The relationship between large herbivore numbers and landscape cover over time is poorly understood. There are two schools of thought: one views large herbivores as relatively passive elements upon the landscape and the other as ecosystem engineers driving vegetation succession. The latter relationship has been used as an argument to support reintroductions of large herbivores onto many landscapes in order to increase vegetation heterogeneity and biodiversity through local-scale disturbance regimes. Most of the research examining the relationship between large herbivores and their impact on landscapes has used extant studies. An alternative approach is to estimate the impact of variations in herbivore populations through time using fossil dung fungal spores and pollen in sedimentary sequences. However, to date, there has been little quantification of fossil dung fungal spore records and their relationship to herbivore numbers, leaving this method open to varied interpretations. In this study, we developed further the dung fungal spore method and determined the relationship between spore abundance in sediments (number cm−2 year−1) and herbivore biomass densities (kg ha−1). To establish this relationship, we used the following: (i) the abundance of Sporormiella spp., Sordaria spp. and Podospora spp. spores in modern sediments from ponds and (ii) weekly counts of contemporary wildlife over a period of 5 years from the rewilded site, Oostvaardersplassen, in the Netherlands. Results from this study demonstrate that there is a highly significant relationship between spore abundance and local biomass densities of herbivores that can be used in the calibration of fossil records. Mammal biomass density (comprising Konik horses, Heck cattle and red deer) predicts in a highly significant way the abundance of all dung fungal spores amalgamated together. This relationship is apparent at a very local scale (<10 m), when the characteristics of the sampled ponds are taken into account (surface area of pond, length of shoreline). In addition, we identify that dung fungal spores are principally transported into ponds by surface run-off from the shores. These results indicate that this method provides a robust quantitative measure of herbivore population size over time.