Moon phase and variation in ambient light conditions can influence predator and prey behaviour. Nocturnal predators locate prey visually, and prey may adjust their activity to minimise their predation risk. Understanding how native mammals in Australia respond to varying phases of the moon and cloud cover (light) enhances knowledge of factors affecting species’ survival and inference regarding ecological and population survey data. Over a two-year period within a fenced conservation reserve, in south-eastern Australia, with reintroduced native marsupial predator and prey species (eastern barred bandicoot, southern brown bandicoot, long-nosed potoroo, rufous bettong, Tasmanian pademelon, brush-tailed rock-wallaby, red-necked wallaby, eastern quoll, spotted-tailed quoll, and naturally occurring swamp wallaby, common brushtail possum, common ringtail possum), we conducted monthly spotlight surveys during different moon phases (full, half and new moon). We found an interaction between cloud cover and moon phase, and an interaction of the two depending on the mammal size and class. Increased activity of prey species corresponded with periods of increasing cloud cover. Predators and medium-sized herbivores were more active during times of low illumination. Our findings suggest that moon phase affects the nocturnal activity of mammal species and that, for prey species, there might be trade-offs between predation risk and foraging. Our findings have implications for: ecological survey design and interpretation of results for mammal populations across moon phases, understanding predator and prey behaviour and interactions in natural and modified (artificial lighting) ecosystems, and potential nocturnal niche partitioning of species.