Kelly M. Harkins

Pre-Columbian mycobacterial genomes reveal seals as a source of New World human tuberculosis

Modern strains of Mycobacterium tuberculosis from the Americas are closely related to those from Europe, supporting the assumption that human tuberculosis was introduced post-contact. This notion, however, is incompatible with archaeological evidence of pre-contact tuberculosis in the New World. Comparative genomics of modern isolates suggests that M. tuberculosis attained its worldwide distribution following human dispersals out of Africa during the Pleistocene epoch, although this has yet to be confirmed with ancient calibration points. Here we present three 1,000-year-old mycobacterial genomes from Peruvian human skeletons, revealing that a member of the M. tuberculosis complex caused human disease before contact. The ancient strains are distinct from known human-adapted forms and are most closely related to those adapted to seals and sea lions. Two independent dating approaches suggest a most recent common ancestor for the M. tuberculosis complex less than 6,000 years ago, which supports a Holocene dispersal of the disease. Our results implicate sea mammals as having played a role in transmitting the disease to humans across the ocean.

Phylogenomic reconstruction supports supercontinent origins for Leishmania.

Leishmania, a genus of parasites transmitted to human hosts and mammalian/reptilian reservoirs by an insect vector, is the causative agent of the human disease complex leishmaniasis. The evolutionary relationships within the genus Leishmania and its origins are the source of ongoing debate, reflected in conflicting phylogenetic and biogeographic reconstructions. This study employs a recently described bioinformatics method, SISRS, to identify over 200,000 informative sites across the genome from newly sequenced and publicly available Leishmania data. This dataset is used to reconstruct the evolutionary relationships of this genus. Additionally, we constructed a large multi-gene dataset, using it to reconstruct the phylogeny and estimate divergence dates for species. We conclude that the genus Leishmania evolved at least 90-100 million years ago, supporting a modified version of the Multiple Origins hypothesis that we call the Supercontinent hypothesis. According to this scenario, separate Leishmania clades emerged prior to, and during, the breakup of Gondwana. Additionally, we confirm that reptile-infecting Leishmania are derived from mammalian forms and that the species that infect porcupines and sloths form a clade long separated from other species. Finally, we firmly place the guinea-pig infecting species, Leishmaniaenriettii, the globally dispersed Leishmaniasiamensis, and the newly identified Australian species from a kangaroo, as sibling species whose distribution arises from the ancient connection between Australia, Antarctica, and South America.

Ancient pathogen genomics: insights into timing and adaptation.

Disease is a major cause of natural selection affecting human evolution, whether through a sudden pandemic or persistent morbidity and mortality. Recent contributions in the field of ancient pathogen genomics have advanced our understanding of the antiquity and nature of human-pathogen interactions through time. Technical advancements have facilitated the recovery, enrichment, and high-throughput sequencing of pathogen and parasite DNA from archived and archaeological remains. These time-stamped genomes are crucial for calibrating molecular clocks to infer the timing of evolutionary events, while providing finer-grain resolution to phylogenetic reconstructions and complex biogeographical patterns. Additionally, genome scale data allow better identification of substitutions linked to adaptations of the pathogen to their human hosts. As methodology continues to improve, ancient genomes of humans and their diverse microbiomes from a range of eras and archaeological contexts will enable population-level ancient analyses in the near future and a better understanding of their co-evolutionary history.

Screening ancient tuberculosis with qPCR: challenges and opportunities

The field of ancient DNA (aDNA) has rapidly accelerated in recent years as a result of new methods in next-generation sequencing, library preparation and targeted enrichment. Such research is restricted, however, by the highly variable DNA preservation within different tissues, especially when isolating ancient pathogens from human remains. Identifying positive candidate samples via quantitative PCR (qPCR) for downstream procedures can reduce reagent costs, increase capture efficiency and maximize the number of sequencing reads of the target. This study uses four qPCR assays designed to target regions within the Mycobacterium tuberculosis complex (MTBC) to examine 133 human skeletal samples from a wide geographical and temporal range, identified by the presence of skeletal lesions typical of chronic disseminated tuberculosis. Given the inherent challenges working with ancient mycobacteria, strict criteria must be used and primer/probe design continually re-evaluated as new data from bacteria become available. Seven samples tested positive for multiple MTBC loci, supporting them as strong candidates for downstream analyses. Using strict and conservative criteria, qPCR remains a fast and effective screening tool when compared with screening by more expensive sequencing and enrichment technologies.

A composite genome approach to identify phylogenetically informative data from next-generation sequencing

BackgroundImprovements in sequencing technology now allow easy acquisition of large datasets; however, analyzing these data for phylogenetics can be challenging. We have developed a novel method to rapidly obtain homologous genomic data for phylogenetics directly from next-generation sequencing reads without the use of a reference genome. This software, called SISRS, avoids the time consuming steps of de novo whole genome assembly, multiple genome alignment, and annotation.ResultsFor simulations SISRS is able to identify large numbers of loci containing variable sites with phylogenetic signal. For genomic data from apes, SISRS identified thousands of variable sites, from which we produced an accurate phylogeny. Finally, we used SISRS to identify phylogenetic markers that we used to estimate the phylogeny of placental mammals. We recovered eight phylogenies that resolved the basal relationships among mammals using datasets with different levels of missing data. The three alternate resolutions of the basal relationships are consistent with the major hypotheses for the relationships among mammals, all of which have been supported previously by different molecular datasets.ConclusionsSISRS has the potential to transform phylogenetic research. This method eliminates the need for expensive marker development in many studies by using whole genome shotgun sequence data directly. SISRS is open source and freely available at https://github.com/rachelss/SISRS/releases.

Pre-Columbian tuberculosis in Tierra del Fuego? Discussion of the paleopathological and molecular evidence

This work contributes to ongoing discussions about the nature of tuberculosis in the Western Hemisphere prior to the time of European contact. Our example, from the extreme south of South America was, at the time of our study, without firm temporal association or molecular characterization. In Tierra del Fuego, Constantinescu (1999) briefly described vertebral bone lesions compatible with TB in an undated skeleton from Myren 1 site (Chile). The remains of Myren are estimated to represent a man between 18 and 23 years old at the time of death. The objectives of this research are to extend this description, to present molecular results, to establish a radiocarbon date, and to report stable isotopic values for the remains. We provide further description of the remains, including tuberculosis-like skeletal pathology. Radiocarbon dating of 640±20 years BP attributes this individual to the precontact fourteenth-fifteenth centuries. Isotopic ratios for nitrogen and carbon from bone collagen suggest a mixed diet. Molecular results were positive for the rpoB quantitative PCR (qPCR) assays but negative for two independent IS6110 and IS1081 qPCR assays. Further testing using genomic methods to target any mycobacteria for specific identification are needed.