Ricardo Fernandes

Epigenetic variation in mangrove plants occurring in contrasting natural environment.

Background Epigenetic modifications, such as cytosine methylation, are inherited in plant species and may occur in response to biotic or abiotic stress, affecting gene expression without changing genome sequence. Laguncularia racemosa, a mangrove species, occurs in naturally contrasting habitats where it is subjected daily to salinity and nutrient variations leading to morphological differences. This work aims at unraveling how CpG-methylation variation is distributed among individuals from two nearby habitats, at a riverside (RS) or near a salt marsh (SM), with different environmental pressures and how this variation is correlated with the observed morphological variation. Principal Findings Significant differences were observed in morphological traits such as tree height, tree diameter, leaf width and leaf area between plants from RS and SM locations, resulting in smaller plants and smaller leaf size in SM plants. Methyl-Sensitive Amplified Polymorphism (MSAP) was used to assess genetic and epigenetic (CpG-methylation) variation in L. racemosa genomes from these populations. SM plants were hypomethylated (14.6% of loci had methylated samples) in comparison to RS (32.1% of loci had methylated samples). Within-population diversity was significantly greater for epigenetic than genetic data in both locations, but SM also had less epigenetic diversity than RS. Frequency-based (GST) and multivariate (βST) methods that estimate population structure showed significantly greater differentiation among locations for epigenetic than genetic data. Co-Inertia analysis, exploring jointly the genetic and epigenetic data, showed that individuals with similar genetic profiles presented divergent epigenetic profiles that were characteristic of the population in a particular environment, suggesting that CpG-methylation changes may be associated with environmental heterogeneity. Conclusions In spite of significant morphological dissimilarities, individuals of L. racemosa from salt marsh and riverside presented little genetic but abundant DNA methylation differentiation, suggesting that epigenetic variation in natural plant populations has an important role in helping individuals to cope with different environments.

Food Reconstruction Using Isotopic Transferred Signals ( FRUITS): A Bayesian Model for Diet Reconstruction

Human and animal diet reconstruction studies that rely on tissue chemical signatures aim at providing estimates on the relative intake of potential food groups. However, several sources of uncertainty need to be considered when handling data. Bayesian mixing models provide a natural platform to handle diverse sources of uncertainty while allowing the user to contribute with prior expert information. The Bayesian mixing model FRUITS (Food Reconstruction Using Isotopic Transferred Signals) was developed for use in diet reconstruction studies. FRUITS incorporates the capability to account for dietary routing, that is, the contribution of different food fractions (e.g. macronutrients) towards a dietary proxy signal measured in the consumer. FRUITS also provides relatively straightforward means for the introduction of prior information on the relative dietary contributions of food groups or food fractions. This type of prior may originate, for instance, from physiological or metabolic studies. FRUITS performance was tested using simulated data and data from a published controlled animal feeding experiment. The feeding experiment data was selected to exemplify the application of the novel capabilities incorporated into FRUITS but also to illustrate some of the aspects that need to be considered when handling data within diet reconstruction studies. FRUITS accurately predicted dietary intakes, and more precise estimates were obtained for dietary scenarios in which expert prior information was included. FRUITS represents a useful tool to achieve accurate and precise food intake estimates in diet reconstruction studies within different scientific fields (e.g. ecology, forensics, archaeology, and dietary physiology).

Macronutrient-based model for dietary carbon routing in bone collagen and bioapatite

Carbon stable isotope ratios (δ13C), measured in human bone collagen (δ13Ccollagen) and bioapatite (δ13Cbioapatite), are commonly used indicators in ancient human diet reconstruction. The underlying assumption is that human tissues broadly reflect the δ13C signal of dietary food sources (δ13Cdiet) plus an isotopic offset. However, interpretation of results may be confounded by the differentiated routing of macronutrients (energy, that is carbohydrates and lipids, and protein) having associated different isotopic signals (δ13Cenergy, δ13Cprotein). Multiple regression analyses were conducted on data from controlled animal feeding experiments compiled by Froehle et al. (J Archaeol Sci 37:2662–2670, 2010). We derived a simple algebraic macronutrient-based model with δ13Cbioapatiteu2009=u200910.1u2009+u2009δ13Cdiet (‰) and δ13Ccollagenu2009=u20094.8u2009+u20090.74 δ13Cproteinu2009+u20090.26 δ13Cenergy (‰). While the established relationship for δ13Cbioapatite is similar to previously known results, the model also suggests that δ13Ccollagen signal contributions originate from surprisingly consistent proportions of protein and energy macronutrients. Given that feeding experiments explore extreme variations in the proportion of diet macronutrients, the applicability of the proposed model and its predictions were tested in a variety of well-known, wild animal and human, natural contexts. Possible biochemical mechanisms explaining these empirical results are discussed.

Dietary history contributes to enterotype-like clustering and functional metagenomic content in the intestinal microbiome of wild mice

Significance Recent investigation of several mammalian hosts suggests that their intestinal bacterial communities display evidence of clusters characterized by differences in specific bacterial taxa, a concept referred to as enterotypes. By performing stable isotope analysis of environmental samples, monitoring communities during dietary shifts, and collecting functional metagenomic sequence data, we provide novel insight into the origins and dynamics of enterotype-like community clustering in wild house mice. Two clusters are present in wild mice, one associated with higher plant-derived and another with animal-derived food intake, which can shift within a period of 1 wk. Remarkably, these clusters display shared characteristics with those present in humans, chimpanzees, and laboratory mice, suggesting ancient shared traits among mammalian bacterial communities. Understanding the origins of gut microbial community structure is critical for the identification and interpretation of potential fitness-related traits for the host. The presence of community clusters characterized by differences in the abundance of signature taxa, referred to as enterotypes, is a debated concept first reported in humans and later extended to other mammalian hosts. In this study, we provide a thorough assessment of their existence in wild house mice using a panel of evaluation criteria. We identify support for two clusters that are compositionally similar to clusters identified in humans, chimpanzees, and laboratory mice, characterized by differences in Bacteroides, Robinsoniella, and unclassified genera belonging to the family Lachnospiraceae. To further evaluate these clusters, we (i) monitored community changes associated with moving mice from the natural to a laboratory environment, (ii) performed functional metagenomic sequencing, and (iii) subjected wild-caught samples to stable isotope analysis to reconstruct dietary patterns. This process reveals differences in the proportions of genes involved in carbohydrate versus protein metabolism in the functional metagenome, as well as differences in plant- versus meat-derived food sources between clusters. In conjunction with wild-caught mice quickly changing their enterotype classification upon transfer to a standard laboratory chow diet, these results provide strong evidence that dietary history contributes to the presence of enterotype-like clustering in wild mice.

Quantitative diet reconstruction of a Neolithic population using a Bayesian mixing model (FRUITS): The case study of Ostorf (Germany)

OBJECTIVESnThe island cemetery site of Ostorf (Germany) consists of individual human graves containing Funnel Beaker ceramics dating to the Early or Middle Neolithic. However, previous isotope and radiocarbon analysis demonstrated that the Ostorf individuals had a diet rich in freshwater fish. The present study was undertaken to quantitatively reconstruct the diet of the Ostorf population and establish if dietary habits are consistent with the traditional characterization of a Neolithic diet.nnnMETHODSnQuantitative diet reconstruction was achieved through a novel approach consisting of the use of the Bayesian mixing model Food Reconstruction Using Isotopic Transferred Signals (FRUITS) to model isotope measurements from multiple dietary proxies (δ13 Ccollagen , δ15 Ncollagen , δ13 Cbioapatite , δ34 Smethione , 14 Ccollagen ). The accuracy of model estimates was verified by comparing the agreement between observed and estimated human dietary radiocarbon reservoir effects.nnnRESULTSnQuantitative diet reconstruction estimates confirm that the Ostorf individuals had a high protein intake due to the consumption of fish and terrestrial animal products. However, FRUITS estimates also show that plant foods represented a significant source of calories. Observed and estimated human dietary radiocarbon reservoir effects are in good agreement provided that the aquatic reservoir effect at Lake Ostorf is taken as reference.nnnCONCLUSIONSnThe Ostorf population apparently adopted elements associated with a Neolithic culture but adapted to available local food resources and implemented a subsistence strategy that involved a large proportion of fish and terrestrial meat consumption. This case study exemplifies the diversity of subsistence strategies followed during the Neolithic. Am J Phys Anthropol 158:325-340, 2015.

Towards the use of radiocarbon as a dietary proxy: Establishing a first wide-ranging radiocarbon reservoir effects baseline for Germany

Radiocarbon reservoir effects (RREs) are observed when the 14C concentration of aquatic reservoirs is lower than the contemporary atmosphere. Within these reservoirs, aquatic species will also have a 14C depleted signal, and humans feeding on these species will show a dietary RRE. Human dietary RREs are often viewed as a problem for the establishment of reliable chronologies. However, they also represent an opportunity to introduce radiocarbon as a dietary proxy when investigating possible past human consumption of aquatic food groups. Here, a synthesis of previously published and new radiocarbon dates on edible aquatic species from central and northern Germany is presented. The samples were collected from modern and archaeological contexts. The goal was to provide an approximate RRE baseline within Germany. The results show that within the German context, local RREs in edible aquatic species are usually large and variable. The variability in local RREs implies that precise quantitative human dietary estimates will most likely not be possible. However, the large values of local RREs allow the use of 14C measured in human bone collagen as an extra dietary proxy that can aid in detecting the consumption of aquatic food groups when traditional isotopic proxies (δ13C and δ15N) do not provide unambiguous estimates.

Mussels with Meat: Bivalve Tissue-Shell Radiocarbon Age Differences and Archaeological Implications

Local reservoir ages are often estimated from the difference between the radiocarbon ages of aquatic material and associated terrestrial samples for which no reservoir effect is expected. Frequently, the selected aquatic material consists of bivalve shells that are typically well preserved in the archaeological record. For instance, large shell middens attest to the importance of mussel consumption at both coastal and inland sites. However, different physiological mechanisms associated with tissue and shell growth may result in differences in reservoir effects between the surviving component (shell) and the component relevant to dietary reservoir effects in consumers (tissue). The current study examines bivalve tissue-shell age differences both from freshwater and marine contexts close to archaeological sites where human consumption of mollusks has been attested. Results exhibited significant 14C age differences between bivalve tissue and shell in a freshwater context. In a marine context, no significant bivalve tissue-shell age differences were observed. The results also showed that riverine and lacustrine shells show large and variable freshwater reservoir effects. The results have important implications for establishing local reservoir effects especially in a freshwater environment. For good a priori knowledge of expected 14C differences in organic and inorganic water, carbon is thus necessary. Furthermore, the high variability in freshwater shell 14C ages implies the need for representative sampling from the archaeological record.

The use of Hand-Held XRF for investigating the composition and corrosion of Roman copper-alloyed artefacts

The study of archaeological artefacts using deployed in situ analytical instruments presents some obvious advantages. Including, obtaining an immediate feedback that can be used to redefine in real-time fieldwork strategies. Ideally analytical field instruments should also have characteristics that limit damage to the studied artefact.Here, we present a case study on the use of a Hand Held XRF (HH XRF) device employed to define the composition of copper-alloyed artefacts retrieved from the Roman military site of Fectio in the vicinity of Vechten (The Netherlands). The goals of the study were to establish artefact preservation status, to investigate artefact elemental composition, and to compare the composition of artefact corrosion layer and uncorroded core.The results showed that the objects were in an overall good preservation state. Decuprification and dezincification represented the probable main corrosion processes resulting in the formation of smooth corrosion layers or patinas. The major elemental composition of the artefacts’ uncorroded cores showed a wide-range variability most likely associated with recycling practices of scrap metal during the 3rd century CE.

A novel tertiary prep-HPLC method for the isolation of single amino acids for AMS-radiocarbon measurement

AMS-radiocarbon measurements of amino acids can potentially provide more reliable radiocarbon dates than bulk collagen analysis. Nonetheless, the applicability of such an approach is often limited by the low-throughput of existing isolation methods and difficulties in determining the contamination introduced during the separation process. A novel tertiary prep-HPLC amino acid isolation method was developed that relies on the combustion of eluted material without requiring any additional chemical steps. Amino acid separation was carried out using a gradient mix of pure water and phosphoric acid with an acetonitrile step in-between runs to remove hydrophobic molecules from the separation column. The amount of contaminant carbon and its 14C content were determined from two-point measurements of collagen samples of known 14C content. The amount of foreign carbon due to the isolation process was estimated at 4±1μg and its 14C content was 0.43±0.01 F14C. Radiocarbon values corrected for carbon contamination have only a minor increase in uncertainties. For Holocene samples, this corresponds to an added uncertainty typically smaller than 10 14Cyears. The developed method can be added to routine AMS measurements without implying significant operational changes and offers a level of measurement uncertainty that is suitable for many archaeological, ecological, environmental, and biological applications.

Investigating the correlation between monthly average temperatures and tithe proxy data from the Low Countries

This paper investigates the correlation between monthly average temperatures and tithe leasing dates in the Low Countries between 1600 and 1810. The information was obtained from manorial tithe leasing records distributed across the Netherlands and Belgium. Similar research in France and in Switzerland relied on annual dates of grape harvest as a temperature proxy and provided a strong correlation between harvest dates and average monthly temperatures. However, the analysis of our data indicates a low correlation between average monthly temperatures, using the Labrijn temperature series, and tithe leasing dates. Possible reasons for this behaviour include soil variety, diversity in crop growth patterns, historical factors, and variety of crop types. The complexity of variables, therefore, does not permit for the usage of tithe leasing dates for ancient temperature reconstruction. Nonetheless, useful information on crop behaviour under extreme climatic conditions was obtained.