Combining target enrichment and Sanger sequencing data to clarify the systematics of the diverse Neotropical butterfly subtribe Euptychiina (Nymphalidae, Satyrinae) Author Espeland, Marianne Author Nakahara, Shinichi Author Zacca, Thamara Author Barbosa, Eduardo P. Author Huertas, Blanca Author Marín, Mario A. Author Lamas, Gerardo Author Benmesbah, Mohamed Author Brévignon, Christian Author Casagrande, Mirna M. Author Fåhraeus, Christer Author Grishin, Nick Author Kawahara, Akito Y. Author Mielke, Olaf H. H. Author Miller, Jacqueline Y. Author Nakamura, Ichiro Author Navas, Vanessa Author Patrusky, Brooke Author Pyrcz, Tomasz W. Author Richards, Lindsay Author Tan, Denise Author Tyler, Stephanie Author Viloria, Angel Author Warren, Andrew D. Author Xiao, Lei Author Freitas, André V. L. Author Willmott, Keith R. text Zoological Research 2023 2023-02-15 2023 1 73 journal article 58039 10.1111/syen.12590 bfb878f3-8a74-46d3-a104-36485c32aaba 7909395 Splendeuptychia clade The ‘ Splendeuptychia clade’ was proposed by Peña et al. (2010) , and was partially recovered by Espeland et al. (2019a) with the addition of Magneuptychia and Cepheuptychia ; the clade also contains those species included within the ‘ Cissia clade’ of Murray and Prowell (2005) . In our present study, this clade comprises the genera Splendeuptychia (9 described species), Huberonympha (monotypic), Stevenaria (3 species), Cepheuptychia (3 species), Omacha (monotypic), Vareuptychia (2 species), Magneuptychia (2 species), Capronnieria (2 species), Caeruleuptychia (26 species), Emeryus (3 species), Colombeia (3 species), Malaveria (17 species), Modestia (8 species), Scriptor (monotypic), Paryphthimoides (12 species), Trico gen. n. (monotypic), Occulta gen.n. (monotypic), Deltaya gen.n. (6 species), and Modica gen.n. (5 species). The clade itself, and all its genera, are well-supported in all trees ( Figures 10 , S 2 and S 3), but the relationships among genera are typically only weakly supported, with a few exceptions. There is relatively good support in both the FULL and 4GENES datasets for a clade including Huberonympha , Stevenaria and Cepheuptychia being the sister group to the remainder of the clade, followed by Trico gen.n . and Occulta , which either form a clade or a grade. There is also good support for a clade consisting of Malaveria , Colombeia , Emeryus , Deltayagen .n. , Scriptor , and Parypthimoides (all trees ≥ 100/95), which, with the exception of Emeryus and Scriptor , which were not included there, were also found in the backbone tree in this study and in Espeland et al. (2019a) . The placement of genera within this clade is, however, rather uncertain, and the positions of Deltaya gen.n. and Emeryus seem especially unstable. In the best FULL tree Deltaya gen.n. is sister to Scriptor ( Figure 10 ), in the second best FULL tree it is sister to Capronnieria , and in both 4GENES trees it is sister to Scriptor + Colombeia , but no placement is well-supported. Emeryus similarly moves around, and in the best FULL tree it is sister to Malaveria + Colombeia (on a very short branch) and in the second best FULL tree it is sister to Modica gen.n . Both 4GENES trees again give different placements of this genus; in the best tree it is sister to Parypthimoides and in the second best tree it is instead sister to a clade consisting of Scriptor , Deltayagen .n. and Modica gen.n. (again on a very short branch). Three genes or more were available for at least some samples of both Deltaya gen. n. and Emeryus , but none of these genera were included in the backbone, so long-branch-attraction might be one of the reasons why these genera move around. More data is needed to possibly resolve these relationships, but the very short branches in the backbonemight make this task difficult. Several recent papers have provided generic revisions and descriptions of new taxa from this clade ( Zacca et al., 2014 ; Zacca, Casagrande, et al., 2017; Zacca, Casagrande, Mielke, Huertas, Barbosa, Freitas, et al., 2020 [ Paryphthimoides ], Zacca, Casagrande, Mielke, Huertas, Espeland, Freitas, et al., 2020 [ Vareuptychia ], Zacca, Casagrande, Mielke, Huertas, Barbosa, Freitas, & Willmott, 2020 [ Emeryus ]; Costa et al., 2016 [ Huberonympha , Stevenaria , Magneuptychia ]; Benmesbah et al., 2018 , 2021 [ Malaveria , Modestia ]; Nakahara, Zacca, et al., 2018 [ Caeruleuptychia ], Nakahara, Lamas, et al., 2020 [ Scriptor ], Nakahara et al., 2023 [ Caeruleuptychia ]; Andrade-Correa et al., 2019 [ Omacha ]; Viloria, 2022 [ Malaveria ]). Andrade-Correa et al. (2019) described Colombeia as a monotypic genus with Euptychia mycalesis as type species, but our molecular data show that species to be a member of a strongly supported clade containing two other species, which we here also transfer to Colombeia ( Colombeia nossis comb.n. and Colombeia hotchkissi comb.n. ). The genus Malaveria was recently described with Euptychia nebulosa as type species ( Benmesbah et al., 2021 ), with three other described species moved into the genus and a further four species described. These last species descriptions highlighted hitherto unrecognized important morphological and molecular variation, but all of the described species are allopatric with respect to their closest relatives, or with only limited evidence for sympatry, and further research is needed to confirm their status. Barbosa et al. (2022) showed that five additional species also form a clade with Malaveria species and are best placed in that genus, and a number of other new species require description. We also provisionally transfer Euptychia argyrospila Butler, 1867 from Yphthimoides to Malaveria ( comb.n. ) since partial DNA barcode data support this placement, while the male genitalia illustrated by Forster (1964) also appear to be similar to those of other Malaveria , but this classification requires confirmation. Most recently, Viloria (2022) described the new genus Rudyphthimoides with a new species, R . iseai , as type species, and also included several species placed by Barbosa et al. (2022) in Malaveria . While it seems likely that Rudyphthimoides forms a clade (containing M . maepius , M . affinis and M . mythra in Figure 10 ), retaining this generic name would restrict the name Malaveria to a very closely related, and weakly supported, clade, and require description of a new monotypic genus for M . grimon . Alternatively, recognizing Malaveria as including M . grimon , Rudyphthimoides , and remaining Malaveria results in a strongly supported clade, separated from other genera by a relatively long branch ( Figure 10 ), which is likely to remain stable. We therefore prefer this latter solution and treat Rudyphthimoides ( syn.n. ) as a synonym of Malaveria . The genus Modestia was recently described with Euptychia modesta as type species ( Benmesbah et al., 2021 ), with the inclusion of one other described species and one newly described species. Molecular data recovered a strongly supported clade containing the three original species of Modestia along with four species previously placed in Magneuptychia ( Lamas, 2004 ) , which are here transferred to Modestia . The genus Nubila was described by Andrade-Correa et al. (2019) with Euptychia gera Hewitson, 1850 as type species, with five additional closely related species transferred from Magneuptychia and Splendeuptychia . These species do indeed form a well-supported clade in our study ( Figure 10 ), which is placed without strong support as sister to a clade containing the recently described Splendeuptychia tupinamba and an undescribed species. Both these clades are sister to a clade containing the type species of Splendeuptychia . Splendeuptychia tupinamba and the undescribed species are phenotypically intermediate between Nubila and the clade containing other described Splendeuptychia , and since morphological data also support a close relationship among these species ( Huertas, 2014 ), we believe the best taxonomic solution is to regard them as a single genus, so we therefore synonymize Nubila syn.n. with Splendeuptychia . Considering the high diversity in the wing pattern, venation, and genitalia exhibited by the members of the ‘ Splendeuptychia clade’, it is difficult to identify possible morphological synapomorphies and distinctive characters for the clade. Most of the species of the clade occur in South America, except for members of Vareuptychia and some Paryphthimoides species that occur in Central America. Habitat preferences of species in this clade range from lowland rainforest and savanna to cloud forest up to 2200 m . Adults of Magneuptychia , Emeryus and Paryphthimoides are commonly attracted to traps baited with rotting fruit ( LourenÇo et al., 2019 ; Melo et al., 2019 ; Santos et al., 2018 ) and some Caeruleuptychia , Paryphthimoides and Splendeuptychia species show hilltopping behaviour ( Benmesbah et al., 2018 ; Nakahara, Zacca, et al., 2018). Known larvae of the species in this clade feed on various genera of Poaceae (e.g. Beccaloni et al., 2008 ; Brown, 1992 ; Corahua-Espinoza, Nakahara, Kabir, et al., 2022; Corahua-Espinoza, Nakahara, Shellman, et al., 2022; DeVries, 1987 ; Janzen & Hallwachs, 2022 ; Núñez-Bustos, 2010 ), with additional records of Cyperaceae and Marantaceae for Vareuptychia themis ( Janzen & Hallwachs, 2022 ) and Heliconiaceae for Modica confusa comb.n. ( Janzen & Hallwachs, 2022 ). For most species of this clade, published information on the natural history, immatures and biology is still unknown, with a few exceptions (Corahua-Espinoza, Nakahara, Kabir, et al., 2022; Corahua-Espinoza, Nakahara, Shellman, et al., 2022; Freitas, 2022 ; Janzen & Hallwachs, 2022 ; Kaminski & Freitas, 2008 ; Nakahara, Hoffman, et al., 2020).