A chromosomal-level genome assembly of Serrognathus titanus Boisduval, 1835 (Coleoptera: Lucanidae).

The Stag beetle (Coleoptera: Lucanidae) is a fascinating group, often considered one of the most primitive within the Scarabaeoidea. They are valuable models for studying beetle evolution. However, the lack of high-quality genomes hinders our understanding of the evolution and ecology of Lucanidae. In this study, we present a chromosome-level genome of Serrognathus titanus by combining PacBio HiFi long reads, Illumina short reads, and Hi-C data. The genome spans 384.07 Mb, with a scaffold N50 size of 75.81 Mb, and most contigs (97.45%, 374.30 Mb) were anchored into six chromosomes. Our BUSCO analysis of the assembly indicates a completeness of 97.6% (n = 1,367), with 92.8% single-copy BUSCOs and 4.8% duplicated BUSCOs identified. Additionally, we found that the genome contains 43.87% (168.50 Mb) repeat elements and identified 14,263 predicted protein-coding genes. The high-quality genome of S. titanus provides valuable genomic information for comprehending the evolution and ecology of Lucanidae.

Chromosome-level genome assembly of marmalade hoverfly Episyrphus balteatus (Diptera: Syrphidae).

Episyrphus balteatus can provide dual ecosystem services including pest control and pollination, which the larvae are excellent predators of aphid pest whereas adults are efficient pollinator. In this study, we assembled a high-quality genome of E. balteatus from northern China geographical population at the chromosome level by using Illumina, PacBio long reads, and Hi-C technologies. The 467.42 Mb genome was obtained from 723 contigs, with a contig N50 of 9.16 Mb and Scaffold N50 of 118.85 Mb, and 90.25% (431.75 Mb) of the assembly was anchored to 4 pseudo-autosomes and one pseudo-heterosome. In total, 14,848 protein-coding genes were annotated, and 95.14% of genes were fully represented in NR, GO, KEGG databases. Besides, we also obtained the mitochondrial genome of E. balteatus of 16, 837 bp in length with 37 typical mitochondrial genes. Overall, this high-quality genome is valuable for evolutionary and genetic studies of E. balteatus and other Syrphidae hoverfly species.

Homing gene drives can transfer rapidly between Anopheles gambiae strains with minimal carryover of flanking sequences.

CRISPR-Cas9 homing gene drives are designed to induce a targeted double-stranded DNA break at a wild type allele ('recipient'), which, when repaired by the host cell, is converted to the drive allele from the homologous ('donor') chromosome. Germline localisation of this process leads to super-Mendelian inheritance of the drive and the rapid spread of linked traits, offering a novel strategy for population control through the deliberate release of drive individuals. During the homology-based DNA repair, additional segments of the recipient chromosome may convert to match the donor, potentially impacting carrier fitness and strategy success. Using Anopheles gambiae strains with variations around the drive target site, here we assess the extent and nature of chromosomal conversion. We show both homing and meiotic drive contribute as mechanisms of inheritance bias. Additionally, over 80% of homing events resolve within 50 bp of the chromosomal break, enabling rapid gene drive transfer into locally-adapted genetic backgrounds.

Chromosomal-level genome assembly of Hylurgus ligniperda: insights into host adaptation and environmental tolerance.

BACKGROUND: Hylurgus ligniperda (Coleoptera: Curculionidae) is a worldwide forest quarantine pest. It is widely distributed, has many host tree species, and possesses strong adaptability. To explore its environmental adaptability and the related molecular mechanisms, we conducted chromosome-level genome sequencing and analyzed the transcriptome under different environmental factors, identifying key expressed genes. RESULTS: We employed PacBio, Illumina, and Hi-C sequencing techniques to assemble a 520 Mb chromosomal-level genome of H. ligniperda, obtaining an N50 of 39.97 Mb across 138 scaffolds. A total of 10,765 protein-coding genes were annotated after repeat masking. Fourteen chromosomes were identified, among which Hyli14 was determined to be the sex chromosome. Survival statistics were tested over various growth periods under high temperature and low humidity conditions. The maximum survival period of adults reached 292 days at 25 °C, 65% relative humidity. In comparison, the maximum survival period was 14 days under 35 °C, 65% relative humidity, and 106 days under 25°C, 40% relative humidity. This indicated that environmental stress conditions significantly reduced adults' survival period. We further conducted transcriptome analysis to screen for potentially influential differentially expressed genes, such as CYP450 and Histone. Subsequently, we performed gene family analysis to gain insights into their functions and interactions, such as CYP450 and Histone. CYP450 genes affected the detoxification metabolism of enzymes in the Cytochrome P450 pathway to adapt to different environments. Histone genes are involved in insect hormone biosynthesis and longevity-regulating pathways in H. ligniperda to adapt to environmental stress. CONCLUSIONS: The genome at the chromosome level of H. ligniperda was assembled for the first time. The mortality of H. ligniperda increased significantly at 35 ℃, 65% RH, and 25 ℃, 40% RH. CYP450 and Histone genes played an important role in response to environmental stress. This genome offers a substantial genetic resource for investigating the molecular mechanisms behind beetle invasion and spread.

The grasshopper genome reveals long-term gene content conservation of the X Chromosome and temporal variation in X Chromosome evolution.

We present the first chromosome-level genome assembly of the grasshopper, Locusta migratoria, one of the largest insect genomes. We use coverage differences between females (XX) and males (X0) to identify the X Chromosome gene content, and find that the X Chromosome shows both complete dosage compensation in somatic tissues and an underrepresentation of testis-expressed genes. X-linked gene content from L. migratoria is highly conserved across seven insect orders, namely Orthoptera, Odonata, Phasmatodea, Hemiptera, Neuroptera, Coleoptera, and Diptera, and the 800 Mb grasshopper X Chromosome is homologous to the fly ancestral X Chromosome despite 400 million years of divergence, suggesting either repeated origin of sex chromosomes with highly similar gene content, or long-term conservation of the X Chromosome. We use this broad conservation of the X Chromosome to test for temporal dynamics to Fast-X evolution, and find evidence of a recent burst evolution for new X-linked genes in contrast to slow evolution of X-conserved genes.

Chromosome-level genome assembly of an oligophagous leaf beetle Ophraella communa (Coleoptera: Chrysomelidae).

The leaf beetle Ophraella communa LeSage (Coleoptera: Chrysomelidae) is an effective biological control agent of the common ragweed. Here, we assembled a chromosome-level genome of the O. communa by combining Illumina, Nanopore, and Hi-C sequencing technologies. The genome size of the final genome assembly is 733.1 Mb, encompassing 17 chromosomes, with an improved contig N50 of 7.05 Mb compared to the original version. Genome annotation reveals 25,873 protein-coding genes, with functional annotations available for 22,084 genes (85.35%). Non-coding sequence annotation identified 204 rRNAs, 626 tRNAs, and 1791 small RNAs. Repetitive elements occupy 414.41 Mb, constituting 57.76% of the genome. This high-quality genome is fundamental for advancing biological control strategies employing O. communa.

Two chromosome-level genome assemblies of galling aphids Slavum lentiscoides and Chaetogeoica ovagalla.

Slavum lentiscoides and Chaetogeoica ovagalla are two aphid species from the subtribe Fordina of Fordini within the subfamily Eriosomatinae, and they produce galls on their primary host plants Pistacia. We assembled chromosome-level genomes of these two species using Nanopore long-read sequencing and Hi-C technology. A 332 Mb genome assembly of S. lentiscoides with a scaffold N50 of 19.77 Mb, including 11,747 genes, and a 289 Mb genome assembly of C. ovagalla with a scaffold N50 of 11.85 Mb, containing 14,492 genes, were obtained. The Benchmarking Universal Single-Copy Orthologs (BUSCO) benchmark of the two genome assemblies reached 93.7% (91.9% single-copy) and 97.0% (95.3% single-copy), respectively. The high-quality genome assemblies in our study provide valuable resources for future genomic research of galling aphids.

A chromosome-level genome assembly of Prosopocoilus inquinatus Westwood, 1848 (Coleoptera: Lucanidae).

Lucanidae (Coleoptera: Scarabaeidae) are fascinating beetles exhibiting significant dimorphism and are widely used as beetle evolutionary study models. However, lacking high-quality genomes prohibits our understanding of Lucanidae. Herein, we proposed a chromosome-level genome assembly of a widespread species, Prosopocoilus inquinatus, combining PacBio HiFi, Illumina, and Hi-C data. The genome size reaches 649.73 Mb, having the scaffold N50 size of 59.50 Mb, and 99.6% (647.13 Mb) of the assembly successfully anchored on 12 chromosomes. The BUSCO analysis of the genome exhibits a completeness of 99.6% (n = 1,367), including 1,362 (98.5%) single-copy BUSCOs and 15 (1.1%) duplicated BUSCOs. The genome annotation identifies that the genome contains 61.41% repeat elements and 13,452 predicted protein-coding genes. This high-quality Lucanidae genome provides treasured genomic information to our knowledge of stag beetles.

Primary cell cultures from the single-chromosome ant Myrmecia croslandi.

The number of chromosomes varies tremendously across species. It is not clear whether having more or fewer chromosomes could be advantageous. The probability of non-disjunction should theoretically decrease with smaller karyotypes, but too long chromosomes should enforce spatial constraint for their segregation during the mitotic anaphase. Here, we propose a new experimental cell system to acquire novel insights into the mechanisms underlying chromosome segregation. We collected the endemic Australian ant Myrmecia croslandi, the only known species with the simplest possible karyotype of a single chromosome in the haploid males (and one pair of chromosomes in the diploid females), since males are typically haploid in hymenopteran insects. Five colonies, each with a queen and a few hundreds of workers, were collected in the Canberra district (Australia), underwent karyotype analysis to confirm the presence of a single pair of chromosomes in worker pupae, and were subsequently maintained in the laboratory in Paris (France). Starting from dissociated male embryos, we successfully conducted primary cell cultures comprised of single-chromosome cells. This could be developed into a unique model that will be of great interest for future genomic and cell biology studies related to mitosis.

Chromosome-level genome assembly and sex chromosome identification of the pink stem borer, Sesamia inferens (Lepidoptera: Noctuidae).

The pink stem borer, Sesamia inferens Walker (Lepidoptera: Noctuidae), is one of the most notorious pest insects of rice and maize crops in the world. Here, we generated a high-quality chromosome-level genome assembly of S. inferens, using a combination of Illumina, PacBio HiFi and Hi-C technologies. The total assembly size was 973.18 Mb with a contig N50 of 33.39 Mb, anchored to 31 chromosomes, revealing a karyotype of 30 + Z. The BUSCO analysis indicated a high completeness of 98.90% (n = 5286), including 5172 (97.8%) single-copy BUSCOs and 58 (1.1%) duplicated BUSCOs. The genome contains 58.59% (564.58 Mb) repeat elements and 26628 predicted protein-coding genes. The chromosome-level genome assembly of S. inferens provides in-depth knowledge and will be a helpful resource for the Lepidoptera and pest control research communities.

Genomic evidence reveals three W-autosome fusions in Heliconius butterflies.

Sex chromosomes are evolutionarily labile in many animals and sometimes fuse with autosomes, creating so-called neo-sex chromosomes. Fusions between sex chromosomes and autosomes have been proposed to reduce sexual conflict and to promote adaptation and reproductive isolation among species. Recently, advances in genomics have fuelled the discovery of such fusions across the tree of life. Here, we discovered multiple fusions leading to neo-sex chromosomes in the sapho subclade of the classical adaptive radiation of Heliconius butterflies. Heliconius butterflies generally have 21 chromosomes with very high synteny. However, the five Heliconius species in the sapho subclade show large variation in chromosome number ranging from 21 to 60. We find that the W chromosome is fused with chromosome 4 in all of them. Two sister species pairs show subsequent fusions between the W and chromosomes 9 or 14, respectively. These fusions between autosomes and sex chromosomes make Heliconius butterflies an ideal system for studying the role of neo-sex chromosomes in adaptive radiations and the degeneration of sex chromosomes over time. Our findings emphasize the capability of short-read resequencing to detect genomic signatures of fusion events between sex chromosomes and autosomes even when sex chromosomes are not explicitly assembled.

The chromosome-level genome assembly of Aphidoletes aphidimyza Rondani (Diptera: Cecidomyiidae).

Aphidoletes aphidimyza is widely recognized as an effective predator of aphids in agricultural systems. However, there is limited understanding of its predation mechanisms. In this study, we generated a high-quality chromosome level of the A. aphidimyza genome by combining PacBio, Illumina, and Hi-C data. The genome has a size of 192.08 Mb, with a scaffold N50 size of 46.85 Mb, and 99.08% (190.35 Mb) of the assembly is located on four chromosomes. The BUSCO analysis of our assembly indicates a completeness of 97.8% (n = 1,367), including 1,307 (95.6%) single-copy BUSCOs and 30 (2.2%) duplicated BUSCOs. Additionally, we annotated a total of 13,073 protein-coding genes, 18.43% (35.40 Mb) repetitive elements, and 376 non-coding RNAs. Our study is the first time to report the chromosome-scale genome for the species of A. aphidimyza. It provides a valuable genomic resource for the molecular study of A. aphidimyza.

Divergent evolution of male-determining loci on proto-Y chromosomes of the housefly.

Houseflies provide a good experimental model to study the initial evolutionary stages of a primary sex-determining locus because they possess different recently evolved proto-Y chromosomes that contain male-determining loci (M) with the same male-determining gene, Mdmd. We investigate M-loci genomically and cytogenetically revealing distinct molecular architectures among M-loci. M on chromosome V (MV) has two intact Mdmd copies in a palindrome. M on chromosome III (MIII) has tandem duplications containing 88 Mdmd copies (only one intact) and various repeats, including repeats that are XY-prevalent. M on chromosome II (MII) and the Y (MY) share MIII-like architecture, but with fewer repeats. MY additionally shares MV-specific sequence arrangements. Based on these data and karyograms using two probes, one derives from MIII and one Mdmd-specific, we infer evolutionary histories of polymorphic M-loci, which have arisen from unique translocations of Mdmd, embedded in larger DNA fragments, and diverged independently into regions of varying complexity.

A Chromosome-Level Genome Assembly and Annotation for the Oecanthus rufescens (Orthoptera: Oecanthidae).

Oecanthus is a genus of cricket known for its distinctive chirping and distributed across major zoogeographical regions worldwide. This study focuses on Oecanthus rufescens, and conducts a comprehensive examination of its genome through genome sequencing technologies and bioinformatic analysis. A high-quality chromosome-level genome of O. rufescens was successfully obtained, revealing significant features of its genome structure. The genome size is 877.9 Mb, comprising ten pseudo-chromosomes and 70 other sequences, with a GC content of 41.38% and an N50 value of 157,110,771 bp, indicating a high level of continuity. BUSCO assessment results demonstrate that the genome's integrity and quality are high (of which 96.8% are single-copy and 1.6% are duplicated). Comprehensive genome annotation was also performed, identifying approximately 310 Mb of repetitive sequences, accounting for 35.3% of the total genome sequence, and discovering 15,481 tRNA genes, 4,082 rRNA genes, and 1,212 other noncoding genes. Furthermore, 15,031 protein-coding genes were identified, with BUSCO assessment results showing that 98.4% (of which 96.3% are single-copy and 1.6% are duplicated) of the genes were annotated.

Chromosome-Level Assembly and Annotation of the Genome of the Endangered Giant Patagonian Bumble Bee Bombus dahlbomii.

This article describes a genome assembly and annotation for Bombus dahlbomii, the giant Patagonian bumble bee. DNA from a single, haploid male collected in Argentina was used for PacBio (HiFi) sequencing, and Hi-C technology was then used to map chromatin contacts. Using Juicer and manual curation, the genome was scaffolded into 18 main pseudomolecules, representing a high-quality, near chromosome-level assembly. The sequenced genome size is estimated at 265 Mb. The genome was annotated based on RNA sequencing data of another male from Argentina, and BRAKER3 produced 15,767 annotated genes. The genome and annotation show high completeness, with >95% BUSCO scores for both the genome and annotated genes (based on conserved genes from Hymenoptera). This genome provides a valuable resource for studying the biology of this iconic and endangered species, as well as for understanding the impacts of its decline and designing strategies for its preservation.

Chromosome-level Genome Assembly of Theretra japonica (Lepidoptera: Sphingidae).

Theretra japonica is an important pollinator and agricultural pest in the family Sphingidae with a wide range of host plants. High-quality genomic resources facilitate investigations into behavioral ecology, morphological and physiological adaptations, and the evolution of genomic architecture. However, chromosome-level genome of T. japonica is still lacking. Here we sequenced and assembled the high-quality genome of T. japonica by combining PacBio long reads, Illumina short reads, and Hi-C data. The genome was contained in 95 scaffolds with an accumulated length of 409.55 Mb (BUSCO calculated a genome completeness of 99.2%). The 29 pseudochromosomes had a combined length of 403.77 Mb, with a mapping rate of 98.59%. The genomic characterisation of T. japonica will contribute to further studies for Sphingidae and Lepidoptera.

Evolutionary patterns and functional effects of 3D chromatin structures in butterflies with extensive genome rearrangements.

Chromosome rearrangements may distort 3D chromatin architectures and thus change gene regulation, yet how 3D chromatin structures evolve in insects is largely unknown. Here, we obtain chromosome-level genomes for four butterfly species, Graphium cloanthus, Graphium sarpedon, Graphium eurypylus with 2n = 30, 40, and 60, respectively, and Papilio bianor with 2n = 60. Together with large-scale Hi-C data, we find that inter-chromosome rearrangements very rarely disrupted the pre-existing 3D chromatin structure of ancestral chromosomes. However, some intra-chromosome rearrangements changed 3D chromatin structures compared to the ancestral configuration. We find that new TADs and subTADs have emerged across the rearrangement sites where their adjacent compartments exhibit uniform types. Two intra-chromosome rearrangements altered Rel and lft regulation, potentially contributing to wing patterning differentiation and host plant choice. Notably, butterflies exhibited chromatin loops between Hox gene cluster ANT-C and BX-C, unlike Drosophila. Our CRISPR-Cas9 experiments in butterflies confirm that knocking out the CTCF binding site of the loops in BX-C affected the phenotypes regulated by Antp in ANT-C, resulting in legless larva. Our results reveal evolutionary patterns of insect 3D chromatin structures and provide evidence that 3D chromatin structure changes can play important roles in the evolution of traits.

W chromosome sequences of two bombycid moths provide an insight into the origin of Fem.

Fem is a W-linked gene that encodes a piRNA precursor, and its product, Fem piRNA, is a master factor of female determination in Bombyx mori. Fem has low similarity to any known sequences, and the origin of Fem remains unclear. So far, two hypotheses have been proposed for the origin of Fem: The first hypothesis is that Fem is an allele of Masc, which assumes that the W chromosome was originally a homologous chromosome of the Z chromosome. The second hypothesis is that Fem arose by the transposition of Masc to the W chromosome. To explore the origin of Fem, we determined the W chromosome sequences of B. mori and, as a comparison, a closely relative bombycid species of Trilocha varians with a Fem-independent sex determination system. To our surprise, although the sequences of W and Z chromosomes show no homology to each other, a few pairs of homologues are shared by W and Z chromosomes, indicating the W chromosome of both species originated from Z chromosome. In addition, the W chromosome of T. varians lacks Fem, while the W chromosome of B. mori has over 100 copies of Fem. The high-quality assembly of the W chromosome of B. mori arose the third hypothesis about the origin of Fem: Fem is a chimeric sequence of multiple transposons. More than half of one transcriptional unit of Fem shows a significant homology to RTE-BovB. Moreover, the Fem piRNA-producing region could correspond to the boundary of the two transposons, gypsy and satellite DNA.

Chromosome segregation during spermatogenesis occurs through a unique center-kinetic mechanism in holocentric moth species.

Precise regulation of chromosome dynamics in the germline is essential for reproductive success across species. Yet, the mechanisms underlying meiotic chromosomal events such as homolog pairing and chromosome segregation are not fully understood in many species. Here, we employ Oligopaint DNA FISH to investigate mechanisms of meiotic homolog pairing and chromosome segregation in the holocentric pantry moth, Plodia interpunctella, and compare our findings to new and previous studies in the silkworm moth, Bombyx mori, which diverged from P. interpunctella over 100 million years ago. We find that pairing in both Bombyx and Plodia spermatogenesis is initiated at gene-rich chromosome ends. Additionally, both species form rod shaped cruciform-like bivalents at metaphase I. However, unlike the telomere-oriented chromosome segregation mechanism observed in Bombyx, Plodia can orient bivalents in multiple different ways at metaphase I. Surprisingly, in both species we find that kinetochores consistently assemble at non-telomeric loci toward the center of chromosomes regardless of where chromosome centers are located in the bivalent. Additionally, sister kinetochores do not seem to be paired in these species. Instead, four distinct kinetochores are easily observed at metaphase I. Despite this, we find clear end-on microtubule attachments and not lateral microtubule attachments co-orienting these separated kinetochores. These findings challenge the classical view of segregation where paired, poleward-facing kinetochores are required for accurate homolog separation in meiosis I. Our studies here highlight the importance of exploring fundamental processes in non-model systems, as employing novel organisms can lead to the discovery of novel biology.

Multiple independent origins of the female W chromosome in moths and butterflies.

Lepidoptera, the most diverse group of insects, exhibit female heterogamy (Z0 or ZW), which is different from most other insects (male heterogamy, XY). Previous studies suggest a single origin of the Z chromosome. However, the origin of the lepidopteran W chromosome remains poorly understood. Here, we assemble the genome from females down to the chromosome level of a model insect (Bombyx mori) and identify a W chromosome of approximately 10.1 megabase using a newly developed tool. In addition, we identify 3593 genes that were not previously annotated in the genomes of B. mori. Comparisons of 21 lepidopteran species (including 17 ZW and four Z0 systems) and three trichopteran species (Z0 system) reveal that the formation of Ditrysia W involves multiple mechanisms, including previously proposed canonical and noncanonical models, as well as a newly proposed mechanism called single-Z turnover. We conclude that there are multiple independent origins of the W chromosome in the Ditrysia (most moths and all butterflies) of Lepidoptera.