Genome Skimming: A Rapid Approach to Gaining Diverse Biological Insights into Multicellular Pathogens.
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ABSTRACT:
SUBMITTER: Denver DR
PROVIDER: S-EPMC4973915 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Developing genomic insights is challenging in nonmodel species for which resources are often scarce and prohibitively costly. Here, we explore the potential of a recently established approach using Pool-seq data to generate a de novo genome assembly for mining exons, upon which Pool-seq data are used to estimate population divergence and diversity. We do this for two pairs of sympatric populations of brown trout (Salmo trutta): one naturally sympatric set of populations and another pair of populations introduced to a common environment. We validate our approach by comparing the results to those from markers previously used to describe the populations (allozymes and individual-based single nucleotide polymorphisms [SNPs]) and from mapping the Pool-seq data to a reference genome of the closely related Atlantic salmon (Salmo salar). We find that genomic differentiation (F ST) between the two introduced populations exceeds that of the naturally sympatric populations (F ST = 0.13 and 0.03 between the introduced and the naturally sympatric populations, respectively), in concordance with estimates from the previously used SNPs. The same level of population divergence is found for the two genome assemblies, but estimates of average nucleotide diversity differ ( π ¯ ≈ 0.002 and π ¯ ≈ 0.001 when mapping to S. trutta and S. salar, respectively), although the relationships between population values are largely consistent. This discrepancy might be attributed to biases when mapping to a haploid condensed assembly made of highly fragmented read data compared to using a high-quality reference assembly from a divergent species. We conclude that the Pool-seq-only approach can be suitable for detecting and quantifying genome-wide population differentiation, and for comparing genomic diversity in populations of nonmodel species where reference genomes are lacking.
| S-EPMC6802065 | biostudies-literature
Project description:Solitons have been observed in various physical phenomena. Here, we show that the distinct characteristics of solitons are present in the mass cell movement of non-chemotactic mutants of the cellular slime mould Dictyostelium discoideum. During starvation, D. discoideum forms multicellular structures that differentiate into spore or stalk cells and, eventually, a fruiting body. Non-chemotactic mutant cells do not form multicellular structures; however, they do undergo mass cell movement in the form of a pulsatile soliton-like structure (SLS). We also found that SLS induction is mediated by adhesive cell-cell interactions. These observations provide novel insights into the mechanisms of biological solitons in multicellular movement.
| S-EPMC3725511 | biostudies-other
Project description:Silks are the long threads at the tips of maize ears onto which pollen land and sperm nuclei travel long distances to fertilize egg cells, giving rise to embryos and seeds; however fungal pathogens also use this route to invade developing grain, causing damaging ear rots with dangerous mycotoxins. This review highlights the importance of silks as the direct highways by which globally important fungal pathogens enter maize kernels. First, the most important silk-entering fungal pathogens in maize are reviewed, including Fusarium graminearum, Fusarium verticillioides, and Aspergillus flavus, and their mycotoxins. Next, we compare the different modes used by each fungal pathogen to invade the silks, including susceptible time intervals and the effects of pollination. Innate silk defences and current strategies to protect silks from ear rot pathogens are reviewed, and future protective strategies and silk-based research are proposed. There is a particular gap in knowledge of how to improve silk health and defences around the time of pollination, and a need for protective silk sprays or other technologies. It is hoped that this review will stimulate innovations in breeding, inputs, and techniques to help growers protect silks, which are expected to become more vulnerable to pathogens due to climate change.
| S-EPMC6313692 | biostudies-literature
Project description:Shallow whole genome sequencing data (genome skimming data) from 12 genera of the tribe Anthospermeae (Rubioideae, Rubiaceae)
| PRJEB62527 | ENA
Project description:ObjectivesAs a result of the introduction of rapid benchtop sequencers, the time required to subculture a bacterial pathogen to extract sufficient DNA for library preparation can now exceed the time to sequence said DNA. We have eliminated this rate-limiting step by developing a protocol to generate DNA libraries for whole-genome sequencing directly from single bacterial colonies grown on primary culture plates.MethodsWe developed our protocol using single colonies of 17 bacterial pathogens responsible for severe human infection that were grown using standard diagnostic media and incubation conditions. We then applied this method to four clinical scenarios that currently require time-consuming reference laboratory tests: full identification and genotyping of salmonellae; identification of blaNDM-1, a highly transmissible carbapenemase resistance gene, in Klebsiella pneumoniae; detection of genes encoding staphylococcal toxins associated with specific disease syndromes; and monitoring of vaccine targets to detect vaccine escape in Neisseria meningitidis.ResultsWe validated our single-colony whole-genome sequencing protocol for all 40 combinations of pathogen and selective, non-selective or indicator media tested in this study. Moreover, we demonstrated the clinical value of this method compared with current reference laboratory tests.ConclusionsThis advance will facilitate the implementation of whole-genome sequencing into diagnostic and public health microbiology.
| S-EPMC3977605 | biostudies-literature
Project description:DNA barcoding is critical to conservation and biodiversity research, yet public reference databases are incomplete. Existing barcode databases are biased toward cytochrome oxidase subunit I (COI) and frequently lack associated voucher specimens or geospatial metadata, which can hinder reliable species assignments. The emergence of metabarcoding approaches such as environmental DNA (eDNA) has necessitated multiple marker techniques combined with barcode reference databases backed by voucher specimens. Reference barcodes have traditionally been generated by Sanger sequencing, however sequencing multiple markers is costly for large numbers of specimens, requires multiple separate PCR reactions, and limits resulting sequences to targeted regions. High-throughput sequencing techniques such as genome skimming enable assembly of complete mitogenomes, which contain the most commonly used barcoding loci (e.g., COI, 12S, 16S), as well as nuclear ribosomal repeat regions (e.g., ITS1&2, 18S). We evaluated the feasibility of genome skimming to generate barcode references databases for marine fishes by assembling complete mitogenomes and nuclear ribosomal repeats. We tested genome skimming across a taxonomically diverse selection of 12 marine fish species from the collections of the National Museum of Natural History, Smithsonian Institution. We generated two sequencing libraries per species to test the impact of shearing method (enzymatic or mechanical), extraction method (kit-based or automated), and input DNA concentration. We produced complete mitogenomes for all non-chondrichthyans (11/12 species) and assembled nuclear ribosomal repeats (18S-ITS1-5.8S-ITS2-28S) for all taxa. The quality and completeness of mitogenome assemblies was not impacted by shearing method, extraction method or input DNA concentration. Our results reaffirm that genome skimming is an efficient and (at scale) cost-effective method to generate all mitochondrial and common nuclear DNA barcoding loci for multiple species simultaneously, which has great potential to scale for future projects and facilitate completing barcode reference databases for marine fishes.
| S-EPMC9359134 | biostudies-literature
Project description:Studies on foraging partitioning in pollinators can provide critical information to the understanding of food-web niche and pollination functions, thus aiding conservation. Metabarcoding based on PCR amplification and high-throughput sequencing has seen increasing applications in characterizing pollen loads carried by pollinators. However, amplification bias across taxa could lead to unpredictable artefacts in estimation of pollen compositions. We examined the efficacy of a genome-skimming method based on direct shotgun sequencing in quantifying mixed pollen, using mock samples (five and 14 mocks of flower and bee pollen, respectively). The results demonstrated a high level of repeatability and accuracy in identifying pollen from mixtures of varied species ratios. All pollen species were detected in all mocks, and pollen frequencies estimated from the number of sequence reads of each species were significantly correlated with pollen count proportions (linear model, R2 = 86.7%, p = 2.2e-16). For >97% of the mixed taxa, pollen proportion could be quantified by sequencing to the correct order of magnitude, even for species which constituted only 0.2% of the total pollen. In addition, DNA extracted from pollen grains equivalent to those collected from a single honeybee corbicula was sufficient for genome-skimming. We conclude that genome-skimming is a feasible approach to identifying and quantifying mixed pollen samples. By providing reliable and sensitive taxon identification and relative abundance, this method is expected to improve our understanding in studies that involve plant-pollinator interactions, such as pollen preference in corbiculate bees, pollen diet analyses and identification of landscape pollen resource use from beehives.
| S-EPMC6900181 | biostudies-literature
Project description:BACKGROUND: Terns (Charadriiformes: Sterninae) are a lineage of cosmopolitan shorebirds with a disputed evolutionary history that comprises several species of conservation concern. As a non-model system in genetics, previous study has left most of the nuclear genome unexplored, and population-level studies are limited to only 15% of the world's species of terns and noddies. Screening of polymorphic nuclear sequence markers is needed to enhance genetic resolution because of supposed low mitochondrial mutation rate, documentation of nuclear insertion of hypervariable mitochondrial regions, and limited success of microsatellite enrichment in terns. Here, we investigated the phylogenetic and population genetic utility for terns and relatives of a variety of nuclear markers previously developed for other birds and spanning the nuclear genome. Markers displaying a variety of mutation rates from both the nuclear and mitochondrial genome were tested and prioritized according to optimal cross-species amplification and extent of genetic polymorphism between (1) the main tern clades and (2) individual Royal Terns (Thalasseus maxima) breeding on the US East Coast. RESULTS: Results from this genome skimming effort yielded four new nuclear sequence-based markers for tern phylogenetics and 11 intra-specific polymorphic markers. Further, comparison between the two genomes indicated a phylogenetic conflict at the base of terns, involving the inclusion (mitochondrial) or exclusion (nuclear) of the Angel Tern (Gygis alba). Although limited mitochondrial variation was confirmed, both nuclear markers and a short tandem repeat in the mitochondrial control region indicated the presence of considerable genetic variation in Royal Terns at a regional scale. CONCLUSIONS: These data document the value of intronic markers to the study of terns and allies. We expect that these and additional markers attained through next-generation sequencing methods will accurately map the genetic origin and species history of this group of birds.
| S-EPMC3292991 | biostudies-literature
Project description:Recent technological advances in long-read high-throughput sequencing and assembly methods have facilitated the generation of annotated chromosome-scale whole-genome sequence data for evolutionary studies; however, generating such data can still be difficult for many plant species. For example, obtaining high-molecular-weight DNA is typically impossible for samples in historical herbarium collections, which often have degraded DNA. The need to fast-freeze newly collected living samples to conserve high-quality DNA can be complicated when plants are only found in remote areas. Therefore, short-read reduced-genome representations, such as target capture and genome skimming, remain important for evolutionary studies. Here, we review the pros and cons of each technique for non-model plant taxa. We provide guidance related to logistics, budget, the genomic resources previously available for the target clade, and the nature of the study. Furthermore, we assess the available bioinformatic analyses, detailing best practices and pitfalls, and suggest pathways to combine newly generated data with legacy data. Finally, we explore the possible downstream analyses allowed by the type of data generated using each technique. We provide a practical guide to help researchers make the best-informed choice regarding reduced genome representation for evolutionary studies of non-model plants in cases where whole-genome sequencing remains impractical.
| S-EPMC10439825 | biostudies-literature