A revisited phylogeography of Nautilus pompilius

The cephalopod genus Nautilus is considered a “living fossil” with a contested number of extant and extinct species, and a benthic lifestyle that limits movement of animals between isolated seamounts and landmasses in the Indo‐Pacific. Nautiluses are fished for their shells, most heavily in the Philippines, and these fisheries have little monitoring or regulation. Here, we evaluate the hypothesis that multiple species of Nautilus (e.g., N. belauensis, N. repertus and N. stenomphalus) are in fact one species with a diverse phenotypic and geologic range. Using mitochondrial markers, we show that nautiluses from the Philippines, eastern Australia (Great Barrier Reef), Vanuatu, American Samoa, and Fiji fall into distinct geographical clades. For phylogenetic analysis of species complexes across the range of nautilus, we included sequences of Nautilus pompilius and other Nautilus species from GenBank from localities sampled in this study and others. We found that specimens from Western Australia cluster with samples from the Philippines, suggesting that interbreeding may be occurring between those locations, or that there is limited genetic drift due to large effective population sizes. Intriguingly, our data also show that nautilus identified in other studies as N. belauensis, N. stenomphalus, or N. repertus are likely N. pompilius displaying a diversity of morphological characters, suggesting that there is significant phenotypic plasticity within N. pompilius.     

Genomic analyses of a “living fossil”: The endangered dove‐tree

Davidia involucrata Baill, also known as the dove‐tree, is a living fossil and an endangered species currently restricted to the mountains of southwestern and central China. It has a beautiful and innovative trait of high horticultural value: two white bracts covering the flower caputila. Here, we report on the chromosome‐scale genome of this species using single‐molecule real‐time long reads and chromosome conformation capture (Hi‐C) techniques. This species has a larger genome size of 1,169 Mb and contains relatively more genes (42,554) than the closely related species Camptotheca acuminata (397 Mb and 31,825 genes). Both species shared one recent whole genome duplication before their divergence. The expansion of the repetitive elements after their divergence contributed greatly to the increase in the genome size of the dove‐tree. Photosynthesis‐related genes were almost absent or showed reduced expression in the bracts of the dove‐tree, while defence‐ and chemical‐related genes increased greatly, highlighting the important roles of the bracts in protecting flowers and attracting pollinators. The effective population size of the dove‐tree continuously decreased during the climate changes of the Quaternary. Such climate sensitivity should be fully considered in conservation efforts for this relict endangered species in the context of continuous climate warming in the future.     

Effective population size is strongly correlated with breeding pond size in the endangered California tiger salamander, <Emphasis Type="Italic">Ambystoma californiense</Emphasis>

Maintaining genetic diversity and population viability in endangered and threatened species is a primary concern of conservation biology. Genetic diversity depends on population connectivity and effective population size (N_e), both of which are often compromised in endangered taxa. While the importance of population connectivity and gene flow has been well studied, investigating effective population sizes in natural systems has received far less attention. However, N_e plays a prominent role in the maintenance of genetic diversity, the prevention of inbreeding depression, and in determining the probability of population persistence. In this study, we examined the relationship between breeding pond characteristics and N_e in the endangered California tiger salamander, Ambystoma californiense. We sampled 203 individuals from 10 breeding ponds on a local landscape, and used 11 polymorphic microsatellite loci to quantify genetic structure, gene flow, and effective population sizes. We also measured the areas of each pond using satellite imagery and classified ponds as either hydrologically-modified perennial ponds or naturally occurring vernal pools, the latter of which constitute the natural breeding habitat for A. californiense. We found no correlation between pond area and heterozygosity or allelic diversity, but we identified a strong positive relationship between breeding pond area and N_e, particularly for vernal pools. Our results provide some of the first empirical evidence that variation in breeding habitat can be associated with differences in N_e and suggest that a more complete understanding of the environmental features that influence N_e is an important component of conservation genetics and management.     

Genetic diversity of small populations: Not always “doom and gloom”?

Is a key theory of evolutionary and conservation biology—that loss of genetic diversity can be predicted from population size—on shaky ground? In the face of increasing human‐induced species depletion and habitat fragmentation, this question and the study of genetic diversity in small populations are paramount to understanding the limits of species’ responses to environmental change and to providing remedies to endangered species conservation. Few empirical studies have investigated to what degree some small populations might be buffered against losses of genetic diversity. Even fewer studies have experimentally tested the potential underlying mechanisms. The study of Schou, Loeschcke, Bechsgaard, Schlotterer, and Kristensen ( 2017 ) in this issue of Molecular Ecology is elegant in combining classic common garden experimentation with population genomics on an iconic experimental model species (Drosophila melanogaster). The authors reveal a slower rate of loss of genetic diversity in small populations under varying thermal regimes than theoretically expected and hence an unexpected retention of genetic diversity. They are further able to hone in on a plausible mechanism: associative overdominance, wherein homozygosity of deleterious recessive alleles is especially disfavoured in genomic regions of low recombination. These results contribute to a budding literature on the varying mechanisms underlying genetic diversity in small populations and encourage further such research towards the effective management and conservation of fragmented or endangered populations.     

Lineage‐specific sequence evolution and exon edge conservation partially explain the relationship between evolutionary rate and expression level in A. thaliana

Rapidly evolving proteins can aid the identification of genes underlying phenotypic adaptation across taxa, but functional and structural elements of genes can also affect evolutionary rates. In plants, the ‘edges’ of exons, flanking intron junctions, are known to contain splice enhancers and to have a higher degree of conservation compared to the remainder of the coding region. However, the extent to which these regions may be masking indicators of positive selection or account for the relationship between dN/dS and other genomic parameters is unclear. We investigate the effects of exon edge conservation on the relationship of dN/dS to various sequence characteristics and gene expression parameters in the model plant Arabidopsis thaliana. We also obtain lineage‐specific dN/dS estimates, making use of the recently sequenced genome of Thellungiella parvula, the second closest sequenced relative after the sister species Arabidopsis lyrata. Overall, we find that the effect of exon edge conservation, as well as the use of lineage‐specific substitution estimates, upon dN/dS ratios partly explains the relationship between the rates of protein evolution and expression level. Furthermore, the removal of exon edges shifts dN/dS estimates upwards, increasing the proportion of genes potentially under adaptive selection. We conclude that lineage‐specific substitutions and exon edge conservation have an important effect on dN/dS ratios and should be considered when assessing their relationship with other genomic parameters.     

The distribution,diversity, and conservation status of Cycas in China

As ancient gymnosperm and woody plants, cycads have survived through dramatic tectonic activities, climate fluctuation, and environmental variations making them of great significance in studying the origin and evolution of flora biodiversity. However, they are among the most threatened plant groups in the world. The principal aim of this review is to outline the distribution, diversity, and conservation status of Cycas in China and provide suggestions for conservation practices. In this review, we describe the taxonomy, distribution, and conservation status of Cycas in China. By comparing Chinese Cycas species with its relatives worldwide, we then discuss the current genetic diversity, genetic differentiation of Cycas, and try to disentangle the potential effects of Quaternary climate changes and topographical events on Cycas. We review conservation practices from both researchers and practitioners for these rare and endangered species. High genetic diversity at the species level and strong genetic differentiation within Cycas have been observed. Most Cycas species in southwest China have experienced population retreats in contrast to the coastal Cycas's expansion during the Quaternary glaciation. Additionally, human activities and habitat fragmentation have pushed these endangered taxa to the brink of extinction. Although numerous efforts have been made to mitigate threats to Cycas survival, implementation and compliance monitoring in protection zones are currently inadequate. We outline six proposals to strengthen conservation measures for Cycas in China and anticipate that these measures will provide guidelines for further research on population genetics as well as conservation biology of not only cycads but also other endangered species worldwide.     

Morphological characterization and phylogenetic relationships of Indochinese box turtles—The Cuora galbinifrons complex

The members of the Indochinese box turtle complex, namely Cuora galbinifrons, Cuora bourreti, and Cuora picturata, rank the most critically endangered turtle species on earth after more than three decades of over‐harvesting for food, traditional Chinese medicine, and pet markets. Despite advances in molecular biology, species boundaries and phylogenetic relationships, the status of the C. galbinifrons complex remains unresolved due to the small number of specimens observed and collected in the field. In this study, we present analyses of morphologic characters as well as mitochondrial and nuclear DNA data to reconstruct the species boundaries and systematic relationships within the C. galbinifrons complex. Based on principal component analysis (PCA) and statistical analysis, we found that phenotypic traits partially overlapped among galbinifrons, bourreti, and picturata, and that galbinifrons and bourreti might be only subspecifically distinct. Moreover, we used the mitochondrial genome, COI, and nuclear gene Rag1 under the maximum likelihood criteria and Bayesian inference criteria to elucidate whether C. galbinifrons could be divided into three separate species or subspecies. We found strong support for a sister relationship between picturata and the other two species, and consequently, we recommend maintaining picturata as a full species, and classifying bourreti and galbinifrons as subspecies of C. galbinifrons. These findings provide evidence for a better understanding of the evolutionary histories of these critically endangered turtles.     

High‐resolution melting of the cytochrome B gene in fecal DNA: A powerful approach for fox species identification of the Lycalopex genus in Chile

Easy, economic, precise species authentication is currently necessary in many areas of research and diagnosis in molecular biology applied to conservation studies of endangered species. Here, we present a new method for the identification of three fox species of the Lycalopex genus in Chile. We developed an assay based on high‐resolution melt analysis of the mitochondrial cytochrome B gene, allowing a simple, low cost, fast, and accurate species determination. To validate the assay applicability for noninvasive samples, we collected fecal samples in the Atacama Desert, finding unexpectedly one species outside of its known distribution range. We conclude that the assay has a potential to become a valuable tool for a standardized genetic monitoring of the Lycalopex species in Chile.     

De novo sequencing and chromosomal‐scale genome assembly of leopard coral grouper,Plectropomus leopardus

The leopard coral grouper, Plectropomus leopardus, belonging to the family Epinephelinae, is a carnivorous coral reef fish widely distributed in tropical and subtropical waters of the Indo‐Pacific. Due to its appealing body appearance and delicious taste, P. leopardus has become a popular commercial fish for aquaculture in many countries. However, the lack of genomic and molecular resources for P. leopardus has hindered study of its biology and genomic breeding programmes. Here we report the de novo sequencing and assembly of the P. leopardus genome using a combination of 10 × Genomics, high‐throughput chromosome conformation capture (Hi‐C) and PacBio long‐read sequencing technologies. The genome assembly has a total length of 881.55 Mb with a scaffold N50 of 34.15 Mb, consisting of 24 pseudochromosome scaffolds. busco analysis showed that 97.2% of the conserved single‐copy genes were retrieved, indicating the assembly was almost entire. We predicted 25,248 protein‐coding genes, among which 96.5% were functionally annotated. Comparative genomic analyses revealed that gene family expansions in P. leopardus were associated with immune‐related pathways. In addition, we identified 5,178,453 single nucleotide polymorphisms based on genome resequencing of 54 individuals. The P. leopardus genome and genomic variation data provide valuable genomic resources for studies of its genetics, evolution and biology. In particular, it is expected to benefit the development of genomic breeding programmes in the farming industry.     

A chromosome-scale assembly of the bilberry genome identifies a complex locus controlling berry anthocyanin composition

Bilberry (Vaccinium myrtillus L.) belongs to the Vaccinium genus, which includes blueberries (Vaccinium spp.) and cranberry (V. macrocarpon). Unlike its cultivated relatives, bilberry remains largely undomesticated, with berry harvesting almost entirely from the wild. As such, it represents an ideal target for genomic analysis, providing comparisons with the domesticated Vaccinium species. Bilberry is prized for its taste and health properties and has provided essential nutrition for Northern European indigenous populations. It contains high concentrations of phytonutrients, with perhaps the most important being the purple colored anthocyanins, found in both skin and flesh. Here, we present the first bilberry genome assembly, comprising 12 pseudochromosomes assembled using Oxford Nanopore (ONT) and Hi-C Technologies. The pseudochromosomes represent 96.6% complete BUSCO genes with an assessed LAI score of 16.3, showing a high conservation of synteny against the blueberry genome. Kmer analysis showed an unusual third peak, indicating the sequenced samples may have been from two individuals. The alternate alleles were purged so that the final assembly represents only one haplotype. A total of 36,404 genes were annotated after nearly 48% of the assembly was masked to remove repeats. To illustrate the genome quality, we describe the complex MYBA locus, and identify the key regulating MYB genes that determine anthocyanin production. The new bilberry genome builds on the genomic resources and knowledge of Vaccinium species, to help understand the genetics underpinning some of the quality attributes that breeding programs aspire to improve. The high conservation of synteny between bilberry and blueberry genomes means that comparative genome mapping can be applied to transfer knowledge about marker-trait association between these two species, as the loci involved in key characters are orthologous.     

Whole‐genome resequencing reveals the pleistocene temporal dynamics of Branchiostoma belcheri and Branchiostoma floridae populations

Global climatic fluctuations governed the ancestral demographic histories of species and contributed to place the current population status into a more extensive ecological and evolutionary context. Genetic variations will leave unambiguous signatures in the patterns of intraspecific genetic variation in extant species since the genome of each individual is an imperfect mosaic of the ancestral genomes. Here, we report the genome sequences of 20 Branchiostoma individuals by whole‐genome resequencing strategy. We detected over 140 million genomic variations for each Branchiostoma individual. In particular, we applied the pairwise sequentially Markovian coalescent (PSMC) method to estimate the trajectories of changes in the effective population size (N_e) of Branchiostoma population during the Pleistocene. We evaluated the threshold of sequencing depth for proper inference of demographic histories using PSMC was ≥25×. The PSMC results highlight the role of historical global climatic fluctuations in the long‐term population dynamics of Branchiostoma. The inferred ancestral N_e of the Branchiostoma belcheri populations from Zhanjiang and Xiamen (China) seawaters was different in amplitude before the first (mutation rate = 3 × 10^?9) or third glaciation (mutation rate = 9 × 10^?9) of the Pleistocene, indicating that the two populations most probably started to evolve in isolation in their respective seas after the first or third glaciation of the Pleistocene. A pronounced population bottleneck coinciding with the last glacial maximum was observed in all Branchiostoma individuals, followed by a population expansion occurred during the late Pleistocene. Species that have experienced long‐term declines may be especially vulnerable to recent anthropogenic activities. Recently, the industrial pollution and the exploitation of sea sand have destroyed the harmonious living environment of amphioxus species. In the future, we need to protect the habitat of Branchiostoma and make full use of these detected genetic variations to facilitate the functional study of Branchiostoma for adaptation to local environments.     

Identifying evolutionarily significant units for conservation of the endangered Malus sieversii using genome‐wide RADseq data

Malus sieversii, a wild progenitor of the domesticated apple, is an endangered species and is assigned second conservation priority by the China Plant Red Data Book. It is urgent to carry out in situ conservation of this species, but previous studies have not identified evolutionarily significant units (ESUs) for conservation management. In this study, we investigated the genetic diversity and relationships of six M. sieversii populations from China using integrated analysis of microsatellite (nSSR) data, genome‐wide SNPs and previous results in order to propose a reasonable conservation management. The results showed that levels of genetic diversity were inconsistently reflected by our nSSR and previous studies, suggesting that indices of genetic diversity are not effective to identify priority conservation areas for M. sieversii. Based on the selection criteria of ESUs for endangered species conservation, ESUs should reflect lineage divergence, geographical separation and different adaptive variation. Our phylogenetic tree based on genome‐wide SNPs yielded a clear relationship of divergent lineages among M. sieversii populations, leading to new different from those of previous studies. Three independent lineages, including the pairs of populations Huocheng‐Yining, Gongliu‐Xinyuan and Tuoli‐Emin, were identified. The geographic distances between populations among the different phylogenetic lineages were much greater than those within the same phylogenetic lineage. A cluster analysis on environmental variables showed that the three independent lineages inhabit different environmental conditions, suggesting that they may have adapted to different environments. Based on the results, we propose that three independent ESUs should be recognized as conservation units for M. sieversii in China.     

Chromosome‐level genome assembly of Triplophysa tibetana,a fish adapted to the harsh high‐altitude environment of the Tibetan Plateau

Triplophysa is an endemic fish genus of the Tibetan Plateau in China. Triplophysa tibetana, which lives at a recorded altitude of ~4,000 m and plays an important role in the highland aquatic ecosystem, serves as an excellent model for investigating high‐altitude environmental adaptation. However, evolutionary and conservation studies of T. tibetana have been limited by scarce genomic resources for the genus Triplophysa. In the present study, we applied PacBio sequencing and the Hi‐C technique to assemble the T. tibetana genome. A 652‐Mb genome with 1,325 contigs with an N50 length of 3.1 Mb was obtained. The 1,137 contigs were further assembled into 25 chromosomes, representing 98.7% and 80.47% of all contigs at the base and sequence number level, respectively. Approximately 260 Mb of sequence, accounting for ~39.8% of the genome, was identified as repetitive elements. DNA transposons (16.3%), long interspersed nuclear elements (12.4%) and long terminal repeats (11.0%) were the most repetitive types. In total, 24,372 protein‐coding genes were predicted in the genome, and ~95% of the genes were functionally annotated via a search in public databases. Using whole genome sequence information, we found that T. tibetana diverged from its common ancestor with Danio rerio ~121.4 million years ago. The high‐quality genome assembled in this work not only provides a valuable genomic resource for future population and conservation studies of T. tibetana, but it also lays a solid foundation for further investigation into the mechanisms of environmental adaptation of endemic fishes in the Tibetan Plateau.     

Genomic investigation of the Chinese alligator reveals wild-extinct genetic diversity and genomic consequences of their continuous decline

Critically endangered species are usually restricted to small and isolated populations. High inbreeding without gene flow among populations further aggravates their threatened condition and reduces the likelihood of their long-term survival. Chinese alligator (Alligator sinensis) is one of the most endangered crocodiles in the world and has experienced a continuous decline over the past c. 1 million years. In order to identify the genetic status of the remaining populations and aid conservation efforts, we assembled the first high-quality chromosome-level genome of Chinese alligator and explored the genomic characteristics of three extant breeding populations. Our analyses revealed the existence of at least three genetically distinct populations, comprising two breeding populations in China (Changxing and Xuancheng) and one breeding population in an American wildlife refuge. The American population does not belong to the last two populations of its native range (Xuancheng and Changxing), thus representing genetic diversity extinct in the wild and provides future opportunities for genetic rescue. Moreover, the effective population size of these three populations has been continuously declining over the past 20 ka. Consistent with this decline, the species shows extremely low genetic diversity, a large proportion of long runs of homozygous fragments, and mutational load across the genome. Finally, to provide genomic insights for future breeding management and conservation, we assessed the feasibility of mixing extant populations based on the likelihood of introducing new deleterious alleles and signatures of local adaptation. Overall, this study provides a valuable genomic resource and important genomic insights into the ecology, evolution, and conservation of critically endangered alligators.     

Chromosome‐level de novo genome assembly of Sarcophaga peregrina provides insights into the evolutionary adaptation of flesh flies

Sarcophaga peregrina is considered to be of great ecological, medical and forensic significance, and has unusual biological characteristics such as an ovoviviparous reproductive pattern and adaptation to feed on carrion. The availability of a high‐quality genome will help to further reveal the mechanisms underlying these charcateristics. Here we present a 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 million years ago. Comparative genomic analysis revealed expanded and positively selected genes related to biological features that aid in clarifying its ovoviviparous reproduction and carrion‐feeding adaptations, such as lipid metabolism, olfactory receptor activity, antioxidant enzymes, proteolysis and serine‐type endopeptidase activity. Protein‐coding genes associated with ovoviparity, such as yolk proteins, transferrin and acid sphingomyelinase, were identified. This study provides a valuable genomic resource for S. peregrina, and sheds insight into further revealing the underlying molecular mechanisms of adaptive evolution.     

A glimpse into the past and predictions for the future: the molecular evolution of the tuberculosis agent

Recent advances in genomics and molecular biology are providing an excellent opportunity to get a glimpse into the past, to examine the present, and to predict the future evolution of pathogenic mycobacteria, and in particular that of Mycobacterium tuberculosis, the agent of human tuberculosis. The recent availability of genome sequences of several Mycobacterium canettii strains, representing evolutionary early‐branching tubercle bacilli, has allowed the genomic and molecular features of the putative ancestor of the M. tuberculosis complex (MTBC) to be reconstituted. Analyses have identified extensive lateral gene transfer and recombination events in M. canettii and/or the MTBC, leading to suggestions of a past environmental reservoir where the ancestor(s) of the tubercle bacilli might have adapted to an intracellular lifestyle. The daily increases in M. tuberculosis genome data and the remaining urgent Public Health problem of tuberculosis make it more important than ever to try and understand the origins and the future evolution of the MTBC. Here we critically discuss a series of questions on gene‐loss, acquisition, recombination, mutation and conservation that have recently arisen and which are key to better understand the outstanding evolutionary success of one of the most widespread and most deadly bacterial pathogens in the history of humankind.     

Using genomic information for management planning of an endangered perennial,Viola uliginosa

Species occupying habitats subjected to frequent natural and/or anthropogenic changes are a challenge for conservation management. We studied one such species, Viola uliginosa, an endangered perennial wetland species typically inhabiting sporadically flooded meadows alongside rivers/lakes. In order to estimate genomic diversity, population structure, and history, we sampled five sites in Finland, three in Estonia, and one each in Slovenia, Belarus, and Poland using genomic SNP data with double‐digest restriction site‐associated DNA sequencing (ddRAD‐seq). We found monophyletic populations, high levels of inbreeding (mean population F_SNP = 0.407–0.945), low effective population sizes (N_e = 0.8–50.9), indications of past demographic expansion, and rare long‐distance dispersal. Our results are important in implementing conservation strategies for V. uliginosa, which should include founding of seed banks, ex situ cultivations, and reintroductions with individuals of proper origin, combined with continuous population monitoring and habitat management.     

Considerations for monitoring population trends of colonial waterbirds using the effective number of breeders and census estimates

Detecting trends in population size fluctuations is a major focus in ecology, evolution, and conservation biology. Populations of colonial waterbirds have been monitored using demographic approaches to determine annual census size (N_a). We propose the addition of genetic estimates of the effective number of breeders (N_b) as indirect measures of the risk of loss of genetic diversity to improve the evaluation of demographics and increase the accuracy of trend estimates in breeding colonies. Here, we investigated which methods of the estimation of N_b are more precise under conditions of moderate genetic diversity, limited sample sizes and few microsatellite loci, as often occurs with natural populations. We used the wood stork as a model species and we offered a workflow that researchers can follow for monitoring bird breeding colonies. Our approach started with simulations using five estimators of N_b and the theoretical results were validated with empirical data collected from breeding colonies settled in the Brazilian Pantanal wetland. In parallel, we estimated census size using a corrected method based on counting active nests. Both in simulations and in natural populations, the approximate Bayesian computation (ABC) and sibship assignment (SA) methods yielded more precise estimates than the linkage disequilibrium, heterozygosity excess, and molecular coancestry methods. In particular, the ABC method performed best with few loci and small sample sizes, while the other estimators required larger sample sizes and at least 13 loci to not underestimate N_b. Moreover, according to our N_b/N_a estimates (values were often ≤0.1), the wood stork colonies evaluated could be facing the loss of genetic diversity. We demonstrate that the combination of genetic and census estimates is a useful approach for monitoring natural breeding bird populations. This methodology has been recommended for populations of rare species or with a known history of population decline to support conservation efforts.