Sex-specific ecology has management implications, but rapid sex-chromosome turnover in fishes hinders development of markers to sex monomorphic species. Here, we use annotated genomes and reduced-representation sequencing data for two Australian percichthyids, the Macquarie perch Macquaria australasica and the golden perch M. ambigua, and whole genome resequencing data for 50 Macquarie perch of each sex, to detect sex-linked loci, identify a candidate sex-determining gene and develop an affordable sexing assay. In-silico pool-seq tests of 1,492,004 Macquarie perch SNP loci revealed that a 275-Kb scaffold, containing the transcription factor SOX1b gene, was enriched for gametologous loci. Within this scaffold, 22 loci were sex-linked in a predominantly XY system, with females being homozygous at all 22, and males being heterozygous at two or more. Seven XY-gametologous loci were within a 146-bp region. Being ~38 Kb upstream of SOX1b, it might act as an enhancer controlling SOX1b transcription in the bipotential gonad that drives gonad differentiation. A PCR-RFLP sexing assay, targeting one of the Y-linked SNPs, tested in 66 known-sex Macquarie perch and two individuals of each sex of three confamilial species, and amplicon sequencing of 400 bp encompassing the 146-bp region, revealed that the few sex-linked positions differ between species and between Macquarie perch populations. This indicates sex-chromosome lability in Percichthyidae, also supported by non-homologous scaffolds containing sex-linked loci for Macquarie- and golden perches. The resources developed here will facilitate genomic research in Percichthyidae. Sex-linked markers will be useful for determining genetic sex in some populations and studying sex chromosome turnover.

The hyper-diverse order Coleoptera comprises a staggering ~25% of known species on Earth. Despite recent breakthroughs in next generation sequencing, there remains a limited representation of beetle diversity in assembled genomes. Most notably, the ground beetle family Carabidae, comprising more than 40,000 described species, has not been studied in a comparative genomics framework using whole genome data. Here we generate a high-quality genome assembly for Nebria riversi, to examine sources of novelty in the genome evolution of beetles, as well as genetic changes associated with specialization to high elevation alpine habitats. In particular, this genome resource provides a foundation for expanding comparative molecular research into mechanisms of insect cold adaptation. Comparison to other beetles shows a strong signature of genome compaction, with N. riversi possessing a relatively small genome (~147 Mb) compared to other beetles, with associated reductions in repeat element content and intron length. Small genome size is not, however, associated with fewer protein-coding genes, and an analysis of gene family diversity shows significant expansions of genes associated with cellular membranes and membrane transport, as well as protein phosphorylation and muscle filament structure. Finally, our genomic analyses show that these high elevation beetles have endosymbiotic Spiroplasma, with several metabolic pathways (e.g. propanoate biosynthesis) that might complement N. riversi, although its role as a beneficial symbiont or as a reproductive parasite remains equivocal.

Schistosomiasis is a neglected tropical disease of humans caused by blood flukes of the genus Schistosoma – the only dioecious parasitic flatworms. Although aspects of sex determination, differentiation and reproduction have been studied in some Schistosoma species, almost nothing is understood for Schistosoma japonicum - the causative agent of schistosomiasis japonica. This relates mainly to a lack of high-quality genomic and transcriptomic resources for this species. As current draft genomes for S. japonicum are highly fragmented, we assembled here a chromosome-level reference genome (seven autosomes, the Z-chromosome and partial W-chromosome), achieving a substantially enhanced gene annotation. Utilising this genome, we discovered that the sex chromosomes of S. japonicum and its congener S. mansoni independently suppressed recombination during evolution, forming four and two ‘strata’, respectively. By exploring the W-chromosome and sex-specific transcriptomes, we identified 35 W-linked genes and 257 female-preferentially transcribed genes (FTGs) and identified a signature for sex determination and differentiation in S. japonicum. These FTGs cluster within autosomes or the Z-chromosome and exhibit a highly dynamic transcription profile during the pairing of female and male schistosomules (advanced juveniles), representing a critical phase for the maturation of the female worms, suggesting distinct layers of regulatory control of gene transcription at this stage of development. Collectively, these data provide a valuable resource for further functional genomic characterisation of S. japonicum, shed light on the evolution of sex chromosomes in this highly virulent human blood fluke and provide a pathway to identify novel targets for development of intervention tools against schistosomiasis.

Revegetation projects face the major challenge of sourcing the optimal plant material. This is often done with limited information about plant performance and increasingly requires to factor resilience to climate change. Functional traits can be used as quantitative indices of plant performance and guide provenancing, but trait values expected under novel conditions are often unkown. To support climate-resilient provenancing efforts, we develop a trait prediction model that integrates the effect of genetic variation with fine-scale temperature variation. We train our model on multiple field plantings of Arabidopsis thaliana and predict two relevant fitness traits -- days-to-bolting and fecundity -- across the species' European range. Prediction accuracies were high for days-to-bolting and moderate for fecundity, with the majority of trait variation explained by temperature differences between plantings. Projection under future climate predicted a decline in fecundity, although this response was heterogeneous across the range. In response, we identified novel genotypes that could be introduced to genetically offset the fitness decay. Our study highlights the value of predictive models to aid seed provenancing and improve the success of revegetation projects.

Karyotypic changes in chromosome number and structure are drivers in the divergent evolution of diverse plant species and lineages. This study aimed to reveal the origins of the unique karyotype (2n = 12) and phylogenetic relationships of the genus Megadenia (Brassicaceae). A high-quality chromosome-scale genome was assembled for Megadenia pygmaea using Nanopore long reads and high-throughput chromosome conformation capture (Hi-C). The assembled genome is 215.2-Mb and is anchored on six pseudo-chromosomes. We annotated a total of 25,607 high-confidence protein-coding genes and corroborated the phylogenetic affinity of Megadenia with the expanded Lineage II, which contains numerous agricultural crops. We dated the divergence of Megadenia from its closest relatives to 27.04 (19.11-36.60) million years ago. A reconstruction of the chromosomal composition of the species was performed based on the de novo assembled genome and comparative chromosome painting analysis. The karyotype structure of M. pygmaea is very similar to the previously inferred Proto-Calepineae Karyotype (PCK; n = 7) of the Brassicaceae Lineage II. However, an end-to-end translocation between two ancestral chromosomes reduced the chromosome number from n = 7 to n = 6, comparable to Megadenia. Our reference genome provides fundamental information for use in horticulture, plant breeding and evolutionary study of this genus.

Despite the increasing accessibility of high-throughput sequencing, obtaining high-quality genomic data on non-model organisms without proximate well-assembled and annotated genomes remains challenging. Here we describe a workflow that takes advantage of distant genomic resources and ingroup transcriptomes to select and jointly enrich long open reading frames (ORFs) and ultraconserved elements (UCEs) from genomic samples for integrative studies of microevolutionary and macroevolutionary dynamics. This workflow is applied to samples of the African unionid bivalve tribe Coelaturini (Parreysiinae) at basin and continent-wide scales. Our results indicate that ORFs are efficiently captured without prior identification of intron-exon boundaries. The enrichment of UCEs was less successful, but nevertheless produced substantial datasets. Exploratory continent-wide phylogenetic analyses with ORF supercontigs (>515,000 parsimony informative sites) resulted in a fully resolved phylogeny, the backbone of which was also retrieved with UCEs (>11,000 informative sites). Variant calling on ORFs and UCEs of Coelaturini from the Malawi Basin produced ~2,000 SNPs per population pair. Estimates of nucleotide diversity and population differentiation were similar for ORFs and UCEs. They were low compared to previous estimates in mollusks, but comparable to those in recently diversifying Malawi cichlids and other taxa at an early stage of speciation. Skimming off-target sequence data from the same enriched libraries of Coelaturini from the Malawi Basin, we reconstructed the maternally-inherited mitogenome, which displays the gene order inferred for the most recent common ancestor of Unionidae. Overall, our workflow and results provide exciting perspectives for integrative genomic studies of microevolutionary and macroevolutionary dynamics in non-model organisms.

Sarcophaga peregrina is usually considered to be of great ecological, medical and forensic significance, and has the biological characteristics such as the ovoviviparous reproductive pattern and adaptation to feed on carrion. However, the underlying mechanisms still remain unsolved by lack of high-quality genome. Here we present de novo–assembled genome at chromosome-scale for S. peregrina. The final assembled genome was 560.31 Mb with contig N50 of 3.84 Mb. Hi-C scaffolding reliably anchored six pseudochromosomes, accounting for 97.76% of the assembled genome. Moreover, 45.70% of repeat elements were identified in the genome. A total of 14,476 protein-coding genes were functionally annotated, accounting for 92.14% of all predicted genes. Phylogenetic analysis indicated that S. peregrina and S. bullata diverged ~7.14 Mya. Comparative genomic analysis revealed expanded and positively selected genes related to biological features that aid in clarifying its ovoviviparous reproduction and necrophagous habit, such as horionic membrane formation and Dorso-ventral axis formation, lipid metabolism, and olfactory receptor activity. This study provides a valuable genomic resource of S. peregrina, and sheds insight into further revealing the underlying molecular mechanisms of adaptive evolution.

Understanding landscape connectivity has become a global priority for mitigating the impact of landscape fragmentation on biodiversity. Link-based methods traditionally rely on relating pairwise genetic distance between individuals or demes to their landscape distance (e.g., geographic distance, cost distance). In this study, we present an alternative to conventional statistical approaches to refine cost surfaces by adapting the Gradient Forest (GF) approach to produce a resistance surface. Used in community ecology, GF is an extension of random forest (RF), and has been implemented in genomic studies to model species genetic offset under future climatic scenarios. By design, this adapted method, resGF, has the ability to handle multiple environmental predicators and is not subjected to traditional assumptions of linear models such as independence, normality and linearity. Using genetic simulations, resGF performance was compared to other published methods. In univariate scenarios, resGF was able to distinguish the true surface contributing to genetic diversity among competing surfaces better than the compared methods. In multivariate scenarios, the GF approach performed similarly to the other RF-based approach using least-cost transect analysis (LCTA). Additionally, two worked examples are provided using two previously published datasets. This machine learning algorithm has the potential to improve our understanding of landscape connectivity and can inform long-term biodiversity conservation strategies.

Interactions of organisms with their environment are complex and environmental regulation at different levels of biological organization is often non-linear. Therefore, the genotype to phenotype continuum requires study at multiple levels of organization. While studies of transcriptome regulation are now common for many species, quantitative studies of environmental effects on proteomes are needed. Here we report the generation of a data-independent acquisition (DIA) assay library that enables simultaneous targeted proteomics of thousands of Oreochromis niloticus kidney proteins using a label- and gel-free workflow that is well suited for ecologically relevant field samples. We demonstrate the usefulness of this DIA assay library by discerning environmental effects on the kidney proteome of O. niloticus. Moreover, we demonstrate that the DIA assay library approach generates data that are complimentary rather than redundant to transcriptomics data. Transcript and protein abundance differences in kidneys of tilapia acclimated to freshwater and brackish water (25 g/kg) were correlated for 2114 unique genes. A high degree of non-linearity in salinity-dependent regulation of transcriptomes and proteomes was revealed suggesting that the regulation of O. niloticus renal function by environmental salinity relies heavily on post-transcriptional mechanisms. The application of functional enrichment analyses using STRING and KEGG to DIA assay datasets is demonstrated by identifying myo-inositol metabolism, antioxidant and xenobiotic functions, and signaling mechanisms as key elements controlled by salinity in tilapia kidneys. The DIA assay library resource presented here can be adopted for other tissues and other organisms to study proteome dynamics during changing ecological contexts.

Palaeolimnological records provide valuable information about how phytoplankton respond to long-term drivers of environmental change. Traditional palaeolimnological tools such as microfossils and pigments are restricted to taxa that leave sub-fossil remains, and a method that can be applied to the wider community is required. Sedimentary DNA (sedDNA), extracted from lake sediment cores, shows promise in palaeolimnology, but validation against data from long-term monitoring of lake water is necessary to enable its development as a reliable record of past phytoplankton communities. To address this need, 18S rRNA gene amplicon sequencing was carried out on lake sediments from a core collected from Esthwaite Water (English Lake District) spanning ~105 years. This sedDNA record was compared with concurrent long-term microscopy-based monitoring of phytoplankton in the surface water. Broadly comparable trends were observed between the datasets, with respect to the diversity and relative abundance and occurrence of chlorophytes, dinoflagellates, ochrophytes and bacillariophytes. Up to 20% of genera identified in the microscopy record were also detected using sedDNA, and sedDNA revealed a previously undetected community of phytoplankton. However, a substantial proportion of genera identified by microscopy were not detected using sedDNA, highlighting the current limitations of the technique that require further development such as reference database coverage. These results suggest that sedDNA can be used as an effective record of past phytoplankton communities, at least over timescales of less than 100 years, but the taphonomic processes which may affect its reliability, such as the extent and rate of deposition and DNA degradation, require further research.

Age is an essential trait for understanding the ecology and management of wildlife. A conventional method of estimating age in wild animals is counting annuli formed in the cementum of teeth. This method has been used in bears despite some disadvantages, such as high invasiveness and the requirement for experienced observers. In this study, we established a novel age estimation method based on DNA methylation levels using blood collected from 49 brown bears of known ages living in both captivity and the wild. We performed bisulfite pyrosequencing and obtained methylation levels at 39 cytosine-phosphate-guanine (CpG) sites adjacent to 12 genes. The methylation levels of CpGs adjacent to four genes showed a significant correlation with age. The best model was based on DNA methylation levels at just four CpG sites adjacent to a single gene, SLC12A5, and it had high accuracy with a mean absolute error of 1.3 years and median absolute error of 1.0 year after leave-one-out cross-validation. This model represents the first epigenetic method of age estimation in brown bears, which provides benefits over tooth-based methods, including high accuracy, less invasiveness, and a simple procedure. Our model has the potential for application to other bear species, which will greatly improve ecological research, conservation, and management.

Age is necessary information for the study of life history of wild animals. A general method to estimate the age of odontocetes is counting dental growth layer groups (GLGs). However, this method is highly invasive as it requires the capture and handling of individuals to collect their teeth. Recently, the development of DNA-based age estimation methods has been actively studied as an alternative to such invasive methods, of which many have used biopsy samples. However, if DNA-based age estimation can be developed from fecal samples, age estimation can be performed without touching or disrupting individuals, thus establishing an entirely non-invasive method. We developed an age estimation model using the methylation rate of two gene regions, GRIA2 and CDKN2A, measured through methylation-sensitive high-resolution melting (MS-HRM) from fecal samples of wild Indo-Pacific bottlenose dolphins (Tursiops aduncus). The age of individuals was known through conducting longitudinal individual identification surveys underwater. Methylation rates were quantified from 36 samples. Both gene regions showed a significant correlation between age and methylation rate. The age estimation model was constructed based on the methylation rates of both genes which achieved sufficient accuracy (after LOOCV: MAE = 5.08, R2 = 0.34) for the ecological studies of the Indo-Pacific bottlenose dolphins, with a lifespan of 40-50 years. This is the first study to report the use of non-invasive fecal samples to estimate the age of marine mammals.

Dispersal is a crucial mechanism to living beings, allowing them to reach new resources such that populations and species can explore new environments. However, directly observing the dispersal mechanisms of widespread species can be costly or even impracticable, which is the case for mangrove trees. The influence of ocean currents on the mangroves’ propagules’ movement has been increasingly evident; however, few studies mechanistically relate the patterns of population distribution with the dispersal by oceanic currents under an integrated framework. Here, we evaluate the role of oceanic currents on dispersal and connectivity of Rhizophora mangle along the Southwest Atlantic. We inferred population genetic structure and migration rates based on single nucleotide polymorphisms, simulated the displacement of propagules along the region and tested our hypotheses with Mantel tests and redundancy analysis. We observed a two populations structure, north and south, which is corroborated by other studies with Rhizophora and other coastal plants. The inferred recent migration rates do not indicate gene flow between the sampled sites. Conversely, long-term migration rates were low across groups and contrasting dispersal patterns within each one, which is consistent with long-distance dispersal events. Our hypothesis tests suggests that both isolation by distance and isolation by oceanography (derived from the oceanic currents) can explain the neutral genetic variation of R. mangle in the region. Our findings expand current knowledge of mangrove connectivity and highlight how the association of molecular methods with oceanographic simulations improve the interpretation power of the dispersal process, which has ecological and evolutionary implications.

Technological advances in DNA sequencing over the last decade now permit the production and curation of large genomic datasets in an increasing number of non-model species. Additionally, this new data provides the opportunity for combining datasets, resulting in larger studies with a broader taxonomic range. Whilst the benefits of new sequencing platforms are obvious, shifts in sequencing technology can also pose challenges for those wishing to combine new sequencing data with data sequenced on older platforms. Here, we outline the types of studies where the use of curated data might be beneficial, and highlight potential biases that might be introduced by combining data from different sequencing platforms. As an example of the challenges associated with combining data across sequencing platforms, we focus on the impact of the shift in Illumina's base calling technology from a four-channel to a two-channel system. We caution that when data is combined from these two systems, erroneous guanine base calls that result from the two-channel chemistry can make their way through a bioinformatic pipeline, eventually leading to inaccurate and potentially misleading conclusions. We also suggest solutions for dealing with such potential artifacts, which make samples sequenced on different sequencing platforms appear more differentiated from one another than they really are. Finally, we stress the importance of archiving tissue samples and the associated sequences for the continued reproducibility and reusability of sequencing data in the face of ever-changing sequencing platform technology.

Fungi form diverse communities and play essential roles in many terrestrial ecosystems, yet there are methodological challenges in taxonomic and phylogenetic placement of fungi from environmental sequences. To address such challenges we investigated spatio-temporal structure of a fungal community using soil metabarcoding with four different sequencing strategies: short amplicon sequencing of the ITS2 region (300–400\ bp) with Illumina MiSeq, Ion Torrent Ion S5, and PacBio RS II, all from the same PCR library, as well as long amplicon sequencing of the full ITS and partial LSU regions (1200–1600\ bp) with PacBio RS II. Resulting community structure and diversity depended more on statistical method than sequencing technology. The use of long-amplicon sequencing enables construction of a phylogenetic tree from metabarcoding reads, which facilitates taxonomic identification of sequences. However, long reads present issues for denoising algorithms in diverse communities. We present a solution that splits the reads into shorter homologous regions prior to denoising, and then reconstructs the full denoised reads. In the choice between short and long amplicons, we suggest a hybrid approach using short amplicons for sampling breadth and depth, and long amplicons to characterize the local species pool for improved identification and phylogenetic analyses.

[Definitive version of this article may be found here] The mitochondrial gene cytochrome-c-oxidase subunit 1 (COI) is useful in many taxa for phylogenetics, population genetics, metabarcoding, and rapid species identifications. However, the phylum Ctenophora (comb jellies) has historically been difficult to study due to divergent mitochondrial sequences and the corresponding inability to amplify COI with degenerate and standard COI ‘barcoding’ primers. As a result, there are very few COI sequences available for ctenophores, despite over 200 described species in the phylum. Here, we designed new primers and amplified the COI fragment from members of all major groups of ctenophores, including many undescribed species. Phylogenetic analyses of the resulting COI sequences revealed high diversity within many groups that was not evident from more conserved 18S rDNA sequences, in particular among the Lobata. The COI phylogenetic results also revealed unexpected community structure within the genus Bolinopsis, suggested new species within the genus Bathocyroe, and supported the ecological and morphological differences of some species such as Lampocteis cruentiventer and similar lobates (Lampocteis sp. ‘V’ stratified by depth, and ‘A’ differentiated by color). The newly described primers reported herein provide important tools to enable researchers to illuminate the diversity of ctenophores worldwide via quick molecular identifications, improve the ability to analyze environmental DNA by improving reference libraries and amplifications, and enable a new breadth of population genetic studies.

Single-nucleotide polymorphism (SNP) analyses are a powerful tool for population genetics, pedigree reconstruction and phenotypic trait mapping. SNPs could also be useful for sexing individuals in species with reduced sexual dimorphism, yet this possibility remains poorly explored. Here, we develop a novel protocol for molecular sexing of birds based on the detection of unique Z- and W-linked SNP markers. Our method is based on the identification of two unique loci, one in each sexual chromosome. Individuals are considered males when they are heterozygotic for the Z-linked SNP and females when they are homozygote for the Z-linked SNP and have the W-linked SNP. We validated the method in the Jackdaw (Corvus monedula), a species whose reduced sexual dimorphism makes it difficult to sex individuals in the wild. We assessed the reliability of the method with 36 individuals of known sex, and found that their sex was correctly assigned in 100% of cases. The sex-linked markers also proved to be widely applicable to discriminate males and females from a sample of 927 genotyped individuals of different maturity stages with an accuracy of 99.5%. Given that SNP markers are increasingly used in quantitative genetic analyses of wild populations, the approach we propose has a great potential to be integrated into broader genetic research programmes without the need of additional sexing techniques.

A high-quality reference genome is necessary to determine the molecular mechanisms underlying important biological phenomena; therefore, in the present study, a chromosome-level genome assembly of the Chinese shrimp Fenneropenaeus chinensis was performed. Muscle of a male shrimp was sequenced using PacBio platform, and assembled by Hi-C technology. The assembled F. chinensis genome was 1,465.32 Mb with contig N50 of 472.84 Kb, including 57.73% repetitive sequences, and was anchored to 43 pseudochromosomes, with scaffold N50 of 36.87 Mb. In total, 25,026 protein-coding genes were predicted. The genome size of F. chinensis showed significant contraction in comparison with that of other penaeid species, which is likely related to migration observed in this species. However, the F. chinensis genome included several expanded gene families related to cellular processes and metabolic processes, and the contracted gene families were associated with virus infection process. The findings signify the adaptation of F. chinensis to the selection pressure of migration and cold environment. Furthermore, the selection signature analysis identified genes associated with metabolism, phototransduction, and nervous system in cultured shrimps when compared with wild population, indicating targeted, artificial selection of growth, vision, and behavior during domestication. The construction of the genome of F. chinensis provided valuable information for the further genetic mechanism analysis of important biological processes, and will facilitate the research of genetic changes during evolution.