RESUMO
Recent excavations of Late Antiquity settlements in the Negev Highlands of southern Israel uncovered a society that established commercial-scale viticulture in an arid environment [D. Fuks et al., Proc. Natl. Acad. Sci. U.S.A. 117, 19780-19791 (2020)]. We applied target-enriched genome-wide sequencing and radiocarbon dating to examine grapevine pips that were excavated at three of these sites. Our analyses revealed centuries long and continuous grape cultivation in the Southern Levant. The genetically diverse pips also provided clues to ancient cultivation strategies aimed at improving agricultural productivity and ensuring food security. Applying genomic prediction analysis, a pip dated to the eighth century CE was determined to likely be from a white grape, to date the oldest to be identified. In a kinship analysis, another pip was found to be descendant from a modern Greek cultivar and was thus linked with several popular historic wines that were once traded across the Byzantine Empire. These findings shed light on historical Byzantine trading networks and on the genetic contribution of Levantine varieties to the classic Aegean landscape.
Assuntos
Vitis , Vinho , História Antiga , Vitis/genética , DNA Antigo , Arqueologia , IsraelRESUMO
BACKGROUND: Social insects vary considerably in their social organization both between and within species. In the California harvester ant, Pogonomyrmex californicus (Buckley 1867), colonies are commonly founded and headed by a single queen (haplometrosis, primary monogyny). However, in some populations in California (USA), unrelated queens cooperate not only during founding (pleometrosis) but also throughout the life of the colony (primary polygyny). The genetic architecture and evolutionary dynamics of this complex social niche polymorphism (haplometrosis vs pleometrosis) have remained unknown. RESULTS: We provide a first analysis of its genomic basis and evolutionary history using population genomics comparing individuals from a haplometrotic population to those from a pleometrotic population. We discovered a recently evolved (< 200 k years), 8-Mb non-recombining region segregating with the observed social niche polymorphism. This region shares several characteristics with supergenes underlying social polymorphisms in other socially polymorphic ant species. However, we also find remarkable differences from previously described social supergenes. Particularly, four additional genomic regions not in linkage with the supergene show signatures of a selective sweep in the pleometrotic population. Within these regions, we find for example genes crucial for epigenetic regulation via histone modification (chameau) and DNA methylation (Dnmt1). CONCLUSIONS: Altogether, our results suggest that social morph in this species is a polygenic trait involving a potential young supergene. Further studies targeting haplo- and pleometrotic individuals from a single population are however required to conclusively resolve whether these genetic differences underlie the alternative social phenotypes or have emerged through genetic drift.
Assuntos
Formigas , Animais , Formigas/genética , Comportamento Social , Genômica , Genoma de Inseto , Polimorfismo Genético , Evolução Biológica , Feminino , California , Evolução MolecularRESUMO
Social insect colonies exhibit colony-level phenotypes such as social immunity and task coordination, which are produced by the individual phenotypes. Mapping the genetic basis of such phenotypes requires associating the colony-level phenotype with the genotypes in the colony. In this paper, we examine alternative approaches to DNA extraction, library construction, and sequencing for genome wide association studies (GWAS) of colony-level traits using a population sample of Cataglyphis niger ants. We evaluate the accuracy of allele frequency estimation from sequencing a pool of individuals (pool-seq) from each colony using either whole-genome sequencing or reduced representation genomic sequencing. Based on empirical measurement of the experimental noise in sequenced DNA pools, we show that reduced representation pool-seq is drastically less accurate than whole-genome pool-seq. Surprisingly, normalized pooling of samples did not result in greater accuracy than un-normalized pooling. Subsequently, we evaluate the power of the alternative approaches for detecting quantitative trait loci (QTL) of colony-level traits by using simulations that account for an environmental effect on the phenotype. Our results can inform experimental designs and enable optimizing the power of GWAS depending on budget, availability of samples and research goals. We conclude that for a given budget, sequencing un-normalized pools of individuals from each colony provides optimal QTL detection power.
Assuntos
Mapeamento Cromossômico/métodos , Estudo de Associação Genômica Ampla/métodos , Metagenômica/métodos , Alelos , Animais , Formigas , Comportamento Animal/fisiologia , Frequência do Gene/genética , Genótipo , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas , Comportamento Social , Sequenciamento Completo do Genoma/métodosRESUMO
BACKGROUND: A major focus of evolutionary biology is the formation of reproductive barriers leading to divergence and ultimately, speciation. Often, it is not clear whether the separation of populations is complete or if there still is ongoing gene flow in the form of rare cases of admixture, known as isolation with migration. Here, we studied the speciation of two fire ant species, Solenopsis invicta and Solenopsis richteri, both native to South America, both inadvertently introduced to North America in the early twentieth century. While the two species are known to admix in the introduced range, in the native range no hybrids were found. RESULTS: We conducted a population genomic survey of native and introduced populations of the two species using reduced representation genomic sequencing of 337 samples. Using maximum likelihood analysis over native range samples, we found no evidence of any gene flow between the species since they diverged. We estimated their time of divergence to 190,000 (100,000-350,000) generations ago. Modelling the demographic history of native and introduced S. invicta populations, we evaluated their divergence times and historic and contemporary population sizes, including the original founder population in North America, which was estimated at 26 (10-93) unrelated singly-mated queens. CONCLUSIONS: We provide evidence for complete genetic isolation maintained between two invasive species in their natïve range, based, for the first time, on large scale genomic data analysis. The results lay the foundations for further studies into different stages in the formation of genetic barriers in dynamic, invasive populations.
Assuntos
Formigas/genética , Especiação Genética , Hibridização Genética , Espécies Introduzidas , Animais , Sequência de Bases , Genética Populacional , Geografia , Modelos Teóricos , América do Norte , Densidade Demográfica , Isolamento Reprodutivo , América do Sul , Especificidade da Espécie , Fatores de TempoRESUMO
The fire ant Solenopsis invicta and its close relatives are highly invasive. Enhanced social cooperation may facilitate invasiveness in these and other invasive ant species. We investigated whether invasiveness in Solenopsis fire ants was accompanied by positive selection on sociobiological traits by applying a phylogenomics approach to infer ancient selection, and a population genomics approach to infer recent and ongoing selection in both native and introduced S. invicta populations. A combination of whole-genome sequencing of 40 haploid males and reduced-representation genomic sequencing of 112 diploid workers identified 1,758,116 and 169,682 polymorphic markers, respectively. The resulting high-resolution maps of genomic polymorphism provide high inference power to test for positive selection. Our analyses provide evidence of positive selection on putative ion channel genes, which are implicated in neurological functions, and on vitellogenin, which is a key regulator of development and caste determination. Furthermore, molecular functions implicated in pheromonal signalling have experienced recent positive selection. Genes with signatures of positive selection were significantly more often those overexpressed in workers compared with queens and males, suggesting that worker traits are under stronger selection than queen and male traits. These results provide insights into selection pressures and ongoing adaptation in an invasive social insect and support the hypothesis that sociobiological traits are under more positive selection than nonsocial traits in such invasive species.
Assuntos
Formigas/genética , Seleção Genética/genética , Animais , Formigas/classificação , Evolução Molecular , Espécies Introduzidas , FilogeniaRESUMO
Genomes of eusocial insects code for dramatic examples of phenotypic plasticity and social organization. We compared the genomes of seven ants, the honeybee, and various solitary insects to examine whether eusocial lineages share distinct features of genomic organization. Each ant lineage contains â¼4000 novel genes, but only 64 of these genes are conserved among all seven ants. Many gene families have been expanded in ants, notably those involved in chemical communication (e.g., desaturases and odorant receptors). Alignment of the ant genomes revealed reduced purifying selection compared with Drosophila without significantly reduced synteny. Correspondingly, ant genomes exhibit dramatic divergence of noncoding regulatory elements; however, extant conserved regions are enriched for novel noncoding RNAs and transcription factor-binding sites. Comparison of orthologous gene promoters between eusocial and solitary species revealed significant regulatory evolution in both cis (e.g., Creb) and trans (e.g., fork head) for nearly 2000 genes, many of which exhibit phenotypic plasticity. Our results emphasize that genomic changes can occur remarkably fast in ants, because two recently diverged leaf-cutter ant species exhibit faster accumulation of species-specific genes and greater divergence in regulatory elements compared with other ants or Drosophila. Thus, while the "socio-genomes" of ants and the honeybee are broadly characterized by a pervasive pattern of divergence in gene composition and regulation, they preserve lineage-specific regulatory features linked to eusociality. We propose that changes in gene regulation played a key role in the origins of insect eusociality, whereas changes in gene composition were more relevant for lineage-specific eusocial adaptations.
Assuntos
Formigas/genética , Genoma de Inseto , Animais , Comportamento Animal , Sítios de Ligação , Sequência Conservada , Metilação de DNA , Evolução Molecular , Regulação da Expressão Gênica , Himenópteros/genética , Proteínas de Insetos/genética , MicroRNAs/genética , Modelos Genéticos , Filogenia , Sequências Reguladoras de Ácido Nucleico , Análise de Sequência de DNA , Comportamento Social , Especificidade da Espécie , Sintenia , Fatores de Transcrição/genéticaRESUMO
The evolution of ants is marked by remarkable adaptations that allowed the development of very complex social systems. To identify how ant-specific adaptations are associated with patterns of molecular evolution, we searched for signs of positive selection on amino-acid changes in proteins. We identified 24 functional categories of genes which were enriched for positively selected genes in the ant lineage. We also reanalyzed genome-wide data sets in bees and flies with the same methodology to check whether positive selection was specific to ants or also present in other insects. Notably, genes implicated in immunity were enriched for positively selected genes in the three lineages, ruling out the hypothesis that the evolution of hygienic behaviors in social insects caused a major relaxation of selective pressure on immune genes. Our scan also indicated that genes implicated in neurogenesis and olfaction started to undergo increased positive selection before the evolution of sociality in Hymenoptera. Finally, the comparison between these three lineages allowed us to pinpoint molecular evolution patterns that were specific to the ant lineage. In particular, there was ant-specific recurrent positive selection on genes with mitochondrial functions, suggesting that mitochondrial activity was improved during the evolution of this lineage. This might have been an important step toward the evolution of extreme lifespan that is a hallmark of ants.
Assuntos
Formigas/genética , Genes Mitocondriais , Proteínas de Insetos/genética , Animais , Formigas/classificação , Bases de Dados Genéticas , Evolução Molecular , Genoma de Inseto , Família Multigênica , Filogenia , Seleção GenéticaRESUMO
Errors in the inferred multiple sequence alignment may lead to false prediction of positive selection. Recently, methods for detecting unreliable alignment regions were developed and were shown to accurately identify incorrectly aligned regions. While removing unreliable alignment regions is expected to increase the accuracy of positive selection inference, such filtering may also significantly decrease the power of the test, as positively selected regions are fast evolving, and those same regions are often those that are difficult to align. Here, we used realistic simulations that mimic sequence evolution of HIV-1 genes to test the hypothesis that the performance of positive selection inference using codon models can be improved by removing unreliable alignment regions. Our study shows that the benefit of removing unreliable regions exceeds the loss of power due to the removal of some of the true positively selected sites.
Assuntos
Modelos Genéticos , Alinhamento de Sequência/métodos , Alinhamento de Sequência/normas , Simulação por Computador , Bases de Dados Genéticas , Evolução Molecular , Genes Virais , HIV-1/genética , Filogenia , Seleção GenéticaRESUMO
The transformer (tra) gene is a key regulator in the signalling hierarchy controlling all aspects of somatic sexual differentiation in Drosophila and other insects. Here, we show that six of the seven sequenced ants have two copies of tra. Surprisingly, the two paralogues are always more similar within species than among species. Comparative sequence analyses indicate that this pattern is owing to the ongoing concerted evolution after an ancestral duplication rather than independent duplications in each of the six species. In particular, there was strong support for inter-locus recombination between the paralogues of the ant Atta cephalotes. In the five species where the location of paralogues is known, they are adjacent to each other in four cases and separated by only few genes in the fifth case. Because there have been extensive genomic rearrangements in these lineages, this suggests selection acting to conserve their synteny. In three species, we also find a signature of positive selection in one of the paralogues. In three bee species where information is available, the tra gene is also duplicated, the copies are adjacent and in at least one species there was recombination between paralogues. These results suggest that concerted evolution plays an adaptive role in the evolution of this gene family.
Assuntos
Evolução Molecular , Duplicação Gênica , Genes de Insetos , Himenópteros/genética , Seleção Genética , Processos de Determinação Sexual , Animais , Teorema de Bayes , Feminino , Himenópteros/crescimento & desenvolvimento , Masculino , Filogenia , Análise de Sequência de DNA , Homologia de Sequência , Especificidade da EspécieRESUMO
In recent years, both homing endonucleases (HEases) and zinc-finger nucleases (ZFNs) have been engineered and selected for the targeting of desired human loci for gene therapy. However, enzyme engineering is lengthy and expensive and the off-target effect of the manufactured endonucleases is difficult to predict. Moreover, enzymes selected to cleave a human DNA locus may not cleave the homologous locus in the genome of animal models because of sequence divergence, thus hampering attempts to assess the in vivo efficacy and safety of any engineered enzyme prior to its application in human trials. Here, we show that naturally occurring HEases can be found, that cleave desirable human targets. Some of these enzymes are also shown to cleave the homologous sequence in the genome of animal models. In addition, the distribution of off-target effects may be more predictable for native HEases. Based on our experimental observations, we present the HomeBase algorithm, database and web server that allow a high-throughput computational search and assignment of HEases for the targeting of specific loci in the human and other genomes. We validate experimentally the predicted target specificity of candidate fungal, bacterial and archaeal HEases using cell free, yeast and archaeal assays.
Assuntos
Endodesoxirribonucleases/metabolismo , Marcação de Genes , Algoritmos , Animais , Archaea/enzimologia , Bactérias/enzimologia , Sequência de Bases , Sequência Conservada , Bases de Dados de Ácidos Nucleicos , Endodesoxirribonucleases/genética , Fungos/enzimologia , Humanos , Modelos Animais , MutaçãoRESUMO
Double-digest Restriction-site Associated DNA sequencing (ddRADseq) is widely used to generate genomic data for non-model organisms in evolutionary and ecological studies. Along with affordable paired-end sequencing, this method makes population genomic analyses more accessible. However, multiple factors should be considered when designing a ddRADseq experiment, which can be challenging for new users. The generated data often suffer from substantial read overlaps and adaptor contamination, severely reducing sequencing efficiency and affecting data quality. Here, we analyse diverse datasets from the literature and carry out controlled experiments to understand the effects of enzyme choice and size selection on sequencing efficiency. The empirical data reveal that size selection is imprecise and has limited efficacy. In certain scenarios, a substantial proportion of short fragments pass below the lower size-selection cut-off resulting in low sequencing efficiency. However, enzyme choice can considerably mitigate inadvertent inclusion of these shorter fragments. A simple model based on these experiments is implemented to predict the number of genomic fragments generated after digestion and size selection, number of SNPs genotyped, number of samples that can be multiplexed and the expected sequencing efficiency. We developed ddgRADer - http://ddgrader.haifa.ac.il/ - a user-friendly webtool and incorporated these calculations to aid in ddRADseq experimental design while optimizing sequencing efficiency. This tool can also be used for single enzyme protocols such as Genotyping-by-Sequencing. Given user-defined study goals, ddgRADer recommends enzyme pairs and allows users to compare and choose enzymes and size-selection criteria. ddgRADer improves the accessibility and ease of designing ddRADseq experiments and increases the probability of success of the first population genomic study conducted in labs with no prior experience in genomics.
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Social isolation negatively affects health, induces detrimental behaviors, and shortens lifespan in social species. Little is known about the mechanisms underpinning these effects because model species are typically short-lived and non-social. Using colonies of the carpenter ant Camponotus fellah, we show that social isolation induces hyperactivity, alters space-use, and reduces lifespan via changes in the expression of genes with key roles in oxidation-reduction and an associated accumulation of reactive oxygen species. These physiological effects are localized to the fat body and oenocytes, which perform liver-like functions in insects. We use pharmacological manipulations to demonstrate that the oxidation-reduction pathway causally underpins the detrimental effects of social isolation on behavior and lifespan. These findings have important implications for our understanding of how social isolation affects behavior and lifespan in general.
Assuntos
Formigas , Animais , Longevidade , Estresse Oxidativo , Isolamento Social , FígadoRESUMO
Hornets are the largest of the social wasps, and are important regulators of insect populations in their native ranges. Hornets are also very successful as invasive species, with often devastating economic, ecological and societal effects. Understanding why these wasps are such successful invaders is critical to managing future introductions and minimising impact on native biodiversity. Critical to the management toolkit is a comprehensive genomic resource for these insects. Here we provide the annotated genomes for two hornets, Vespa crabro and Vespa velutina. We compare their genomes with those of other social Hymenoptera, including the northern giant hornet Vespa mandarinia. The three hornet genomes show evidence of selection pressure on genes associated with reproduction, which might facilitate the transition into invasive ranges. Vespa crabro has experienced positive selection on the highest number of genes, including those putatively associated with molecular binding and olfactory systems. Caste-specific brain transcriptomic analysis also revealed 133 differentially expressed genes, some of which are associated with olfactory functions. This report provides a spring-board for advancing our understanding of the evolution and ecology of hornets, and opens up opportunities for using molecular methods in the future management of both native and invasive populations of these over-looked insects.
Assuntos
Vespas , Animais , Vespas/genética , Espécies Introduzidas , ReproduçãoRESUMO
Evaluating the accuracy of multiple sequence alignment (MSA) is critical for virtually every comparative sequence analysis that uses an MSA as input. Here we present the GUIDANCE web-server, a user-friendly, open access tool for the identification of unreliable alignment regions. The web-server accepts as input a set of unaligned sequences. The server aligns the sequences and provides a simple graphic visualization of the confidence score of each column, residue and sequence of an alignment, using a color-coding scheme. The method is generic and the user is allowed to choose the alignment algorithm (ClustalW, MAFFT and PRANK are supported) as well as any type of molecular sequences (nucleotide, protein or codon sequences). The server implements two different algorithms for evaluating confidence scores: (i) the heads-or-tails (HoT) method, which measures alignment uncertainty due to co-optimal solutions; (ii) the GUIDANCE method, which measures the robustness of the alignment to guide-tree uncertainty. The server projects the confidence scores onto the MSA and points to columns and sequences that are unreliably aligned. These can be automatically removed in preparation for downstream analyses. GUIDANCE is freely available for use at http://guidance.tau.ac.il.
Assuntos
Alinhamento de Sequência/métodos , Software , Proteínas do Vírus da Imunodeficiência Humana/química , Internet , Análise de Sequência de Proteína , Proteínas Virais Reguladoras e Acessórias/químicaRESUMO
Multiple sequence alignment (MSA) is the basis for a wide range of comparative sequence analyses from molecular phylogenetics to 3D structure prediction. Sophisticated algorithms have been developed for sequence alignment, but in practice, many errors can be expected and extensive portions of the MSA are unreliable. Hence, it is imperative to understand and characterize the various sources of errors in MSAs and to quantify site-specific alignment confidence. In this paper, we show that uncertainties in the guide tree used by progressive alignment methods are a major source of alignment uncertainty. We use this insight to develop a novel method for quantifying the robustness of each alignment column to guide tree uncertainty. We build on the widely used bootstrap method for perturbing the phylogenetic tree. Specifically, we generate a collection of trees and use each as a guide tree in the alignment algorithm, thus producing a set of MSAs. We next test the consistency of every column of the MSA obtained from the unperturbed guide tree with respect to the set of MSAs. We name this measure the "GUIDe tree based AligNment ConfidencE" (GUIDANCE) score. Using the Benchmark Alignment data BASE benchmark as well as simulation studies, we show that GUIDANCE scores accurately identify errors in MSAs. Additionally, we compare our results with the previously published Heads-or-Tails score and show that the GUIDANCE score is a better predictor of unreliably aligned regions.
Assuntos
Algoritmos , Sequência de Aminoácidos , Sequência de Bases , Alinhamento de Sequência/métodos , Análise de Sequência de DNA/métodos , Animais , Simulação por Computador , Bases de Dados Factuais , Drosophila melanogaster/genética , Dados de Sequência Molecular , Filogenia , Curva ROC , SoftwareRESUMO
Inteins are selfish genetic elements that disrupt the sequence of protein-coding genes and are excised post-translationally. Most inteins also contain a HEN (homing endonuclease) domain, which is important for their horizontal transmission. The present review focuses on the evolution of inteins and their nested HENs, and highlights several unsolved questions that could benefit from molecular genetic approaches. Such approaches can be well carried out in halophilic archaea, which are naturally intein-rich and have highly developed genetic tools for their study. In particular, the fitness effects of harbouring an intein/HEN can be tested in direct competition assays, providing additional insights that will improve current evolutionary models.
Assuntos
Endonucleases/química , Endonucleases/genética , Evolução Molecular , Inteínas/genética , Archaea/genética , Archaea/metabolismo , Endonucleases/metabolismo , MutaçãoRESUMO
Colony social organization of multiple Solenopsis fire ant species is determined by a supergene with two haplotypes SB and Sb, which are similar to X/Y sex chromosomes. The ancestral monogyne (single-queen) social form has been associated with homozygous SB/SB queens, while queens in colonies with the derived polygyne (multi-queen) social structure are heterozygous SB/Sb. By comparing 14 Solenopsis invicta genomes and the outgroup S. fugax, we dated the formation of the supergene to 1.1 (0.7-1.6) million years ago, much older than previous estimates, and close to the estimated time of speciation of the two socially polymorphic species S. invicta and S. richteri. We also used 12 S. invicta and S. richteri genomes to compare the evolutionary distances between these species and the distances between the social haplotypes, and found them to be similar. A phylogenetic analysis suggested that the monophyletic Sb clade is more closely related to S. richteri SB haplotypes than to S. invicta SB haplotypes. We conclude that the formation of the supergene occurred concomitantly with the process of speciation of the Solenopsis socially-polymorphic clade, and hypothesize that the Sb variant first arouse in one incipiently-speciating population and then introgressed into the other populations or species.
Assuntos
Formigas/genética , Formigas/fisiologia , Comportamento Animal , Especiação Genética , Comportamento Social , Alelos , Animais , Cruzamentos Genéticos , Genoma de Inseto , Haplótipos , Heterozigoto , Funções Verossimilhança , Modelos Genéticos , Filogenia , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único , Recombinação Genética , Cromossomo X , Cromossomo YRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
Diverse invertebrate taxa including all 200,000 species of Hymenoptera (ants, bees, wasps and sawflies) have a haplodiploid sex determination system, where females are diploid and males are haploid. Thus, hymenopteran genome projects can make use of DNA from a single haploid male sample, which is assumed advantageous for genome assembly. For the purpose of gene annotation, transcriptome sequencing is usually conducted using RNA from a pool of individuals. We conducted a comparative analysis of genome and transcriptome assembly and annotation methods, using genetic sources of different ploidy: (1) DNA from a haploid male or a diploid female (2) RNA from the same haploid male or a pool of individuals. We predicted that the use of a haploid male as opposed to a diploid female will simplify the genome assembly and gene annotation thanks to the lack of heterozygosity. Using DNA and RNA from the same haploid individual is expected to provide better confidence in transcript-to-genome alignment, and improve the annotation of gene structure in terms of the exon/intron boundaries. The haploid genome assemblies proved to be more contiguous, with both contig and scaffold N50 size at least threefold greater than their diploid counterparts. Completeness evaluation showed mixed results. The SOAPdenovo2 diploid assembly was missing more genes than the haploid assembly. The SPAdes diploid assembly had more complete genes, but a higher level of duplicates, and a greatly overestimated genome size. When aligning the two transcriptomes against the male genome, the male transcriptome gave 2-3% more complete transcripts than the pool transcriptome for genes with comparable expression levels in both transcriptomes. However, this advantage disappears in the final results of the gene annotation pipeline that incorporates evidence from homologous proteins. The RNA pool is still required to obtain the full transcriptome with genes that are expressed in other life stages and castes. In conclusion, the use of a haploid source material for a de novo genome project provides a substantial advantage to the quality of the genome draft and the use of RNA from the same haploid individual for transcriptome to genome alignment provides a minor advantage for genes that are expressed in the adult male.
Assuntos
Diploide , Perfilação da Expressão Gênica/métodos , Genoma de Inseto/genética , Genômica/métodos , Haploidia , Insetos/genética , Animais , Mapeamento de Sequências Contíguas , Feminino , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Insetos/classificação , Masculino , Anotação de Sequência Molecular , RNA/genética , RNA/metabolismoRESUMO
In social insects, due to considerable polyphenism as well as high level of hybridization, the delimitation of species can be challenging. The genus Cataglyphis presents a high level of diversification, making it an excellent model with which to study evolutionary paths. Israel appears to be a "hot spot" for recent speciation in this genus. Although previous studies have described multiple species of Cataglyphis in Israel, a recent genetic study has questioned the existence of some of these historically described species. The present study focuses on an apparent species complex, the C. niger species complex which includes C. niger, C. savigyi, and C. drusus that are distinguishable by their mitochondrial DNA (and therefore named mitotypes) but not by their nuclear DNA. Using a multi-method approach (genetics, chemistry and behavior), we show that these mitotypes also differ in their social structures and are readily distinguishable by their cuticular hydrocarbons profiles. While most populations of the different mitotypes are allopatric, at our study site they are sympatric, but nonetheless maintain the observed differences between them. This raises the evolutionary question: Are these incipient species that have diverged with gene flow, or is this a case of social and chemical polymorphism that is maintained within a single species? Unveiling the interplay between social polyphenism and species segregation is at the core of evolutionary biology.