<p>Artificial light at night (ALAN) negatively affects a broad range of animal species, with severe implications for conservation policy development and strategic planning globally. Birds are one of the most widely used ecological indicator groups in monitoring environmental changes. However, most studies examining the effects of ALAN are focused on diurnal bird species. It would therefore be necessary to study these effects in more detail on species with at least crepuscular or nocturnal activity, since they may be more vulnerable. We investigated the effects of illumination on the nest box occupancy of the western barn owl (<i>Tyto alba</i>; hereafter barn owl) and tawny owl (<i>Strix aluco</i>) in illuminated vs. unilluminated church towers and the reproductive output in nest boxes in these towers by comparing the numbers of eggs and chicks fledged. We found reduced breeding presence in illuminated towers in both species but no difference in reproduction parameters for either of the species. Our results underscore that light pollution has a negative consequence on the nest box occupancy of barn owls and tawny owls due to reducing breeding site suitability. This raises the threat that artificial light at night may hinder the conservation of such nocturnal bird species whose reproduction may be increasingly connected to human settlements in the future.</p>