Ancient Mycobacterium leprae genome reveals medieval English red squirrels as animal leprosy host.

Leprosy, one of the oldest recorded diseases in human history, remains prevalent in Asia, Africa, and South America, with over 200,000 cases every year.1,2 Although ancient DNA (aDNA) approaches on the major causative agent, Mycobacterium leprae, have elucidated the disease's evolutionary history,3,4,5 the role of animal hosts and interspecies transmission in the past remains unexplored. Research has uncovered relationships between medieval strains isolated from archaeological human remains and modern animal hosts such as the red squirrel in England.6,7 However, the time frame, distribution, and direction of transmissions remains unknown. Here, we studied 25 human and 12 squirrel samples from two archaeological sites in Winchester, a medieval English city well known for its leprosarium and connections to the fur trade. We reconstructed four medieval M. leprae genomes, including one from a red squirrel, at a 2.2-fold average coverage. Our analysis revealed a phylogenetic placement of all strains on branch 3 as well as a close relationship between the squirrel strain and one newly reconstructed medieval human strain. In particular, the medieval squirrel strain is more closely related to some medieval human strains from Winchester than to modern red squirrel strains from England, indicating a yet-undetected circulation of M. leprae in non-human hosts in the Middle Ages. Our study represents the first One Health approach for M. leprae in archaeology, which is centered around a medieval animal host strain, and highlights the future capability of such approaches to understand the disease's zoonotic past and current potential.

Investigation of a Medieval Pilgrim Burial Excavated from the Leprosarium of St Mary Magdalen Winchester, UK.

We have examined the remains of a Pilgrim burial from St Mary Magdalen, Winchester. The individual was a young adult male, aged around 18-25 years at the time of death. Radiocarbon dating showed the remains dated to the late 11th-early 12th centuries, a time when pilgrimages were at their height in Europe. Several lines of evidence in connection with the burial suggested this was an individual of some means and prestige. Although buried within the leprosarium cemetery, the skeleton showed only minimal skeletal evidence for leprosy, which was confined to the bones of the feet and legs. Nonetheless, molecular testing of several skeletal elements, including uninvolved bones all showed robust evidence of DNA from Mycobacterium leprae, consistent with the lepromatous or multibacillary form of the disease. We infer that in life, this individual almost certainly suffered with multiple soft tissue lesions. Genotyping of the M.leprae strain showed this belonged to the 2F lineage, today associated with cases from South-Central and Western Asia. During osteological examination it was noted that the cranium and facial features displayed atypical morphology for northern European populations. Subsequently, geochemical isotopic analyses carried out on tooth enamel indicated that this individual was indeed not local to the Winchester region, although it was not possible to be more specific about their geographic origin.

Mycobacterium leprae genomes from a British medieval leprosy hospital: towards understanding an ancient epidemic.

BACKGROUND: Leprosy has afflicted humankind throughout history leaving evidence in both early texts and the archaeological record. In Britain, leprosy was widespread throughout the Middle Ages until its gradual and unexplained decline between the 14th and 16th centuries. The nature of this ancient endemic leprosy and its relationship to modern strains is only partly understood. Modern leprosy strains are currently divided into 5 phylogenetic groups, types 0 to 4, each with strong geographical links. Until recently, European strains, both ancient and modern, were thought to be exclusively type 3 strains. However, evidence for type 2 strains, a group normally associated with Central Asia and the Middle East, has recently been found in archaeological samples in Scandinavia and from two skeletons from the medieval leprosy hospital (or leprosarium) of St Mary Magdalen, near Winchester, England. RESULTS: Here we report the genotypic analysis and whole genome sequencing of two further ancient M. leprae genomes extracted from the remains of two individuals, Sk14 and Sk27, that were excavated from 10th-12th century burials at the leprosarium of St Mary Magdalen. DNA was extracted from the surfaces of bones showing osteological signs of leprosy. Known M. leprae polymorphisms were PCR amplified and Sanger sequenced, while draft genomes were generated by enriching for M. leprae DNA, and Illumina sequencing. SNP-typing and phylogenetic analysis of the draft genomes placed both of these ancient strains in the conserved type 2 group, with very few novel SNPs compared to other ancient or modern strains. CONCLUSIONS: The genomes of the two newly sequenced M. leprae strains group firmly with other type 2F strains. Moreover, the M. leprae strain most closely related to one of the strains, Sk14, in the worldwide phylogeny is a contemporaneous ancient St Magdalen skeleton, vividly illustrating the epidemic and clonal nature of leprosy at this site. The prevalence of these type 2 strains indicates that type 2F strains, in contrast to later European and associated North American type 3 isolates, may have been the co-dominant or even the predominant genotype at this location during the 11th century.

Genome-wide comparison of medieval and modern Mycobacterium leprae.

Leprosy was endemic in Europe until the Middle Ages. Using DNA array capture, we have obtained genome sequences of Mycobacterium leprae from skeletons of five medieval leprosy cases from the United Kingdom, Sweden, and Denmark. In one case, the DNA was so well preserved that full de novo assembly of the ancient bacterial genome could be achieved through shotgun sequencing alone. The ancient M. leprae sequences were compared with those of 11 modern strains, representing diverse genotypes and geographic origins. The comparisons revealed remarkable genomic conservation during the past 1000 years, a European origin for leprosy in the Americas, and the presence of an M. leprae genotype in medieval Europe now commonly associated with the Middle East. The exceptional preservation of M. leprae biomarkers, both DNA and mycolic acids, in ancient skeletons has major implications for palaeomicrobiology and human pathogen evolution.

Detection and strain typing of ancient Mycobacterium leprae from a medieval leprosy hospital.

Nine burials excavated from the Magdalen Hill Archaeological Research Project (MHARP) in Winchester, UK, showing skeletal signs of lepromatous leprosy (LL) have been studied using a multidisciplinary approach including osteological, geochemical and biomolecular techniques. DNA from Mycobacterium leprae was amplified from all nine skeletons but not from control skeletons devoid of indicative pathology. In several specimens we corroborated the identification of M. leprae with detection of mycolic acids specific to the cell wall of M. leprae and persistent in the skeletal samples. In five cases, the preservation of the material allowed detailed genotyping using single-nucleotide polymorphism (SNP) and multiple locus variable number tandem repeat analysis (MLVA). Three of the five cases proved to be infected with SNP type 3I-1, ancestral to contemporary M. leprae isolates found in southern states of America and likely carried by European migrants. From the remaining two burials we identified, for the first time in the British Isles, the occurrence of SNP type 2F. Stable isotope analysis conducted on tooth enamel taken from two of the type 3I-1 and one of the type 2F remains revealed that all three individuals had probably spent their formative years in the Winchester area. Previously, type 2F has been implicated as the precursor strain that migrated from the Middle East to India and South-East Asia, subsequently evolving to type 1 strains. Thus we show that type 2F had also spread westwards to Britain by the early medieval period.

A contextual study of the medieval hospital and cemetery of St Mary Magdalen, Winchester, England.

This paper examines the osteoarchaeological evidence for leprosy in 38 skeletons excavated from the north cemetery of the hospital of St Mary Magdalen, Winchester (founded by the late 11th century) between 2009 and 2011. This cemetery, to the north of the medieval chapel, represents a discrete burial area, separate from the main, more recent cemetery to the south. The analysis indicates skeletal evidence for leprosy in over 85% (33) of the burials. This is therefore a much larger percentage than has previously been recorded for British material. The skeletal remains also provide evidence for amputation, possible palliative care as well as a pilgrim burial. Overall work at Winchester represents the most extensive excavation of an early leper hospital with accompanying cemetery to date, providing a unique opportunity for the cross-examination of skeletal and contextual data. Therefore the St Mary Magdalen cemetery is discussed in reference to such issues as the status of leper hospitals and social perceptions of hospital inmates in the medieval period.