A novel NGS library preparation method to characterize native termini of fragmented DNA.

Biological and chemical DNA fragmentation generates DNA molecules with a variety of termini, including blunt ends and single-stranded overhangs. We have developed a Next Generation Sequencing (NGS) assay, XACTLY, to interrogate the termini of fragmented DNA, information traditionally lost in standard NGS library preparation methods. Here we describe the XACTLY method, showcase its sensitivity and specificity, and demonstrate its utility in in vitro experiments. The XACTLY assay is able to report relative abundances of all lengths and types (5' and 3') of single-stranded overhangs, if present, on each DNA fragment with an overall accuracy between 80-90%. In addition, XACTLY retains the sequence of each native DNA molecule after fragmentation and can capture the genomic landscape of cleavage events at single nucleotide resolution. The XACTLY assay can be applied as a novel research and discovery tool for fragmentation analyses and in cell-free DNA

Correlating Bayesian date estimates with climatic events and domestication using a bovine case study

The tribe Bovini contains a number of commercially and culturally important species, such as cattle. Understanding their evolutionary time scale is important for distinguishing between post-glacial and domestication-associated population expansions, but estimates of bovine divergence times have been hindered by a lack of reliable calibration points. We present a Bayesian phylogenetic analysis of 481 mitochondrial D-loop sequences, including 228 radiocarbon-dated ancient DNA sequences, using a multi-demographic coalescent model. By employing the radiocarbon dates as internal calibrations, we co-estimate the bovine phylogeny and divergence times in a relaxed-clock framework. The analysis yields evidence for significant population expansions in both taurine and zebu cattle, European aurochs and yak clades. The divergence age estimates support domestication-associated expansion times (less than 12kyr) for the major haplogroups of cattle. We compare the molecular and palaeontological estimates for the Bison-Bos divergence

Is amino acid racemization a useful tool for screening for ancient DNA in bone?

Many rare and valuable ancient specimens now carry the scars of ancient DNA research, as questions of population genetics and phylogeography require larger sample sets. This fuels the demand for reliable techniques to screen for DNA preservation prior to destructive sampling. Only one such technique has been widely adopted: the extent of aspartic acid racemization (AAR). The kinetics of AAR are believed to be similar to the rate of DNA depurination and therefore a good measure of the likelihood of DNA survival. Moreover, AAR analysis is only minimally destructive. We report the first comprehensive test of AAR using 91 bone and teeth samples from temperate and high-latitude sites that were analysed for DNA. While the AAR range of all specimens was low (0.02-0.17), no correlation was found between the extent of AAR and DNA amplification success. Additional heating experiments and surveys of the literature indicated that d/l Asx is low in bones until almost all the collagen is lost. This is because aspartic acid is retained in the bone within the constrained environment of the collagen triple helix, where it cannot racemize for steric reasons. Only if the helix denatures to soluble gelatin can Asx racemize readily, but this soluble gelatine is readily lost in most burial environments. We conclude that Asx d/l is not a useful screening technique for ancient DNA from bone

Sedimentary DNA and molecular evidence for early human occupation of the Faroe Islands

The Faroe Islands, a North Atlantic archipelago between Norway and Iceland, were settled by Viking explorers in the mid-9th century CE. However, several indirect lines of evidence suggest earlier occupation of the Faroes by people from the British Isles. Here, we present sedimentary ancient DNA and molecular fecal biomarker evidence from a lake sediment core proximal to a prominent archaeological site in the Faroe Islands to establish the earliest date for the arrival of people in the watershed. Our results reveal an increase in fecal biomarker concentrations and the first appearance of sheep DNA at 500 CE (95% confidence interval 370-610 CE), pre-dating Norse settlements by 300 years. Sedimentary plant DNA indicates an increase in grasses and the disappearance of woody plants, likely due to livestock grazing. This provides unequivocal evidence for human arrival and livestock disturbance in the Faroe Islands centuries before Viking settlement in the 9th century

Sedimentary DNA and Molecular Evidence for Early Human Occupation of the Faroe Islands

The Faroe Islands, a North Atlantic archipelago between Norway and Iceland, were settled by Viking explorers in the mid-9th century CE. However, several indirect lines of evidence suggest earlier occupation of the Faroes by people from the British Isles. Here, we present sedimentary ancient DNA and molecular fecal biomarker evidence from a lake sediment core proximal to a prominent archaeological site in the Faroe Islands to establish the earliest date for the arrival of people in the watershed. Our results reveal an increase in fecal biomarker concentrations and the first appearance of sheep DNA at 500 CE (95% confidence interval 370-610 CE), pre-dating Norse settlements by 300 years. Sedimentary plant DNA indicates an increase in grasses and the disappearance of woody plants, likely due to livestock grazing. This provides unequivocal evidence for human arrival and livestock disturbance in the Faroe Islands centuries before Viking settlement in the 9th century

Synonymous Substitution Rates Predict HIV Disease Progression as a Result of Underlying Replication Dynamics

Upon HIV transmission, some patients develop AIDS in only a few months, while others remain disease free for 20 or more years. This variation in the rate of disease progression is poorly understood and has been attributed to host genetics, host immune responses, co-infection, viral genetics, and adaptation. Here, we develop a new “relaxed-clock” phylogenetic method to estimate absolute rates of synonymous and nonsynonymous substitution through time. We identify an unexpected association between the synonymous substitution rate of HIV and disease progression parameters. Since immune activation is the major determinant of HIV disease progression, we propose that this process can also determine viral generation times, by creating favourable conditions for HIV replication. These conclusions may apply more generally to HIV evolution, since we also observed an overall low synonymous substitution rate for HIV-2, which is known to be less pathogenic than HIV-1 and capable of tempering the detrimental effects of immune activation. Humoral immune responses, on the other hand, are the major determinant of nonsynonymous rate changes through time in the envelope gene, and our relaxed-clock estimates support a decrease in selective pressure as a consequence of immune system collapse

Is amino acid racemization a useful tool for screening for ancient DNA in bone?

Many rare and valuable ancient specimens now carry the scars of ancient DNA research, as questions of population genetics and phylogeography require larger sample sets. This fuels the demand for reliable techniques to screen for DNA preservation prior to destructive sampling. Only one such technique has been widely adopted: the extent of aspartic acid racemization (AAR). The kinetics of AAR are believed to be similar to the rate of DNA depurination and therefore a good measure of the likelihood of DNA survival. Moreover, AAR analysis is only minimally destructive. We report the first comprehensive test of AAR using 91 bone and teeth samples from temperate and high-latitude sites that were analysed for DNA. While the AAR range of all specimens was low (0.02 -0.17), no correlation was found between the extent of AAR and DNA amplification success. Additional heating experiments and surveys of the literature indicated that D/L Asx is low in bones until almost all the collagen is lost. This is because aspartic acid is retained in the bone within the constrained environment of the collagen triple helix, where it cannot racemize for steric reasons. Only if the helix denatures to soluble gelatin can Asx racemize readily, but this soluble gelatine is readily lost in most burial environments. We conclude that Asx D/L is not a useful screening technique for ancient DNA from bone

Isotopic evidence for long-distance connections of the AD thirteenth century Promontory caves occupants

Las cuevas Promontory (Utah) y la Cueva Franktown (Colorado) contienen registros de alta fidelidad en cuanto a las ocupaciones de corto término de grupos humanos con cultura material conectada al Subártico/Planicies del Norte. Esta investigación utiliza excremento, pelo, piel de cuero y colágeno del hueso de bisonte provenientes de los sitios Promontory y Franktown para establecer una base de datos isotópicos locales y variabilidad isotópica de carbono e identifica la piel de cuero de procedencia lejana. La envoltura de tobillo de un mocasín recuperado de la Cueva 1 de Promontory tuvo un valor δ13C que indica un componente C4 considerable en la dieta del animal, lo que es un resultado distinto a las dietas C3 que se identificaron a partir del análisis de 171 muestras de bisonte provenientes de Promontory y el norte de Utah. Aplicamos una combinación única de análisis isotópico de tejidos múltiples, mapas isotópicos (“isoscapes”) de carbono, ADN antiguo (identificación de sexo y especie), tasa de remodelación de tejidos, contextos arqueológicos y la ecología del bisonte para demonstrar que el valor alto δ13C probablemente no es un resultado debido al consumo de plantas locales, movilidad del bisonte o comercio. En cambio, la piel de cuero del bisonte fue probablemente adquirida a través de viajes de larga distancia hacia zonas con pastos C4 abundantes muy al sur o al este. El conocimiento extenso sobre el paisaje obtenido a través de asociaciones de larga distancia debió permitir a los habitantes de las cuevas Promontory tomar decisiones bien informadas sobre las direcciones y rutas de movimiento para realizar un cambio territorial, el cual parece haber ocurrido a finales del sigo XIII.The Promontory caves (Utah) and Franktown Cave (Colorado) contain high-fidelity records of short-term occupations by groups with material culture connections to the Subarctic/Northern Plains. This research uses Promontory and Franktown bison dung, hair, hide, and bone collagen to establish local baseline carbon isotopic variability and identify leather from a distant source. The ankle wrap of one Promontory Cave 1 moccasin had a δ13C value that indicates a substantial C4 component to the animal's diet, unlike the C3 diets inferred from 171 other Promontory and northern Utah bison samples. We draw on a unique combination of multitissue isotopic analysis, carbon isoscapes, ancient DNA (species and sex identification), tissue turnover rates, archaeological contexts, and bison ecology to show that the high δ13C value was not likely a result of local plant consumption, bison mobility, or trade. Instead, the bison hide was likely acquired via long-distance travel to/from an area of abundant C4 grasses far to the south or east. Expansive landscape knowledge gained through long-distance associations would have allowed Promontory caves inhabitants to make well-informed decisions about directions and routes of movement for a territorial shift, which seems to have occurred in the late thirteenth century

Ancient papillomavirus-host co-speciation in Felidae

The evolutionary rate of feline papillomaviruses is inferred from the phylogenetic analysis of their hosts, providing evidence for long-term virus-host co-speciatio

Rapid range shifts and megafaunal extinctions associated with late Pleistocene climate change

Large-scale changes in global climate at the end of the Pleistocene significantly impacted ecosystems across North America. However, the pace and scale of biotic turnover in response to both the Younger Dryas cold period and subsequent Holocene rapid warming have been challenging to assess because of the scarcity of well dated fossil and pollen records that covers this period. Here we present an ancient DNA record from Hall's Cave, Texas, that documents 100 vertebrate and 45 plant taxa from bulk fossils and sediment. We show that local plant and animal diversity dropped markedly during Younger Dryas cooling, but while plant diversity recovered in the early Holocene, animal diversity did not. Instead, five extant and nine extinct large bodied animals disappeared from the region at the end of the Pleistocene. Our findings suggest that climate change affected the local ecosystem in Texas over the Pleistocene-Holocene boundary, but climate change on its own may not explain the disappearance of the megafauna at the end of the Pleistocene.This study was supported by the Australian Research Council Discovery Project DP160104473, Forrest Research Foundation (to F.V.S.); field work in 2016 was funded by Stafford Research LLC