Near chromosome-level and highly repetitive genome assembly of the snake pipefish Entelurus aequoreus (Syngnathiformes: Syngnathidae).

The snake pipefish, Entelurus aequoreus (Linnaeus, 1758), is a northern Atlantic fish inhabiting open seagrass environments that recently expanded its distribution range. Here, we present a highly contiguous, near chromosome-scale genome of E. aequoreus. The final assembly spans 1.6 Gbp in 7,391 scaffolds, with a scaffold N50 of 62.3 Mbp and L50 of 12. The 28 largest scaffolds (>21 Mbp) span 89.7% of the assembly length. A BUSCO completeness score of 94.1% and a mapping rate above 98% suggest a high assembly completeness. Repetitive elements cover 74.93% of the genome, one of the highest proportions identified in vertebrates. Our demographic modeling identified a peak in population size during the last interglacial period, suggesting the species might benefit from warmer water conditions. Our updated snake pipefish assembly is essential for future analyses of the morphological and molecular changes unique to the Syngnathidae.

Genomic analysis reveals limited hybridization among three giraffe species in Kenya.

BACKGROUND: In the speciation continuum, the strength of reproductive isolation varies, and species boundaries are blurred by gene flow. Interbreeding among giraffe (Giraffa spp.) in captivity is known, and anecdotal reports of natural hybrids exist. In Kenya, Nubian (G. camelopardalis camelopardalis), reticulated (G. reticulata), and Masai giraffe sensu stricto (G. tippelskirchi tippelskirchi) are parapatric, and thus, the country might be a melting pot for these taxa. We analyzed 128 genomes of wild giraffe, 113 newly sequenced, representing these three taxa. RESULTS: We found varying levels of Nubian ancestry in 13 reticulated giraffe sampled across the Laikipia Plateau most likely reflecting historical gene flow between these two lineages. Although comparatively weaker signs of ancestral gene flow and potential mitochondrial introgression from reticulated into Masai giraffe were also detected, estimated admixture levels between these two lineages are minimal. Importantly, contemporary gene flow between East African giraffe lineages was not statistically significant. Effective population sizes have declined since the Late Pleistocene, more severely for Nubian and reticulated giraffe. CONCLUSIONS: Despite historically hybridizing, these three giraffe lineages have maintained their overall genomic integrity suggesting effective reproductive isolation, consistent with the previous classification of giraffe into four species.

Seascape Genomics and Phylogeography of the Sailfish (Istiophorus platypterus).

Permeable phylogeographic barriers characterize the vast open ocean, boosting gene flow and counteracting population differentiation and speciation of widely distributed and migratory species. However, many widely distributed species consists of distinct populations throughout their distribution, evidencing that our understanding of how the marine environment triggers population and species divergence are insufficient. The sailfish is a circumtropical and highly migratory billfish that inhabits warm and productive areas. Despite its ecological and socioeconomic importance as a predator and fishery resource, the species is threatened by overfishing, requiring innovative approaches to improve their management and conservation status. Thus, we presented a novel high-quality reference genome for the species and applied a seascape genomics approach to understand how marine environmental features may promote local adaptation and how it affects gene flow between populations. We delimit two populations between the Atlantic and Indo-Western Pacific oceans and detect outlier loci correlated with sea surface temperature, salinity, oxygen, and chlorophyll concentrations. However, the most significant explanatory factor that explains the differences between populations was isolation by distance. Despite recent population drops, the sailfish populations are not inbred. For billfishes in general, genome-wide heterozygosity was found to be relatively low compared to other marine fishes, evidencing the need to counteract overfishing effects. In addition, in a climate change scenario, management agencies must implement state-of-the-art sequencing methods, consider our findings in their management plans, and monitor genome-wide heterozygosity over time to improve sustainable fisheries and the long-term viability of its populations.

A chromosome-scale reference genome assembly of the great sand eel, Hyperoplus lanceolatus.

Despite increasing sequencing efforts, numerous fish families still lack a reference genome, which complicates genetic research. One such understudied family is the sand lances (Ammodytidae, literally: "sand burrower"), a globally distributed clade of over 30 fish species that tend to avoid tidal currents by burrowing into the sand. Here, we present the first annotated chromosome-level genome assembly of the great sand eel (Hyperoplus lanceolatus). The genome assembly was generated using Oxford Nanopore Technologies long sequencing reads and Illumina short reads for polishing. The final assembly has a total length of 808.5 Mbp, of which 97.1% were anchored into 24 chromosome-scale scaffolds using proximity-ligation scaffolding. It is highly contiguous with a scaffold and contig N50 of 33.7 and 31.3 Mbp, respectively, and has a BUSCO completeness score of 96.9%. The presented genome assembly is a valuable resource for future studies of sand lances, as this family is of great ecological and commercial importance and may also contribute to studies aiming to resolve the suprafamiliar taxonomy of bony fishes.

A Chromosome-Scale Genome Assembly of the Okapi (Okapia johnstoni).

The okapi (Okapia johnstoni), or forest giraffe, is the only species in its genus and the only extant sister group of the giraffe within the family Giraffidae. The species is one of the remaining large vertebrates surrounded by mystery because of its elusive behavior as well as the armed conflicts in the region where it occurs, making it difficult to study. Deforestation puts the okapi under constant anthropogenic pressure, and it is currently listed as "Endangered" on the IUCN Red List. Here, we present the first annotated de novo okapi genome assembly based on PacBio continuous long reads, polished with short reads, and anchored into chromosome-scale scaffolds using Hi-C proximity ligation sequencing. The final assembly (TBG_Okapi_asm_v1) has a length of 2.39 Gbp, of which 98% are represented by 28 scaffolds > 3.9 Mbp. The contig N50 of 61 Mbp and scaffold N50 of 102 Mbp, together with a BUSCO score of 94.7%, and 23 412 annotated genes, underline the high quality of the assembly. This chromosome-scale genome assembly is a valuable resource for future conservation of the species and comparative genomic studies among the giraffids and other ruminants.

Conservation Genomics of Two Threatened Subspecies of Northern Giraffe: The West African and the Kordofan Giraffe.

Three of the four species of giraffe are threatened, particularly the northern giraffe (Giraffa camelopardalis), which collectively have the smallest known wild population estimates. Among the three subspecies of the northern giraffe, the West African giraffe (Giraffa camelopardalis peralta) had declined to 49 individuals by 1996 and only recovered due to conservation efforts undertaken in the past 25 years, while the Kordofan giraffe (Giraffa camelopardalis antiquorum) remains at <2300 individuals distributed in small, isolated populations over a large geographical range in Central Africa. These combined factors could lead to genetically depauperated populations. We analyzed 119 mitochondrial sequences and 26 whole genomes of northern giraffe individuals to investigate their population structure and assess the recent demographic history and current genomic diversity of West African and Kordofan giraffe. Phylogenetic and population structure analyses separate the three subspecies of northern giraffe and suggest genetic differentiation between populations from eastern and western areas of the Kordofan giraffe's range. Both West African and Kordofan giraffe show a gradual decline in effective population size over the last 10 ka and have moderate genome-wide heterozygosity compared to other giraffe species. Recent inbreeding levels are higher in the West African giraffe and in Kordofan giraffe from Garamba National Park, Democratic Republic of Congo. Although numbers for both West African and some populations of Kordofan giraffe have increased in recent years, the threat of habitat loss, climate change impacts, and illegal hunting persists. Thus, future conservation actions should consider close genetic monitoring of populations to detect and, where practical, counteract negative trends that might develop.

Population analysis of retrotransposons in giraffe genomes supports RTE decline and widespread LINE1 activity in Giraffidae.

BACKGROUND: The majority of structural variation in genomes is caused by insertions of transposable elements (TEs). In mammalian genomes, the main TE fraction is made up of autonomous and non-autonomous non-LTR retrotransposons commonly known as LINEs and SINEs (Long and Short Interspersed Nuclear Elements). Here we present one of the first population-level analysis of TE insertions in a non-model organism, the giraffe. Giraffes are ruminant artiodactyls, one of the few mammalian groups with genomes that are colonized by putatively active LINEs of two different clades of non-LTR retrotransposons, namely the LINE1 and RTE/BovB LINEs as well as their associated SINEs. We analyzed TE insertions of both types, and their associated SINEs in three giraffe genome assemblies, as well as across a population level sampling of 48 individuals covering all extant giraffe species. RESULTS: The comparative genome screen identified 139,525 recent LINE1 and RTE insertions in the sampled giraffe population. The analysis revealed a drastically reduced RTE activity in giraffes, whereas LINE1 is still actively propagating in the genomes of extant (sub)-species. In concert with the extremely low activity of the giraffe RTE, we also found that RTE-dependent SINEs, namely Bov-tA and Bov-A2, have been virtually immobile in the last 2 million years. Despite the high current activity of the giraffe LINE1, we did not find evidence for the presence of currently active LINE1-dependent SINEs. TE insertion heterozygosity rates differ among the different (sub)-species, likely due to divergent population histories. CONCLUSIONS: The horizontally transferred RTE/BovB and its derived SINEs appear to be close to inactivation and subsequent extinction in the genomes of extant giraffe species. This is the first time that the decline of a TE family has been meticulously analyzed from a population genetics perspective. Our study shows how detailed information about past and present TE activity can be obtained by analyzing large-scale population-level genomic data sets.

Whole-genome analysis of giraffe supports four distinct species.

Species is the fundamental taxonomic unit in biology and its delimitation has implications for conservation. In giraffe (Giraffa spp.), multiple taxonomic classifications have been proposed since the early 1900s.1 However, one species with nine subspecies has been generally accepted,2 likely due to limited in-depth assessments, subspecies hybridizing in captivity,3,4 and anecdotal reports of hybrids in the wild.5 Giraffe taxonomy received new attention after population genetic studies using traditional genetic markers suggested at least four species.6,7 This view has been met with controversy,8 setting the stage for debate.9,10 Genomics is significantly enhancing our understanding of biodiversity and speciation relative to traditional genetic approaches and thus has important implications for species delineation and conservation.11 We present a high-quality de novo genome assembly of the critically endangered Kordofan giraffe (G. camelopardalis antiquorum)12 and a comprehensive whole-genome analysis of 50 giraffe representing all traditionally recognized subspecies. Population structure and phylogenomic analyses support four separately evolving giraffe lineages, which diverged 230-370 ka ago. These lineages underwent distinct demographic histories and show different levels of heterozygosity and inbreeding. Our results strengthen previous findings of limited gene flow and admixture among putative giraffe species6,7,9 and establish a genomic foundation for recognizing four species and seven subspecies, the latter of which should be considered as evolutionary significant units. Achieving a consensus over the number of species and subspecies in giraffe is essential for adequately assessing their threat level and will improve conservation efforts for these iconic taxa.

Education in the genomics era: Generating high-quality genome assemblies in university courses.

Recent advances in genome sequencing technologies have simplified the generation of genome data and reduced the costs for genome assemblies, even for complex genomes like those of vertebrates. More practically oriented genomic courses can prepare university students for the increasing importance of genomic data used in biological and medical research. Low-cost third-generation sequencing technology, along with publicly available data, can be used to teach students how to process genomic data, assemble full chromosome-level genomes, and publish the results in peer-reviewed journals, or preprint servers. Here we outline experiences gained from 2 master's-level courses and discuss practical considerations for teaching hands-on genome assembly courses.

Improving the Chromosome-Level Genome Assembly of the Siamese Fighting Fish (Betta splendens) in a University Master's Course.

Ever decreasing costs along with advances in sequencing and library preparation technologies enable even small research groups to generate chromosome-level assemblies today. Here we report the generation of an improved chromosome-level assembly for the Siamese fighting fish (Betta splendens) that was carried out during a practical university master's course. The Siamese fighting fish is a popular aquarium fish and an emerging model species for research on aggressive behavior. We updated the current genome assembly by generating a new long-read nanopore-based assembly with subsequent scaffolding to chromosome-level using previously published Hi-C data. The use of ∼35x nanopore-based long-read data sequenced on a MinION platform (Oxford Nanopore Technologies) allowed us to generate a baseline assembly of only 1,276 contigs with a contig N50 of 2.1 Mbp, and a total length of 441 Mbp. Scaffolding using the Hi-C data resulted in 109 scaffolds with a scaffold N50 of 20.7 Mbp. More than 99% of the assembly is comprised in 21 scaffolds. The assembly showed the presence of 96.1% complete BUSCO genes from the Actinopterygii dataset indicating a high quality of the assembly. We present an improved full chromosome-level assembly of the Siamese fighting fish generated during a university master's course. The use of ∼35× long-read nanopore data drastically improved the baseline assembly in terms of continuity. We show that relatively in-expensive high-throughput sequencing technologies such as the long-read MinION sequencing platform can be used in educational settings allowing the students to gain practical skills in modern genomics and generate high quality results that benefit downstream research projects.

Chromosome-level genome assembly of a benthic associated Syngnathiformes species: the common dragonet, Callionymus lyra.

Background: The common dragonet, Callionymus lyra, is one of three Callionymus species inhabiting the North Sea. All three species show strong sexual dimorphism. The males show strong morphological differentiation, e.g., species-specific colouration and size relations, while the females of different species have few distinguishing characters. Callionymus belongs to the 'benthic associated clade' of the order Syngnathiformes. The 'benthic associated clade' so far is not represented by genome data and serves as an important outgroup to understand the morphological transformation in 'long-snouted' syngnatiformes such as seahorses and pipefishes. Findings: Here, we present the chromosome-level genome assembly of C. lyra. We applied Oxford Nanopore Technologies' long-read sequencing, short-read DNBseq, and proximity-ligation-based scaffolding to generate a high-quality genome assembly. The resulting assembly has a contig N50 of 2.2 Mbp and a scaffold N50 of 26.7 Mbp. The total assembly length is 568.7 Mbp, of which over 538 Mbp were scaffolded into 19 chromosome-length scaffolds. The identification of 94.5% complete BUSCO genes indicates high assembly completeness. Additionally, we sequenced and assembled a multi-tissue transcriptome with a total length of 255.5 Mbp that was used to aid the annotation of the genome assembly. The annotation resulted in 19,849 annotated transcripts and identified a repeat content of 27.7%. Conclusions: The chromosome-level assembly of C. lyra provides a high-quality reference genome for future population genomic, phylogenomic, and phylogeographic analyses.

Phylogeographic history of South American populations of the silky anteater Cyclopes didactylus (Pilosa: Cyclopedidae)

Abstract Cyclopes didactylus, commonly called silky anteater, is the smallest and least studied of the anteaters. It is an arboreal species occurring in rainforests, ranging from southern Mexico to Central and South America, with an apparently disjoint distribution between Amazon and Atlantic rainforests in Brazil. Although seven subspecies are recognized, little is known about its geographical variation. Thus, to evaluate the population dynamics and evolutionary history of the South American silky anteater, we analyzed 1542 bp sequences of the mitochondrial control region (CR), COI and Cyt-b genes of 32 individuals. Haplotype network, AMOVA and molecular dating analyses were performed and identified seven geographic clusters. The split of lineages separating Cyclopedidae (Cyclopes) and Myrmecophagidae (Myrmecophaga and Tamandua genera) was estimated around 41 million years ago (mya), and the intraspecific lineage diversification of C. didactylus began in the Miocene around 13.5 mya, likely in southwestern Amazonia. Tectonic and climatic events that took place in South America during the Tertiary and Quaternary seem to have influenced the evolutionary history of the species at different levels. This is the first study to investigate the population dynamics and phylogeography of the silky anteater, which contributes to a better comprehension of the biogeography of South America.

Phylogeographic history of South American populations of the silky anteater Cyclopes didactylus (Pilosa: Cyclopedidae).

Cyclopes didactylus, commonly called silky anteater, is the smallest and least studied of the anteaters. It is an arboreal species occurring in rainforests, ranging from southern Mexico to Central and South America, with an apparently disjoint distribution between Amazon and Atlantic rainforests in Brazil. Although seven subspecies are recognized, little is known about its geographical variation. Thus, to evaluate the population dynamics and evolutionary history of the South American silky anteater, we analyzed 1542 bp sequences of the mitochondrial control region (CR), COI and Cyt-b genes of 32 individuals. Haplotype network, AMOVA and molecular dating analyses were performed and identified seven geographic clusters. The split of lineages separating Cyclopedidae (Cyclopes) and Myrmecophagidae (Myrmecophaga and Tamandua genera) was estimated around 41 million years ago (mya), and the intraspecific lineage diversification of C. didactylus began in the Miocene around 13.5 mya, likely in southwestern Amazonia. Tectonic and climatic events that took place in South America during the Tertiary and Quaternary seem to have influenced the evolutionary history of the species at different levels. This is the first study to investigate the population dynamics and phylogeography of the silky anteater, which contributes to a better comprehension of the biogeography of South America.