Mind the gap! Integrating taxonomic approaches to assess ant diversity at the southern extreme of the Atlantic Forest

Abstract

Understanding patterns of species diversity relies on accurate taxonomy which can only be achieved by long-term natural history research and the use of complementary information to establish species boundaries among cryptic taxa. We used DNA barcoding to characterize the ant diversity of Iguazú National Park (INP), a protected area of the Upper Paraná Atlantic Forest ecoregion, located at the southernmost extent of this forest. We assessed ant diversity using both cytochrome c oxidase subunit 1 (COI) sequences and traditional morphological approaches, and compared the results of these two methods. We successfully obtained COI sequences for 312 specimens belonging to 124 species, providing a DNA barcode reference library for nearly 50% of the currently known ant fauna of INP. Our results support a clear barcode gap for all but two species, with a mean intraspecific divergence of 0.72%, and an average congeneric distance of 17.25%. Congruently, the library assembled here was useful for the discrimination of the ants of INP and allowed us to link unidentified males and queens to their worker castes. To detect overlooked diversity, we classified the DNA barcodes into Molecular Operational Taxonomic Units (MOTUs) using three different clustering algorithms, and compared their number and composition to that of reference species identified based on morphology. The MOTU count was always higher than that of reference species regardless of the method, suggesting that the diversity of ants at INP could be between 6% and 10% higher than currently recognized. Lastly, our survey contributed with 78 new barcode clusters to the global DNA barcode reference library, and added 36 new records of ant species for the INP, being 23 of them new citations for Argentina.

Keywords: Argentina; DNA barcoding; Formicidae; Iguazú National Park; species delimitation.

Figure 1

Neighbor‐joining (NJ) tree of 312 COI sequences of Iguazú National Park ants computed with a K2P substitution model (30 high‐quality sequences for specimens that were not identified to species were not included). Symbols next to the terminals indicate when a species was split (filled) or merged (blank) by RESL (circles), TCS (squares), or ABGD (stars). Numbers above the node correspond to NJ/ML (maximum likelihood) bootstrap support values based on 1,000 pseudoreplicates

Figure 2

Frequency distribution of genetic distances within species and among congeneric species

Figure 3

Barcode gap analysis for 65 species of ants with two or more individuals. Each individual is represented by a point, and the distance to the furthest heterospecific is plotted against the minimum distance to the nearest neighbor. The vertical dashed line shows the 95th percentile of all intraspecific distances (5.75%), while the horizontal one corresponds to the lower 5% of congeneric distances (13.25%). Points below the diagonal (1:1 relationship) correspond to Ectatomma edentatum and Neoponera crenata (see text for more details)

Figure 4

Number of MOTUs obtained for each clustering delimitation methodology. Dashed line represents the number of identified species (124).

Figure 5

Percentages of MATCHES, SPLITS AND MERGES for the different clustering methods discussed in the text based on the correspondence between reference species and MOTUs boundaries.

Figure 6

One of the new records of Iguazú National Park: the arboreal termite hunter, Cylindromyrmex brasiliensis