Hidden biodiversity in Neotropical streams: DNA barcoding uncovers high endemicity of freshwater macroinvertebrates at small spatial scales

RESEARCH ARTICLE Hidden biodiversity in Neotropical streams: DNA barcoding uncovers high endemicity of freshwater macroinvertebrates at small spatial scales Luis F. De Leo´ nID1,2,3*, Aydee´ CornejoID4, Ronnie G. Gavila´nID5,6, Celestino AguilarID2,3 1 Department of Biology, University of Massachusetts Boston, Boston, MA, United States of America, 2 Centro de Biodiversidad y Descubrimiento de Drogas, Instituto de Investigaciones Cientı´ficas y Servicios de Alta Tecnologı´a (INDICASAT AIP), Panama´, Repu´ blica de Panama´ , 3 Smithsonian Tropical Research Institute, Balboa, Panama´ , 4 Instituto Conmemorativo Gorgas de Estudios de la Salud, Panama´, Repu´blica de Panama´, 5 Centro Nacional de Salud Pu´blica, Instituto Nacional de Salud, Lima, Peru´, 6 Escuela Profesional de Medicina Humana, Universidad Privada San Juan Bautista, Lima, Peru´ * luis.deleonreyna@umb.edu Abstract Aquatic macroinvertebrates play a crucial role in freshwater ecosystems, but their diversity remains poorly known, particularly in the tropics. This “taxonomic void” limits our under- standing of biodiversity patterns and processes in freshwater ecosystems, and the scale at which they operate. We used DNA barcoding to estimate lineage diversity (and the diversity of unique haplotypes) in 224 specimens of freshwater macroinvertebrates at a small spatial scale within the Panama Canal Watershed (PCW). In addition, we compiled available bar- coding data to assess macroinvertebrate diversity at a broader spatial scale spanning the

Isthmus of Panama. Consistently across two species delimitation algorithms (i.e., ABGD and GMYC), we found high lineage diversity within the PCW, with ~ 100–106 molecular operational taxonomic units (MOTUs) across 168 unique haplotypes. We also found a high lineage diversity along the Isthmus of Panama, but this diversity peaked within the PCW. However, our rarefaction/extrapolation approach showed that this diversity remains under- sampled. As expected, these results indicate that the diversity of Neotropical freshwater macroinvertebrates is higher than previously thought, with the possibility of high endemicity even at narrow spatial scales. Consistent with previous work on aquatic insects and other freshwater taxa in this region, geographic isolation is likely a main factor shaping these pat- terns of diversity. However, other factors such as habitat variability and perhaps local adap- tation might be reshaping these patterns of diversity at a local scale. Although further research is needed to better understand the processes driving diversification in freshwater macroinvertebrates, we suggest that Neotropical streams hold a high proportion of hidden biodiversity. Understanding this diversity is crucial in the face of increasing human disturbance. PLOS ONE PLOS ONE | https://doi.org/10.1371/journal.pone.0231683 August 7, 2020 1 / 13 a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 OPEN ACCESS Citation: De Leo´n LF, Cornejo A, Gavila´n RG, Aguilar C (2020) Hidden biodiversity in Neotropical streams: DNA barcoding uncovers high endemicity

of freshwater macroinvertebrates at small spatial scales. PLoS ONE 15(8): e0231683. https://doi. org/10.1371/journal.pone.0231683 Editor: Michael A. Chadwick, King’s College London, UNITED KINGDOM Received: March 28, 2020 Accepted: July 8, 2020 Published: August 7, 2020 Copyright: © 2020 De Leo´n et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: COI sequences associated with this study are available in GenBank (https://www.ncbi.nlm.nih.gov) under accession numbers KX039451 - KX039650 and KU980966 - KU981004. Data are also available on the Barcode of Life Data System (BOLD systems, under project code INVPA: http://www.boldsystems.org/). Taxonomic and molecular diversity data have been provided in Supporting information (S1 Table). Funding: Financial support was provided by a grant from the Consortium for the Barcode of Life to L. F.