Genome Skimming Research Articles

Genetic characterization of a group of commercial African timber species: From genomics to barcoding.

In the last decades, illegal logging has posed a serious threat for the integrity of forest ecosystems and for biodiversity conservation in tropical Africa. Although international treaties and regulatory plans have been implemented to reduce illegal logging, much of the total timber volume is harvested and traded illegally from tropical African forest regions. As a result, the development and the application of analytical tools to enhance the traceability and the identification of wood and related products is critical to enforce international regulations. Among available techniques, DNA barcoding is a promising approach for the molecular identification of plant species. However, although it has been used successfully for the discrimination of animal species, no set of genetic markers is available for the universal identification of plant species. In this work, we firstly characterized the genetic diversity of 17 highly-valuable African timber species from five genera (Afzelia, Guibourtia, Leplea, Milicia, Tieghemella) across their distribution ranges in West and Central Africa using the genome skimming approach in order to reconstruct their chloroplast genomes and nuclear ribosomal DNA. Next, we identified single-nucleotide polymorphisms (SNPs) for the discrimination of closely-related species. In this way, we successfully developed and tested novel species-specific genetic barcodes for species identification.

Phylogenetic relationship and comparative analysis of the main Bupleuri Radix species in China.

Bupleuri Radix (Chaihu) is a famous traditional Chinese medicine derived from Bupleurum, Apiaceae. The origin of cultivated Chaihu germplasm in China is unclear, which has led to unstable Chaihu quality. In this study, we reconstructed the phylogeny of the main Chaihu germplasm species in China and identified potential molecular markers to authenticate its origin. Three Bupleurum species (eight individuals), B. bicaule, B. chinense, and B. scorzonerifolium, were selected for genome skimming. Published genomes from B. falcatum and B. marginatum var. stenophyllum were used for comparative analysis. Sequences of the complete plastid genomes were conserved with 113 identical genes ranging from 155,540 to 155,866 bp in length. Phylogenetic reconstruction based on complete plastid genomes resolved intrageneric relationships of the five Bupleurum species with high support. Conflicts between the plastid and nuclear phylogenies were observed, which were mainly ascribed to introgressive hybridization. Comparative analysis showed that noncoding regions of the plastomes had most of the variable sequences. Eight regions (atpF-atpH, petN-psbM, rps16-psbK, petA-psbJ, ndhC-trnV/UAC and ycf1) had high divergence values in Bupleurum species and could be promising DNA barcodes for Chaihu authentication. A total of seven polymorphic cpSSRs and 438 polymorphic nSSRs were detected across the five Chaihu germplasms. Three photosynthesis-related genes were under positive selection, of which accD reflected the adaptation fingerprint of B. chinense to different ecological habitats. Our study provides valuable genetic information for phylogenetic investigation, germplasm authentication, and molecular breeding of Chaihu species.

Testing species relationships and delimitation in the Amazonian hyperdominantAstrocaryumsectionHuicungo(Arecaceae) using chloroplast data from genome skimming

AbstractHyperdominant trees in Amazonia account for half of the individual trees (>10 cm dbh) in the forest, and thus play a crucial role in ecosystem dynamics. However, several of these widespread hyperdominant species may be complexes hiding cryptic diversity that can affect species richness estimates and conservation priorities. Here, we study the intraspecific variation ofAstrocaryum murumuru(Arecaceae), a keystone and hyperdominant species in Amazonia, also known asAstrocaryumsect.Huicungo, a complex of 15 understory to subcanopy palm species. Using chloroplast DNA from genome skimming (>66 kbp alignment) in a Bayesian framework, we present evidence thatA. sect.Huicungorepresents three separately evolving lineages, suggesting that the section is not a single hyperdominant species, and that the 15 morphology‐based species may be an over‐representation. Genome skimming chloroplast data did not fully resolve the species‐level phylogenetic relationships inA. sect.Huicungomostly because of gene discordance and the paraphyly of most species. Contrary to a previous nuclear‐based phylogenetic analysis, the chloroplast genomic data did not recoverA. sect.Huicungomonophyletic, but yielded monophyly in an increased number of species (six) in the complex. Interspecific phylogenetic relationships showed a geographic pattern, and the traditional morphology‐based classification was not supported. Our phylogenomic results are discussed in light of earlier phylogeographical studies using Sanger sequencing. Our findings show the utility of genome skimming data in species delimitation analyses to uncover intraspecific variation of hyperdominant species in Amazonia, the largest evergreen tropical forest.

Plastid phylogenomics and molecular evolution of Thismiaceae (Dioscoreales).

Species in Thismiaceae can no longer photosynthesize, and instead obtain carbon from soil fungi. Here we infer Thismiaceae phylogeny using plastid genome data, and also characterize the molecular evolution of this genome. We assembled five Thismiaceae plastid genomes from genome skimming data, adding to previously published data for phylogenomic inference. We investigated plastid-genome structural changes, considering locally colinear blocks (LCBs). We also characterized possible shifts in selection pressure in retained genes by considering changes in ω, the ratio of non-synonymous to synonymous changes. Thismiaceae experienced two major pulses of gene loss around the early diversification of the family, with subsequent scattered gene losses across descendent lineages. In addition to massive size reduction, Thismiaceae plastid genomes experienced occasional inversions, and there likely were two independent losses of the plastid inverted repeat (IR) region. Retained plastid genes remain under generally strong purifying selection (ω << 1), with significant and sporadic weakening or strengthening in several instances. The bifunctional trnE-UUC gene of Thismia huangii likely retains a secondary role in heme biosynthesis, despite a probable loss of functionality in protein translation. Several cis-spliced group IIA introns are retained, despite the loss of the plastid intron maturase, matK. We infer that most gene losses in Thismiaceae occurred early and rapidly, following the initial loss of photosynthesis in its stem lineage. As a species-rich, fully mycoheterotrophic lineage, Thismiaceae provides a model system for uncovering the unique and divergent ways in which plastid genomes evolve in heterotrophic plants. This article is protected by copyright. All rights reserved.

Phylogenomics and morphological evolution of the mega-diverse genus Artemisia (Asteraceae: Anthemideae): implications for its circumscription and infrageneric taxonomy.

Artemisia is a mega-diverse genus consisting of ~400 species. Despite its medicinal importance and ecological significance, a well-resolved phylogeny for global Artemisia, a natural generic delimitation and infrageneric taxonomy remain missing, owing to the obstructions from limited taxon sampling and insufficient information on DNA markers. Its morphological characters, such as capitulum, life form and leaf, show marked variations and are widely used in its infrageneric taxonomy. However, their evolution within Artemisia is poorly understood. Here, we aimed to reconstruct a well-resolved phylogeny for global Artemisia via a phylogenomic approach, to infer the evolutionary patterns of its key morphological characters and to update its circumscription and infrageneric taxonomy. We sampled 228 species (258 samples) of Artemisia and its allies from both fresh and herbarium collections, covering all the subgenera and its main geographical areas, and conducted a phylogenomic analysis based on nuclear single nucleotide polymorphisms (SNPs) obtained from genome skimming data. Based on the phylogenetic framework, we inferred the possible evolutionary patterns of six key morphological characters widely used in its previous taxonomy. The genus Kaschgaria was revealed to be nested in Artemisia with strong support. A well-resolved phylogeny of Artemisia consisting of eight highly supported clades was recovered, two of which were identified for the first time. Most of the previously recognized subgenera were not supported as monophyletic. Evolutionary inferences based on the six morphological characters showed that different states of these characters originated independently more than once. The circumscription of Artemisia is enlarged to include the genus Kaschgaria. The morphological characters traditionally used for the infrageneric taxonomy of Artemisia do not match the new phylogenetic tree. They experienced a more complex evolutionary history than previously thought. We propose a revised infrageneric taxonomy of the newly circumscribed Artemisia, with eight recognized subgenera to accommodate the new results.

Discovering the Repeatome of Five Species Belonging to the Asteraceae Family: A Computational Study.

Genome divergence by repeat proliferation and/or loss is a process that plays a crucial role in species evolution. Nevertheless, knowledge of the variability related to repeat proliferation among species of the same family is still limited. Considering the importance of the Asteraceae family, here we present a first contribution towards the metarepeatome of five Asteraceae species. A comprehensive picture of the repetitive components of all genomes was obtained by genome skimming with Illumina sequence reads and by analyzing a pool of full-length long terminal repeat retrotransposons (LTR-REs). Genome skimming allowed us to estimate the abundance and variability of repetitive components. The structure of the metagenome of the selected species was composed of 67% repetitive sequences, of which LTR-REs represented the bulk of annotated clusters. The species essentially shared ribosomal DNA sequences, whereas the other classes of repetitive DNA were highly variable among species. The pool of full-length LTR-REs was retrieved from all the species and their age of insertion was established, showing several lineage-specific proliferation peaks over the last 15-million years. Overall, a large variability of repeat abundance at superfamily, lineage, and sublineage levels was observed, indicating that repeats within individual genomes followed different evolutionary and temporal dynamics, and that different events of amplification or loss of these sequences may have occurred after species differentiation.

The phylogeny and evolutionary ecology of hoverflies (Diptera: Syrphidae) inferred from mitochondrial genomes

Hoverflies (Diptera: Syrphidae) are a diverse group of pollinators and a major research focus in ecology, but their phylogenetic relationships remain incompletely known. Using a genome skimming approach we generated mitochondrial genomes for 91 species, capturing a wide taxonomic diversity of the family. To reduce the required amount of input DNA and overall cost of the library construction, sequencing and assembly was conducted on mixtures of specimens, which raises the problem of chimera formation of mitogenomes. We present a novel chimera detection test based on gene tree incongruence, but identified only a single mitogenome of chimeric origin. Together with existing data for a final set of 127 taxa, phylogenetic analysis on nucleotide and amino acid sequences using Maximum Likelihood and Bayesian Inference revealed a basal split of Microdontinae from all other syrphids. The remainder consists of several deep clades assigned to the subfamily Eristalinae in the current classification, including a clade comprising the subfamily Syrphinae (plus Pipizinae). These findings call for a re-definition of subfamilies, but basal nodes had insufficient support to fully justify such action. Molecular-clock dating placed the origin of the Syrphidae crown group in the mid-Cretaceous while the Eristalinae-Syrphinae clade likely originated near the K/Pg boundary. Transformation of larval life history characters on the tree suggests that Syrphidae initially had sap feeding larvae, which diversified greatly in diet and habitat association during the Eocene and Oligocene, coinciding with the diversification of angiosperms and the evolution of various insect groups used as larval host, prey, or mimicry models. Mitogenomes proved to be a powerful phylogenetic marker for studies of Syrphidae at subfamily and tribe levels, allowing dense taxon sampling that provided insight into the great ecological diversity and rapid evolution of larval life history traits of the hoverflies.

Testing and using complete plastomes for authentication of medicinal Polygonatum species (Asparagaceae)

Polygonati Rhizoma, a traditional Chinese medicine important to the pharmaceutical and food industries, is frequently adulterated with morphologically similar congeneric species, posing quality and effectiveness issues for the products. Accordingly, the accurate and sensitive authentication of medicinal Polygonatum species is vital to the standardization of Polygonati Rhizoma products. In view of the difficulties in morphological analysis and the use of standard DNA barcodes for authentication of medicinal Polygonatum species, an alternative authentication method is needed. Based on inter- and intraspecies sampling, phylogenetic and genetic distance analyses were performed to evaluate the discriminative power of complete plastomes for the identification of Polygonatum species. The results indicate that the genetic criteria of complete plastomes are in good correspondence with species boundaries and therefore can be used for unambiguous identification of medicinal Polygonatum species and morphologically similar congeners, which verifies the theoretical prediction that the application of genome skimming to generate complete plastomes as molecular tags can serve as an effective and accurate method to detect whether Polygonati Rhizoma is adulterated with congeneric species. Additionally, in this study, a taxonomically well-curated and high-resolution reference plastome superbarcode library for Polygonatum was developed for the first time, providing a valuable genetic resource for the application of genome skimming to monitor and control the production and trade of Polygonati Rhizoma to guarantee the effectiveness of downstream products. The findings also provide new insights to clarify the taxonomic ambiguities of the generic circumscription of Polygonatum and Heteropolygonatum, as well as the species delimitation of Polygonatum cyrtonema, Polygonatum nodosum, and Polygonatum punctatum, which will contribute to proper exploitation and conservation of Polygonatum species.

The complete mitochondrial genome and phylogenetic characterization of two putative subspecies of golden jackal (Canis aureus cruesemanni and Canis aureus moreotica)

The golden jackal (Canis aureus) is a canid species found across southern Eurasia. Several subspecies of this animal have been genetically studied in regions such as Europe, the Middle East, and India. However, one subspecies that lacks current research is the Indochinese jackal (Canis aureus cruesemanni), which is primarily found in Southeast Asia. Using a genome skimming approach, we assembled the first complete mitochondrial genome for an Indochinese jackal from Thailand. To expand the number of available Canis aureus mitogenomes, we also assembled and sequenced the first complete mitochondrial genome of a golden jackal from Turkey, representing the C. a. moreotica subspecies. The mitogenomes contained 37 annotated genes and are 16,729 bps (C. a. cruesemanni) and 16,669 bps (C. a. moreotica) in length. Phylogenetic analysis with 26 additional canid mitogenomes and analyses of a cytochrome b gene-only data set together support the Indochinese jackal as a distinct and early-branching lineage among golden jackals, thereby supporting its recognition as a possible subspecies. These analyses also demonstrate that the golden jackal from Turkey is likely not a distinct lineage due to close genetic relationships with golden jackals from India and Israel.

Genome skimming elucidates the evolutionary history of Octopoda

Phylogenies for Octopoda have, until now, been based on morphological characters or a few genes. Here we provide the complete mitogenomes and the nuclear 18S and 28S ribosomal genes of twenty Octopoda specimens, comprising 18 species of Cirrata and Incirrata, representing 13 genera and all five putative families of Cirrata (Cirroctopodidae, Cirroteuthidae, Grimpoteuthidae, Opisthoteuthidae and Stauroteuthidae) and six families of Incirrata (Amphitretidae, Argonautidae, Bathypolypodidae, Eledonidae, Enteroctopodidae, and Megaleledonidae) which were assembled using genome skimming. Phylogenetic trees were built using Maximum Likelihood and Bayesian Inference with several alignment matrices. All mitochondrial genomes had the ‘typical’ genome composition and gene order previously reported for octopodiforms, except Bathypolypus ergasticus, which appears to lack ND5, two tRNA genes that flank ND5 and two other tRNA genes. Argonautoidea was revealed as sister to Octopodidae by the mitochondrial protein-coding gene dataset, however, it was recovered as sister to all other incirrate octopods with strong support in an analysis using nuclear rRNA genes. Within Cirrata, our study supports two existing classifications suggesting neither is likely in conflict with the true evolutionary history of the suborder. Genome skimming is useful in the analysis of phylogenetic relationships within Octopoda; inclusion of both mitochondrial and nuclear data may be key.

Phylogenomics of Aralia sect. Aralia (Araliaceae): Signals of hybridization and insights into its species delimitations and intercontinental biogeography

Genome-scale data have significantly increased the number of informative characters for phylogenetic analyses and recent studies have also revealed widespread phylogenomic discordance in many plant lineages. Aralia sect. Aralia is a small plant lineage (14 spp.) of the ginseng family Araliaceae with a disjunct distribution between eastern Asia (11 spp.) and North America (3 spp.). We herein employ sequences of hundreds of nuclear loci and the complete plastomes using targeted sequence capture and genome skimming to reconstruct the phylogenetic and biogeographic history of this section. We detected substantial conflicts among nuclear genes, yet different analytical strategies generated largely congruent topologies from the nuclear data. Significant cytonuclear discordance was detected, especially concerning the positions of the three North American species. The phylogenomic results support two intercontinental disjunctions: (1) Aralia californica of western North America is sister to the eastern Asian clade consisting of A. cordata and A. continentalis in the nuclear tree, and (2) the eastern North American A. racemosa forms a clade with A. bicrenata from southwestern North America, and the North American A. racemosa - A. bicrenata clade is then sister to the eastern Asian clade consisting of A. glabra (Japan), A. fargesii (C China), and A. apioides and A. atropurpurea (the Hengduan Mountains). Aralia cordata is supported to be disjunctly distributed in Japan, Taiwan, the Ulleung island of Korea, and in Central, Southwest and South China, and Aralia continentalis is redefined with a narrower distribution in Northeast China, eastern Russia and peninsular Korea.

Phylogeny and Historical Biogeography of the East Asian Clematis Group, Sect. Tubulosae, Inferred from Phylogenomic Data.

The evolutionary history of Clematis section Tubulosae, an East Asian endemic lineage, has not been comprehensively studied. In this study, we reconstruct the phylogeny of this section with a complete sampling using a phylogenomic approach. The genome skimming method was applied to obtain the complete plastome sequence, the nuclear ribosomal DNA (nrDNA), and the nuclear SNPs data for phylogenetic reconstruction. Using a Bayesian molecular clock approach and ancestral range reconstruction, we reconstruct biogeographical history and discuss the biotic and abiotic factors that may have shaped the distribution patterns of the section. Both nuclear datasets better resolved the phylogeny of the sect. Tubulosae than the plastome sequence. Sect. Tubulosae was resolved as a monophyletic group sister to a clade mainly containing species from the sect. Clematis and sect. Aspidanthera. Within sect. Tubulosae, two major clades were resolved by both nuclear datasets. Two continental taxa, C. heracleifolia and C. tubulosa var. ichangensis, formed one clade. One continental taxon, C. tubulosa, and all the other species from Taiwan island, the Korean peninsula, and the Japanese archipelago formed the other clade. Molecular dating results showed that sect. Tubulosae diverged from its sister clade in the Pliocene, and all the current species diversified during the Pleistocene. Our biogeographical reconstruction suggested that sect. Tubulosae evolved and began species diversification, most likely in mainland China, then dispersed to the Korean peninsula, and then expanded its range through the Japanese archipelago to Taiwan island. Island species diversity may arise through allopatric speciation by vicariance events following the range fragmentation triggered by the climatic oscillation and sea level change during the Pleistocene epoch. Our results highlight the importance of climatic oscillation during the Pleistocene to the spatial-temporal diversification patterns of the sect. Tubulosae.

Deciphering complex reticulate evolution of Asian Buddleja (Scrophulariaceae): insights into the taxonomy and speciation of polyploid taxa in the Sino-Himalayan region.

Species of the genus Buddleja in Asia are mainly distributed in the Sino-Himalayan region and form a challenging taxonomic group, with extensive hybridization and polyploidization. A phylogenetic approach to unravelling the history of reticulation in this lineage will deepen our understanding of the speciation in biodiversity hotspots. For this study, we obtained 80 accessions representing all the species in the Asian Buddleja clade, and the ploidy level of each taxon was determined by flow cytometry analyses. Whole plastid genomes, nuclear ribosomal DNA, single nucleotide polymorphisms and a large number of low-copy nuclear genes assembled from genome skimming data were used to investigate the reticulate evolutionary history of Asian Buddleja. Complex cytonuclear conflicts were detected through a comparison of plastid and species trees. Gene tree incongruence was also analysed to detect any reticulate events in the history of this lineage. Six hybridization events were detected, which are able to explain the cytonuclear conflict in Asian Buddleja. Furthermore, PhyloNet analysis combining species ploidy data indicated several allopolyploid speciation events. A strongly supported species tree inferred from a large number of low-copy nuclear genes not only corrected some earlier misinterpretations, but also indicated that there are many Asian Buddleja species that have been lumped mistakenly. Divergent time estimation shows two periods of rapid diversification (8-10 and 0-3 Mya) in the Asian Buddleja clade, which might coincide with the final uplift of the Hengduan Mountains and Quaternary climate fluctuations, respectively. This study presents a well-supported phylogenetic backbone for the Asian Buddleja species, elucidates their complex and reticulate evolutionary history and suggests that tectonic activity, climate fluctuations, polyploidization and hybridization together promoted the diversification of this lineage.

Giant Fern Genomes Show Complex Evolution Patterns: A Comparative Analysis in Two Species of Tmesipteris (Psilotaceae).

Giant genomes are rare across the plant kingdom and their study has focused almost exclusively on angiosperms and gymnosperms. The scarce genetic data that are available for ferns, however, indicate differences in their genome organization and a lower dynamism compared to other plant groups. Tmesipteris is a small genus of mainly epiphytic ferns that occur in Oceania and several Pacific Islands. So far, only two species with giant genomes have been reported in the genus, T. tannensis (1C = 73.19 Gbp) and T. obliqua (1C = 147.29 Gbp). Low-coverage genome skimming sequence data were generated in these two species and analyzed using the RepeatExplorer2 pipeline to identify and quantify the repetitive DNA fraction of these genomes. We found that both species share a similar genomic composition, with high repeat diversity compared to taxa with small (1C < 10 Gbp) genomes. We also found that, in general, characterized repetitive elements have relatively high heterogeneity scores, indicating ancient diverging evolutionary trajectories. Our results suggest that a whole genome multiplication event, accumulation of repetitive elements, and recent activation of those repeats have all played a role in shaping these genomes. It will be informative to compare these data in the future with data from the giant genome of the angiosperm Paris japonica, to determine if the structures observed here are an emergent property of massive genomic inflation or derived from lineage specific processes.

Using low-coverage whole genome sequencing (genome skimming) to delineate three introgressed species of buffalofish (Ictiobus)

Consumption of buffalofish has been sporadically associated with Haff disease-like illnesses involving sudden onset muscle pain and weakness due to skeletal muscle rhabdomyolysis, but determination of precisely which species are associated with these illnesses has been impeded by a lack of species-specific DNA-based markers. Here, three closely related species of buffalofish native to the Mississippi River Basin (Ictiobus bubalus, Ictiobus cyprinellus and Ictiobus niger) that have previously proven genetically indistinguishable using both mitochondrial and nuclear single-locus sequencing were reliably discriminated using low-coverage whole genome sequencing (‘genome skimming’). Using 44 specimens representing the three species collected from the mid/upper (Missouri) and lower (Louisiana) regions of the species’ native ranges, the SISRS (Site Identification from Short Read Sequences) bioinformatics pipeline was adapted to (1) identify over 620Mbp of putatively homologous nuclear sequence data and (2) isolate over 140,000 single-nucleotide polymorphisms (SNPs) that supported accurate species delimitation, all without the use of a reference genome or annotation data. These sites were used to classify Ictiobus spp. samples with genome-skim data, along with a larger set (n = 67) where ultraconserved elements (UCEs) were sequenced. Analyses of whole mitochondrial data revealed more limited signal. Nearly all samples matched their purported species based on morphologic identification, but two Missouri samples morphologically identified as I. niger grouped with samples of I. bubalus, albeit with significant enrichment of I. niger SNPs. To our knowledge this is the first report of a DNA-based tool to reliably discriminate these three morphologically distinct species.

Phylogenetic relationships of the Amblyomma cajennense complex (Acari: Ixodidae) at mitogenomic resolution

The genus Amblyomma is the third most diverse in the number of species within the Ixodidae, with practically half of its species distributed in the Americas, though there are also species occurring in Africa, Asia, and Australia. Within the genus, there are several species complexes with veterinary and public health importance. The Amblyomma cajennense complex, in the Americas, is represented by six species with a wide distribution, from Texas to northern Argentina. We combined two sequencing techniques to generate complete mitogenomes of species belonging to the Amblyomma cajennense complex: genome skimming and long-range PCRs sequencing methods. Thus, we generated seven new mitochondrial genomes for all species of the Amblyomma cajennense complex, except for Amblyomma interandinum. Genetic distances between the mitogenomes corroborate the clear differentiation between the five species of the Amblyomma cajennense complex. The phylogenetic relationships of these species had previously been evaluated by combining partial nuclear and mitochondrial genes and here these relationships are corroborated with a more robust framework of data, which demonstrates that the conjunction of mitochondrial and nuclear partial genes can resolve close relationships when entire genes or genomes are unavailable. The gene order, structure, composition, and length are stable across these mitogenomes, and they share the general characteristics of Metastriata. Future studies should increase the number of available mitogenomes for this genus, especially for those species from the Indo-Pacific region and Africa, by means of a better understanding of their relationships and evolutionary process.

Mining various genomic resources to resolve old alpha-taxonomy questions: A test of the species hypothesis of the Proteocephalus longicollis species complex (Cestoda: Platyhelminthes) from salmonid fishes

High-throughput sequencing strategies became commonly employed to study non-model parasites, but the corresponding genomes and transcriptomes were seldom mined following the original publication. Similar to the data generated with genome skimming techniques based on shallow-depth shotgun genomes, various genomic and transcriptomic resources can be screened for useful molecular phylogenetic markers traditionally characterised with Sanger sequencing. Here, we provide an example of a strategy using reduced-representation genomic as well as transcriptomic data to obtain broad insights into the molecular diversity of the cestode Proteocephalus longicollis, a common parasite of salmonids distributed throughout the Holarctic region. We extract popular mitochondrial and nuclear ribosomal markers from various genomic resources for hundreds of parasite specimens from multiple European whitefish populations and compare those with Proteocephalus representatives from other species of salmonids and various geographical regions. In contrast with the previous morphology-based assessments, molecular phylogeny reveals a high degree of genetic divergence between Proteocephalus isolates from different salmonids, contrastingly low genetic differentiation within the parasite’s populations hosted by the European whitefish (Coregonus lavaretus species complex), and a sister species relationship of Proteocephalus from European whitefish and Proteocephalus percae, a parasite of European perch (Perca fluviatilis). Proteocephalus spp. from North American lake whitefish, brown trout and Arctic charr each formed clearly distinct lineages. These results advance our understanding of the interrelationships of the Proteocephalus-aggregate, a well-recognized clade of Holarctic freshwater fish proteocephalids, and support resurrection of some of the nominal species of Proteocephalus, including Proteocephalus exiguus La Rue, 1911 from North American coregonids and Proteocephalus fallax La Rue, 1911 from European C. lavaretus, reserving Proteocephalus longicollis (Zeder, 1800) exclusively for parasites of Salmo trutta.

Evaluation of genome skimming to detect and characterise human and livestock helminths

The identification of gastrointestinal helminth infections of humans and livestock almost exclusively relies on the detection of eggs or larvae in faeces, followed by manual counting and morphological characterisation to differentiate species using microscopy-based techniques. However, molecular approaches based on the detection and quantification of parasite DNA are becoming more prevalent, increasing the sensitivity, specificity and throughput of diagnostic assays. High-throughput sequencing, from single PCR targets through to the analysis of whole genomes, offers significant promise towards providing information-rich data that may add value beyond traditional and conventional molecular approaches; however, thus far, its utility has not been fully explored to detect helminths in faecal samples. In this study, low-depth whole genome sequencing, i.e. genome skimming, has been applied to detect and characterise helminth diversity in a set of helminth-infected human and livestock faecal material. The strengths and limitations of this approach are evaluated using three methods to characterise and differentiate metagenomic sequencing data based on (i) mapping to whole mitochondrial genomes, (ii) whole genome assemblies, and (iii) a comprehensive internal transcribed spacer 2 (ITS2) database, together with validation using quantitative PCR (qPCR). Our analyses suggest that genome skimming can successfully identify most single and multi-species infections reported by qPCR and can provide sufficient coverage within some samples to resolve consensus mitochondrial genomes, thus facilitating phylogenetic analyses of selected genera, e.g. Ascaris spp. Key to this approach is both the availability and integrity of helminth reference genomes, some of which are currently contaminated with bacterial and host sequences. The success of genome skimming of faecal DNA is dependent on the availability of vouchered sequences of helminths spanning both taxonomic and geographic diversity, together with methods to detect or amplify minute quantities of parasite nucleic acids in mixed samples.

Chloroplast genome skimming of a potential agroforestry species Melia dubia. Cav and its comparative phylogenetic analysis with major Meliaceae members.

The online version contains supplementary material available at 10.1007/s13205-022-03447-1.

Meliaceae genomes provide insights into wood development and limonoids biosynthesis.

Meliaceae is a useful plant family owing to its high-quality timber and its many limonoids that have pharmacological and biological activities. Although some genomes of Meliaceae species have been reported, many questions regarding their unique family features, namely wood quality and natural products, have not been answered. In this study, we provide the whole-genome sequence of Melia azedarach comprising 237.16 Mb with a contig N50 of 8.07 Mb, and an improved genome sequence of Azadirachta indica comprising 223.66 Mb with a contig N50 of 8.91 Mb. Moreover, genome skimming data, transcriptomes and other published genomes were comprehensively analysed to determine the genes and proteins that produce superior wood and valuable limonoids. Phylogenetic analysis of chloroplast genomes, single-copy gene families and single-nucleotide polymorphisms revealed that Meliaceae should be classified into two subfamilies: Cedreloideae and Melioideae. Although the Meliaceae species did not undergo additional whole-genome duplication events, the secondary wall biosynthetic genes of the woody Cedreloideae species, Toona sinensis, expanded significantly compared to those of A. indica and M. azedarach, especially in downstream transcription factors and cellulose/hemicellulose biosynthesis-related genes. Moreover, expanded special oxidosqualene cyclase catalogues can help diversify Sapindales skeletons, and the clustered genes that regulate terpene chain elongation, cyclization and modification would support their roles in limonoid biosynthesis. The expanded clans of terpene synthase, O-methyltransferase and cytochrome P450, which are mainly derived from tandem duplication, are responsible for the different limonoid classes among the species. These results are beneficial for further investigations of wood development and limonoid biosynthesis.