Resolving Relationships among the Megadiverse Butterflies and Moths with a Novel Pipeline for Anchored Phylogenomics

Abstract

The advent of next-generation sequencing technology has allowed for thecollection of large portions of the genome for phylogenetic analysis. Hybrid enrichment and transcriptomics are two techniques that leverage next-generation sequencing and have shown much promise. However, methods for processing hybrid enrichment data are still limited. We developed a pipeline for anchored hybrid enrichment (AHE) read assembly, orthology determination, contamination screening, and data processing for sequences flanking the target "probe" region. We apply this approach to study the phylogeny of butterflies and moths (Lepidoptera), a megadiverse group of more than 157,000 described species with poorly understood deep-level phylogenetic relationships. We introduce a new, 855 locus AHE kit for Lepidoptera phylogenetics and compare resulting trees to those from transcriptomes. The enrichment kit was designed from existing genomes, transcriptomes, and expressed sequence tags and was used to capture sequence data from 54 species from 23 lepidopteran families. Phylogenies estimated from AHE data were largely congruent with trees generated from transcriptomes, with strong support for relationships at all but the deepest taxonomic levels. We combine AHE and transcriptomic data to generate a new Lepidoptera phylogeny, representing 76 exemplar species in 42 families. The tree provides robust support for many relationships, including those among the seven butterfly families. The addition of AHE data to an existing transcriptomic dataset lowers node support along the Lepidoptera backbone, but firmly places taxa with AHE data on the phylogeny. Combining taxa sequenced for AHE with existing transcriptomes and genomes resulted in a tree with strong support for (Calliduloidea $+$ Gelechioidea $+$ Thyridoidea) $+$ (Papilionoidea $+$ Pyraloidea $+$ Macroheterocera). To examine the efficacy of AHE at a shallow taxonomic level, phylogenetic analyses were also conducted on a sister group representing a more recent divergence, the Saturniidae and Sphingidae. These analyses utilized sequences from the probe region and data flanking it, nearly doubled the size of the dataset; resulting trees supported new phylogenetics relationships, especially within the Saturniidae and Sphingidae (e.g., Hemarina derived in the latter). We hope that our data processing pipeline, hybrid enrichment gene set, and approach of combining AHE data with transcriptomes will be useful for the broader systematics community.

Anchored hybrid enrichment provides new insights into the phylogeny and evolution of longhorned beetles (Cerambycidae)

“…Probes for Coleoptera were designed using methods similar to those used for a recently published probe set for Diptera (Young et al, 2016) and Lepidoptera (Breinholt et al, 2017). Specifically, we obtained nucleotide alignments of 4485 protein-coding genes for 13 insect species from Niehuis et al (2012).…”

“…Bi et al, 2012;Kawahara & Breinholt, 2014;Misof et al, 2014;Kjer et al, 2015;Lei & Dong, 2016) or target enrichment (e.g. Faircloth et al, 2012;Lemmon et al, 2012;Smith et al, 2014;Brandley et al, 2015;Eytan et al, 2015;Prum et al, 2015;Hamilton et al, 2016;Young et al, 2016;Breinholt et al, 2017), to generate large-scale phylogenomic datasets for use in phylogeny reconstruction, particularly in groups for which relationships have been difficult to resolve using smaller samples of molecular data (see Lemmon & Lemmon, 2013 for a review of alternative approaches). WGS produces the most data, but is the most expensive of these approaches and also carries a high bioinformatic and computational burden.…”

A global checklist of the Bombycoidea (Insecta: Lepidoptera)

“…The morphology-based phylogenetic studies of Minet (1991) and Minet (1994) were seminal for the modern classification of Bombycoidea. Subsequent molecular studies proposed many new intrafamilial backbone phylogenies of Bombycoidea (e.g., Barber et al 2015, Breinholt et al 2017, Kawahara and Barber 2015, Kawahara et al 2009, Regier et al 2008a, Zwick 2008, Zwick et al 2011) and the higher classification of the superfamily has changed significantly, but some parts remain inadequately resolved. At lower levels, there have been only a relatively small number of phylogenetic studies focusing on particular genera (e.g., Ylla et al 2005, Rubinoff and Le Roux 2008, Kawahara et al 2013, Ponce et al 2014, Rubinoff et al 2017), while new species descriptions continue to accumulate at a very high pace.…”

“…The new tribes, subtribes and genera described by Cooper (2002) are here treated as junior synonyms. We also present a new higher classification of Sphingidae, based on Kawahara et al (2009), Barber and Kawahara (2013) and a more recent phylogenomic study by Breinholt et al (2017), as well as a reviewed genus and species level classification, as documented by Kitching (2018).…”

A stable phylogenomic classification of Travunioidea (Arachnida, Opiliones, Laniatores) based on sequence capture of ultraconserved elements

“…The stability of any phylogenetics-based classification relies upon high confidence and support for internal relationships. In other animal groups, genomic- or subgenomic-scale approaches have produced phylogenies with generally higher nodal support and have resolved difficult relationships (e.g., Blaimer et al 2015, Garrison et al 2016, Hamilton et al 2016, Baca et al 2017, Branstetter et al 2017, Breinholt et al 2017, Hedin et al 2018). In this paper we utilize DNA sequences derived from capture of ultraconserved elements (UCEs; Faircloth 2017) to reconstruct phylogenomic relationships within Travunioidea.…”