A 14th century CE Brucella melitensis genome and the recent expansion of the Western Mediterranean clade.

Brucellosis is a disease caused by the bacterium Brucella and typically transmitted through contact with infected ruminants. It is one of the most common chronic zoonotic diseases and of particular interest to public health agencies. Despite its well-known transmission history and characteristic symptoms, we lack a more complete understanding of the evolutionary history of its best-known species-Brucella melitensis. To address this knowledge gap we fortuitously found, sequenced and assembled a high-quality ancient B. melitensis draft genome from the kidney stone of a 14th-century Italian friar. The ancient strain contained fewer core genes than modern B. melitensis isolates, carried a complete complement of virulence genes, and did not contain any indication of significant antimicrobial resistances. The ancient B. melitensis genome fell as a basal sister lineage to a subgroup of B. melitensis strains within the Western Mediterranean phylogenetic group, with a short branch length indicative of its earlier sampling time, along with a similar gene content. By calibrating the molecular clock we suggest that the speciation event between B. melitensis and B. abortus is contemporaneous with the estimated time frame for the domestication of both sheep and goats. These results confirm the existence of the Western Mediterranean clade as a separate group in the 14th CE and suggest that its divergence was due to human and ruminant co-migration.

Examining pathogen DNA recovery across the remains of a 14th century Italian friar (Blessed Sante) infected with Brucella melitensis.

OBJECTIVE: To investigate variation in ancient DNA recovery of Brucella melitensis, the causative agent of brucellosis, from multiple tissues belonging to one individual MATERIALS: 14 samples were analyzed from the mummified remains of the Blessed Sante, a 14 th century Franciscan friar from central Italy, with macroscopic diagnosis of probable brucellosis. METHODS: Shotgun sequencing data from was examined to determine the presence of Brucella DNA. RESULTS: Three of the 14 samples contained authentic ancient DNA, identified as belonging to B. melitensis. A genome (23.81X depth coverage, 0.98 breadth coverage) was recovered from a kidney stone. Nine of the samples contained reads classified as B. melitensis (7-169), but for many the data quality was insufficient to withstand our identification and authentication criteria. CONCLUSIONS: We identified significant variation in the preservation and abundance of B. melitensis DNA present across multiple tissues, with calcified nodules yielding the highest number of authenticated reads. This shows how greatly sample selection can impact pathogen identification. SIGNIFICANCE: Our results demonstrate variation in the preservation and recovery of pathogen DNA across tissues. This study highlights the importance of sample selection in the reconstruction of infectious disease burden and highlights the importance of a holistic approach to identifying disease. LIMITATIONS: Study focuses on pathogen recovery in a single individual. SUGGESTIONS FOR FURTHER RESEARCH: Further analysis of how sampling impacts aDNA recovery will improve pathogen aDNA recovery and advance our understanding of disease in past peoples.

Twenty-first century bioarchaeology: Taking stock and moving forward.

This article presents outcomes from a Workshop entitled "Bioarchaeology: Taking Stock and Moving Forward," which was held at Arizona State University (ASU) on March 6-8, 2020. Funded by the National Science Foundation (NSF), the School of Human Evolution and Social Change (ASU), and the Center for Bioarchaeological Research (CBR, ASU), the Workshop's overall goal was to explore reasons why research proposals submitted by bioarchaeologists, both graduate students and established scholars, fared disproportionately poorly within recent NSF Anthropology Program competitions and to offer advice for increasing success. Therefore, this Workshop comprised 43 international scholars and four advanced graduate students with a history of successful grant acquisition, primarily from the United States. Ultimately, we focused on two related aims: (1) best practices for improving research designs and training and (2) evaluating topics of contemporary significance that reverberate through history and beyond as promising trajectories for bioarchaeological research. Among the former were contextual grounding, research question/hypothesis generation, statistical procedures appropriate for small samples and mixed qualitative/quantitative data, the salience of Bayesian methods, and training program content. Topical foci included ethics, social inequality, identity (including intersectionality), climate change, migration, violence, epidemic disease, adaptability/plasticity, the osteological paradox, and the developmental origins of health and disease. Given the profound changes required globally to address decolonization in the 21st century, this concern also entered many formal and informal discussions.

Dataset of oxygen, carbon, and strontium isotope values from the Imperial Roman site of Velia (ca. 1st-2nd c. CE), Italy.

The oxygen (δ18Ocarbonate), strontium (87Sr/86Sr), and previously unpublished carbon (δ13Ccarbonate) isotope data presented herein from the Imperial Roman site of Velia (ca. 1st to 2nd c. CE) were obtained from the dental enamel of human permanent second molars (M2). In total, the permanent M2s of 20 individuals (10 male and 10 female) were sampled at the Museo delle Civiltà in Rome (formerly the Museo Nazionale Preistorico Etnografico "L. Pigorini") and were subsequently processed and analysed at McMaster University. A subsample of teeth (n=5) was initially subjected to Fourier transform infrared spectroscopy (FTIR) analysis to assess for diagenetic alteration through calculation of crystallinity index (CI) values. Subsequently, tooth enamel was analysed for δ13Ccarbonate and δ18Ocarbonate (VPDB) using a VG OPTIMA Isocarb isotope ratio mass spectrometer (IRMS) at McMaster Research for Stable Isotopologues (MRSI), and 87Sr/86Sr was measured by dynamic multi-collection using a thermal ionization mass spectrometer (TIMS) in the School of Geography and Earth Sciences. The dental enamel isotope data presented represent the first δ18O, δ13Ccarbonate, and 87Sr/86Sr values analysed from Imperial Roman Campania to date, providing data of use for comparative analyses of δ18O, δ13C, and 87Sr/86Sr values within the region and for assisting in documenting human mobility in archaeological contexts. Full interpretation of the δ18O and 87Sr/86Sr data presented here is provided in "Imperial Roman mobility and migration at Velia (1st to 2nd c. CE) in southern Italy" [1].

Gastrointestinal infection in Italy during the Roman Imperial and Longobard periods: A paleoparasitological analysis of sediment from skeletal remains and sewer drains.

OBJECTIVE: This study aims to investigate parasitic infection in Italy during the Roman period (27 BCE-476 CE) and subsequent Longobard (Lombard) period (6th-8th CE). MATERIALS: Sediment samples from drains and burials from Roman Imperial-period sites in Italy (Lucus Feroniae, Oplontis, Vacone, and Vagnari), Late Antique and Longobard-period burials at Selvicciola (ca. 4th-8th CE), and Longobard-period burials at Vacone and Povegliano Veronese. METHODS: Microscopy was used to identify helminth eggs and enzyme-linked immunosorbent assay (ELISA) to detect protozoan antigens. RESULTS: Roundworm and whipworm were found in pelvic sediment from Roman-period burials, while roundworm and the protozoan Giardia duodenalis were identified in Roman-period drains. In pelvic sediment from the Late Antique through Longobard periods, roundworm and Taenia tapeworm eggs were identified. CONCLUSIONS: Fecal-oral parasites were found throughout Imperial Roman Italy, suggesting that gastrointestinal infections caused a significant disease burden. In the Longobard period we see continuity in transmission of fecal-oral parasites, and the appearance of zoonotic parasites acquired from eating undercooked meat. SIGNIFICANCE: A wealth of information exists about certain diseases in the Roman period, but relatively little is known about intestinal parasites in Italy during the Roman and Longobard periods. This is the first evidence for Giardia in Roman period Italy, and for any parasites in the Longobard period in Italy. LIMITATIONS: Low egg concentrations and lack of controls for some samples makes it difficult to differentiate true infections from environmental contamination in some cases. SUGGESTIONS FOR FUTURE RESEARCH: Continual study of samples from Roman and Longobard period Italy.

Quantifying cortical bone in fragmentary archeological second metacarpals.

OBJECTIVES: Skeletal variation in cortical bone thickness is an indicator of bone quality and health in archeological populations. Second metacarpal radiogrammetry, which measures cortical thickness at the shaft midpoint, is traditionally used to evaluate bone loss in bioarcheological and some clinical contexts. However fragmentary elements are regularly omitted because the midpoint cannot be determined. This methodological limitation reduces sample sizes and biases them against individuals prone to fracture, such as older individuals with low bone mass. This study introduces a new technique for measuring cortical bone in second metacarpals, the "Region of Interest" (ROI) method, which quantifies bone in archeological remains with less-than-ideal preservation while accounting for cortical heterogeneity. MATERIALS AND METHODS: The ROI method was adapted from digital X-ray radiogrammetry (DXR), a clinical method used to estimate bone mineral density, and tested using second metacarpals from Middenbeemster, Netherlands, a 19th century known age and sex skeletal collection. The ROI method quantifies cortical bone area within a 1.9 cm-long, mid-diaphyseal region, standardized for body size differences using total area (CAIROI ). CAIROI values were compared to traditional radiogrammetric cortical indices (CI) to assess the method's ability to identify age-related bone loss. RESULTS: CAIROI values have high intra- and interobserver replicability and are strongly and significantly correlated with CI values for both males (r[n = 39] = 0.906, p = 0.000) and females (r[n = 58] = 0.925, p = 0.000). CONCLUSION: The ROI method complements traditional radiogrammetry analyses and provides a reliable way to quantify cortical bone in incomplete second metacarpals, thereby maximizing sample sizes, allowing patterns in bone acquisition and loss to be more comprehensively depicted in archeological assemblages.

Maintaining mobility after fracture: A biomechanical analysis of fracture consequences at the Roman Sites of Ancaster (UK) and Vagnari (Italy).

OBJECTIVE: This study uses biomechanical data from tibiae to investigate the functional consequences of lower limb fractures. Adults with malunited fractures are hypothesized to have experienced altered mobility, indicated by asymmetric tibial cross-sectional geometries (CSG). MATERIALS: Ninety-three adults from Roman (1st to 4th centuries CE) Ancaster, UK and Vagnari, Italy (Ancaster n = 16 adults with lower limb fracture:53 without fracture; Vagnari n = 5:19) METHODS: Biplanar radiographs were used to quantify and compare tibial CSG properties and asymmetries between individuals with and without fractures to femora, tibiae, and/or fibulae. The amount of angulation, rotation, and overlap, indicative of linear deformity, were measured for each fracture. Individuals who loaded their fractured leg differently than their opposite, uninjured leg were identified using outlying amounts of CSG asymmetry. RESULTS: Two Ancaster individuals had poorly aligned fractures. None of the Ancaster or Vagnari individuals with lower limb fractures had CSG properties or asymmetries outside the calculated normal ranges. CONCLUSIONS: Regardless of how a fracture healed, individuals at Ancaster and Vagnari generally resumed mobility after trauma whenever possible. SIGNIFICANCE: This research contributes information about injury recovery and suggests that resilient behaviors and persistent mobility may have been valued or required responses to fracture in the study communities. This work advises that impairment should not be inferred based solely on the appearance of lesions. LIMITATIONS: Site, sex, and age patterns in injury recovery are not evaluated due to sample size limitations. SUGGESTIONS FOR FURTHER RESEARCH: Biomechanical assessments of post-traumatic function in varied cultural contexts are advised in order to further characterize the impact that physical and social factors have on injury recovery.

Analysis of patterning in the occurrence of skeletal lesions used as indicators of vitamin D deficiency in subadult and adult skeletal remains.

Paleopathological investigations of conditions linked to vitamin D deficiency have increased in the last twenty years, and a suite of skeletal lesions has been established to aid in the diagnosis of vitamin D deficiency disease in subadults and adults. This paper analyzes the occurrence of these lesions in a large skeletal series comprising 3541 Roman period individuals (1st-6th century AD). Sixteen lesions reported in rickets in subadults, and 13 associated with residual rickets and osteomalacia in adults, were analyzed. Among subadults, there were clear associations among post-cranial lesions. Porotic cranial changes were associated with each other, but not with post-cranial lesions. A range of conditions could have produced the cranial lesions. There was a general paucity of correlations between indicators found in adults, and the difficulty in recording bending deformities was clear. Pseudofractures appear to provide a useful means of investigating osteomalacia in adults. In general, a simple algorithmic approach using presence or absence of lesions is unlikely to provide an adequate means of diagnosing vitamin D deficiency in paleopathology. Knowledge and consideration of the underlying physiological mechanisms involved in lesion formation, combined with individual judgement, will be required to differentially diagnose cases.

Mapping the origins of Imperial Roman workers (1st-4th century CE) at Vagnari, Southern Italy, using 87 Sr/86 Sr and δ18 O variability.

OBJECTIVES: We obtained the oxygen and strontium isotope composition of teeth from Roman period (1st to 4th century CE) inhabitants buried in the Vagnari cemetery (Southern Italy), and present the first strontium isotope variation map of the Italian peninsula using previously published data sets and new strontium data. We test the hypothesis that the Vagnari population was predominantly composed of local individuals, instead of migrants originating from abroad. MATERIALS AND METHODS: We analyzed the oxygen (18 O/16 O) and strontium (87 Sr/86 Sr) isotope composition of 43 teeth. We also report the 87 Sr/86 Sr composition of an additional 13 molars, 87 Sr/86 Sr values from fauna (n = 10), and soil (n = 5) samples local to the area around Vagnari. The 87 Sr/86 Sr variation map of Italy uses 87 Sr/86 Sr values obtained from previously published data sources from across Italy (n = 199). RESULTS: Converted tooth carbonate (δ18 ODW ) and 87 Sr/86 Sr data indicate that the majority of individuals buried at Vagnari were local to the region. ArcGIS bounded Inverse Distance Weighting (IDW) interpolation of the pan-Italian 87 Sr/86 Sr data set approximates the expected 87 Sr/86 Sr range of Italy's geological substratum, producing the first strontium map of the Italian peninsula. DISCUSSION: Results suggest that only 7% of individuals buried at Vagnari were born elsewhere and migrated to Vagnari, while the remaining individuals were either local to Vagnari (58%), or from the southern Italian peninsula (34%). Our results are consistent with the suggestion that Roman Imperial lower-class populations in southern Italy sustained their numbers through local reproduction measures, and not through large-scale immigration from outside the Italian peninsula.

Age estimation in older adults: Use of pulp/tooth ratios calculated from tooth sections.

OBJECTIVES: Accurate age estimates are foundational for bioarchaeological research, yet the ability to accurately age older adult skeletons remains elusive. This study uses a new version of pulp/tooth area calculations to investigate chronological age of older archaeological individuals. MATERIALS AND METHODS: Pulp/tooth area ratios were calculated on modern control teeth (n = 10) that were first radiographed and then sectioned for comparative analysis. Pulp/tooth area ratios were determined on sectioned teeth using ImageJ software for: (a) modern individuals of known age (n = 26); (b) individuals from Belleville, Ontario, Canada (1821-1874) with documented age (n = 50); and (c) Belleville individuals with skeletally estimated age (n = 122). RESULTS: Calculations from tooth sections on modern teeth (n = 10) resulted in a mean absolute error (MAE) of ±3.9 years, whereas the radiographic method for the same teeth had an MAE of ±14.45 years. Results indicate that sectioned pulp/tooth area ratios are a significant predictor of chronological age (p < .005), with MAEs of ±4.53 years for Belleville and ±3.77 years for modern individuals. There were no statistically significant differences in age estimations between modern and archaeological individuals, or with respect to tooth type, sex, or intra/inter-observer estimations. DISCUSSION: This study provides a new more accurate method for estimating age-at-death, particularly for individuals in the 50+ age category. Sectioning the teeth and directly measuring exposed pulp chambers results in age estimations that were within ±4.15 years for both modern and archaeological individuals, thus presenting a method that will enhance the ability to age older individuals.

Plasmodium falciparum malaria in 1st-2nd century CE southern Italy.

The historical record attests to the devastation malaria exacted on ancient civilizations, particularly the Roman Empire [1]. However, evidence for the presence of malaria during the Imperial period in Italy (1st-5th century CE) is based on indirect sources, such as historical, epigraphic, or skeletal evidence. Although these sources are crucial for revealing the context of this disease, they cannot establish the causative species of Plasmodium. Importantly, definitive evidence for the presence of malaria is now possible through the implementation of ancient DNA technology. As malaria is presumed to have been at its zenith during the Imperial period [1], we selected first or second molars from 58 adults from three cemeteries from this time: Isola Sacra (associated with Portus Romae, 1st-3rd century CE), Velia (1st-2nd century CE), and Vagnari (1st-4th century CE). We performed hybridization capture using baits designed from the mitochondrial (mtDNA) genomes of Plasmodium spp. on a prioritized subset of 11 adults (informed by metagenomic sequencing). The mtDNA sequences generated provided compelling phylogenetic evidence for the presence of P. falciparum in two individuals. This is the first genomic data directly implicating P. falciparum in Imperial period southern Italy in adults.

Investigating Residential History Using Stable Hydrogen and Oxygen Isotopes of Human Hair and Drinking Water.

The relationship between isotopic signals in human hair and geographic region has potential forensic applications for identifying unknown individuals' place of recent residence. This study analyzes δ(2) H and δ(18) O isotopes in residential tap water and bulk hair samples from 17 volunteers representing 12 locations in Ontario, Canada. There is a strong correlation (R(2) = 0.9) between δ(2) H and δ(18) O values of the water samples. In contrast, the δ(2) H and δ(18) O values of the hair samples are weakly correlated (R(2) = 0.3), and the greater variability in the data is linked to dietary factors. This study demonstrates that the δ(2) H and δ(18) O values of hair and drinking water can be used to help identify potential place of residence in forensic cases, particularly in relation to proximity to large bodies of water such as the Great Lakes, but interpretations are complicated by the contribution of both water and diet to δ(2) H and δ(18) O values in hair.

You are not what you eat during physiological stress: Isotopic evaluation of human hair.

Variation in δ(13) C and δ(15) N values can be assessed to understand not only diet, but also the influence of physiological factors on an individual. The metabolic balance of an individual can impact isotopic signals in tissues that are forming during the periods of metabolic stress. Fluctuating δ(15) N values are associated with physiological stressors that alter an individual's metabolism such as infection, injury, or pregnancy. This study examines variation in δ(13) C and δ(15) N values along sequentially segmented hair in both modern and archaeological individuals. Subjects with an observable skeletal pathology, known chronic illness, or evidence of pregnancy were compared with controls exhibiting no evidence of physiological stress. The results on hair samples from individuals from 19(th) century Belleville, Ontario, four modern cadavers (two with cancer and two sudden deaths), and two living pregnant women indicate that δ(15) N values are approximately 1‰ higher in individuals with a pathological condition (e.g., infection, fracture, or cancer) and are 1‰ lower during pregnancy, whereas δ(13) C values show less variability. Higher nitrogen values may represent the recycling of nitrogen derived from the breakdown of existing proteins in the body (catabolism), whereas lower δ(15) N values are related to increased utilization of dietary and urea nitrogen for tissue synthesis during pregnancy. These findings suggest that short-term fluctuations of δ(15) N values may be the result of changes in an individual's metabolic balance, and that metabolic imbalance poses a confounding factor to ancient dietary studies when using rapidly growing tissues such as hair.

Isotopic and dental evidence for infant and young child feeding practices in an imperial Roman skeletal sample.

This study integrates isotopic, palaeopathological, and historical evidence to investigate infant and young child feeding practices in a Roman period (1st to 3rd centuries AD) skeletal sample from the Isola Sacra necropolis (Rome, Italy). Stable isotope analysis of bone collagen from 37 rib samples indicates that transitional feeding began by the end of the first year and weaning occurred by 2-2.5 years of age. Both delta(15)N and delta(13)C data clearly show the trophic level effect associated with breastfeeding. Childhood diet is investigated using dental pathology data in the deciduous dentitions of 78 individuals aged between 1 and 12 years. The presence of calculus, caries, and tooth wear in young children suggests that individuals were provided complementary foods and other items that impacted their dental health at an early age. The isotopic and dental data are generally consistent with the historical evidence from the Roman period with respect to the general timetable of weaning and the character of complementary foods. This is the first study to integrate isotopic and deciduous dental pathology data to explore infant and young child feeding practices in the Roman world.

Isotopic evidence for age-related immigration to imperial Rome.

Oxygen stable isotope ratios (delta(18)O) have been determined in carbonate in paired first and third molar teeth from individuals (N = 61) who lived in the town of Portus Romae ("Portus") and who were buried in the necropolis of Isola Sacra (First to Third centuries AD) near Rome, Italy. We compare these analyses with data for deciduous teeth of modern Roman children. Approximately one-third of the archaeological sample has first molar (M1) values outside the modern range, implying a large rate of population turnover at that time, consistent with historical data. Delta (18)O(ap) values suggest that a group within the sample migrated to the area before the third molar (M3) crown had completely formed (i.e., between 10 and 17.5 years of age). This is the first quantitative assessment of population mobility in Classical antiquity. This study demonstrates that migration was not limited to predominantly single adult males, as suggested by historical sources, but rather a complex phenomenon involving families. We hypothesize that migrants most likely came from higher elevations to the East and North of Rome. One individual with a higher delta(18)O value may have come (as a child) from an area isotopically similar to North Africa.

Isotopic evidence for age-related variation in diet from Isola Sacra, Italy.

This study examines collagen (N=105) and apatite (N=65) data from an Imperial Roman skeletal sample from the necropolis of Isola Sacra (Rome, Italy). This paper explores correlations between the isotopic composition of bone samples and the inferred age and sex of these individuals (aged 5--45+ years). The collagen of males, and older individuals in general, was significantly enriched in (15)N but not (13)C. Bone carbonate was somewhat depleted in (13)C in some older individuals, suggesting increased consumption of olive oil and possibly wine. Subadults (>5 years) in the sample appear to have consumed an almost exclusively terrestrial diet. This study demonstrates a clear trend in dietary patterns between adult age groups, as well as between adults and children within a population.