Phylogenomics resolves major relationships and reveals significant diversification rate shifts in the evolution of silk moths and relatives.

BACKGROUND: Silkmoths and their relatives constitute the ecologically and taxonomically diverse superfamily Bombycoidea, which includes some of the most charismatic species of Lepidoptera. Despite displaying spectacular forms and diverse ecological traits, relatively little attention has been given to understanding their evolution and drivers of their diversity. To begin to address this problem, we created a new Bombycoidea-specific Anchored Hybrid Enrichment (AHE) probe set and sampled up to 571 loci for 117 taxa across all major lineages of the Bombycoidea, with a newly developed DNA extraction protocol that allows Lepidoptera specimens to be readily sequenced from pinned natural history collections. RESULTS: The well-supported tree was overall consistent with prior morphological and molecular studies, although some taxa were misplaced. The bombycid Arotros Schaus was formally transferred to Apatelodidae. We identified important evolutionary patterns (e.g., morphology, biogeography, and differences in speciation and extinction), and our analysis of diversification rates highlights the stark increases that exist within the Sphingidae (hawkmoths) and Saturniidae (wild silkmoths). CONCLUSIONS: Our study establishes a backbone for future evolutionary, comparative, and taxonomic studies of Bombycoidea. We postulate that the rate shifts identified are due to the well-documented bat-moth "arms race". Our research highlights the flexibility of AHE to generate genomic data from a wide range of museum specimens, both age and preservation method, and will allow researchers to tap into the wealth of biological data residing in natural history collections around the globe.

Complete mitochondrial genome of Saturnia jonasii (Lepidoptera: Saturniidae): Genomic comparisons and phylogenetic inference among Bombycoidea.

The complete mitochondrial genome (mitogenome) of Saturnia jonasii (Lepidoptera: Saturniidae) was sequenced and compared to those of 19 other bombycoid species. Furthermore, the mitogenome sequences were used to infer phylogenetic relationships among bombycoid species. The 15,261-bp Saturnia jonasii mitogenome contained the typical sets of genes and gene arrangements found in majority of Lepidoptera. All Bombycoidea species, including Saturnia jonasii, have a 15-33-bp spacer sequence at the trnS2-ND1 junction. The phylogenetic reconstruction of bombycoid species consistently and strongly supported monophylies of the families, Saturniidae, Bombycidae, and Sphingidae, based on Bayesian inference (BI) and maximum-likelihood (ML) methods. Among these families, the Bombycidae and Sphingidae species consistently showed a sister relationship, regardless of data partitions; the BI method strongly supported this relationship, whereas it was moderately supported using the ML method.

Analysis on the Complete Mitochondrial Genome of Andraca theae (Lepidoptera: Bombycoidea).

The bombycid moth, Andraca theae (Matsumura) (Lepidoptera: Bombycoidea) is an important pest of tea in southeastern China. In the present study, the mitochondrial genome (mitogenome) of A. theae was amplified by polymerase chain reaction and sequenced. The complete mitogenome of A. theae, encoding 37 genes, was 15,737 bp in length (Genbank no. KX365419), and consisted of 13 protein-coding genes (PCGs), 22 tRNA genes, 2 ribosomal RNA genes and an adenine (A) + thymine (T)-rich region (AT-rich region). The gene order of A. theae mitogenome was typical for Lepidoptera mitogenomes. Except for cox1, which started with CGA, all other 12 PCGs started with ATN. Eleven of the 13 PCGs ended with TAA, expect for cox1 and cox2, which ended with a single T. The maximum likelihood method and the Bayesian method were used to analyze the phylogenetic relationship among 22 representative bombycoid species with a matrix consisting of the 13 PCGs of the mitogenomes of the 22 species. The topological structures of the two phylogenetic trees we constructed were almost identical, with the results indicating that the bombycid species, including A. theae, clustered into a single clade with a bootstrap value of 58% and a posterior probability of 0.98. The phylogenetic relationship among the Bombycoidea species analyzed was Lasiocampidae + (Bombycidae + (Saturniidae + Sphingidae)) which was supported by a high bootstrap value of 100% and a posterior probability of 1.00.

Long read genome assembly of Automeris io (Lepidoptera: Saturniidae) an emerging model for the evolution of deimatic displays.

Automeris moths are a morphologically diverse group with 135 described species that have a geographic range that spans from the New World temperate zone to the Neotropics. Many Automeris have elaborate hindwing eyespots that are thought to deter or disrupt the attack of potential predators, allowing the moth time to escape. The Io moth (Automeris io), known for its striking eyespots, is a well-studied species within the genus and is an emerging model system to study the evolution of deimatism. Existing research on the eyespot pattern development will be augmented by genomic resources that allow experimental manipulation of this emerging model. Here, we present a high-quality, PacBio HiFi genome assembly for Io moth to aid existing research on the molecular development of eyespots and future research on other deimatic traits. This 490 Mb assembly is highly contiguous (N50 = 15.78 mbs) and complete (benchmarking universal single-copy orthologs = 98.4%). Additionally, we were able to recover orthologs of genes previously identified as being involved in wing pattern formation and movement.

An updated checklist of the wild silkmoths (Lepidoptera, Saturniidae) of Colombia.

In recent years, the study of wild silkmoths (Lepidoptera: Saturniidae) has increased exponentially due to the intense sampling effort and the use of molecular evidence for species delimitation, which led to the description of numerous new taxa especially from the Neotropic. Given these rapid advances, the checklist of the Colombian Saturniidae needs to be updated to cover the taxonomy, distribution, and diversity of these moths in the country. After an extensive review of literature, data repositories, and collections, an updated and comprehensive list of Saturniidae from Colombia is presented, including their occurrence status in each Colombian department. The checklist includes 7 subfamilies, 55 genera, and 790 taxa (766 in species rank) of Saturniidae in Colombia. Current distribution data show that the genus Winbrechlinia, the subgenusDarylesia, 379 species, and 18 subspecies are endemic to Colombia. Moreover, a dichotomic key to the Colombian subfamilies is provided. A few taxonomic changes are proposed based on a thorough taxonomic revision of the Colombian taxa. This revision also addresses the issue of outdated species names reported in the first checklist of Colombian Saturniidae (Amarillo-Suárez 2000) and excludes old records of taxa that are considered dubious for Colombia based on new evidence. By presenting an updated list of Colombian species, including the newly described taxa, this study aims at eliminating confusion stemming from outdated names and provides a useful resource for researching and conservating Saturniidae in Colombia. We wish to offer a common reference for future studies on the biodiversity and biogeography of moths in the Neotropical realm.

Characterization of the complete mitochondrial genome sequence of Smerinthus caecus (Lepidoptera: Sphingidae).

In this study, we sequenced and analyzed the complete mitogenome of Smerinthus caecus Ménétriés, 1857. The mitogenome of S. caecus is a circular structure, and 15,363 bp long in size and encodes 13 protein-coding genes (PCGs), two ribosomal RNA genes (rRNAs), 22 transfer RNA genes (tRNAs), and a control region (CR). An extremely high AT bias of 79.2% was found in the nucleotide composition of mitogenome. Most of the PCGs used ATN as the start codon and TAA or TAG as the stop codon, which is similar to most other insect mitogenomes, except cox1, which starts with CGA. The phylogeny of Smerinthinae was reconstructed using a maximum-likelihood method, a total of 33 mitogenomes were sampled for phylogenetic analyses. The subfamily Langiinae was selected as outgroup. The results confirmed the position of S. caecus in the Smerinthinae, in which Smerinthus caecus was placed as the sister taxon to Smerinthus planus, then to Laothoe amurensis.

Next-generation sequencing yields the complete mitochondrial genome of Rhodoprasina callantha (Lepidoptera: Sphingidae) and its evolutionary status.

In this study, we sequenced and analyzed the complete mitochondrial genome of Rhodoprasina callantha Jordan, 1929. The complete mitochondrial genome sequence of R. callantha was 15,343 bp in size and encoded 13 protein-coding genes (PCGs), two ribosomal RNA genes (rRNA), 22 transfer RNA genes (tRNA) and one control region (CR). The nucleotide composition of mitogenome was highly biased to A and T. Most protein-coding genes (PCGs) initiate with the standard start codon of ATN and terminate with the typical stop codon TAA/TAG. The phylogeny of Sphingidae based on nucleotide sequences of PCGs recovered the monophyly of subfamilies of Sphingidae with high support values. Langinae was the first subfamily diverged in Sphingidae, which was in accordance with previous study. R. callantha was the member of subfamily Smerinthinae, and closely related to the genus Marumba.

The complete mitochondrial genome sequence of the hawkmoth, Ambulyx tobii (Lepidoptera: Sphingidae) and phylogenetic analysis.

In this study, we sequenced and analyzed the complete mitochondrial genome of Ambulyx tobii to compare mitochondrial genome structures and reconstruct phylogenetic relationships. The complete mitochondrial genome sequence of A. tobii is circular, 15,343 bp in size and encodes 13 protein-coding genes (PCGs), two ribosomal RNA genes (rRNAs), 22 transfer RNA genes (tRNAs), and a control region (CR). Nucleotide composition is highly biased toward A + T nucleotides (81.2%). Most PCGs initiate with the standard start codon of ATN and terminate with the typical stop codon TAA/TAG. Phylogenetic analyses were performed using both 13 PCGs and whole mitochondrial genomes showed that A. tobii is closely related to A. substrigilis.

Sequencing and analysis of the complete mitochondrial genome of Laothoe amurensis sinica (Lepidoptera: Sphingidae) from China and its phylogenetic analysis.

The complete mitochondrial genome (mitogenome) of Laothoe amurensis sinica (Rothschild & Jordan, 1903) was sequenced. The L. amurensis sinica mitogenome is circular, double-stranded, with length of 15,341 bp. Gene content, gene order and orientation are all typical of Sphingidae. Nucleotide composition is highly biased toward A + T nucleotides (79.2%). Most of 13 protein-coding genes (PCGs) initiate with the standard start codon of ATN and terminate with the typical stop codon TAA/TAG or incomplete T. Phylogenetic analyses based on the maximum-likelihood (ML) on the W-IQ-Tree web server showed that L. amurensis sinica have a close relationship to the lineage formed by Clanis bilineata and Leucophlebia lineata.

Phylogenetic relationship and characterization of the complete mitochondrial genome sequence of Smerinthus planus (Lepidoptera: Sphingidae).

In this study, the complete mitochondrial genome of Smerinthus planus Walker, 1856 was sequenced and analyzed. This mitochondrial genome is circular, 15,375 bp long, and includes 37 typical metazoan mitochondrial genes (13 protein-coding genes (PCGs), two ribosomal RNA genes, and 22 transfer RNA genes) and an A + T-rich region. Nucleotide composition is highly biased toward A + T nucleotides (80.1%). All 13 PCGs initiate with the standard start codon of ATN and terminate with the typical stop codon TAA/TAG. Phylogenetic analyses were performed using amino acids of 13 PCGs which shows that S. planus is closely related to Barbourion lemaii.

Complete mitochondrial genome of Kentrochrysalis streckeri (Lepidoptera: Sphingidae) and phylogenetic analysis.

In this study, we sequenced the complete mitogenome of Kentrochrysalis streckeri (Staudinger, 1880). The complete mitogenome sequence of K. streckeri is circular, 15,253 bp in size and contains 13 protein-coding genes (PCGs), two ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, and a control region (CR). Nucleotide composition was A + T biased, and all the PCGs exhibited a positive AT-skew, which was reflected in the nucleotide composition, codon, and amino acid usage. Most PCGs start with ATG or ATT and stop with TAA. However, COX1 gene starts with CGA and three genes (COX1, COX2, NAD5) use the incomplete stop codon T. Phylogenetic analyses showed that the relationship (K. streckeri+((Manduca sexta+Sphinx morio)+(Psilogramma increta+(Psilogramma menephron+Notonagemia analis scribae)))).

The Primary Complete Mitochondrial Genome of the Lappet Moth Brahmophthalma hearseyi (Lepidoptera: Brahmaeidae) and Related Phylogenetic Analysis.

Background: Brahmophthalma hearseyi (Lepidoptera: Brahmaeidae) is widely distributed across China. Its larvae damage the leaves of many plants such as those belonging to the Oleaceae family, causing significant economic losses and seriously affecting the survival and reproduction of Cervus nippon; however, genetic data for this species are scarce. Methods: The complete mitochondrial genome (mitogenome) of B. hearseyi was sequenced using long-PCR and primer-walking methods. Phylogenetic analysis that was based on 13 PCGs and two rRNAs was carried out using the neighbor-joining and Bayesian interference methods. Results: The mitogenome is a typical circular molecule that is made up of 15,442 bp, which includes 13 protein-coding genes (PCGs), 2 ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, and an A + T-rich region (456 bp). All of the PCGs, except for COX1 and COX2, start with ATN codons. COX2 and ND5 use the incomplete termination codon T, and 11 other PCGs use the typical stop codon TAA. All tRNA genes, except for trnS1 and trnS2, display a typical cloverleaf structure; trnS1 lacks the "DHU" arm, whereas trnS2 exhibits two mismatched base pairs in the anticodon stem. Phylogenetic analysis showed that B. hearseyi is clustered into Brahmaeidae, and the phylogenetic relationships are (Brahmaeidae + Lasiocampidae) + (Bombycidae + (Sphingidae + Saturniidae)). Conclusions: This study provides the first mitogenomic resources for the Brahmaeidae.

A global checklist of the Bombycoidea (Insecta: Lepidoptera).

BACKGROUND: Bombycoidea is an ecologically diverse and speciose superfamily of Lepidoptera. The superfamily includes many model organisms, but the taxonomy and classification of the superfamily has remained largely in disarray. Here we present a global checklist of Bombycoidea. Following Zwick (2008) and Zwick et al. (2011), ten families are recognized: Anthelidae, Apatelodidae, Bombycidae, Brahmaeidae, Carthaeidae, Endromidae, Eupterotidae, Phiditiidae, Saturniidae and Sphingidae. The former families Lemoniidae and Mirinidae are included within Brahmaeidae and Endromidae respectively. The former bombycid subfamilies Oberthueriinae and Prismostictinae are also treated as synonyms of Endromidae, and the former bombycine subfamilies Apatelodinae and Phitditiinae are treated as families. NEW INFORMATION: This checklist represents the first effort to synthesize the current taxonomic treatment of the entire superfamily. It includes 12,159 names and references to their authors, and it accounts for the recent burst in species and subspecies descriptions within family Saturniidae (ca. 1,500 within the past 10 years) and to a lesser extent in Sphingidae (ca. 250 species over the same period). The changes to the higher classification of Saturniidae proposed by Nässig et al. (2015) are rejected as premature and unnecessary. 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).

Complete mitochondrial genome of the hawkmoth Notonagemia analis scribae (Lepidoptera: Sphingidae).

Currently, the mitochondrial genome of only two species of Sphingidae have been completely sequenced. For the phylogenetic study of Bombycoidea (including Bombycidae, Saturniidae and Sphingidae) using mitochondrial genomes (mitogenomes), more species are required as a basis for future research. In the present study, we sequenced the complete mitogenome of the hawkmoth, Notonagemia analis scribae (Lepidoptera: Sphingidae), to enrich the Sphingidae database. The length of the N. a. scribae genome was 15,303 bp with a typical set of genes (13 protein-coding genes [PCGs], 2 rRNA genes, and 22 tRNA genes), and one major non-coding A + T-rich region. The COI gene had a CGA start codon, which is the start codon for this gene in the majority of lepidopteran species, whereas other PCGs began with ATN codons. A 318-bp A + T-rich region harbored the blocks of conserved sequences that are typically found in lepidopteran insects, excluding a poly-A stretch, which is typically found at the end of the A + T-rich region. Phylogenetic analysis using the 13 PCGs indicated that N. a. scribae grouped together with two within-familial species, Sphinx morio and Manduca sexta, with the highest nodal support both by Bayesian inference and maximum-likelihood methods, forming the Sphingidae monophyletic group.

Phylotranscriptomics: saturated third codon positions radically influence the estimation of trees based on next-gen data.

Recent advancements in molecular sequencing techniques have led to a surge in the number of phylogenetic studies that incorporate large amounts of genetic data. We test the assumption that analyzing large number of genes will lead to improvements in tree resolution and branch support using moths in the superfamily Bombycoidea, a group with some interfamilial relationships that have been difficult to resolve. Specifically, we use a next-gen data set that included 19 taxa and 938 genes (∼1.2M bp) to examine how codon position and saturation might influence resolution and node support among three key families. Maximum likelihood, parsimony, and species tree analysis using gene tree parsimony, on different nucleotide and amino acid data sets, resulted in largely congruent topologies with high bootstrap support compared with prior studies that included fewer loci. However, for a few shallow nodes, nucleotide and amino acid data provided high support for conflicting relationships. The third codon position was saturated and phylogenetic analysis of this position alone supported a completely different, potentially misleading sister group relationship. We used the program RADICAL to assess the number of genes needed to fix some of these difficult nodes. One such node originally needed a total of 850 genes but only required 250 when synonymous signal was removed. Our study shows that, in order to effectively use next-gen data to correctly resolve difficult phylogenetic relationships, it is necessary to assess the effects of synonymous substitutions and third codon positions.

Esfingofauna (Lepidoptera: Sphingidae) do Parque Nacional do Jaú e arredores, Amazonas, Brasil

A survey of Sphingidae fauna (Lepidoptera, Sphingidae) were studied at Parque Nacional do Jaú and surroundings, Amazonas, Brazil during july/1993 to june/1996. The collects were made using a mixed mercury vapor lamp and black light UV on a white sheet in periods of 12 consecutive hours. 79 hawkmoths species, 25 genera in 2362 specimens were collected with predominance of the species of tribe Dilophonotini (45 species), followed by Sphingini (13), Macroglossini (8), Philampelini (7) and Ambulycini (6). Data are compared with others areas of brazilian Amazonia: municipal district of Itacoatiara (AM), Ilha de Maracá and Pacaraima (RR). It is presented a list of species of hawk moths including 16 new register for the State of Amazonas and equal for all localities were collected.

A fauna de esfingídeos (Lepidoptera, Sphingidae) foi estudada no Parque Nacional do Jaú e arredores, Amazonas, Brasil, durante o período de julho de 1993 a junho de 1996. O método empregado para as coletas foi luz mista de mercúrio e luz negra UV montadas sobre um lençol branco em períodos de 12 horas consecutivas. Foram coletados 2362 espécimes de 25 gêneros e 79 espécies, com a predominância de espécies das tribos Dilophonotini (45 espécies), seguida por Sphingini (13 spp.), Macroglossini (8 spp.), Philampelini (7 spp.) e Ambulycini (6 spp.). Os dados são comparados a outras áreas da Amazônia brasileira: Município de Itacoatiara (AM), Ilha de Maracá e Pacaraima (RR). Apresenta-se uma lista das espécies de esfingídeos incluindo 16 registros novos para o Estado do Amazonas e igualmente para todas as localidades em que foram coletadas.