Mitogenomes of the four Agathymus holotypes collected 55 years ago

Giant-Skipper butterflies from the genus Agathymus (family Hesperiidae) are unusual as their caterpillars feed inside Agave leaves. Relationships among Agathymus taxa and their names (i.e., if they are species, subspecies, or synonyms) are poorly understood due to phenotypic similarity. DNA sequences are promising to clarify the taxonomic questions, but it is challenging to sequence name-bearing types that are usually old specimens with poorly preserved DNA. Using next generation sequencing, we assembled mitochondrial genomes of four Agathymus mariae group holotype specimens collected more than 55 years ago and housed pinned and dry in the American Museum of Natural History (New York, NY). We compared the holotype mitogenomes to those we obtained from fresh A. mariae specimens and the sister species Agathymus micheneri. All but A. micheneri mitogenomes were highly similar to each other (more than 99% identity), suggesting that the four names chinatiensis, lajitaensis, rindgei and gilberti proposed by H. A. Freeman in 1964 may not refer to species-level taxa. The mitogenomes grouped eastern populations (rindgei and gilberti) together and apart from the western populations (nominal mariae, chinatiensis and lajitaensis). Mexican A. micheneri differs by about 2.5% (about 5% in the COI barcode region) from A. mariae, and is likely to be a distinct species.