Body size, nuptial pad size and hormone levels: potential non-destructive biomarkers of reproductive health in wild toads (Bufo bufo).

Amphibians are declining and fertility/fecundity are major drivers of population stability. The development of non-destructive methods to assess reproductive health are needed as destructive measures are fundamentally at odds with conservation goals for declining species. We investigated the utility of body size, nuptial pad size and forelimb width as non-destructive biomarkers of internal reproductive physiology, by analysing correlations with commonly used destructive methods in adult male toads (Bufo bufo) from a low human impact and a high human impact site. Principal component analyses revealed that size was the most important variable for explaining inter-individual differences in other measured endpoints, both non-destructive and destructive, except for hormone levels and nuptial pad, which were independent of size. Toads from the LI and the HI site differed in almost all of the measured endpoints; this was largely driven by the significantly smaller size of toads from the HI site. Correlational analyses within sites revealed that size was correlated with several reproductive endpoints in toads from the HI site but not the LI site, indicating a possible limiting effect of size on reproductive physiology. Intersex was observed in 33% of toads from the HI site and incidence was not related to any other measured endpoint. In conclusion, we provide evidence that size is associated with reproductive physiology and that nuptial pad/hormone levels have potential as additional markers due to their independence from size. We also show that human activities can have a negative effect on reproductive physiology of the common toad.