Abstract Fireflies and their fascinating luminous courtships have inspired centuries of scientific study. Today firefly luciferase is widely used in biotechnology, but the evolutionary origin of their bioluminescence 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. 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 but contentious hypothesis of parallel gains of bioluminescence. Our analyses support two independent gains of bioluminescence between fireflies and click-beetles, and provide new insights into the genes, chemical defenses, and symbionts that evolved alongside their luminous lifestyle. One Sentence Summary: Comparative analyses of the first linkage-group-resolution genomes of fireflies and related bioluminescent beetles address long-standing questions of the origin and evolution of bioluminescence and its associated traits.
Abstract Firefly flashes are well-known visual signals used by these insects to find, identify, and choose mates. However, many firefly species have lost the ability to produce light as adults. These “unlighted” species generally lack developed adult light organs, are diurnal rather than nocturnal, and are believed to use volatile pheromones acting over a distance to locate mates. While cuticular hydrocarbons, which may function in mate recognition at close range, have been examined for a handful of the over 2000 extant firefly species, no volatile pheromone has ever been identified. In this study, using coupled gas chromatography - electroantennographic detection, we detected a single female-emitted compound that elicited antennal responses from wild-caught male winter fireflies, Photinus corrusca . The compound was identified as (1 S )- exo -3-hydroxycamphor (hydroxycamphor). In field trials at two sites across the species’ eastern North American range, large numbers of male P. corrusca were attracted to synthesized hydroxycamphor, verifying its function as a volatile sex attractant pheromone. Males spent more time in contact with lures treated with synthesized hydroxycamphor than those treated with solvent only in laboratory two-choice assays. Further, using single sensillum recordings, we characterized a pheromone-sensitive odorant receptor neuron in a specific olfactory sensillum on male P. corrusca antennae and demonstrated its sensitivity to hydroxycamphor. Thus, this study has identified the first volatile pheromone and its corresponding sensory neuron for any firefly species, and provides a tool for monitoring P. corrusca populations for conservation, and further inquiry into the chemical and cellular bases for sexual communication among fireflies.
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.
Identifying the basis of phenotypic variation is a key objective of genetics. This work has been mostly limited to model systems with a plethora of genetic manipulation and functional characterization tools. With the development of high-throughput sequencing and new computational tools, it is possible to identify candidate genes related to phenotypic variation in non-model organisms. Fireflies are excellent for studying phenotypic variation because of their diverse and well-characterized behaviors. Most adult fireflies emit a single mating flash pattern and do not eat. In contrast, adult females of many species in the genus Photuris employ multiple flash patterns and prey upon mate-seeking males of other firefly species. To investigate the genetic basis for this variation, we used comparative transcriptomics to identify positively selected genes between a predatory firefly, Photuris sp., and a non-predatory relative, Photuris frontalis, controlling for genes generally under selection in fireflies by comparing to a Photinus firefly. Nine gene families were identified under positive selection in the predatory versus non-predatory Photuris comparison, including genes involved in digestion, detoxification, vision, reproduction, and neural processes. These results generate intriguing hypotheses about the genetic basis for insect behavior and highlight the utility of comparative transcriptomic tools to investigate complex behaviors in non-model systems.
Most organisms are dependent on sensory cues from their environment for survival and reproduction. Fireflies (Coleoptera: Lampyridae) represent an ideal system for studying sensory niche adaptation due to many species relying on bioluminescent communication; as well as a diversity of ecologies. Here; using transcriptomics; we examine the phototransduction pathway in this non-model organism; and provide some of the first evidence for positive selection in the phototransduction pathway beyond opsins in beetles. Evidence for gene duplications within Lampyridae are found in inactivation no afterpotential C and inactivation no afterpotential D. We also find strong support for positive selection in arrestin-2; inactivation no afterpotential D; and transient receptor potential-like; with weak support for positive selection in guanine nucleotide-binding protein G(q) subunit alpha and neither inactivation nor afterpotential C. Taken with other recent work in flies; butterflies; and moths; this represents an exciting new avenue of study as we seek to further understand diversification and constraint on the phototransduction pathway in light of organism ecology.
Abstract The firefly Photinus pyralis inhabits a wide range of latitudinal and ecological niches, with populations living from temperate to tropical habitats. Despite its broad distribution, its demographic history is unknown. In this study, we modelled and inferred different demographic scenarios for North American populations of P. pyralis , which were collected from Texas to New Jersey. We used a combination of A BC techniques (for multi‐population/colonization analyses) and likelihood inference ( dadi, StairwayPlot2 , PoMo) for single‐population demographic inference, which proved useful with our RAD data. We uncovered that the most ancestral North American population lays in Texas, which further colonized the Central region of the US and more recently the North Eastern coast. Our study confidently rejects a demographic scenario where the North Eastern populations colonized more southern populations until reaching Texas. To estimate the age of divergence between of P. pyralis , which provides deeper insights into the history of the entire species, we assembled a multi‐locus phylogenetic data covering the genus Photinus. We uncovered that the phylogenetic node leading to P. pyralis lies at the end of the Miocene. Importantly, modelling the demographic history of North American P. pyralis serves as a null model of nucleotide diversity patterns in a widespread native insect species, which will serve in future studies for the detection of adaptation events in this firefly species, as well as a comparison for future studies of other North American insect taxa.
Previous research suggests that fireflies (Coleoptera: Lampyridae) are susceptible to commonly used insecticides. In the United States, there has been a rapid and widespread adoption of neonicotinoid insecticides, predominantly used as seed coatings on large-acreage crops like corn, soy, and cotton. Neonicotinoid insecticides are persistent in soil yet mobile in water, so they have potential to contaminate firefly habitats both in and adjacent to application sites. As a result, fireflies may be at high risk of exposure to neonicotinoids, possibly jeopardizing this already at-risk group of charismatic insects.To assess the sensitivity of fireflies to neonicotinoids, we exposed larvae of Photuris versicolor complex and Photinus pyralis to multiple levels of clothianidin-treated soil and monitored feeding behavior, protective soil chamber formation, intoxication, and mortality.Pt. versicolor and Pn. pyralis larvae exhibited long-term intoxication and mortality at concentrations above 1,000 ng g-1 soil (1 ppm). Under sub-lethal clothianidin exposure, firefly larvae fed less and spent less time in protective soil chambers, two behavioral changes that could decrease larval survival in the wild.Both firefly species demonstrated sub-lethal responses in the lab to clothianidin exposure at field-realistic concentrations, although Pt. versicolor and Pn. pyralis appeared to tolerate higher clothianidin exposure relative to other soil invertebrates and beetle species. While these two firefly species, which are relatively widespread in North America, appear somewhat tolerant of neonicotinoid exposure in a laboratory setting, further work is needed to extend this conclusion to wild populations, especially in rare or declining taxa.
Abstract Fireflies (Coleoptera: Lampyridae) consist of over 2,000 described extant species. A well-resolved phylogeny of fireflies is important for the study of their population genetics, bioluminescence, evolution, and conservation. We used a recently published anchored hybrid enrichment dataset (AHE; 436 loci for 88 Lampyridae species and 10 outgroup species) and state-of-the-art statistical methods (the fossilized birth-death-range process implemented in a Bayesian framework) to estimate a time-calibrated phylogeny of Lampyridae. Unfortunately, estimating calibrated phylogenies using AHE and the latest and most robust time-calibration strategies is not possible because of computational constraints. As a solution, we subset the full dataset by applying three different strategies: (i) using the most complete loci, (ii) using the most homogeneous loci, and (iii) using the loci with the highest accuracy to infer the well established Photinus clade. The estimated topology using the three data subsets agreed on almost all major clades and only showed minor discordance within less supported nodes. The estimated divergence times overlapped for all nodes that are shared between the topologies. Thus, divergence time estimation is robust as long as the topology inference is robust and any well selected data subset suffices. Additionally, we observed an un-expected amount of gene tree discordance between the 436 AHE loci. Our assessment of model adequacy showed that standard phylogenetic substitution models are not adequate for any of the 436 AHE loci which is likely to bias phylogenetic inferences. We performed a simulation study to explore the impact of (a) incomplete lineage sorting, (b) uniformly distributed and systematic missing data, and (c) systematic bias in the position of highly variable and conserved sites. For our simulated data, we observed less gene tree variation which shows that the empirically observed amount of gene tree discordance for the AHE dataset is unexpected and needs further investigation.
The mutational patterns of large tandem arrays of short sequence repeats remain largely unknown, despite observations of their high levels of variation in sequence and genomic abundance within and between species. Many factors can influence the dynamics of tandem repeat evolution; however, their evolution has only been examined over a limited phylogenetic sample of taxa. Here, we use publicly available whole-genome sequencing data of 85 haploid mutation accumulation lines derived from six geographically diverse Chlamydomonas reinhardtii isolates to investigate genome-wide mutation rates and patterns in tandem repeats in this species. We find that tandem repeat composition differs among ancestral strains, both in genome-wide abundance and presence/absence of individual repeats. Estimated mutation rates (repeat copy number expansion and contraction) were high, averaging 4.3×10−4 per generation per single unit copy. Although orders of magnitude higher than other types of mutation previously reported in C. reinhardtii, these tandem repeat mutation rates were one order of magnitude lower than what has recently been found in Daphnia pulex, even after correcting for lower overall genome-wide satellite abundance in C. reinhardtii. Most high-abundance repeats were related to others by a single mutational step. Correlations of repeat copy number changes within genomes revealed clusters of closely related repeats that were strongly correlated positively or negatively, and similar patterns of correlation arose independently in two different mutation accumulation experiments. Together, these results paint a dynamic picture of tandem repeat evolution in this unicellular alga.
