Publications

2018
McKenna DD, Clarke D, Anderson R, Astrin J, Brown S, Chamorro L, Davis S, de Medeiros BAS, del Rio M, Haran J, et al. Morphological and Molecular Perspectives on the Phylogeny, Evolution, and Classification of Weevils (Coleoptera: Curculionoidea): Proceedings from the 2016 International Weevil Meeting. Diversity. 2018;10 :64.
de Medeiros BAS, Farrell BD. Whole-genome amplification in double-digest RADseq results in adequate libraries but fewer sequenced loci. PeerJP [Internet]. 2018;(6) :e5089. DOI (full text)Abstract
Whole-genome amplification by multiple displacement amplification (MDA) is a promising technique to enable the use of samples with only limited amount of DNA for the construction of RAD-seq libraries. Previous work has shown that, when the amount of DNA used in the MDA reaction is large, double-digest RAD-seq (ddRAD) libraries prepared with amplified genomic DNA result in data that are indistinguishable from libraries prepared directly from genomic DNA. Based on this observation, here we evaluate the quality of ddRAD libraries prepared from MDA-amplified genomic DNA when the amount of input genomic DNA and the coverage obtained for samples is variable. By simultaneously preparing libraries for five species of weevils (Coleoptera, Curculionidae), we also evaluate the likelihood that potential contaminants will be encountered in the assembled dataset. Overall, our results indicate that MDA may not be able to rescue all samples with small amounts of DNA, but it does produce ddRAD libraries adequate for studies of phylogeography and population genetics even when conditions are not optimal. We find that MDA makes it harder to predict the number of loci that will be obtained for a given sequencing effort, with some samples behaving like traditional libraries and others yielding fewer loci than expected. This seems to be caused both by stochastic and deterministic effects during amplification. Further, the reduction in loci is stronger in libraries with lower amounts of template DNA for the MDA reaction. Even though a few samples exhibit substantial levels of contamination in raw reads, the effect is very small in the final dataset, suggesting that filters imposed during dataset assembly are important in removing contamination. Importantly, samples with strong signs of contamination and biases in heterozygosity were also those with fewer loci shared in the final dataset, suggesting that stringent filtering of samples with significant amounts of missing data is important when assembling data derived from MDA-amplified genomic DNA. Overall, we find that the combination of MDA and ddRAD results in high-quality datasets for population genetics as long as the sequence data is properly filtered during assembly.
Kim SI, de Medeiros BAS, Byun B-K, Lee S, Kang J-H, Lee B, Farrell BD. West meets East: How do rainforest beetles become circum-Pacific? Evolutionary origin of Callipogon relictus and allied species (Cerambycidae: Prioninae) in the New and Old Worlds. Molecular Phylogenetics and Evolution [Internet]. 2018;125 :163-176. DOI (full text)Abstract
The longhorn beetle genus Callipogon Audinet-Serville represents a small group of large wood-boring beetles whose distribution pattern exhibits a unique trans-Pacific disjunction between the East Asian temperate rainforest and the tropical rainforest of the Neotropics. To understand the biogeographic history underlying this circum-Pacific disjunct distribution, we reconstructed a molecular phylogeny of the subfamily Prioninae with extensive sampling of Callipogon using multilocus sequence data of 99 prionine and four parandrine samples (ingroups), together with two distant outgroup species. Our sampling of Callipogon includes 18 of the 24 currently accepted species, with complete representation of all species in our focal subgenera. Our phylogenetic analyses confirmed the purported affinity between the Palearctic Callipogon relictus and its Neotropical congeners. Furthermore, based on molecular dating under the fossilized birth-death (FBD) model with comprehensive fossil records and probabilistic ancestral range reconstructions, we estimated the crown group Callipogon to have originated in the Paleocene circa 60 million years ago (Ma) across the Neotropics and Eastern Palearctics. The divergence between the Palearctic C. relictus and its Neotropical congeners is explained as the result of a vicariance event following the demise of boreotropical forest across Beringia at the Eocene-Oligocene boundary. As C. relictus represents the unique relictual species that evidentiates the lineage's expansive ancient distribution, we evaluated its conservation importance through species distribution modelling. Though we estimated a range expansion for C. relictus by 2050, we emphasize a careful implementation of conservation programs towards the protection of primary forest across its current habitats, as the species remains highly vulnerable to habitat disturbance.
De-la-Mora M, Piñero D, Oyama K, Farrell BD, Magallón S, Núñez-Farfán J. Evolution of Trichobaris (Curculionidae) in relation to host plants: Geometric morphometrics, phylogeny and phylogeography. Molecular Phylogenetics and Evolution [Internet]. 2018;124 (July 2018) :37-49. DOI (full text)Abstract
The family Curculionidae (Coleoptera), the “true” weevils, have diversified tightly linked to the evolution of flowering plants. Here, we aim to assess diversification at a lower taxonomic level. We analyze the evolution of the genus Trichobaris in association with their host plants. Trichobaris comprises eight to thirteen species; their larvae feed inside the fruits of Datura spp. or inside the stem of wild and cultivated species of Solanaceae, such as potato, tobacco and tomato. We ask the following questions: (1) does the rostrum of Trichobaris species evolve according to the plant tissue used to oviposit, i.e., shorter rostrum to dig in stems and longer to dig in fruits? and (2) does Trichobaris diversify mainly in relation to the use of Datura species? For the first question, we estimated the phylogeny of Trichobaris based on four gene sequences (nuclear 18S and 28S rRNA genes and mitochondrial 16S rRNA and COI genes). Then, we carried out morphogeometric analyses of the Trichobaris species using 75 landmarks. For the second question, we calibrated a COI haplotype phylogeny using a constant rate of divergence to infer the diversification time of Trichobaris species, and we traced the host plant species on the haplotype network. We performed an ancestral state reconstruction analysis to infer recent colonization events and conserved associations with host plant species. We found that ancestral species in the Trichobaris phylogeny use the stem of Solanum plants for oviposition and display weak sexual dimorphism of rostrum size, whereas other, more recent species of Trichobaris display sexual dimorphism in rostrum size and use the fruits of Datura species, and a possible reversion to use the stem of Solanaceae was detected in one Trichobaris species. The use of Datura species by Trichobaris species is widely distributed on haplotype networks and restricted to Trichobaris species that originated ca. 5 ± 1.5 Ma. Given that the origin of Trichobaris is estimated to be ca. 6 ± 1.5 Ma, it is likely that Datura has played a role in its diversification.
Shin S, Clarke DJ, Lemmon AR, Lemmon EM, Aitken AL, Haddad S, Farrell BD, Marvaldi AE, Oberprieler RG, McKenna DD. Phylogenomic Data Yield New and Robust Insights into the Phylogeny and Evolution of Weevils. Molecular Biology and Evolution [Internet]. 2018;35 (4) :823-836. DOI (full text)Abstract
The phylogeny and evolution of weevils (the beetle superfamily Curculionoidea) has been extensively studied, but many relationships, especially in the large family Curculionidae (true weevils; > 50,000 species), remain uncertain. We used phylogenomic methods to obtain DNA sequences from 522 protein-coding genes for representatives of all families of weevils and all subfamilies of Curculionidae. Most of our phylogenomic results had strong statistical support, and the inferred relationships were generally congruent with those reported in previous studies, but with some interesting exceptions. Notably, the backbone relationships of the weevil phylogeny were consistently strongly supported, and the former Nemonychidae (pine flower snout beetles) were polyphyletic, with the subfamily Cimberidinae (here elevated to Cimberididae) placed as sister group of all other weevils. The clade comprising the sister families Brentidae (straight-snouted weevils) and Curculionidae was maximally supported and the composition of both families was firmly established. The contributions of substitution modeling, codon usage and/or mutational bias to differences between trees reconstructed from amino acid and nucleotide sequences were explored. A reconstructed timetree for weevils is consistent with a Mesozoic radiation of gymnosperm-associated taxa to form most extant families and diversification of Curculionidae alongside flowering plants—first monocots, then other groups—beginning in the Cretaceous.
2017
Laurindo da Silva F, Farrell BD. Non-biting midges (Diptera: Chironomidae) research in South America: subsidizing biogeographic hypotheses. Annales de Limnologie - International Journal of Limnology [Internet]. 2017;53 :111-128. DOI (full text)Abstract
We present data on Chironomidae (Insecta: Diptera) collected in South America together with results on the mitochondrial DNA diversity within selected megadiverse genera. This work is part of an on-going project on the ancient origin of South American biodiversity using non-biting midges. Collections were made at 42 localities, in March 2014 and February 2015, in a diverse array of habitats, including small streams, rivers, ponds, lakes and bays. In total, 3196 representatives of six subfamilies were collected. Sixty-one genera were identified, containing at least 211 species. The subfamilies Chironominae and Orthocladiinae predominated in all samples. Tanypodinae were often present, but rarely in large numbers. Except for Podonomus pepinellii, reported from Brazil, Podonominae were collected in a few localities in Argentina (Arroyo Lopez, and Arroyo Gutierrez and Gutierrez Lake) and Chile (Llanquihue Lake). Prodiamesinae were only recorded in Chile. Analysis of DNA barcode sequences using neighbor-joining estimation supported 66 species within the selected genera. The chironomid fauna of South America includes multiple genera with worldwide distributions, with Australian, Nearctic and Neotropical components.
de Medeiros BAS, Barghini A, Vanin SA. Streetlights attract a broad array of beetle species. Revista Brasileira de Entomologia [Internet]. 2017;61 (1) :74 - 79. DOI (full text)Abstract
Light pollution on ecosystems is a growing concern, and knowledge about the effects of outdoor lighting on organisms is crucial to understand and mitigate impacts. Here we build up on a previous study to characterize the diversity of all beetles attracted to different commonly used streetlight set ups. We find that lights attract beetles from a broad taxonomic and ecological spectrum. Lights that attract a large number of insect individuals draw an equally high number of insect species. While there is some evidence for heterogeneity in the preference of beetle species to different kinds of light, all species are more attracted to some light radiating ultraviolet. The functional basis of this heterogeneity, however, is not clear. Our results highlight that control of ultraviolet radiation in public lighting is important to reduce the number and diversity of insects attracted to lights.
deMedieros_streetlights_beetles.pdf
Haddad S, Shin S, Lemmon AR, Lemmon EM, Svacha P, Farrell BD, Ślipinski A, Windsor D, McKenna DD. Anchored hybrid enrichment provides new insights into the phylogeny and evolution of longhorned beetles (Cerambycidae). Systematic Entomology [Internet]. 2017. DOI (full text)Abstract
Cerambycidae is a species-rich family of mostly wood-feeding (xylophagous) beetles containing nearly 35,000 known species. The higher-level phylogeny of Cerambycidae has never been robustly reconstructed using molecular phylogenetic data or a comprehensive sample of higher taxa, and its internal relationships and evolutionary history remain the subjects of ongoing debate. We reconstructed the higher-level phylogeny of Cerambycidae using phylogenomic data from 522 single copy nuclear genes, generated via anchored hybrid enrichment. Our taxon sample included exemplars of all families and 23/30 subfamilies of Chrysomeloidea, with a focus on the large family Cerambycidae. Our results reveal a monophyletic Cerambycidae sensu stricto in all but one analysis, and a polyphyletic Cerambycidae sensu lato. When monophyletic, Cerambycidae sensu stricto was sister to the family Disteniidae. Relationships among the subfamilies of Cerambycidae sensu stricto were also recovered with strong statistical support except for Cerambycinae being made paraphyletic by Dorcasomus (Dorcasominae) in the nucleotide (but not amino acid) trees. Most other chrysomeloid families represented by more than one terminal taxon – Chrysomelidae, Disteniidae, Vesperidae, and Orsodacnidae – were monophyletic, but Megalopodidae was rendered paraphyletic by Cheloderus (Oxypeltidae). Our study corroborates some relationships within Chrysomeloidea that were previously inferred from morphological data, while also reporting several novel relationships. The present work thus provides a robust framework for future, more deeply taxon-sampled, phylogenetic and evolutionary studies of the families and subfamilies of Cerambycidae sensu lato and other Chrysomeloidea.
Peris D, Pérez-de la Fuente R, Peñalver E, Delclòs X, Barrón E, Labandeira CC. False Blister Beetles and the Expansion of Gymnosperm-Insect Pollination Modes before Angiosperm Dominance. Current Biology [Internet]. 2017;27 :897–904. DOI (full text)
2016
Pérez-de la Fuente R, Delclòs X, Peñalver E, Engel MS. A defensive behavior and plant-insect interaction in Early Cretaceous amber - The case of the immature lacewing Hallucinochrysa diogenesi. Arthropod Structure and Development [Internet]. 2016;45 (2) :133–139. DOI (full text)Abstract

Amber holds special paleobiological significance due to its ability to preserve direct evidence of biotic interactions and animal behaviors for millions of years. Here we review the finding of Hallucinochrysa diogenesi Pérez-de la Fuente, Delclòs, Peñalver and Engel, 2012, a morphologically atypical larva related to modern green lacewings (Insecta: Neuroptera) that was described in Early Cretaceous amber from the El Soplao outcrop (northern Spain). The fossil larva is preserved with a dense cloud of fern trichomes that corresponds to the trash packet the insect gathered and carried on its back for camouflaging and shielding, similar to that which is done by its extant relatives. This finding supports the prominent role of wildfires in the paleoecosystem and provides direct evidence of both an ancient planteinsect interaction and an early acquisition of a defensive behavior in an insect lineage. Overall, the fossil of H. diogenesi showcases the potential that the amber record offers to reconstruct not only the morphology of fossil arthropods but, more remarkably, their lifestyles and ecological relationships.

Farrell BD. The Biology of Consciousness. ReVista : Harvard Review of Latin America. [Internet]. 2016;16 (1) :26–29. Web version Biology of Consciousness (PDF)
Farrell BD. The Biology of Culture. ReVista : Harvard Review of Latin America [Internet]. 2016;16 (1) :2. Web version Biology of Culture (PDF)
Wiedenbrug S, Laurindo da Silva F. Diplosmittia caribensis, a new Orthocladiinae (Diptera: Chironomidae) from the Dominican Republic. Zootaxa [Internet]. 2016;4103 (1) :71–74. DOIAbstract

The genus Diplosmittia was erected by Sæther (1981) based on Diplosmittia harrisoni from St. Lucia and St. Vincent in the British West Indies. Prior to the present study the genus comprised nine species, all except D. carinata Sæther were known only from Neotropical Region (Ashe & O'Connor, 2012). During sampling in the surroundings of a highly organic polluted river, in the National Botanical Garden in Santo Domingo, Dominican Republic, the present second author collected several imagines of Diplosmittia that did not fit any taxon treated in the recent review of the genus (Pinho et al. 2009). In the present paper, the male of this new species is described and illustrated. Alcohol-preserved specimens were dissected and slide mounted in Euparal. Morphological terminology and abbreviations follow Sæther (1980) and Epler (1988). Measurement are taken according to Epler (1988). The holotype is deposited in the entomological collection of the Museum of Comparative Zoology (MCZ), Harvard University, USA and paratypes are deposited in the Museo Nacional de Historia Natural (MNHN), Dominican Republic and Zoologische Staatssammlung München (ZSM), Germany.

wiedenbrug_laurindo_da_silva_2016.pdf
Farrell BD. Music and Nature: From Humpbacks to Harvard. ReVista : Harvard Review of Latin America. 2016;15 (2) :2-4. Music & Nature 2016 (PDF)
2015
Peñalver E, Arillo A, Pérez-de la Fuente R, Riccio ML, Delclòs X, Barrón E, Grimaldi DA. Long-Proboscid Flies as Pollinators of Cretaceous Gymnosperms. Current Biology [Internet]. 2015;25 (14) :1917–1923. DOIAbstract

The great evolutionary success of angiosperms has traditionally been explained, in part, by the partnership of these plants with insect pollinators [1-6]. The main approach to understanding the origins of this pervasive relationship has been study of the pollinators of living cycads, gnetaleans, and basal angiosperms [7]. Among the most morphologically specialized living pollinators are diverse, long-proboscid flies. Early such flies include the brachyceran family Zhangsolvidae, previously known only as compression fossils from the Early Cretaceous of China and Brazil. It belongs to the infraorder Stratiomyomorpha, a group that includes the flower-visiting families Xylomyidae and Stratiomyidae. New zhangsolvid specimens in amber from Spain (ca. 105 mega-annum [Ma]) and Myanmar (100 Ma) reveal a detailed proboscis structure adapted to nectivory. Pollen clumped on a specimen from Spain is Exesipollenites, attributed to a Mesozoic gymnosperm, most likely the Bennettitales. Late Mesozoic scorpionflies with a long proboscis have been proposed as specialized pollinators of various extinct gymnosperms, but pollen has never been observed on or in their bodies [8]. The new discovery is a very rare co-occurrence of pollen with its insect vector and provides substantiating evidence that other long-proboscid Mesozoic insects were gymnosperm pollinators. Evidence is thus now gathering that visitors and probable pollinators of early anthophytes, or seed plants, involved some insects with highly specialized morphological adaptations, which has consequences for interpreting the reproductive modes of Mesozoic gymnosperms and the significance of insect pollination in angiosperm success.

Laurindo da Silva F, Trivinho-strixino S, Silva Neubern de Oliveira C. Chironomidae types in the reference collection of the Laboratory of Ecology of Aquatic Insects, São Carlos, Brazil. CHIRONOMUS Journal of Chironomidae Research. 2015 :20–29.Abstract

The Chironomidae (Insecta: Diptera) type collection at the Laboratory of Ecology of Aquatic Insects (LEIA - UFSCar) is reviewed. It comprises 103 primary types, as well as 95 paratypes, mostly resulting from research by S. Trivinho-Strixino and G. Strixino. Notes updating the taxonomic status are provided for some species.

Laurindo da Silva F, Wiedenbrug S, Farrell BD. A Preliminary Survey of the Non-biting Midges (Diptera: Chironomidae) of the Dominican Republic. CHIRONOMUS Journal of Chironomidae Research [Internet]. 2015;(28) :12–19. DOI (full text)Abstract

Chironomidae (Diptera) are among the most diverse and widespread aquatic insects, with roughly 5,500 described species. However, prior to the present work, no species of Chironomidae had been documented from the island of Hispaniola. Collections of non-biting midges, with emphasis on the lotic fauna, were made in the Dominican Republic during July of 2015. In total, 578 specimens belonging to 27 genera and at least 44 species within the subfamilies Chironominae (20 taxa), Orthocladiinae (16 taxa) and Tanypodinae (8 taxa) were found. The subfamilies Chironominae and Orthocladiinae predominated. Polypedilum was the most widespread and diverse genus of Chironominae. Metriocnemus were collected in bromeliad tanks. The chironomid fauna in Dominican Republic includes multiple genera with worldwide distributions, including Holarctic and Neotropical components.

DR Non-biting Midges 2015 (PDF)
McKenna DD, Wild AL, Kanda K, Bellamy CL, Beutel RG, Caterino MS, Farnum CW, Hawks DC, Ivie MA, Jameson ML, et al. The beetle tree of life reveals that Coleoptera survived end-Permian mass extinction to diversify during the Cretaceous terrestrial revolution. Systematic Entomology [Internet]. 2015;40 (4) :835-880. DOI (full text)Abstract

Here we present a phylogeny of beetles (Insecta: Coleoptera) based on DNA sequence data from eight nuclear genes, including six single-copy nuclear protein-coding genes, for 367 species representing 172 of 183 extant families. Our results refine existing knowledge of relationships among major groups of bee- tles. Strepsiptera was confirmed as sister to Coleoptera and each of the suborders of Coleoptera was recovered as monophyletic. Interrelationships among the subor- ders, namely Polyphaga (Adephaga (Archostemata, Myxophaga)), in our study differ from previous studies. Adephaga comprised two clades corresponding to Hydrade- phaga and Geadephaga. The series and superfamilies of Polyphaga were mostly monophyletic. The traditional Cucujoidea were recovered in three distantly related clades. Lymexyloidea was recovered within Tenebrionoidea. Several of the series and superfamilies of Polyphaga received moderate to maximal clade support in most analyses, for example Buprestoidea, Chrysomeloidea, Coccinelloidea, Cucujiformia, Curculionoidea, Dascilloidea, Elateroidea, Histeroidea and Hydrophiloidea. However, 836 D. D. McKenna et al.
many of the relationships within Polyphaga lacked compatible resolution under maximum-likelihood and Bayesian inference, and/or lacked consistently strong nodal support. Overall, we recovered slightly younger estimated divergence times than pre- vious studies for most groups of beetles. The ordinal split between Coleoptera and Strepsiptera was estimated to have occurred in the Early Permian. Crown Coleoptera appeared in the Late Permian, and only one or two lineages survived the end-Permian mass extinction, with stem group representatives of all four suborders appearing by the end of the Triassic. The basal split in Polyphaga was estimated to have occurred in the Triassic, with the stem groups of most series and superfamilies originating during the Triassic or Jurassic. Most extant families of beetles were estimated to have Creta- ceous origins. Overall, Coleoptera experienced an increase in diversification rate com- pared to the rest of Neuropteroidea. Furthermore, 10 family-level clades, all in suborder Polyphaga, were identified as having experienced significant increases in diversification rate.These include most beetle species with phytophagous habits, but also several groups not typically or primarily associated with plants. Most of these groups originated in the Cretaceous, which is also when a majority of the most species-rich beetle families first appeared. An additional 12 clades showed evidence for significant decreases in diversi- fication rate. These clades are species-poor in theModern fauna, but collectively exhibit diverse trophic habits. The apparent success of beetles, as measured by species numbers, may result from their associations with widespread and diverse substrates – especially plants, but also including fungi, wood and leaf litter – but what facilitated these associ- ations in the first place or has allowed these associations to flourish likely varies within and between lineages.Our results provide a uniquelywell-resolved temporal and phylo- genetic framework for studying patterns of innovation and diversification in Coleoptera, and a foundation for further sampling and resolution of the beetle tree of life.

Kim SI, Farrell BD. Phylogeny of world stag beetles (Coleoptera: Lucanidae) reveals a Gondwanan origin of Darwin’s stag beetle. Molecular Phylogenetics and Evolution [Internet]. 2015;86 :35-48. DOI (full text)Abstract

Stag beetles (family Lucanidae Latreille, 1804) are one of the earliest branching lineages of scarab beetles that are characterized by the striking development of the male mandibles. Despite stag beetles’ popularity among traditional taxonomists and amateur collectors, there has been almost no study of lucanid relationships and evolution. Entomologists, including Jeannel (1942), have long recognized resemblance between the austral stag beetles of the tribes Chiasognathini, Colophonini, Lamprimini, Pholidotini, Rhyssonotini, and Streptocerini, but this hypothesis of their close relationship across the continents has never been tested. To gain further insight into lucanid phylogeny and biogeography, we reconstructed the first molecular phylogeny of world stag beetles using DNA sequences from mitochondrial 16S rDNA, nuclear 18S and 28S rDNA, and the nuclear protein-coding (NPC) gene wingless for 93 lucanid species representing all extant subfamilies and 24 out of the 27 tribes, together with 14 representative samples of other early branching scarabaeoid families and two staphyliniform beetle families as outgroups. Both Bayesian inference (BI) and maximum likelihood inference (MLI) strongly supported the monophyly of Lucanidae sensu lato that includes Diphyllostomatidae. Within Lucanidae sensu stricto, the subfamilies Lucaninae and Lampriminae appeared monophyletic under both methods of phylogenetic inferences; however, Aesalinae and Syndesinae were found to be polyphyletic. A time-calibrated phylogeny based on five fossil data estimated the origin of crown group Lucanidae as circa 160 million years ago (MYA). Divergence between the Neotropical and Australasian groups of the Chiasognathini was estimated to be circa 47 MYA, with the South African Colophonini branching off from the ancient Chiasognathini lineage around 87 MYA. Another Gondwanan relationship was recovered between the Australasian Eucarteria and the Neotropical Casignetus, which diverged circa 58 MYA. Lastly, as Jeannel’s hypothesis predicted, divergence within Lampriminae between the Australasian Lamprima and the Neotropical Streptocerus was estimated to be circa 37 MYA. The split of these lineages were generally concordant with the pattern of continental break-up of the super-continent Gondwana, and our biogeographic reconstructions based on the dispersal-extinction-cladogenesis model (DEC) corroborate our view that the divergences in these austral lineages were caused by vicariance events following the Gondwanan break-up. In addition, the phylogenetic position and geographic origin of the Hawaiian genus Apterocyclus was revealed for the first time. Overall, our results provide the framework toward studying lucanid relationships and divergence time estimates, which allowed for more accurate biogeographic explanations and discussions on ancestral lucanids and the evolutionary origin of the enlarged male mandibles.

Rykken JJ, Farrell BD. Developing a pollinator monitoring program for Boston Harbor Islands National Recreation Area: A pilot study. Natural Resource Report NPS/BOHA/NRR—2015/950 [Internet]. 2015. Full text from NPS

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