Opsin proteins are fundamental components of animal vision whose structure largely determines the sensitivity of visual pigments to different wavelengths of light. Surprisingly little is known about opsin evolution in beetles, even though they are the most species rich animal group on Earth and exhibit considerable variation in visual system sensitivities. We reveal the patterns of opsin evolution across 62 beetle species and relatives. Our results show that the major insect opsin class (SW) that typically confers sensitivity to "blue" wavelengths was lost ~300 million years ago, before the origin of modern beetles. We propose that UV and LW opsin gene duplications have restored the potential for trichromacy (three separate channels for colour vision) in beetles up to 12 times and more specifically, duplications within the UV opsin class have likely led to the restoration of "blue" sensitivity up to 10 times. This finding reveals unexpected plasticity within the insect visual system and highlights its remarkable ability to evolve and adapt to the available light and visual cues present in the environment.At the molecular level, the wavelength sensitivity of an animal photoreceptor is determined by the photopigment, comprising an opsin protein bound to a light-absorbing chromophore. Insects commonly possess three opsin proteins (UV, SW and LW) that form photopigments maximally sensitive to ultraviolet (~350 nm), blue (~440 nm) and green (~530 nm) wavelengths, respectively. As insect opsin genes form distinct phylogenetic clades according to their spectral class (UV, SW or LW) the sensitivity ranges of an insect visual system can usually be estimated by the complement of opsin genes present. In some insects photopigment sensitivity has extended outside of this range into the violet (~420 nm) and red (>600 nm) region of the light spectrum through duplications of the SW 1,2 and LW opsins 2,3

Fireflies (Lampyridae Rafinesque) are a diverse family of beetles which exhibit an array of morphologies including varying antennal and photic organ features. Due in part to their morphological diversity, the classification within the Lampyridae has long been in flux. Here we use an anchored hybrid enrichment approach to reconstruct the most extensive molecular phylogeny of Lampyridae to date (436 loci and 98 taxa) and use this phylogeny to evaluate the higher-level classification of the group. None of the currently recognized subfamilies were recovered as monophyletic with high support. We propose several classification changes supported by both phylogenetic and morphological evidence: 1) Pollaclasis Newman, Vestini McDermott (incl. Vesta Laporte, Dodacles Olivier, Dryptelytra Laporte, and Ledocas Olivier), Photoctus McDermott, and Araucariocladus Silveira & Mermudes are transferred to Lampyridae incertae sedis, 2) Psilocladinae Mcdermott, 1964status novum is reestablished for the genus Psilocladus Blanchard, 3) Lamprohizini Kazantsev, 2010 is elevated to Lamprohizinae Kazantsev, 2010status novum and Phausis LeConte is transferred to Lamprohizinae, 4) Memoan Silveira and Mermudes is transferred to Amydetinae Olivier, and 5) Scissicauda McDermott is transferred to Lampyrinae Rafinesque.

Fireflies and their luminous courtships have inspired centuries of scientific study. Today firefly luciferase is widely used in biotechnology, but the evolutionary origin of bioluminescence within beetles remains unclear. To shed light on this long-standing question, we sequenced the genomes of two firefly species that diverged over 100 million-years-ago: the North American Photinus pyralis and Japanese Aquatica lateralis. To compare bioluminescent origins, we also sequenced the genome of a related click beetle, the Caribbean Ignelater luminosus, with bioluminescent biochemistry near-identical to fireflies, but anatomically unique light organs, suggesting the intriguing hypothesis of parallel gains of bioluminescence. Our analyses support independent gains of bioluminescence in fireflies and click beetles, and provide new insights into the genes, chemical defenses, and symbionts that evolved alongside their luminous lifestyle.

Fireflies are some of the most captivating organisms on the planet. They have a rich history as subjects of scientific study, especially in relation to their bioluminescent behavior. Yet, the phylogenetic relationships of fireflies are still poorly understood. Here, we present the first total evidence approach to reconstruct lampyrid phylogeny using both a molecular matrix from six loci and an extensive morphological matrix. Using this phylogeny we test the hypothesis that adult bioluminescence evolved after the origin of the firefly clade. The ancestral state of adult bioluminescence is recovered as non-bioluminescent with one to six gains and five to ten subsequent losses. The monophyly of the family, as well as the subfamilies is also tested. Ototretinae, Cyphonocerinae, Luciolinae (incl. Pristolycus), Amydetinae, "cheguevarinae" sensu Jeng 2008, and Photurinae are highly supported as monophyletic. With the exception of four taxa, Lampyrinae is also recovered as monophyletic with high support. Based on phylogenetic and morphological data Lamprohiza, Phausis, and Lamprigera are transferred to Lampyridae incertae sedis.

SummaryFireflies are among the best-studied of the bioluminescent organisms. Despite longterm interest in the biochemistry, neurobiology, and evolution of firefly flash signals and the widespread biotechnological applications of firefly luciferase, only a limited set of genes related to this complex trait have been described. To investigate the genetic basis of firefly bioluminescence, we generated a high-quality reference genome for the Big Dipper firefly Photinus pyralis, from which the first laboratory luciferase was cloned, using long-read (PacBio), short-read (Illumina), and Hi-C sequencing technologies. To facilitate comparative genomics, we also generated short-read genome assemblies for a Japanese firefly Aquatica lateralis and a bioluminescent click beetle, Ignelater luminosus. Analyses of these genomic datasets supports at least two independent gains of luminescence in beetles, and provides new insights into the evolution of beetle bioluminescence and chemical defenses that likely co-evolved over their 100 million years of evolution.