Identification of plant DNA sequences from bat faecal samples
We initially filtered all sequences for quality and excluded low quality sequences where the PHRED score was <30 as indexed on the Barcode of Life Data Management System (BOLD) (Ratnasingham & Hebert, 2007). We compared the obtained rbc L and ITS2 sequences with the reference libraries of GenBank and BOLD using the BLAST algorithm with default search parameters (Altschul, Gish, Miller, Myers, & Limpman, 1990) and the combined BLAST and Hidden Markov Model methods implemented by the BOLD server (Ratnasingham & Hebert, 2007). For each reference database (BOLD, GenBank), we assigned query sequences to taxon based on highest percentage similarity, and considered a threshold of ≥97% to be a reliable assignment (Lamb, Winsley, Piper, Freidrich, & Siciliano, 2016). When there was an agreement between species-level matches for both markers (rbc L and ITS2) in both databases, with at least one match >97%, we assigned to the level of species. In cases where the query matched with equal similarity to multiple taxa of the same genus, we assigned the taxon to the level of the genus only, and similarly we used the same approach to assign query sequences to the level of the family. Where rbc L and ITS2 sequences matched different species from different genera, both at >97%, we concluded that two taxa were present in the sample and therefore assigned to both genera. Query sequences that did not show significant similarity to a reference were excluded from the analysis.
To corroborate our species assignments, for each candidate genus match, we reconstructed a gene phylogeny in which we included our query sequences together with all available reference sequences from species of the same genus present in BOLD that are also known to occur in Costa Rica. Sequences from rbc L and ITS2 of each plant genus were aligned with ClustalW (Larkin et al. 2007) in BioEdit v7.2.5 (Hall, 1999). For each alignment we ran a model selection test to check which would be the best method to build the phylogenetic tree based on the lowest BIC value. We ran model selection and built the phylogenetic trees using MEGA 6.06 (Tamura, Stecher, Peterson, Filipski, & Kumar, 2013). These phylogenies (not shown) recovered paraphyletic groupings for some species, perhaps through a lack of reference material, and therefore such species assignments were considered unreliable. To address this issue, we took a conservative approach and reduced all data to genus-level designations and repeated our analyses to check for consistency of results (see Supplementary material).
The identification of plant DNA sequences from bat faecal samples during the normal year relied on GenBank and BOLD, with the exception of thetrn H-psb A region which was not searchable within BOLD (see Clare et al. 2019 for more details) for our purposes we used the assignments as given in Clare et al. (2019).