Using portable X-ray fluorescence elemental analysis to explore porous skeletal lesions: Interplay of sex, age at death, and cause of death.

OBJECTIVES: Search for possible associations between bone elemental concentration and the presence of porous skeletal lesions (PSLs), considering the sex, age, and cause of death (COD) of the individuals. MATERIALS AND METHODS: The sample comprised 107 non-adult individuals (56 females, 51 males) aged 0-20 (x̄ = 13.2, SD = 5.8) from the Coimbra and Lisbon Identified Skeletal Collections. Cribra cranii, orbitalia, humeralis, and femoralis were recorded as present/absent, and elemental concentrations were assessed by portable x-ray fluorescence (pXRF). A multivariate statistical approach was applied. RESULTS: Well-preserved skeletons with minimal diagenesis showed no sex-related elemental variations or PSL associations. In contrast, age-at-death correlated with elevated Ca, P, Sr, and Pb levels. Cribra cranii increased with age while other cribra declined post-adolescence. Higher concentrations of Fe and lower of S were linked to cribra cranii. Respiratory infections as COD increased the odds of expressing cribra femoralis (OR = 5.25, CI = 1.25-15.14), cribra cranii (OR = 2.91, CI = 0.97-8.69), and cribra orbitalia (OR = 2.76, CI = 1.06-7.24). DISCUSSION: Feasible pXRF results and low cribra intraobserver error assure replicability. Elevated Ca, P, and Sr in older individuals may relate to skeletal growth, while increased Pb suggests bioaccumulation. Cribra's increase with age reflects different rates of marrow conversion and bone remodeling. Higher Fe and lower S in individuals with cribra cranii possibly reflects poor nutrition, early alcohol use, and sideroblastic anemia, aligning with 19th-20th-century Portugal's living conditions. Respiratory infections increased cribra expression, revealing intricate interplays among inflammation, anemia(s), marrow expansion, and diet. This research highlights a complex scenario and blazes a new path for cribra interpretation.

The Role of Fe, S, P, Ca, and Sr in Porous Skeletal Lesions: A Study on Non-adult Individuals Using pXRF.

Portable X-ray fluorescence is a new tool in the study of human bone. This research aims to investigate if variations in bone elemental concentrations are related with porous skeletal lesions (PSLs). One hundred well-preserved non-adult skeletons aged 0-11 years were selected from the archaeological site Convent of São Domingos, Lisbon (18th-19th century). Measuring a standard reference material and calculating the technical error of measurement assured elemental data reliability. Moreover, measuring soil samples excluded possible contamination of bones with elements from the soil, except for Pb. Additionally, the Ca/P ratio indicates maintenance of bone integrity. Cribra cranii, orbitalia, humeralis, and femoralis were recorded as present/absent, and the estimated intra-/inter-observer errors were low. The multivariate analysis found higher odds of having cribra orbitalia (OR = 1.76; CI = 0.97-3.20) and cribra femoralis (OR = 1.42; CI = 0.73-2.74) in individuals with lower Fe and higher S. Furthermore, higher levels of P, Ca, and Sr increased the odds of individuals developing cribra femoralis (OR = 2.30; CI = 1.23-4.29). Age also correlated with increased odds of exhibiting cribra orbitalia (OR = 1.86; CI = 0.94-3.68), cribra femoralis (OR = 6.97; CI = 2.78-17.45), and cribra humeralis (OR = 8.32; CI = 2.71-25.60). These findings suggest a shared etiology for these three cribras, contrasting with the higher Fe levels in individuals with cribra cranii. Lower Fe and higher S levels in individuals with cribra suggest a complex etiology, possibly involving conditions like megaloblastic or chronic disease anemia(s). Age-related elemental changes support the hypothesis that age influences cribra frequencies. This study highlights PSL complexity and opens new avenues for research.

Elemental analysis using portable X-ray fluorescence: Guidelines for the study of dry human bone.

OBJECTIVE: X-ray fluorescence (XRF) is a non-destructive technique that measures the elemental concentration of different materials, including human bone. Recently, it began to be applied to paleopathological studies due to the development of portable devices and their relative ease of use. However, the lack of uniform procedures hampers comparability and reproducibility. This paper aims to provide guidelines for an efficient and standardized evaluation of bone elemental composition with a portable XRF (pXRF) device. MATERIALS: This technical note is based on the application of the Thermo Scientific Niton XL3t 900 GOLDD+. METHODS: This work includes suggestions for the choice and preparation of human bone samples, both from archaeological context and documented collections, and methodological procedures in pXRF setup, such as choice of calibration, assessment of accuracy, and analysis run time. Additionally, recommendations for data validation and statistical analysis are also included. CONCLUSIONS: This technique has great potential in paleopathology since bone chemical variations may be associated with different pathological conditions, environmental contamination (e.g., lead), and/or administered treatments, such as mercury. Following an expected increase in the number of studies, it is essential to establish good practices that allow results from different researchers to be comparable. SIGNIFICANCE: X-ray fluorescence is a non-destructive technique that measures small concentrations (ppm) of elements from magnesium (12Mg) through bismuth (83Bi). LIMITATIONS: pXRF does not detect elements lighter than Mg, and its lower energy excitation penetrates less than other techniques. SUGGESTIONS FOR FURTHER RESEARCH: Other research groups should test these guidelines and comment on their usefulness and replicability.

Differential diagnosis of a diffuse sclerosis in an identified male skull (early 20th century Coimbra, Portugal): A multimethodological approach for the identification of osteosclerotic dysplasias in skeletonized individuals.

OBJECTIVE: This work aims to discuss the difficulties in diagnosing osteosclerotic changes in skeletonized individuals and to raise awareness of osteosclerotic dysplasias as a group of rare ancient diseases. MATERIALS: The skull of a 62-year-old male individual from the International Exchange Skull Collection, curated by the University of Coimbra, who died in 1928 presenting albuminous nephritis (Bright disease)/uraemia as the registered cause of death. METHODS: The skull was macroscopically and radiologically examined and bone elemental analysis was investigated. The genealogy and medical records of the individual were also searched. RESULTS: The lesions are in accordance with an osteosclerotic process possibly pointing to osteosclerosis, osteosclerotic metaphyseal dysplasia, or dysosteosclerosis, but osteoclasia with hyperphosphatasia, endosteal hyperostosis, sclerosteosis, or osteopathia striata with cranial sclerosis cannot be ruled out. CONCLUSIONS: Representativeness of the skeleton is a crucial feature in diagnosing rare diseases and, to avoid a misdiagnosis, the final diagnosis should include a group of diseases rather than a definite disease. SIGNIFICANCE: Difficulties in diagnosing rare diseases are discussed and best approaches in the study osteosclerotic dysplasias in skeletonized individuals are offered in the light of current clinical knowledge. LIMITATIONS: The absence of the postcranial skeleton and of pathognomonic lesions associated with osteosclerotic dysplasias limits diagnosis. Although rare diseases often have a genetic basis, specific genetic testing for the diagnosis of rare diseases in paleopathological cases are not yet available. SUGGESTIONS FOR FURTHER RESEARCH: Future genetic studies might help narrow down the diagnosis.

Elemental Composition in Female Dry Femora Using Portable X-Ray Fluorescence (pXRF): Association with Age and Osteoporosis.

Pathophysiological conditions can modify the skeletal chemical concentration. This study analyzes the elemental composition in two anatomical regions from dry femoral bone using a portable X-Ray Fluorescence (pXRF) and evaluates its impact in the bone mineral density (BMD). The left femora of 97 female skeletons (21-95 years old individuals) from the Coimbra Identified Skeletal Collection were studied. Diagenetic biases were discarded at the outset and BMD was determined with Dual-energy X-ray absorptiometry. Chemical measurements were performed at the midpoint of the femoral neck and at the midshaft using a pXRF device, and comparisons were made considering the age and the BMD values. Only elements with a Technical Measurement Error ≤ 5% were selected: P, S, Ca, Fe, Zn, As, Sr, Pb and the Ca/P ratio. Statistically significant differences were found between regions, with higher concentrations of P, Ca, Zn and S at the midshaft, and the Ca/P ratio at the femoral neck. The concentration of P is higher in individuals < 50 years, while S and Ca/P ratio increase in individuals ≥ 50 years. The decrease of P with age can be simultaneously related to the decline of its concentration in osteoporosis. Decreased BMD is also associated with higher levels of S and Pb. Osteoporosis enhances the absorption of osteolytic elements in specific locations. This fast and non-destructive technique has proved effective for the comprehension of chemical changes related to bone mass loss. This study highlights the potential of identified skeletal collections to improve the knowledge about bone fragility.

High-Quality Draft Genome Sequences of Three Cyanobacteria Isolated from the Limestone Walls of the Old Cathedral of Coimbra, Portugal.

The recently described species Myxacorys almedinensis and two other cyanobacteria were isolated from the limestone walls of the Old Cathedral of Coimbra, Portugal (UNESCO World Heritage Site). The high-quality genome sequences presented here will be essential for characterization purposes and description of the novel taxa.

High-Quality Draft Genome Sequence of the Microcolonial Black Fungus Aeminium ludgeri DSM 106916.

Aeminium ludgeri is an extremotolerant microcolonial black fungus isolated from a biodeteriorated limestone art piece in the Old Cathedral of Coimbra, Portugal (a UNESCO World Heritage Site). The high-quality draft genome sequence of Aeminium ludgeri presented here represents the first sequenced genome for both the recently described fungal family Aeminiaceae and the genus Aeminium.

Description of Aeminiaceae fam. nov., Aeminium gen. nov. and Aeminiumludgeri sp. nov. (Capnodiales), isolated from a biodeteriorated art-piece in the Old Cathedral of Coimbra, Portugal.

When colonizing stone monuments, microcolonial black fungi are considered one of the most severe and resistant groups of biodeteriorating organisms, posing a very difficult challenge to conservators and biologists working with cultural heritage preservation. During an experimental survey aimed to isolate fungi from a biodeteriorated limestone art piece in the Old Cathedral of Coimbra, Portugal (a UNESCO World Heritage Site), an unknown microcolonial black fungus was retrieved. The isolated fungus was studied through a complete examination based on multilocus phylogeny of a combined dataset of ITS rDNA, LSU and rpb2, in conjunction with morphological, physiological, and ecological characteristics. This integrative analysis allows for the description of a new family, Aeminiaceae fam. nov., a new genus Aeminium gen. nov., and a new species, Aeminiumludgeri sp. nov., in the order Capnodiales.