Supplementary Table 1. Sanger sequencing and NCBI BLAST results for exemplar amplicons from each treatment of the proof-of concept study. From: Using Noninvasive Genetic Sampling to Survey Rare Butterfly Populations

Supplementary Table 1. Sanger sequencing and NCBI BLAST results for exemplar amplicons from each treatment of the proof-of concept study. From: Using Noninvasive Genetic Sampling to Survey Rare Butterfly Populations

Gracillariidae are one of the most diverse families of internally feeding insects, and many species are economically important. Study of this family has been hampered by lack of a robust and comprehensive phylogeny. In the present paper, we sequenced up to 22 genes in 96 gracillariid species, representing all previously recognized subfamilies and genus groups, plus 20 outgroups representing other families and superfamilies. Following objective identification and removal of two rogue taxa, two datasets were constructed: dataset 1, which included 12 loci totalling 9927 bp for 94 taxa, and dataset 2, which supplemented dataset 1 with 10 additional loci for 10 taxa, for a total of 22 loci and 16 167 bp. Maximum likelihood analyses strongly supported the monophyly of Gracillariidae and most previously recognized subfamilies and genus groups. On this basis, we propose a new classification consisting of eight subfamilies, four of which are newly recognized or resurrected: Acrocercopinae Kawahara & Ohshima subfam. n.; Gracillariinae Stainton; Lithocolletinae Stainton; Marmarinae Kawahara & Ohshima subfam. n.; Oecophyllembiinae Réal & Balachowsky; Parornichinae Kawahara & Ohshima subfam. n.; Ornixolinae Kuznetzov & Baryshnikova stat. rev.; and Phyllocnistinae Zeller. The subfamily Gracillariinae is restricted to the monophyletic group comprising Gracillaria Haworth and closely related genera. We also formally transfer Acrocercops scriptulata Meyrick to Ornixolinae and use the name Diphtheroptila Vári, creating Diphtheroptila scriptulata comb. n. An exploratory mapping of larval host-use traits on the phylogeny shows strong conservation of modes of leaf mining but much higher lability of associations with host plant orders and families, suggesting that host shifts could play a significant role in gracillariid diversification.

The hawkmoth genus Manduca is a diverse group of very large, conspicuous moths that has served as an important model across many biological disciplines. Two species in particular, the tobacco hornworm (Manduca sexta) and the tomato hornworm (Manduca quinquemaculatus) have been researched extensively. Studies across biological fields have referred to these two species as being closely related or even sister species, but the extent to which these two model organisms are related remains largely unknown. We conducted a comprehensive multi-gene phylogenetic analysis of Manduca, based on both an ML and Bayesian framework, which resulted in a monophyletic Manduca but only when two other genera, Dolba and Euryglottis are included. We tentatively conclude that the sister group to Manduca sexta comprises the Caribbean M. afflicta and M. johanni, and the sister lineage to this clade includes M. quinquemaculatus and the Hawaiian M. blackburni. Thus, M. sexta and M. quinquemaculatus are closely related, but are not sister species. Biogeographical analyses reveal an ancestral center of diversification in Central America, and Manduca appears to have subsequently colonized North and South America. Our phylogeny provides an important foundation for comparative studies of two model organisms and their relatives.