Second-pandemic strain of Vibrio cholerae from the Philadelphia cholera outbreak of 1849.

In the 19th century, there were several major cholera pandemics in the Indian subcontinent, Europe, and North America. The causes of these outbreaks and the genomic strain identities remain a mystery. We used targeted high-throughput sequencing to reconstruct the Vibrio cholerae genome from the preserved intestine of a victim of the 1849 cholera outbreak in Philadelphia, part of the second cholera pandemic. This O1 biotype strain has 95 to 97% similarity with the classical O395 genome, differing by 203 single-nucleotide polymorphisms (SNPs), lacking three genomic islands, and probably having one or more tandem cholera toxin prophage (CTX) arrays, which potentially affected its virulence. This result highlights archived medical remains as a potential resource for investigations into the genomic origins of past pandemics.

Ancient whole genome enrichment using baits built from modern DNA.

We report metrics from complete genome capture of nuclear DNA from extinct mammoths using biotinylated RNAs transcribed from an Asian elephant DNA extract. Enrichment of the nuclear genome ranged from 1.06- to 18.65-fold, to an apparent maximum threshold of ∼80% on-target. This projects an order of magnitude less costly complete genome sequencing from long-dead organisms, even when a reference genome is unavailable for bait design.

Comparison of methods in the recovery of nucleic acids from archival formalin-fixed paraffin-embedded autopsy tissues.

Archival formalin-fixed paraffin-embedded (FFPE) human tissue collections are typically in poor states of storage across the developing world. With advances in biomolecular techniques, these extraordinary and virtually untapped resources have become an essential part of retrospective epidemiological studies. To successfully use such tissues in genomic studies, scientists require high nucleic acid yields and purity. In spite of the increasing number of FFPE tissue kits available, few studies have analyzed their applicability in recovering high-quality nucleic acids from archived human autopsy samples. Here we provide a study involving 10 major extraction methods used to isolate total nucleic acid from FFPE tissues ranging in age from 3 to 13years. Although all 10 methods recovered quantifiable amounts of DNA, only 6 recovered quantifiable RNA, varying considerably and generally yielding lower DNA concentrations. Overall, we show quantitatively that TrimGen's WaxFree method and our in-house phenol-chloroform extraction method recovered the highest yields of amplifiable DNA, with considerable polymerase chain reaction (PCR) inhibition, whereas Ambion's RecoverAll method recovered the most amplifiable RNA.

A molecular portrait of maternal sepsis from Byzantine Troy.

Pregnancy complications are poorly represented in the archeological record, despite their importance in contemporary and ancient societies. While excavating a Byzantine cemetery in Troy, we discovered calcified abscesses among a woman's remains. Scanning electron microscopy of the tissue revealed 'ghost cells', resulting from dystrophic calcification, which preserved ancient maternal, fetal and bacterial DNA of a severe infection, likely chorioamnionitis. Gardnerella vaginalis and Staphylococcus saprophyticus dominated the abscesses. Phylogenomic analyses of ancient, historical, and contemporary data showed that G. vaginalis Troy fell within contemporary genetic diversity, whereas S. saprophyticus Troy belongs to a lineage that does not appear to be commonly associated with human disease today. We speculate that the ecology of S. saprophyticus infection may have differed in the ancient world as a result of close contacts between humans and domesticated animals. These results highlight the complex and dynamic interactions with our microbial milieu that underlie severe maternal infections.

Estimation of Gene Insertion/Deletion Rates with Missing Data.

Lateral gene transfer is an important mechanism for evolution among bacteria. Here, genome-wide gene insertion and deletion rates are modeled in a maximum-likelihood framework with the additional flexibility of modeling potential missing data. The performance of the models is illustrated using simulations and a data set on gene family phyletic patterns from Gardnerella vaginalis that includes an ancient taxon. A novel application involving pseudogenization/genome reduction magnitudes is also illustrated, using gene family data from Mycobacterium spp. Finally, an R package called indelmiss is available from the Comprehensive R Archive Network at https://cran.r-project.org/package=indelmiss, with support documentation and examples.

Ancient pathogen DNA in archaeological samples detected with a Microbial Detection Array.

Ancient human remains of paleopathological interest typically contain highly degraded DNA in which pathogenic taxa are often minority components, making sequence-based metagenomic characterization costly. Microarrays may hold a potential solution to these challenges, offering a rapid, affordable, and highly informative snapshot of microbial diversity in complex samples without the lengthy analysis and/or high cost associated with high-throughput sequencing. Their versatility is well established for modern clinical specimens, but they have yet to be applied to ancient remains. Here we report bacterial profiles of archaeological and historical human remains using the Lawrence Livermore Microbial Detection Array (LLMDA). The array successfully identified previously-verified bacterial human pathogens, including Vibrio cholerae (cholera) in a 19th century intestinal specimen and Yersinia pestis ("Black Death" plague) in a medieval tooth, which represented only minute fractions (0.03% and 0.08% alignable high-throughput shotgun sequencing reads) of their respective DNA content. This demonstrates that the LLMDA can identify primary and/or co-infecting bacterial pathogens in ancient samples, thereby serving as a rapid and inexpensive paleopathological screening tool to study health across both space and time.

Comment on "DNA from pre-Clovis human coprolites in Oregon, North America".

Gilbert et al. (Reports, 9 May 2008, p. 786) analyzed DNA from radiocarbon-dated paleofecal remains from Paisley Cave, Oregon, which ostensibly demonstrate a human presence in North America predating the well-established Clovis complex. We question the authenticity of their DNA results and argue that in the absence of intact stratigraphy and diagnostic artifacts, and in view of carbon isotope anomalies, the radiocarbon dates of the oldest specimens are unreliable.