Monitoring source water quality in rivers and catchments remains a key priority not only for catchment managers, but also for water utilities. Here, we applied a best-practice approach to characterise pathogens from the genus Cryptosporidium in rivers used as a drinking water source. Sampling during high flow conditions caused by rainfall run-off determined not only the concentration of Cryptosporidium in water, but also identified the species present and the fraction which remained capable of forming an infection - which to date, has rarely been reported in the literature. The results demonstrate a positive and significant correlation between Cryptosporidium and flow (ρ = 0.756) and turbidity (ρ = 0.631) for all rainfall-runoff events, despite variable source water pathogen concentrations. Twelve Cryptosporidium species/genotypes were identified using molecular techniques, with most Cryptosporidium species detected associated with native and non-native wildlife (comprising 70 % of detections), while livestock-derived Cryptosporidium was detected less frequently (28 % of detections). Importantly, only 3/41 of water samples detected the presence of C. parvum (7 %) and no C. hominis was detected (0 %). Cell culture assays were used to measure the fraction of oocysts and gave an overall infectivity fraction of 3.1 %. To our knowledge, this is the first account which has quantified density, oocyst infectivity and foci species identity from a single environmental water sample.