The cuticular wax composition and crystal coverage of leaves and petals differ in a consistent manner between plant species.

Both leaves and petals are covered in a cuticle, which itself contains and is covered by cuticular waxes. The waxes perform various roles in plants' lives, and the cuticular composition of leaves has received much attention. To date, the cuticular composition of petals has been largely ignored. Being the outermost boundary between the plant and the environment, the cuticle is the first point of contact between a flower and a pollinator, yet we know little about how plant-pollinator interactions shape its chemical composition. Here, we investigate the general structure and composition of floral cuticular waxes by analysing the cuticular composition of leaves and petals of 49 plant species, representing 19 orders and 27 families. We show that the flowers of plants from across the phylogenetic range are nearly devoid of wax crystals and that the total wax load of leaves in 90% of the species is higher than that of petals. The proportion of alkanes is higher, and the chain lengths of the aliphatic compounds are shorter in petals than in leaves. We argue these differences are a result of adaptation to the different roles leaves and petals play in plant biology.

Increasing concentration of pure micro- and macro-LDPE and PP plastic negatively affect crop biomass, nutrient cycling, and microbial biomass.

Over the last 50 years, the intense use of agricultural plastic in the form of mulch films has led to an accumulation of plastic in soil, creating a legacy of plastic in agricultural fields. Plastic often contains additives, however it is still largely unknown how these compounds affect soil properties, potentially influencing or masking effects of the plastic itself. Therefore, the aim of this study was to investigate the effects of pure plastics of varying sizes and concentrations, to improve our understanding of plastic-only interactions within soil-plant mesocosms. Maize (Zea mays L.) was grown over eight weeks following the addition of micro and macro low-density polyethylene and polypropylene at increasing concentrations (equivalent to 1, 10, 25, and 50 years mulch film use) and the effects of plastic on key soil and plant properties were measured. We found the effect of both macro and microplastic on soil and plant health is negligible in the short-term (1 to <10 years). However, ≥ 10 years of plastic application for both plastic types and sizes resulted in a clear negative effect on plant growth and microbial biomass. This study provides vital insight into the effect of both macro and microplastics on soil and plant properties.

Volatile organic compounds (VOCs) as a rapid means for assessing the source of coprolites.

The odor of rehydrated coprolites can be used as an informal means of fecal identification. To date, the analysis of volatiles emitted by coprolites from different sources has not been attempted, and the possibility of utilizing volatile organic compounds (VOCs) as fecal biomarkers unexplored. VOCs released by coprolites from the Paisley Caves, were analyzed using solid-phase microextraction (SPME), to assess the variance of results from different coprolites (carnivores, herbivores, or humans). Coprolites from carnivores can be clearly distinguished from those produced by herbivores and humans; these latter two are separated to a lesser degree. Eight discriminatory compounds differentiated between the coprolite sources, and their identities were verified using reference standards. Coprolites and their associated sediments could not be differentiated between using this method, suggesting leaching of VOCs into the burial matrix. This work provides an alternative, more rapid way to assess coprolite origin.

Mechanisms of nitrogen transfer in a model clover-ryegrass pasture: a 15N-tracer approach.

Purpose: Nitrogen (N) transfer from white clover (Trifolium repens cv.) to ryegrass (Lolium perenne cv.) has the potential to meet ryegrass N requirements. This study aimed to quantify N transfer in a mixed pasture and investigate the influence of the microbial community and land management on N transfer. Methods: Split root 15N-labelling of clover quantified N transfer to ryegrass via exudation, microbial assimilation, decomposition, defoliation and soil biota. Incorporation into the microbial protein pool was determined using compound-specific 15N-stable isotope probing approaches. Results: N transfer to ryegrass and soil microbial protein in the model system was relatively small, with one-third arising from root exudation. N transfer to ryegrass increased with no microbial competition but soil microbes also increased N transfer via shoot decomposition. Addition of mycorrhizal fungi did not alter N transfer, due to the source-sink nature of this pathway, whilst weevil grazing on roots decreased microbial N transfer. N transfer was bidirectional, and comparable on a short-term scale. Conclusions: N transfer was low in a model young pasture established from soil from a permanent grassland with long-term N fertilisation. Root exudation and decomposition were major N transfer pathways. N transfer was influenced by soil biota (weevils, mycorrhizae) and land management (e.g. grazing). Previous land management and the role of the microbial community in N transfer must be considered when determining the potential for N transfer to ryegrass. Supplementary Information: The online version contains supplementary material available at 10.1007/s11104-022-05585-0.

Conserved signalling components coordinate epidermal patterning and cuticle deposition in barley.

Faced with terrestrial threats, land plants seal their aerial surfaces with a lipid-rich cuticle. To breathe, plants interrupt their cuticles with adjustable epidermal pores, called stomata, that regulate gas exchange, and develop other specialised epidermal cells such as defensive hairs. Mechanisms coordinating epidermal features remain poorly understood. Addressing this, we studied two loci whose allelic variation causes both cuticular wax-deficiency and misarranged stomata in barley, identifying the underlying genes, Cer-g/ HvYDA1, encoding a YODA-like (YDA) MAPKKK, and Cer-s/ HvBRX-Solo, encoding a single BREVIS-RADIX (BRX) domain protein. Both genes control cuticular integrity, the spacing and identity of epidermal cells, and barley's distinctive epicuticular wax blooms, as well as stomatal patterning in elevated CO2 conditions. Genetic analyses revealed epistatic and modifying relationships between HvYDA1 and HvBRX-Solo, intimating that their products participate in interacting pathway(s) linking epidermal patterning with cuticular properties in barley. This may represent a mechanism for coordinating multiple adaptive features of the land plant epidermis in a cultivated cereal.

Intestinal parasites in the Neolithic population who built Stonehenge (Durrington Walls, 2500 BCE).

Durrington Walls was a large Neolithic settlement in Britain dating around 2500 BCE, located very close to Stonehenge and likely to be the campsite where its builders lived during its main stage of construction. Nineteen coprolites recovered from a midden and associated pits at Durrington Walls were analysed for intestinal parasite eggs using digital light microscopy. Five (26%) contained helminth eggs, 1 with those of fish tapeworm (likely Dibothriocephalus dendriticus) and 4 with those of capillariid nematodes. Analyses of bile acid and sterol from these 5 coprolites show 1 to be of likely human origin and the other 4 to likely derive from dogs. The presence of fish tapeworm reveals that the Neolithic people who gathered to feast at Durrington Walls were at risk of infection from eating raw or undercooked freshwater fish. When the eggs of capillariids are found in the feces of humans or dogs it normally indicates that the internal organs (liver, lung or intestines) of animals with capillariasis have been eaten, and eggs passed through the gut without causing disease. Their presence in multiple coprolites provides new evidence that internal organs of animals were consumed. These novel findings improve our understanding of both parasitic infection and dietary habits associated with this key Neolithic ceremonial site.

Further Biochemical Profiling of Hypholoma fasciculare Metabolome Reveals Its Chemogenetic Diversity.

Natural products with novel chemistry are urgently needed to battle the continued increase in microbial drug resistance. Mushroom-forming fungi are underutilized as a source of novel antibiotics in the literature due to their challenging culture preparation and genetic intractability. However, modern fungal molecular and synthetic biology tools have renewed interest in exploring mushroom fungi for novel therapeutic agents. The aims of this study were to investigate the secondary metabolites of nine basidiomycetes, screen their biological and chemical properties, and then investigate the genetic pathways associated with their production. Of the nine fungi selected, Hypholoma fasciculare was revealed to be a highly active antagonistic species, with antimicrobial activity against three different microorganisms: Bacillus subtilis, Escherichia coli, and Saccharomyces cerevisiae. Genomic comparisons and chromatographic studies were employed to characterize more than 15 biosynthetic gene clusters and resulted in the identification of 3,5-dichloromethoxy benzoic acid as a potential antibacterial compound. The biosynthetic gene cluster for this product is also predicted. This study reinforces the potential of mushroom-forming fungi as an underexplored reservoir of bioactive natural products. Access to genomic data, and chemical-based frameworks, will assist the development and application of novel molecules with applications in both the pharmaceutical and agrochemical industries.

Determination of the δ2 H values of high molecular weight lipids by high-temperature gas chromatography coupled to isotope ratio mass spectrometry.

RATIONALE: The hydrogen isotopic composition of lipids (δ2 Hlipid ) is widely used in food science and as a proxy for past hydrological conditions. Determining the δ2 H values of large, well-preserved triacylglycerides and other microbial lipids, such as glycerol dialkyl glycerol tetraether (GDGT) lipids, is thus of widespread interest but has so far not been possible due to their low volatility which prohibits analysis by traditional gas chromatography/pyrolysis/isotope ratio mass spectrometry (GC/P/IRMS). METHODS: We determined the δ2 H values of large, polar molecules and applied high-temperature gas chromatography (HTGC) methods on a modified GC/P/IRMS system. The system used a high-temperature 7-m GC column, and a glass Y-splitter for low thermal mass. Methods were validated using authentic standards of large, functionalised molecules (triacylglycerides, TGs), purified standards of GDGTs. The results were compared with δ2 H values determined by high-temperature elemental analyser/pyrolysis/isotope ratio mass spectrometry (HTEA/P/IRMS), and subsequently applied to the analysis of GDGTs in a sample from a methane seep and a Welsh peat. RESULTS: The δ2 H values of TGs agreed within error between HTGC/P/IRMS and HTEA/IRMS, with HTGC/P/IRMS showing larger errors. Archaeal lipid GDGTs with up to three cyclisations could be analysed: the δ2 H values were not significantly different between methods with standard deviations of 5 to 6 ‰. When environmental samples were analysed, the δ2 H values of isoGDGTs were 50 ‰ more negative than those of terrestrial brGDGTs. CONCLUSIONS: Our results indicate that the HTGC/P/IRMS method developed here is appropriate to determine the δ2 H values of TGs, GDGTs with up to two cyclisations, and potentially other high molecular weight compounds. The methodology will widen the current analytical window for biomarker and food light stable isotope analyses. Moreover, our initial measurements suggest that bacterial and archaeal GDGT δ2 H values can record environmental and ecological conditions.

Pre-Clovis occupation of the Americas identified by human fecal biomarkers in coprolites from Paisley Caves, Oregon.

When and how people first settled in the Americas is an ongoing area of research and debate. The earliest sites typically only contain lithic artifacts that cannot be directly dated. The lack of human skeletal remains in these early contexts means that alternative sources of evidence are needed. Coprolites, and the DNA contained within them, are one such source, but unresolved issues concerning ancient DNA taphonomy and potential for contamination make this approach problematic. Here, we use fecal lipid biomarkers to demonstrate unequivocally that three coprolites dated to pre-Clovis are human, raise questions over the reliance on DNA methods, and present a new radiocarbon date on basketry further supporting pre-Clovis human occupation.

Temperature Driven Membrane Lipid Adaptation in Glacial Psychrophilic Bacteria.

Bacteria inhabiting non-polar glaciers are exposed to large variations in temperature, which significantly affects the fluidity of bacterial cell membranes. In order to maintain normal functions of the cell membranes, psychrophilic bacteria adapt by changing the composition of cell membrane fatty acids. However, information on the exact pattern of cell membrane adaptability in non-polar low-temperature habitats is scarce. In the present study, 42 bacterial strains were isolated from the Ghulmet, Ghulkin, and Hopar glaciers of the Hunza Valley in the Karakoram Mountain Range, Pakistan and their cell membrane fatty acid distributions studied, using gas chromatography/mass spectrometry (GC-MS) for the analysis of fatty acid methyl esters (FAMEs) liberated by acid-catalyzed methanolysis. Furthermore, Gram-negative and Gram-positive groups were grown under different temperature settings (5, 15, 25, and 35°C) in order to determine the effect of temperature on cell membrane (CM) fatty acid distribution. The analyses identified the major groups of cell membrane fatty acids (FA) as straight-chain monounsaturated fatty acids (n-MUFAs) and branched fatty acids (br-FAs), accounting for more than 70% of the fatty acids analyzed. The distribution of br-FAs and n-FAs in bacterial cell membranes was significantly affected by temperature, with the level of br-FAs decreasing relative to n-FAs with increasing temperature. Notably, the production of polyunsaturated fatty acids (PUFAs) was only seen at lower temperatures. This study contributes to understanding, for the first time, the role of br-FAs in the maintenance of cell membrane fluidity of bacteria inhabiting non-polar habitats.

Birch bark tar in early Medieval England - Continuity of tradition or technological revival?

Birch bark tar is a manufactured product with a history of production and use that reaches back to the Palaeolithic. Its sticky, water resistant and biocidal properties mean that it has a wide range of applications, for example, as a multipurpose adhesive, sealant and in medicine. Archaeological evidence for birch bark tar in the old world covers a broad geographic range from the UK to the Baltic and from the Mediterranean to Scandinavia. In the east and north of this range there is continuity of use to modern times but in western Europe and the British Isles the use of birch bark tar has generally been viewed as limited to prehistory, with gradual displacement by pine tars during the Roman period. Here, we report new finds of birch bark tar from two early Medieval sites in the east of England. Analysis by HT-GC/MS to identify the tars also revealed fatty material, possibly added to modify the tar. The different contexts of the finds point to diverse applications of the material: in one case perhaps a medicine, the other associated with a ceramic container, possibly used for processing the tar. The results present the first identification of birch bark tar from early Medieval archaeological contexts in the UK. Together they indicate a later period of use for birch bark tar in the UK than has been previously observed and raise the question of whether this indicates evidence of a longer continuity of use than hitherto recognised or a later reintroduction of the technology in the Medieval period, in which case the similarities between the find sites, both early Anglo-Saxon cemeteries with comparable assemblages of grave goods, may be significant.

Cell membrane fatty acid and pigment composition of the psychrotolerant cyanobacterium Nodularia spumigena CHS1 isolated from Hopar glacier, Pakistan.

In the present study, cyanobacterium isolate CHS1 isolated from Hopar glacier, Pakistan, was analyzed for the first time for cell membrane fatty acids and production of pigments. Sequencing of the 16-23S intergenetic region confirmed identification of the isolate CHS1 as Nodularia spumigena. All chlorophyll and carotenoid pigments were quantified using high-performance liquid chromatography and experiments to test tolerance against a range of physico-chemical conditions were conducted. Likewise, the fatty acid profile of the cell membrane CHS1 was analyzed using gas chromatography and mass spectroscopy. The cyanobacterium isolate CHS1 demonstrated tolerance to 8 g/L% NaCl, 35°C and pH 5-9. The characteristic polyunsaturated fatty acid (PUFA) of isolate CHS1, C18:4, was observed in fatty acid methyl esters (FAMEs) extracted from the cell membrane. CHS1 was capable of producing saturated fatty acids (SFA) (e.g., C16:0), monounsaturated fatty acids (MUFA) (e.g., C18:1) and polyunsaturated fatty acids (e.g., C20:5) in the cell membrane. In this study, we hypothesize that one mechanism of cold adaptation displayed by isolate CHS1 is the accumulation of high amounts of PUFA in the cell membrane.

Intestinal parasites at the Late Bronze Age settlement of Must Farm, in the fens of East Anglia, UK (9th century B.C.E.).

Little is known about the types of intestinal parasites that infected people living in prehistoric Britain. The Late Bronze Age archaeological site of Must Farm was a pile-dwelling settlement located in a wetland, consisting of stilted timber structures constructed over a slow-moving freshwater channel. At excavation, sediment samples were collected from occupation deposits around the timber structures. Fifteen coprolites were also hand-recovered from the occupation deposits; four were identified as human and seven as canine, using fecal lipid biomarkers. Digital light microscopy was used to identify preserved helminth eggs in the sediment and coprolites. Eggs of fish tapeworm (Diphyllobothrium latum and Diphyllobothrium dendriticum), Echinostoma sp., giant kidney worm (Dioctophyma renale), probable pig whipworm (Trichuris suis) and Capillaria sp. were found. This is the earliest evidence for fish tapeworm, Echinostoma worm, Capillaria worm and the giant kidney worm so far identified in Britain. It appears that the wetland environment of the settlement contributed to establishing parasite diversity and put the inhabitants at risk of infection by helminth species spread by eating raw fish, frogs or molluscs that flourish in freshwater aquatic environments, conversely the wetland may also have protected them from infection by certain geohelminths.

Cretaceous dinosaur bone contains recent organic material and provides an environment conducive to microbial communities.

Fossils were thought to lack original organic molecules, but chemical analyses show that some can survive. Dinosaur bone has been proposed to preserve collagen, osteocytes, and blood vessels. However, proteins and labile lipids are diagenetically unstable, and bone is a porous open system, allowing microbial/molecular flux. These 'soft tissues' have been reinterpreted as biofilms. Organic preservation versus contamination of dinosaur bone was examined by freshly excavating, with aseptic protocols, fossils and sedimentary matrix, and chemically/biologically analyzing them. Fossil 'soft tissues' differed from collagen chemically and structurally; while degradation would be expected, the patterns observed did not support this. 16S rRNA amplicon sequencing revealed that dinosaur bone hosted an abundant microbial community different from lesser abundant communities of surrounding sediment. Subsurface dinosaur bone is a relatively fertile habitat, attracting microbes that likely utilize inorganic nutrients and complicate identification of original organic material. There exists potential post-burial taphonomic roles for subsurface microorganisms.

The chances of establishing a real-world Jurassic Park are slim. During the fossilization process, biological tissues degrade over millions of years, with some types of molecules breaking down faster than others. However, traces of biological material have been found inside some fossils. While some researchers believe these could be the remains of ancient proteins, blood vessels, and cells, traditionally thought to be among the least stable components of bone, others think that they have more recent sources. One hypothesis is that they are in fact biofilms formed by bacteria. To investigate the source of the biological material in fossil bone, Saitta et al. performed a range of analyses on the fossilized bones of a horned dinosaur called Centrosaurus. The bones were carefully excavated in a manner to reduce contamination, and the sediment the bones had been embedded in was also tested for comparison. Saitta et al. found no evidence of ancient dinosaur proteins. However, the fossils contained more organic carbon, DNA, and certain amino acids than the sediment surrounding them. Most of these appeared to have a very recent source. Sequencing the genetic material revealed that the fossil had become a habitat for an unusual community of microbes that is not found in the surrounding sediment or above ground. These buried microbes may have evolved unique ways to thrive inside fossils. Future work could investigate how these unusual organisms live and whether the communities vary in different parts of the world.

Lipid profiling and analytical discrimination of seven cereals using high temperature gas chromatography coupled to high resolution quadrupole time-of-flight mass spectrometry.

Minor lipids in cereals (such as phytosterols and alkylresorcinols) can be important for human nutrition and/or be used as biomarkers for cereal intake. However, the analysis of cereal lipids is very challenging due to the complex lipidome comprising several hundred individual compounds present over a wide range of concentrations. Here we present a method for the profiling of cereal lipids using high temperature gas chromatography coupled to high resolution mass spectrometry (GC/Q-TOF MS). The method was used to investigate the lipid profiles of 77 samples of bread wheat, spelt, einkorn, emmer, barley, rye and oats. Distinct differences in the patterns of alkylresorcinols, free and conjugated sterols and tocopherols between the cereals could be observed. Furthermore, traces of tocomonoenols and diunsaturated and methyl-alkylresorcinols (not previously reported in cereals) could be detected. Finally, the lipid patterns in the cereals could be used to separate the cereals by Principal Component Analysis.

Saccharomyces cerevisiae Atf1p is an alcohol acetyltransferase and a thioesterase in vitro.

The alcohol-O-acyltransferases are bisubstrate enzymes that catalyse the transfer of acyl chains from an acyl-coenzyme A (CoA) donor to an acceptor alcohol. In the industrial yeast Saccharomyces cerevisiae this reaction produces acyl esters that are an important influence on the flavour of fermented beverages and foods. There is also a growing interest in using acyltransferases to produce bulk quantities of acyl esters in engineered microbial cell factories. However, the structure and function of the alcohol-O-acyltransferases remain only partly understood. Here, we recombinantly express, purify and characterize Atf1p, the major alcohol acetyltransferase from S. cerevisiae. We find that Atf1p is promiscuous with regard to the alcohol cosubstrate but that the acyltransfer activity is specific for acetyl-CoA. Additionally, we find that Atf1p is an efficient thioesterase in vitro with specificity towards medium-chain-length acyl-CoAs. Unexpectedly, we also find that mutating the supposed catalytic histidine (H191) within the conserved HXXXDG active site motif only moderately reduces the thioesterase activity of Atf1p. Our results imply a role for Atf1p in CoA homeostasis and suggest that engineering Atf1p to reduce the thioesterase activity could improve product yields of acetate esters from cellular factories. © 2017 The Authors. Yeast published by John Wiley & Sons, Ltd.

Fossilization of melanosomes via sulfurization.

Fossil melanin granules (melanosomes) are an important resource for inferring the evolutionary history of colour and its functions in animals. The taphonomy of melanin and melanosomes, however, is incompletely understood. In particular, the chemical processes responsible for melanosome preservation have not been investigated. As a result, the origins of sulfur-bearing compounds in fossil melanosomes are difficult to resolve. This has implications for interpretations of original colour in fossils based on potential sulfur-rich phaeomelanosomes. Here we use pyrolysis gas chromatography mass spectrometry (Py-GCMS), fourier transform infrared spectroscopy (FTIR) and time of flight secondary ion mass spectrometry (ToF-SIMS) to assess the mode of preservation of fossil microstructures, confirmed as melanosomes based on the presence of melanin, preserved in frogs from the Late Miocene Libros biota (NE Spain). Our results reveal a high abundance of organosulfur compounds and non-sulfurized fatty acid methyl esters in both the fossil tissues and host sediment; chemical signatures in the fossil tissues are inconsistent with preservation of phaeomelanin. Our results reflect preservation via the diagenetic incorporation of sulfur, i.e. sulfurization (natural vulcanization), and other polymerization processes. Organosulfur compounds and/or elevated concentrations of sulfur have been reported from melanosomes preserved in various invertebrate and vertebrate fossils and depositional settings, suggesting that preservation through sulfurization is likely to be widespread. Future studies of sulfur-rich fossil melanosomes require that the geochemistry of the host sediment is tested for evidence of sulfurization in order to constrain interpretations of potential phaeomelanosomes and thus of original integumentary colour in fossils.

7000 year-old tuberculosis cases from Hungary - Osteological and biomolecular evidence.

This study derives from the macroscopic analysis of a Late Neolithic population from Hungary. Remains were recovered from a tell settlement at Hódmezovásárhely-Gorzsa from graves within the settlement as well as pits, ditches, houses and as stray finds. One of the most important discoveries from these remains was evidence of tuberculosis. Pathological analysis of the seventy-one individuals revealed numerous cases of infections and non-specific stress indicators on juveniles and adults, metabolic diseases on juveniles, and evidence of trauma and mechanical changes on adults. Several cases showed potential signs of tuberculosis and further analyses were undertaken, including biomolecular studies. The five individuals were all very young adults and included a striking case of Hypertrophic Pulmonary Osteopathy (HPO) with rib changes, one case with resorptive lesions on the vertebrae, two cases with hypervascularisation on the vertebrae and periosteal remodelling on the ribs, and one case with abnormal blood vessel impressions and a possible lesion on the endocranial surface of the skull. The initial macroscopic diagnosis of these five cases was confirmed by lipid biomarker analyses, and three of them were corroborated by DNA analysis. At present, these 7000-year-old individuals are among the oldest palaeopathological and palaeomicrobiological cases of tuberculosis worldwide.

Morphological and biomolecular evidence for tuberculosis in 8th century AD skeletons from Bélmegyer-Csömöki domb, Hungary.

Macromorphological analysis of skeletons, from 20 selected graves of the 8th century AD Bélmegyer-Csömöki domb, revealed 19 cases of possible skeletal tuberculosis. Biomolecular analyses provided general support for such diagnoses, including the individual without pathology, but the data did not show coherent consistency over the range of biomarkers examined. Amplification of ancient DNA fragments found evidence for the Mycobacterium tuberculosis complex DNA only in five graves. In contrast, varying degrees of lipid biomarker presence were recorded in all except two of the skeletons, though most lipid components appeared to be somewhat degraded. Mycobacterial mycolic acid biomarkers were absent in five cases, but the weak, possibly degraded profiles for the remainder were smaller and inconclusive for either tuberculosis or leprosy. The most positive lipid biomarker evidence for tuberculosis was provided by mycolipenic acid, with 13 clear cases, supported by five distinct possible cases. Combinations of mycocerosic acids were present in all but three graves, but in one case a tuberculosis-leprosy co-infection was indicated. In two specimens with pathology, no lipid biomarker evidence was recorded, but one of these specimens provided M. tuberculosis complex DNA fragments.

A migration-driven model for the historical spread of leprosy in medieval Eastern and Central Europe.

Leprosy was rare in Europe during the Roman period, yet its prevalence increased dramatically in medieval times. We examined human remains, with paleopathological lesions indicative of leprosy, dated to the 6th-11th century AD, from Central and Eastern Europe and Byzantine Anatolia. Analysis of ancient DNA and bacterial cell wall lipid biomarkers revealed Mycobacterium leprae in skeletal remains from 6th-8th century Northern Italy, 7th-11th century Hungary, 8th-9th century Austria, the Slavic Greater Moravian Empire of the 9th-10th century and 8th-10th century Byzantine samples from Northern Anatolia. These data were analyzed alongside findings published by others. M. leprae is an obligate human pathogen that has undergone an evolutionary bottleneck followed by clonal expansion. Therefore M. leprae genotypes and sub-genotypes give information about the human populations they have infected and their migration. Although data are limited, genotyping demonstrates that historical M. leprae from Byzantine Anatolia, Eastern and Central Europe resembles modern strains in Asia Minor rather than the recently characterized historical strains from North West Europe. The westward migration of peoples from Central Asia in the first millennium may have introduced different M. leprae strains into medieval Europe and certainly would have facilitated the spread of any existing leprosy. The subsequent decline of M. leprae in Europe may be due to increased host resistance. However, molecular evidence of historical leprosy and tuberculosis co-infections suggests that death from tuberculosis in leprosy patients was also a factor.