We used Massively Parallel High-Throughput Sequencing to obtain genetic data from a 145-year old holotype specimen of the flying lizard, Draco cristatellus. Obtaining genetic data from this holotype was necessary to resolve an otherwise intractable taxonomic problem involving the status of this species relative to closely related sympatric Draco species that cannot otherwise be distinguished from one another on the basis of museum specimens. Initial analyses suggested that the DNA present in the holotype sample was so degraded as to be unusable for sequencing. However, we used a specialized extraction procedure developed for highly degraded ancient DNA samples and MiSeq shotgun sequencing to obtain just enough low-coverage mitochondrial DNA (721 base pairs) to conclusively resolve the species status of the holotype as well as a second known specimen of this species. The holotype was prepared before the advent of formalin-fixation and therefore was most likely originally fixed with ethanol and never exposed to formalin. Whereas conventional wisdom suggests that formalin-fixed samples should be the most challenging for DNA sequencing, we propose that evaporation during long-term alcohol storage and consequent water-exposure may subject older ethanol-fixed museum specimens to hydrolytic damage. If so, this may pose an even greater challenge for sequencing efforts involving historical samples.

Targeted enrichment of >1,000 ultraconserved elements and divergence time analysis resolves relationships among 120 major acanthomorph lineages and provides a new timescale for acanthomorph radiation in the wake of the K–Pg boundary.

Plasmodium falciparum is a significant human pathogen, particularly in the historical context of the ancient Mediterranean region. The causative species of malaria are “invisible” in the historical record, while malaria as a disease entity is indirectly supported by evidence from literary works (e.g., the Hippocratic Corpus, Celsus’ De Medicina) and non-specific skeletal pathological responses. Although ancient DNA may demonstrate the presence of a pathogen, there remain theoretical and methodological challenges in contextualizing such molecular evidence. Here we present a framework to explore the biosocial context of malaria in 1st–4th c. CE central-southern Italy using genomic, literary, epidemiological, and archaeological evidence to highlight relationships between the Plasmodium parasite, human hosts, Anopheles vector, and environment. By systematically integrating these evidentiary sources, our approach highlights the importance of disease ecology (e.g., climate and landscape) and human-environment interactions (e.g., land use patterns, such as agriculture or infrastructure activities) that differentially impact the potential scope of malaria in the past.

High throughput sequencing (HTS) has been used for a number of years in the field of paleogenomics to facilitate the recovery of small DNA fragments from ancient specimens. Recently, these techniques have also been applied in forensics, where they have been used for the recovery of mitochondrial DNA sequences from samples where traditional PCR-based assays fail because of the very short length of endogenous DNA molecules. Here, we describe the biological sexing of a ~4000-year-old Egyptian mummy using shotgun sequencing and two established methods of biological sex determination (RX and RY), by way of mitochondrial genome analysis as a means of sequence data authentication. This particular case of historical interest increases the potential utility of HTS techniques for forensic purposes by demonstrating that data from the more discriminatory nuclear genome can be recovered from the most damaged specimens, even in cases where mitochondrial DNA cannot be recovered with current PCR-based forensic technologies. Although additional work remains to be done before nuclear DNA recovered via these methods can be used routinely in operational casework for individual identification purposes, these results indicate substantial promise for the retrieval of probative individually identifying DNA data from the most limited and degraded forensic specimens.

Premise of the Study The development of pipelines for locus discovery has spurred the use of target enrichment for plant phylogenomics. However, few studies have compared pipelines from locus discovery and bait design, through validation, to tree inference. We compared three methods within Leguminosae (Fabaceae) and present a workflow for future efforts. Methods Using 30 transcriptomes, we compared Hyb-Seq, MarkerMiner, and the Yang and Smith (Y&S) pipelines for locus discovery, validated 7501 baits targeting 507 loci across 25 genera via Illumina sequencing, and inferred gene and species trees via concatenation- and coalescent-based methods. Results Hyb-Seq discovered loci with the longest mean length. MarkerMiner discovered the most conserved loci with the least flagged as paralogous. Y&S offered the most parsimony-informative sites and putative orthologs. Target recovery averaged 93% across taxa. We optimized our targeted locus set based on a workflow designed to minimize paralog/ortholog conflation and thus present 423 loci for legume phylogenomics. Conclusions Methods differed across criteria important for phylogenetic marker development. We recommend Hyb-Seq as a method that may be useful for most phylogenomic projects. Our targeted locus set is a resource for future, community-driven efforts to reconstruct the legume tree of life.

The Caribbean was one of the last parts of the Americas to be settled by humans, but how and when the islands were first occupied remains a matter of debate. Ancient DNA can help answering these questions, but the work has been hampered by poor DNA preservation. We report the genome sequence of a 1,000-year-old Lucayan Taino individual recovered from the site of Preacher’s Cave in the Bahamas. We sequenced her genome to 12.4-fold coverage and show that she is genetically most closely related to present-day Arawakan speakers from northern South America, suggesting that the ancestors of the Lucayans originated there. Further, we find no evidence for recent inbreeding or isolation in the ancient genome, suggesting that the Lucayans had a relatively large effective population size. Finally, we show that the native American components in some present-day Caribbean genomes are closely related to the ancient Taino, demonstrating an element of continuity between precontact populations and present-day Latino populations in the Caribbean.

We resolve a longstanding question regarding the kinship of two high-status Egyptians from the 12th Dynasty, Nakht-Ankh and Khnum-Nakht, whose mummies were discovered in 1907 by Egyptian workmen directed by Flinders Petrie and Ernest Mackay. Although their coffin inscriptions indicate that Nakht-Ankh and Khnum-Nakht were brothers, when the mummies were unwrapped in 1908 the skeletal morphologies were found to be quite different, suggesting an absence of family relationship. We extracted ancient DNA from the teeth of the two mummies and, following hybridization capture of the mitochondrial and Y chromosome fractions, sequenced the DNA by a next generation method. Analysis of single nucleotide polymorphisms showed that both Nakht-Ankh and Khnum-Nakht belonged to mitochondrial haplotype M1a1, suggesting a maternal relationship. The Y chromosome sequences were less complete but showed variations between the two mummies, indicating that Nakht-Ankh and Khnum-Nakht had different fathers. Our study emphasizes the importance of kinship in ancient Egypt, and represents the first successful typing of both mitochondrial and Y chromosomal DNA in Egyptian mummies.