The Paisley Caves in Oregon record the oldest directly dated human remains (DNA) in the Western Hemisphere. More than 100 high-precision radiocarbon dates show that deposits containing artifacts and ...coprolites ranging in age from 12,450 to 2295 ¹⁴C years ago are well stratified. Western Stemmed projectile points were recovered in deposits dated to 11,070 to 11,340 ¹⁴C years ago, a time contemporaneous with or preceding the Clovis technology. There is no evidence of diagnostic Clovis technology at the site. These two distinct technologies were parallel developments, not the product of a unilinear technological evolution. "Blind testing" analysis of coprolites by an independent laboratory confirms the presence of human DNA in specimens of pre-Clovis age. The colonization of the Americas involved multiple technologically divergent, and possibly genetically divergent, founding groups.
Research involving ancient DNA (aDNA) has experienced a true technological revolution in recent years through advances in the recovery of aDNA and, particularly, through applications of ...high-throughput sequencing. Formerly restricted to the analysis of only limited amounts of genetic information, aDNA studies have now progressed to whole-genome sequencing for an increasing number of ancient individuals and extinct species, as well as to epigenomic characterization. Such advances have enabled the sequencing of specimens of up to 1 million years old, which, owing to their extensive DNA damage and contamination, were previously not amenable to genetic analyses. In this Review, we discuss these varied technical challenges and solutions for sequencing ancient genomes and epigenomes.
Domesticated maize evolved from wild teosinte under human influences in Mexico beginning around 9000 years before the present (yr B.P.), traversed Central America by ~7500 yr B.P., and spread into ...South America by ~6500 yr B.P. Landrace and archaeological maize genomes from South America suggest that the ancestral population to South American maize was brought out of the domestication center in Mexico and became isolated from the wild teosinte gene pool before traits of domesticated maize were fixed. Deeply structured lineages then evolved within South America out of this partially domesticated progenitor population. Genomic, linguistic, archaeological, and paleoecological data suggest that the southwestern Amazon was a secondary improvement center for partially domesticated maize. Multiple waves of human-mediated dispersal are responsible for the diversity and biogeography of modern South American maize.
The ability to inexpensively describe taxonomic diversity is critical in this era of rapid climate and biodiversity changes. The recent genome-skimming approach extends current barcoding practices ...beyond short markers by applying low-pass sequencing and recovering whole organelle genomes computationally. This approach discards the nuclear DNA, which constitutes the vast majority of the data. In contrast, we suggest using all unassembled reads. We introduce an assembly-free and alignment-free tool, Skmer, to compute genomic distances between the query and reference genome skims. Skmer shows excellent accuracy in estimating distances and identifying the closest match in reference datasets.
Freshwater ecosystems are among the most endangered habitats on Earth, with thousands of animal species known to be threatened or already extinct. Reliable monitoring of threatened organisms is ...crucial for data‐driven conservation actions but remains a challenge owing to nonstandardized methods that depend on practical and taxonomic expertise, which is rapidly declining. Here, we show that a diversity of rare and threatened freshwater animals—representing amphibians, fish, mammals, insects and crustaceans—can be detected and quantified based on DNA obtained directly from small water samples of lakes, ponds and streams. We successfully validate our findings in a controlled mesocosm experiment and show that DNA becomes undetectable within 2 weeks after removal of animals, indicating that DNA traces are near contemporary with presence of the species. We further demonstrate that entire faunas of amphibians and fish can be detected by high‐throughput sequencing of DNA extracted from pond water. Our findings underpin the ubiquitous nature of DNA traces in the environment and establish environmental DNA as a tool for monitoring rare and threatened species across a wide range of taxonomic groups.
See also the Perspective by Lodge et al
Metabarcoding of environmental samples on second‐generation sequencing platforms has rapidly become a valuable tool for ecological studies. A fundamental assumption of this approach is the reliance ...on being able to track tagged amplicons back to the samples from which they originated. In this study, we address the problem of sequences in metabarcoding sequencing outputs with false combinations of used tags (tag jumps). Unless these sequences can be identified and excluded from downstream analyses, tag jumps creating sequences with false, but already used tag combinations, can cause incorrect assignment of sequences to samples and artificially inflate diversity. In this study, we document and investigate tag jumping in metabarcoding studies on Illumina sequencing platforms by amplifying mixed‐template extracts obtained from bat droppings and leech gut contents with tagged generic arthropod and mammal primers, respectively. We found that an average of 2.6% and 2.1% of sequences had tag combinations, which could be explained by tag jumping in the leech and bat diet study, respectively. We suggest that tag jumping can happen during blunt‐ending of pools of tagged amplicons during library build and as a consequence of chimera formation during bulk amplification of tagged amplicons during library index PCR. We argue that tag jumping and contamination between libraries represents a considerable challenge for Illumina‐based metabarcoding studies, and suggest measures to avoid false assignment of tag jumping‐derived sequences to samples.
With the advent of DNA sequencing‐based techniques, the way we detect and measure biodiversity is undergoing a radical shift. There is also an increasing awareness of the need to employ intuitively ...meaningful diversity measures based on unified statistical frameworks, so that different results can be easily interpreted and compared. This article aimed to serve as a guide to implementing biodiversity assessment using the general statistical framework developed around Hill numbers into the analysis of systems characterized using DNA sequencing‐based techniques (e.g., diet, microbiomes and ecosystem biodiversity). Specifically, we discuss (a) the DNA‐based approaches for defining the types upon which diversity is measured, (b) how to weight the importance of each type, (c) the differences between abundance‐based versus incidence‐based approaches, (d) the implementation of phylogenetic information into diversity measurement, (e) hierarchical diversity partitioning, (f) dissimilarity and overlap measurement and (g) how to deal with zero‐inflated, insufficient and biased data. All steps are reproduced with real data to also provide step‐by‐step bash and R scripts to enable straightforward implementation of the explained procedures.
Metabarcoding of environmental samples has many challenges and limitations that require carefully considered laboratory and analysis workflows to ensure reliable results. We explore how decisions ...regarding study design, laboratory set‐up, and bioinformatic processing affect the final results, and provide guidelines for reliable study of environmental samples.
We evaluate the performance of four primer sets targeting COI and 16S regions characterizing arthropod diversity in bat faecal samples, and investigate how metabarcoding results are affected by parameters including: (1) number of PCR replicates per sample, (2) sequencing depth, (3) PCR replicate processing strategy (i.e. either additively, by combining the sequences obtained from the PCR replicates, or restrictively, by only retaining sequences that occur in multiple PCR replicates for each sample), (4) minimum copy number for sequences to be retained, (5) chimera removal, and (6) similarity thresholds for Operational Taxonomic Unit (OTU) clustering. Lastly, we measure within‐ and between‐taxa dissimilarities when using sequences from public databases to determine the most appropriate thresholds for OTU clustering and taxonomy assignment.
Our results show that the use of multiple primer sets reduces taxonomic biases and increases taxonomic coverage. Taxonomic profiles resulting from each primer set are principally affected by how many PCR replicates are carried out per sample and how sequences are filtered across them, the sequence copy number threshold and the OTU clustering threshold. We also report considerable diversity differences between PCR replicates from each sample. Sequencing depth increases the dissimilarity between PCR replicates unless the bioinformatic strategies to remove allegedly artefactual sequences are adjusted according to the number of analysed sequences. Finally, we show that the appropriate identity thresholds for OTU clustering and taxonomy assignment differ between markers.
Metabarcoding of complex environmental samples ideally requires (1) investigation of whether more than one primer sets targeting the same taxonomic group is needed to offset primer biases, (2) more than one PCR replicate per sample, (3) bioinformatic processing of sequences that balance diversity detection with removal of artefactual sequences, and (4) empirical selection of OTU clustering and taxonomy assignment thresholds tailored to each marker and the obtained taxa.
Antimicrobial resistance (AMR) is a growing threat to human and animal health. However, in aquatic animals-the fastest growing food animal sector globally-AMR trends are seldom documented, ...particularly in Asia, which contributes two-thirds of global food fish production. Here, we present a systematic review and meta-analysis of 749 point prevalence surveys reporting antibiotic-resistant bacteria from aquatic food animals in Asia, extracted from 343 articles published in 2000-2019. We find concerning levels of resistance to medically important antimicrobials in foodborne pathogens. In aquaculture, the percentage of antimicrobial compounds per survey with resistance exceeding 50% (P50) plateaued at 33% 95% confidence interval (CI) 28 to 37% between 2000 and 2018. In fisheries, P50 decreased from 52% 95% CI 39 to 65% to 22% 95% CI 14 to 30%. We map AMR at 10-kilometer resolution, finding resistance hotspots along Asia's major river systems and coastal waters of China and India. Regions benefitting most from future surveillance efforts are eastern China and India. Scaling up surveillance to strengthen epidemiological evidence on AMR and inform aquaculture and fisheries interventions is needed to mitigate the impact of AMR globally.
The exploitation of non-invasive samples has been widely used in genetic monitoring of terrestrial species. In aquatic ecosystems, non-invasive samples such as feces, shed hair or skin, are less ...accessible. However, the use of environmental DNA (eDNA) has recently been shown to be an effective tool for genetic monitoring of species presence in freshwater ecosystems. Detecting species in the marine environment using eDNA potentially offers a greater challenge due to the greater dilution, amount of mixing and salinity compared with most freshwater ecosystems. To determine the potential use of eDNA for genetic monitoring we used specific primers that amplify short mitochondrial DNA sequences to detect the presence of a marine mammal, the harbor porpoise, Phocoena phocoena, in a controlled environment and in natural marine locations. The reliability of the genetic detections was investigated by comparing with detections of harbor porpoise echolocation clicks by static acoustic monitoring devices. While we were able to consistently genetically detect the target species under controlled conditions, the results from natural locations were less consistent and detection by eDNA was less successful than acoustic detections. However, at one site we detected long-finned pilot whale, Globicephala melas, a species rarely sighted in the Baltic. Therefore, with optimization aimed towards processing larger volumes of seawater this method has the potential to compliment current visual and acoustic methods of species detection of marine mammals.
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