of Technology (AIT)

Lead Applicant

Neil J Rowan

Project Abstract

Aim and rationale: the aim of this timely and pressing multi-disciplinary study was toinvestigate development and optimization of a high-intensity pulsed UV light (PUV) sourcefor the novel disinfection of Cryptosporidium parvum oocysts suspended in water. Therationale and need for same is due to the fact that C. parvum is an enteric coccidian parasitethat is recognised as a frequent cause of illness in humans due to its resistance todisinfection with chlorine at concentrations typically applied in drinking water treatmentplants. Development of alternative methods of Cryptosporidium disinfection for waterapplications (such as UV) has been hindered by numerous factors including the uncertaintysurrounding efficacy of in vitro surrogate viability assays to quantify oocyst survivors posttreatment and the lack of critical data focusing on congruence of in vitro cell culture and invivo animal based infectivity bioassays to evaluate efficacy of using conventional andemerging disinfection techniques.Methods and Results: due to numerous factors including (but not limited to) the lack ofavailability of oocysts due to the fact that this parasite has a complex life cycle that requiresuse of an animal host (with obvious ethical implications) initial findings identified criticalinter-related factors affecting the destruction of other relevant bacteria and yeast undervarying operational conditions. These studies were used to inform subsequent treatmentsusing the more recalcitrant C. parvum. Albeit extensive in scope and depth the latter studyproved essential in terms of limiting use of animals for important infectivity studies theformer element being factored into a successful animal licence application that wasawarded by the Department of Health and Children for this purpose. The in vitro HCT-8cell culture combined with quantitative (real-time) PCR was then compared to the SCIDmouseinfectivity bioassay to evaluate these PUV operating conditions in order to reduce oreliminate C. parvum oocyst infectivity when treated in water. Each treated dose of oocystswas inoculated into 3 replicates of 4 mice or 8 cell culture wells. Infections post UVtreatments were detected by immunofluorescence (IF) microscopy and by quantitative PCRin cell culture and by IF staining of faeces and by haematoxylin and eosin staining ofintestinal villi in mice. There was good agreement between using cell culture-qPCR and themouse assay for determining reduction or elimination of C. parvum infectivity as aconsequence of varying UV operating conditions. Reduction in infectivity depended on theintensity of lamp discharge energy or UV dose (mJ/cm2) applied amount of pulsing andpopulation size of oocysts (P 0.05). Use of surrogate vital staining over-estimatedsurvivoris post-UV treatments compared to use of infectivity assays (P 0.05).Incorporation of humic acid at a concentration above that found in surface water (i.e. 10ppm) did not significantly affect PUV disinfection capability irrespective of parameterstested (P 0.05). It took ca. 3.9 times longer to eliminate C. parvum infectivity usingcontinuous low-pressure monochromatic light at a UV dose of 327 mJ/cm2 compared totreating similar populations of oocysts with PUV at 4.6 mJ/cm2. Use of ATP technique wasshown to be an inferior technique for evaluating PUV disinfection performance comparedto use of cell culture i- qPCR or using the mouse-model. Studies revealed an inability of C.parvum to photo-repair post exposure to lethal doses of conventional (L-P) or PUV. Anextensive battery of eco-toxicity testing revealed that PUV treatment of water containingdifferent pre-determined levels of C. parvum oocysts were safe to release into theenvironment and were of satisfactory potability quality.

Grant Approved

�50,000.00

Research Hub

Natural Environment

Research Theme

Protecting our Water Resources

Start Date

01/10/2006

Initial Projected Completion Date

n/a