This dataset and associated code files support all results reported in Lunn, et al., 2023 (https://doi.org/10.1101/2023.09.06.556454), where we found: Prediction and management of zoonotic pathogen spillover requires an understanding of infection dynamics within reservoir host populations. Transmission risk is often assessed using prevalence of infected hosts, with infection status based on the presence of genomic material. However, detection of viral genomic material alone does not necessarily indicate the presence of infectious virus, which could decouple prevalence from transmission risk. We undertook a multi-faceted investigation of Hendra virus shedding in Pteropus bats, combining insights from virus isolation, viral load proxies, viral prevalence, and longitudinal patterns of shedding, from 6,151 samples. In addition to seasonal and interannual fluctuation in prevalence, we found evidence for periodic shifts in the distribution of viral loads. The proportion of bats shedding high viral loads was higher during peak prevalence periods during which spillover events were observed, and lower during non-peak periods when there were no spillovers. We suggest that prolonged periods of low viral load and low prevalence reflect prolonged shedding of non-infectious RNA, or viral loads that are insufficient or unlikely to overcome dose barriers to spillover infection. These findings show that incorporating viral load (or proxies of viral load), into longitudinal studies of virus excretion will better inform predictions of spillover risk than prevalence alone.