Alexie Papanicolaou

Butterfly genome reveals promiscuous exchange of mimicry adaptations among species

The evolutionary importance of hybridization and introgression has long been debated. Hybrids are usually rare and unfit, but even infrequent hybridization can aid adaptation by transferring beneficial traits between species. Here we use genomic tools to investigate introgression in Heliconius, a rapidly radiating genus of neotropical butterflies widely used in studies of ecology, behaviour, mimicry and speciation. We sequenced the genome of Heliconius melpomene and compared it with other taxa to investigate chromosomal evolution in Lepidoptera and gene flow among multiple Heliconius species and races. Among 12,669 predicted genes, biologically important expansions of families of chemosensory and Hox genes are particularly noteworthy. Chromosomal organization has remained broadly conserved since the Cretaceous period, when butterflies split from the Bombyx (silkmoth) lineage. Using genomic resequencing, we show hybrid exchange of genes between three co-mimics, Heliconius melpomene, Heliconius timareta and Heliconius elevatus, especially at two genomic regions that control mimicry pattern. We infer that closely related Heliconius species exchange protective colour-pattern genes promiscuously, implying that hybridization has an important role in adaptive radiation.

The i5K Initiative: Advancing Arthropod Genomics for Knowledge, Human Health, Agriculture, and the Environment

Insects and their arthropod relatives including mites, spiders, and crustaceans play major roles in the worlds terrestrial, aquatic, and marine ecosystems. Arthropods compete with humans for food and transmit devastating diseases. They also comprise the most diverse and successful branch of metazoan evolution, with millions of extant species. Here, we describe an international effort to guide arthropod genomic efforts, from species prioritization to methodology and informatics. The 5000 arthropod genomes initiative (i5K) community met formally in 2012 to discuss a roadmap for sequencing and analyzing 5000 high-priority arthropods and is continuing this effort via pilot projects, the development of standard operating procedures, and training of students and career scientists. With university, governmental, and industry support, the i5K Consortium aspires to deliver sequences and analytical tools for each of the arthropod branches and each of the species having beneficial and negative effects on humankind.

Heliconius wing patterns: an evo-devo model for understanding phenotypic diversity

Evolutionary Developmental Biology aims for a mechanistic understanding of phenotypic diversity, and present knowledge is largely based on gene expression and interaction patterns from a small number of well-known model organisms. However, our understanding of biological diversification depends on our ability to pinpoint the causes of natural variation at a micro-evolutionary level, and therefore requires the isolation of genetic and developmental variation in a controlled genetic background. The colour patterns of Heliconius butterflies (Nymphalidae: Heliconiinae) provide a rich suite of naturally occurring variants with striking phenotypic diversity and multiple taxonomic levels of variation. Diversification in the genus is well known for its dramatic colour-pattern divergence between races or closely related species, and for Müllerian mimicry convergence between distantly related species, providing a unique system to study the development basis of colour-pattern evolution. A long history of genetic studies has showed that pattern variation is based on allelic combinations at a surprisingly small number of loci, and recent developmental evidence suggests that pattern development in Heliconius is different from the eyespot determination of other butterflies. Fine-scale genetic mapping studies have shown that a shared toolkit of genes is used to produce both convergent and divergent phenotypes. These exciting results and the development of new genomic resources make Heliconius a very promising evo-devo model for the study of adaptive change.

ButterflyBase: a platform for lepidopteran genomics

With over 100 000 species and a large community of evolutionary biologists, population ecologists, pest biologists and genome researchers, the Lepidoptera are an important insect group. Genomic resources [expressed sequence tags (ESTs), genome sequence, genetic and physical maps, proteomic and microarray datasets] are growing, but there has up to now been no single access and analysis portal for this group. Here we present ButterflyBase (http://www.butterflybase.org), a unified resource for lepidopteran genomics. A total of 273 077 ESTs from more than 30 different species have been clustered to generate stable unigene sets, and robust protein translations derived from each unigene cluster. Clusters and their protein translations are annotated with BLAST-based similarity, gene ontology (GO), enzyme classification (EC) and Kyoto encyclopaedia of genes and genomes (KEGG) terms, and are also searchable using similarity tools such as BLAST and MS-BLAST. The database supports many needs of the lepidopteran research community, including molecular marker development, orthologue prediction for deep phylogenetics, and detection of rapidly evolving proteins likely involved in host–pathogen or other evolutionary processes. ButterflyBase is expanding to include additional genomic sequence, ecological and mapping data for key species.

Synteny and Chromosome Evolution in the Lepidoptera: Evidence from Mapping in Heliconius melpomene

The extent of conservation of synteny and gene order in the Lepidoptera has been investigated previously only by comparing a small subset of linkage groups between the moth Bombyx mori and the butterfly Heliconius melpomene. Here we report the mapping of 64 additional conserved genes in H. melpomene, which contributed 47 markers to a comparative framework of 72 orthologous loci spanning all 21 H. melpomene chromosomes and 27 of the 28 B. mori chromosomes. Comparison of the maps revealed conserved synteny across all chromosomes for the 72 loci, as well as evidence for six cases of chromosome fusion in the Heliconius lineage that contributed to the derived 21-chromosome karyotype. Comparisons of gene order on these fused chromosomes revealed two instances of colinearity between H. melpomene and B. mori, but also one instance of likely chromosomal rearrangement. B. mori is the first lepidopteran species to have its genome sequenced, and the finding that there is conserved synteny and gene order among Lepidoptera indicates that the genomic tools developed in B. mori will be broadly useful in other species.

Characterization of a hotspot for mimicry: assembly of a butterfly wing transcriptome to genomic sequence at the HmYb/Sb locus

The mimetic wing patterns of Heliconius butterflies are an excellent example of both adaptive radiation and convergent evolution. Alleles at the HmYb and HmSb loci control the presence/absence of hindwing bar and hindwing margin phenotypes respectively between divergent races of Heliconius melpomene, and also between sister species. Here, we used fine‐scale linkage mapping to identify and sequence a BAC tilepath across the HmYb/Sb loci. We also generated transcriptome sequence data for two wing pattern forms of H. melpomene that differed in HmYb/Sb alleles using 454 sequencing technology. Custom scripts were used to process the sequence traces and generate transcriptome assemblies. Genomic sequence for the HmYb/Sb candidate region was annotated both using the MAKER pipeline and manually using transcriptome sequence reads. In total, 28 genes were identified in the HmYb/Sb candidate region, six of which have alternative splice forms. None of these are orthologues of genes previously identified as being expressed in butterfly wing pattern development, implying previously undescribed molecular mechanisms of pattern determination on Heliconius wings. The use of next‐generation sequencing has therefore facilitated DNA annotation of a poorly characterized genome, and generated hypotheses regarding the identity of wing pattern at the HmYb/Sb loci.

Identification and characterization of three chemosensory receptor families in the cotton bollworm Helicoverpa armigera

BackgroundChemosensory receptors including olfactory receptors (ORs), gustatory receptors (GRs) and ionotropic receptors (IRs) play a central role in sensing chemical signals and guiding insect behaviours, and are potential target genes in insect pest control. The cotton bollworm Helicoverpa armigera is one of the most destructive pest species that can feed on over 200 different plant species. This diversity of host plants is likely linked to a complex chemosensory system. Here we built on previous work to characterize crucial chemosensory tissues linked to environmental interactions including larval antennae, larval mouthparts and larval fat bodies, as well as male and female adult heads, male and female adult tarsi, and female abdomens.ResultsUsing transcriptome sequencing, Trinity RNA-seq assemblies and extensive manual curation, we identified a total of 91 candidate chemosensory receptors (60 candidate ORs, 10 GRs and 21 IRs). Thirty-five of these candidates present full-length transcripts. First, we performed in silico differential expression analysis on different sequenced tissues. Further, we created extensive expression profiles using reverse transcription (RT)-PCR on a variety of adult and larval stages. We found that the expression profile of HarmOR51 was limited to adult male antenna suggesting a role in mating that was further supported by a phylogenetic analysis clustering it into the pheromone receptor clade. HarmOR51 in calcium imaging analysis did not show responses to either of the two H. armigera sex pheromone components (Z9-16:Ald or Z11-16:Ald) inviting a future detailed study. In addition, we found four novel HarmORs (OR1, 53, 54 and 58) that appeared to be larvae-antennal specific. Finally, our expression profiling showed that four “divergent” HarmIRs (IR2, 7d.1, 7d.2 and 7d.3) were expressed in both adult and larval antennae, suggesting a functional divergence from their Drosophila homologues.ConclusionsThis study explored three chemoreceptor superfamily genes using a curated transcriptomic approach coupled with extensive expression profiling and a more limited functional characterization. Our results have now provided an extensive resource for investigating the chemoreceptor complement of this insect pest, and meanwhile allow for targeted experiments to identify potential molecular targets for pest control and to investigate insect-plant interactions.

Butterfly genomics eclosing

Technological and conceptual advances of the last decade have led to an explosion of genomic data and the emergence of new research avenues. Evolutionary and ecological functional genomics, with its focus on the genes that affect ecological success and adaptation in natural populations, benefits immensely from a phylogenetically widespread sampling of biological patterns and processes. Among those organisms outside established model systems, butterflies offer exceptional opportunities for multidisciplinary research on the processes generating and maintaining variation in ecologically relevant traits. Here we highlight research on wing color pattern variation in two groups of Nymphalid butterflies, the African species Bicyclus anynana (subfamily Satyrinae) and species of the South American genus Heliconius (subfamily Heliconiinae), which are emerging as important systems for studying the nature and origins of functional diversity. Growing genomic resources including genomic and cDNA libraries, dense genetic maps, high-density gene arrays, and genetic transformation techniques are extending current gene mapping and expression profiling analysis and enabling the next generation of research questions linking genes, development, form, and fitness. Efforts to develop such resources in Bicyclus and Heliconius underscore the general challenges facing the larger research community and highlight the need for a community-wide effort to extend ongoing functional genomic research on butterflies.

Blue: correcting sequencing errors using consensus and context

MOTIVATION Bioinformatics tools, such as assemblers and aligners, are expected to produce more accurate results when given better quality sequence data as their starting point. This expectation has led to the development of stand-alone tools whose sole purpose is to detect and remove sequencing errors. A good error-correcting tool would be a transparent component in a bioinformatics pipeline, simply taking sequence data in any of the standard formats and producing a higher quality version of the same data containing far fewer errors. It should not only be able to correct all of the types of errors found in real sequence data (substitutions, insertions, deletions and uncalled bases), but it has to be both fast enough and scalable enough to be usable on the large datasets being produced by current sequencing technologies, and work on data derived from both haploid and diploid organisms. RESULTS This article presents Blue, an error-correction algorithm based on k-mer consensus and context. Blue can correct substitution, deletion and insertion errors, as well as uncalled bases. It accepts both FASTQ and FASTA formats, and corrects quality scores for corrected bases. Blue also maintains the pairing of reads, both within a file and between pairs of files, making it compatible with downstream tools that depend on read pairing. Blue is memory efficient, scalable and faster than other published tools, and usable on large sequencing datasets. On the tests undertaken, Blue also proved to be generally more accurate than other published algorithms, resulting in more accurately aligned reads and the assembly of longer contigs containing fewer errors. One significant feature of Blue is that its k-mer consensus table does not have to be derived from the set of reads being corrected. This decoupling makes it possible to correct one dataset, such as small set of 454 mate-pair reads, with the consensus derived from another dataset, such as Illumina reads derived from the same DNA sample. Such cross-correction can greatly improve the quality of small (and expensive) sets of long reads, leading to even better assemblies and higher quality finished genomes. AVAILABILITY AND IMPLEMENTATION The code for Blue and its related tools are available from http://www.bioinformatics.csiro.au/Blue. These programs are written in C# and run natively under Windows and under Mono on Linux.

Genomic tools and cDNA derived markers for butterflies

The Lepidoptera have long been used as examples in the study of evolution, but some questions remain difficult to resolve due to a lack of molecular genetic data. However, as technology improves, genomic tools are becoming increasingly available to tackle unanswered evolutionary questions. Here we have used expressed sequence tags (ESTs) to develop genetic markers for two Müllerian mimic species, Heliconius melpomene and Heliconius erato. In total 1363 ESTs were generated, representing 330 gene objects in H. melpomene and 431 in H. erato. User‐friendly bioinformatic tools were used to construct a nonredundant database of these putative genes (available at http://www.heliconius.org), and annotate them with blast similarity searches, InterPro matches and Gene Ontology terms. This database will be continually updated with EST sequences for the Papilionideae as they become publicly available, providing a tool for gene finding in the butterflies. Alignments of the Heliconius sequences with putative homologues derived from Bombyx mori or other public data sets were used to identify conserved PCR priming sites, and develop 55 markers that can be amplified from genomic DNA in both H. erato and H. melpomene. These markers will be used for comparative linkage mapping in Heliconius and will have applications in other phylogenetic and genomic studies in the Lepidoptera.