Biotic interactions are crucial for determining the structure and dynamics of communities; however, direct measurement of these interactions can be challenging in terms of time and resources, especially when numerous species are involved. Inferring species interactions from species co-occurrence patterns is increasingly being used; however, recent studies have highlighted some limitations. To our knowledge, no attempt has been made to test the accuracy of the existing methods for detecting mutualistic interactions in terrestrial ecosystems. In this study, we compiled two literature-based, long-term datasets of interactions between butterflies and herbaceous plant species in two regions of Germany and compared them with observational abundance and presence/absence data collected within a year in the same regions. We tested how well the species associations generated by three different co-occurrence analysis methods matched those of empirically measured mutualistic associations using sensitivity and specificity analyses and compared the strength of associations. We also checked whether flower abundance data (instead of plant abundance data) increased the accuracy of the co-occurrence models and validated our results using empirical flower visitation data. The results revealed that, although all methods exhibited low sensitivity, our implementation of the Relative Interaction Intensity index with pairwise null models performed the best, followed by the probabilistic method and Spearman's rank correlation method. However, empirical data showed a significant number of interactions that were not detected using co-occurrence methods. Incorporating flower abundance data did not improve sensitivity but enhanced specificity in one region. Further analysis demonstrated incongruence between the predicted co-occurrence associations and actual interaction strengths, with many pairs exhibiting high interaction strength but low co-occurrence or vice versa. These findings underscore the complexity of ecological dynamics and highlight the limitations of current co-occurrence methods for accurately capturing species interactions.
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