To investigate the effect of UV light on Cryptosporidium parvum and Cryptosporidium hominis oocysts in vitro, we exposed intact oocysts to 4-, 10-, 20-, and 40-mJ ⅐ cm ؊2 doses of UV irradiation. Thymine dimers were detected by immunofluorescence microscopy using a monoclonal antibody against cyclobutyl thymine dimers (anti-TDmAb). Dimer-specific fluorescence within sporozoite nuclei was confirmed by colocalization with the nuclear fluorogen 4,6-diamidino-2-phenylindole (DAPI). Oocyst walls were visualized using either commercial fluorescein isothiocyanate-labeled anti-Cryptosporidium oocyst antibodies (FITC-CmAb) or Texas Redlabeled anti-Cryptosporidium oocyst antibodies (TR-CmAb). The use of FITC-CmAb interfered with TD detection at doses below 40 mJ ⅐ cm ؊2 . With the combination of anti-TDmAb, TR-CmAb, and DAPI, dimer-specific fluorescence was detected in sporozoite nuclei within oocysts exposed to 10 to 40 mJ ⅐ cm ؊2 of UV light. Similar results were obtained with C. hominis. C. parvum oocysts exposed to 10 to 40 mJ ⅐ cm ؊2 of UV light failed to infect neonatal mice, confirming that results of our anti-TD immunofluorescence assay paralleled the outcomes of our neonatal mouse infectivity assay. These results suggest that our immunofluorescence assay is suitable for detecting DNA damage in C. parvum and C. hominis oocysts induced following exposure to UV light.UV irradiation as a sterilization technology in the water and food industry is effective in killing contaminating organisms, such as viruses, bacteria, fungal spores, and parasites (8). The major DNA lesions caused by UV light are cyclobutyl pyrimidine dimers (CPD), which are responsible for UV-induced cytotoxicity and mutagenicity in living cells and microorganisms. UV-induced damage consists of chemical base modifications, whereby two adjacent pyrimidine residues (cytosine or thymine) form a dimer (thymine dimer [TD], cytosine dimer, or thymine-cytosine heterodimer) and 6-4 photoproducts. The formation of these lesions in genomic DNA inhibits normal replication and transcription of DNA and results in the inactivation of cells (12). However, UV-induced DNA lesions in living human epidermal cells (21) and in some unicellular microorganisms (16) can be repaired by one or more mechanisms. These mechanisms include the enzyme-dependent nucleotide excision repair, also named dark repair, and the light-dependent reaction known as photoreactivation. Dark repair and photoreactivation enable UV-inactivated microorganisms to recover, which can reduce the efficiency of UV inactivation (20).Cryptosporidium parvum oocysts have the ability to carry out photoreactivation and dark repair at the genomic level (13, 16). Also, nucleotide excision repair genes in C. parvum and Cryptosporidium hominis have been identified. However, UV inactivation of Cryptosporidium oocysts is irreversible, despite the presence of these UV repair genes (18). Rochelle et al. (19) demonstrated the accumulation of cyclobutane TDs in the genome of C. parvum oocysts exposed to increasing dosage...