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The presence of scattered prehistoric human bones in caves and sinkholes is common in many regions of Iberia. These are usually interpreted as erratic elements coming from burial contexts, usually collective associations. These burial contexts are very frequent in karst areas of the Iberian Peninsula since the Early Neolithic, mostly in the Late Neolithic, and Copper Age, while findings from earlier chronologies are much more unusual. In this work, we present partial remains of a human skull from the Mesolithic period, recovered from a cave in the Strait of Gibraltar area. Although there is no conclusive evidence pointing to a dismantled burial context, this constitutes an isolated find, where its final location appears to be consistent with gravitational fall followed by water transportation., We are grateful to members of the caving groups GES Casares (Casares Underground Exploration Group) and SE Mainake (Speleo-Excursionist Society), and the town Council of Casares for their help throughout the work of survey and archaeological research inside Sima Hedionda II/IV. The frontal bone SHII/IV/7001-7 (known as Cecilín) was first located by Carlos Pintos Zanca, Juan Antonio Rodriguez Alarcón, Rafael Beltran, and Pascal Boutery de la SE Mainake. CT scan was carried out in the Clinical Veterinary Hospital of the University of Cordoba. We would like to thank especially Dr Manuel Novales Durán (Department of Animal Medicine and Surgery, UCO) for his help in the scanning process. We would also appreciate the help of Alvaro Perea (Ergonidámica Clínica, Córdoba) in composing Figures 5 and 6 using the OsiriX DICOM Wiever software. Dating Beta-424650 was funded by an internal research grant from La Trobe University. The “Archaeological Survey Project in Sima Hedionda II/IV, Casares (Málaga)” was partly self-financed by the team members themselves, and also partly (Technical equipment, transportation, other datings, maintenance, and accommodation) with the support of the Casares Town Council. Thanks to the efforts of the last institution, this cave is now gated and protected. This work has been carried out in the framework of the Project “Archaeobiology of the Neolithic of the Southern Iberian Peninsula” (NeArqBioSI) A-HUM-460-UGR18 by Consejería de Economía, Conocimiento, Empresas y Universidad. FEDER Programme – Junta de Andalucía-Universidad de Granada. Call 2019. SPG is Juan de la Cierva Incorporación (IJC2019-038830-I) funded by MCIN/AEI/10.13039/501100011033. We would like to thank the anonymous reviewers for their thoughtful comments and efforts in improving our manuscript.

This research was funded by the Institute of Health Carlos III (PI13/01559), including the European Regional Development Fund (FEDER) and the Regional Health Council of Andalusia (Spain) (PI-0405-2014)., This study was conducted in accordance with the Declaration of Helsinki, and the protocol was approved by the Ethics Committee of “Consejería de Salud y Familias, Junta de Andalucía” (PI-0405-2014). and “Consejería de Igualdad, Salud y Políticas Sociales, Junta de Andalucía” (PI13/01559)We follow the standards described in Andalusian and Spanish laws of personal data protection and biomedical research for the treatment of information and biological samples of human origin., Women were informed of all study procedures and gave their informed consent for inclusion before they participated in the study., The authors thank the team of the i-Diet software for their support in the estimation of daily energy and nutrient intake. Likewise, a special mention to the pregnant women who participated in this study and the health professionals from El Poniente Hospital, Almeria., Identification of causal factors that influence fetal growth and anthropometry at birth is of great importance as they provide information about increased risk of disease throughout life. The association between maternal genetic polymorphism MTHFR(677)C>T and anthropometry at birth has been widely studied because of its key role in the one-carbon cycle. MTHFR(677) CT and TT genotypes have been associated with a greater risk of low birth weight, especially in case of deficient intake of folic acid during pregnancy. This study aimed to analyze the association between the maternal MTHFR(677)C>T genetic polymorphism and anthropometry at birth in a population with adequate folate consumption. We included 694 mother-newborn pairs from a prospective population-based birth cohort in Spain, in the Genetics, Early life enviroNmental Exposures and Infant Development in Andalusia (GENEIDA) project. Women were genotyped for MTHFR(677)C>T SNP by Q-PCR using TaqMan (c) probes. Relevant maternal and newborn information was obtained from structured questionnaires and medical records. Results showed that maternal MTHFR(677)C>T genotype was associated with newborn anthropometry. Genotypes CT or CT/TT showed statistically significant associations with increased or decreased risk of large-for-gestational-age (LGA) or small-for-gestational-age (SGA) based on weight and height, depending on the newborn's sex, as well as with SGA in premature neonates. The relationships between this maternal genotype and anthropometry at birth remained despite an adequate maternal folate intake., Instituto de Salud Carlos III PI13/01559, European Commission, Regional Health Council of Andalusia (Spain) PI-0405-2014

This study presents the isotopic ratios, radiocarbon datings, and anthropological analyses of five (N = 5) Early Copper Age individuals from two archaeological sites (Arruzafa and Iglesia Antigua de Alcolea) of the Middle Guadalquivir Valley near the city of Córdoba (Spain). Four had been buried in the same grave, possibly in a single event or in a very short time lapse. The collagen residue analyses of the individuals offer δC values ranging between −20.08 and −18.4 and δN values between 8.57 and 11.15. The findings indicate that the infant and the elderly had, respectively, the richest and poorest animal protein diets, the first likely as a result of nursing. The combined study of these five individuals offers an interesting approach to a small segment of population of the farming communities occupying this territory at the beginning of the Copper Age (3200–3000 cal BC)., Consejería de Innovación y Empresa-Junta de Andalucía, Grant/Award Number: HUM-061658

Background. Despite the widespread use of organophosphate (OP) pesticides, information on predictors of children's exposure to such pesticides is scarce. Objective. To assess exposure to OP pesticides in children 3-11 year-old living in agricultural communities and urban areas from Andalusia (Southern Spain), and to identify the main determinants of exposure. Methods. A longitudinal study was conducted in children 3-11-year-old children living in agricultural communities and urban areas from the provinces of Almeria, Granada and Huelva (Andalusia, Spain) between 2010 and 2011. Urinary levels of six dialkylphosphate (DAP) metabolites were measured by UHPLC-QqQ-MS/MS at the periods of low and high pesticide use in the agriculture (LPU and HPU, respectively). Information on sociodemographic characteristics, parental occupation, residential history, lifestyle and diet, among other relevant factors, was obtained from questionnaires administered to the mothers. Results. A total of 559 and 597 children participated in LPU and HPU periods, respectively. The proportion of urine samples below LOD was 67.4% for ΣDMs (sum of dimethyl metabolites), 77% for ΣDEs (sum of diethyl metabolites) and 58.5% for ΣDAPs (sum of total dialkylphosphate metabolites) in LPU period, and 50.4% for ΣDMs, 65.3% for ΣDEs and 43.9% for ΣDAPs in HPU period. Significantly greater urinary ΣDAP, ΣDM and ΣDE levels were observed in HPU relative to LPU period. Maternal schooling years, proximity of the house to crops or greenhouses, use of insecticides at home, spraying the garden with pesticides, storage of pesticides at home, house cleaning frequency, as well as child's frequency of bath/shower, were found to be the major predictors of urinary levels of ΣDAP. Likewise, not washing fruit and vegetables before consumption and banana consumption were also identified as determinants of the exposure levels. Conclusions. Urinary levels of metabolites of OP pesticides found in this study were relatively lower compared to similar studies. DAP levels were significantly increased in HPU period. Maternal schooling years and variables related to residential environment and home exposures were identified as the most relevant determinants of DAP metabolites. Regarding diet, banana consumption and not washing fruit before consumption were also identified as determinants of the exposure levels. This study contributes to improve our knowledge on the main sources and determinants of children exposure to OPS, and given that children are more vulnerable than adults this information is essential to reduce children exposure and protect their health.

Background: Epidemiological data are not currently used in the risk assessment of chemical substances in a systematic and consistent manner. However, systematic reviews (SRs) could be useful for risk assessment as they appraise and synthesize the best epidemiological knowledge available. Objectives: To conduct a comprehensive literature search of SRs pertaining to pesticide exposure and various neurological outcomes, namely neurodevelopmental abnormalities, Parkinson's disease (PD) and Alzheimer's disease (AD), and to assess the potential contribution of SRs to the risk assessment process. Search methods and selection criteria: Search was conducted in PubMed and Web of Science databases and articles were selected if the following inclusion criteria were met: being a SR, published until April 2015 and without language restrictions. Data collection and analysis: For each neurological outcome, two review authors independently screened the search results for included studies. Data were extracted and summarized in two tables according to 16 criteria. Disagreements were resolved by discussion. Main results: The total number of studies identified in the first search was 65, 304 and 108 for neurodevelopment, PD and AD, respectively. From them, 8, 10 and 2 met the defined inclusion criteria for those outcomes, respectively. Overall, results suggest that prenatal exposure to organophosphates is associated with neurodevelopmental disturbances in preschool and school children. In contrast, postnatal exposures failed to show a clear effect across cohort studies. Regarding PD, 6 SRs reported statistically significant combined effect size estimates, with OR/RR ranging between 1.28 and 1.94. As for AD, 2 out of the 8 original articles included in the SRs found significant associations, with OR of 2.39 and 4.35, although the quality of the data was rather low. Conclusions: The critical appraisal of the SRs identified allowed for discussing the implications of SRs for risk assessment, along with the identification of gaps and limitations of current epidemiological studies that hinder their use for risk assessment. Recommendations are proposed to improve studies for this purpose. In particular, harmonized quantitative data (expressed in standardized units) would allow a better interpretation of results and would facilitate direct comparison of data across studies. Outcomes should be also harmonized for an accurate and reproducible measurement of adverse effects. Appropriate SRs and quantitative synthesis of the evidence should be performed regularly for a continuous update of the risk factors on health outcomes and to determine, if possible, dose–response curves for risk assessment. Keywords: Systematic review, Pesticides, Risk assessment, Neurodevelopment, Parkinson's disease, Alzheimer's disease, Neurodegenerative diseases

This study aimed to assess the association between exposure to chromium and neuropsychological development among children. A cross-sectional study was conducted with 393 children aged 6–11 years old randomly selected from State-funded schools in two provinces in Southern Spain (Almeria and Huelva), in 2010 and 2012. Chromium levels in urine and hair samples were analyzed by inductively coupled plasma mass spectrometry with an octopole reaction system. Neuropsychological development was evaluated using the Wechsler Intelligence Scale for Children-Fourth Edition (WISC-IV) and three computerized tests from the Behavioural Assessment and Research System (BARS): Reaction Time Test (RTT), Continuous Performance Test (CPT) and Selective Attention Test (SAT). Multivariable linear regression models adjusted for potential confounders, including heavy metals, were applied to examine the association between chromium levels and neuropsychological outcomes. A 10-fold increase in urine chromium levels was associated with a decrease of 5.99 points on the WISC-IV Full-Scale IQ (95% CI: 11.98 to −0.02). Likewise, a 10-fold increase in urine chromium levels in boys was associated with a decrease of 0.03 points in the percentage of omissions (95% CI: 0.0 to 0.05) in the SAT, with an increase of 68.35 points in latency (95% CI: 6.60 to 130.12) in the RTT, and with an increase in the number of trials with latencies > 1000 ms (β = 37.92; 95% CI: 2.73 to 73.12) in the RTT. An inverse significant association was detected between chromium levels in hair and latency in the SAT in boys (β = −50.53; 95% CI: 86.86 to −14.22) and girls (β = −55.95; 95% CI: 78.93 to −32.97). Excluding trials with latencies >1000 ms in the RTT increased latency scores by 29.36 points in boys (95% CI: 0.17 to 58.57), and 39.91 points in girls (95% CI: 21.25 to 58.59). This study is the first to show the detrimental effects of postnatal chromium exposure on neuropsychological development in school-aged children.

Polymorphisms in genes encoding xenobiotic-metabolizing enzymes (XME) are important parameters accounting for the wide inter-individual variability to environmental exposures. Paraoxonase-1 (PON1), butyrylcholinesterase (BChE) and Cytochrome-P450 constitute major classes of XME involved in the detoxification of pesticide chemicals, in particular organophosphates. This study explored the allelic frequency, linkage disequilibrium and haplotype analysis of ten common polymorphic variants of seven key genes involved in organophosphate metabolism (BCHE-K, BCHE-A, PON1 Q192R, PON1 L55M, PON1 -108C/T, CYP2C19 G681A, CYP2D6 G1846A, CYP3AP1 -44G/A, GSTM1∗0 and GSTT1∗0) in a children population living near an intensive agriculture area in Spain. It was hypothesized that individuals with unfavorable combinations of gene variants will be more susceptible to adverse effects from organophosphate exposure. Genomic DNA from 496 healthy children was isolated and amplified by PCR. Hydrolysis probes were used for the detection of eight specific SNPs and two copy number variants (CNVs) by using TaqMan® Assay-based real-time PCR. Frequencies of SNPs and CNVs in the target genes were in Hardy-Weinberg equilibrium and broadly consistent with European populations. Linkage disequilibrium was found between the three PON1 genetic polymorphisms studied and between BCHE-K and BCHE-A. The adverse genotype combination (unusual BCHE variants, PON1 55MM/-108TT and null genotype for both GSTM1 and GSTT1) potentially conferring a greater genetic risk from exposure to organophosphates was observed in 0.2% of our study population. This information allows broadening our knowledge about differential susceptibility toward environmental toxicants and may be helpful for further research to understand the inter-individual toxicokinetic variability in response to organophosphate pesticides exposure.

SUMMARYBonamia ostreaeis a protozoan parasite that infects the European flat oysterOstrea edulis, causing systemic infections and resulting in massive mortalities in populations of this valuable bivalve species. In this work, we have characterizedB. ostreaeactin genes and used their sequences for a phylogenetic analysis. Design of different primer sets was necessary to amplify the central coding region of actin genes ofB. ostreae. Characterization of the sequences and their amplification in different samples demonstrated the presence of 2 intragenomic actin genes inB. ostreae, without any intron. The phylogenetic analysis placedB. ostreaein a clade withMinchinia tapetis,Minchinia teredinisandHaplosporidium costaleas its closest relatives, and demonstrated that the paralogous actin genes found inBonamiaresulted from a duplication of the original actin gene after theBonamiaorigin.

Background Information on exposure levels to organochlorine compounds (OCs) in child population is limited, despite their greater vulnerability to the adverse health effects of these chemicals. Objective To determine serum concentrations of 10 OCs (including organochlorine pesticides and polychlorinated biphenyls –PCBs–) in children living in agricultural communities from Almeria (South-Eastern Spain), and to identify the main predictors of exposure related to socio-economic characteristics, diet and lifestyle. Methods A cross-sectional study was conducted on 133 children aged 6–11 years selected from public schools of the study area. OCs compounds were determined in serum samples by GC/ECD. Anthropometric measures were obtained during sample collection. Information on sociodemographic characteristics, parental occupation, residential history, lifestyle and frequency of food consumption, among other relevant factors, was obtained by questionnaires administered to the mothers. Results Geometric means of serum concentrations (ng/ml) were 0.11 for β-hexachloro-cyclohexane (β-HCH), 0.09 for endosulfan, 0.20 for endosulfan-ether, 0.51 for hexachorobenzene (HCB), 0.08 for mirex, 0.06 for oxychlordane, 0.36 for p,p'-DDE, 0.20 for PCB 138, 0.36 for PCB 153, and 0.45 for PCB 180. Percentage of samples above the limit of detection (0.05 ppb) ranged from 32 (β-HCH) to 100 (HCB). A high variability in OC levels depending on the compound was observed between our results and others found in similar studies carried out in children. Variables related to fish consumption were found to be the major dietary determinant of PCB 138, p,p´-DDE, endosulfan-α, β-HCH, mirex and oxychlordane levels. Conclusions Children participating in this study showed detectable levels of many OC, despite these compounds are no longer used. Their presence in children serum can be explained by their high lipophilicity and environmental persistence, leading to contamination of fatty food. In this line, fish consumption seemed to be the most relevant determinant of OC levels found in our study.

Introduction: The interaction between postnatal organophosphate exposure and children PON1 genetic polymorphisms on neuropsychological performance was evaluated in scholar children living in agricu...

Several studies have suggested that exposure to DDT may be related to changes in thyroid hormone levels in animals and humans, even though results across studies are inconsistent. The aim of this study was to assess the association between exposure to p,p'-DDE (a stable metabolite of DDT) and serum levels of thyroid hormones in floriculture workers. A longitudinal study was conducted on 136 male subjects from the States of Mexico and Morelos, Mexico, who were occupationally exposed to pesticides, during agricultural periods of high (rainy season) and low (dry season) levels of pesticide application. Using a structured questionnaire, a survey was carried out on socio-demographic characteristics, anthropometry, clinical history, alcohol and tobacco consumption, residential chemical exposure, and occupational history. Blood and urine samples were collected to determine serum levels of TSH, total T3, total T4, and p,p'-DDE, and metabolites of organophosphate pesticides (OP), respectively. The analysis of the associations between p,p'-DDE levels and thyroid hormone profile adjusting by potential confounding variables including urinary OP metabolites was carried out using multivariate generalized estimating equation (GEE) models. Our results showed that the geometric means of p,p'-DDE levels were 6.17 ng/ml and 4.71 ng/ml in the rainy and dry seasons, respectively. We observed positive associations between the serum levels of p,p'-DDE and those of total T3 (β=0.01, 95% CI: -0.009, 0.03), and total T4 (β=0.08, 95% CI:0.03, 0.14) and negative but no significant changes in TSH in male floricultural workers, supporting the hypothesis that acts as thyroid disruptor in humans.

Objectives p,p9-Dichlorodiphenyldichloroethene (p,p9-DDE) acts as an androgen receptor antagonist, however data regarding its hormonal effects in men are limited. The objective of this study was to evaluate the association between serum levels of p,p9-DDE and reproductive hormone profile in Mexican male flower growers. Methods A longitudinal study was carried out in a population of men working in the production of flowers and ornamental plants in two Mexican states during July–October 2004 (rainy season) and December 2004–May 2005 (dry season). A questionnaire including information on socioeconomic characteristics, tobacco and alcohol use, presence of chronic and acute diseases, occupational history and anthropometry was used and blood and urine samples were obtained. Serum levels of p,p9-DDE were analysed by gas chromatography; FSH, LH, testosterone, oestradiol, inhibin B and prolactin levels were measured by enzymatic immunoassay. Urinary levels of dialkylphosphates (DAPs) were analysed by gas chromatography. Associations between serum levels of p,p9-DDE and male reproductive hormones (both transformed to their natural logarithm) were evaluated using multivariate generalised estimating equation (GEE) models. Results Median p,p9-DDE levels were 677.2 ng/g lipid (range 9.4–12 696.5) during the rainy season and 626.7 ng/g lipid (range 9.4–13 668.1) during the dry season. After adjusting for potential confounders (age, body mass index, state of residence and DAPs), p,p9-DDE levels were negatively associated with prolactin (β=−0.04; 95% CI −0.07 to −0.008) and testosterone (β=0.04; 95% CI −0.08 to 0.005) and positively with inhibin B (β=0.11; 95% CI 0.02 to 0.21). Conclusion These results indicate that p,p9-DDE can affect hypothalamic–pituitary–gonadal axis function in humans.

Bonamia ostreae is an intracellular protozoan which is recognized as a cause of mortality in European populations of flat oysters (Ostrea edulis). Based on the recent characterization of actin genes of B. ostreae, specific primers were designed for real-time PCR using SYBR® Green chemistry. Specificity was demonstrated by the unique melting temperature peak observed in positive samples and by the lack of amplification in samples of oysters infected by closely related parasites, including Bonamia exitiosa. A calibration curve using a cloned template was defined to estimate copy number. The assay had a 6 log- dynamic range, mean inter- and intra-assay variation coefficients of

In this paper we have attempted to clarify the taxonomy and nomenclature of thirteen taxa of the genus Cortinarius subgenus Telamonia (sections Hydrocybe, Fraternii) well represented in the southwestern Mediterranean area of Europe (C. atrocoeruleus, C. bombycinus, C casimiri, C. contrarius, C. decipiens, C. fraternus, C. gallurae, C. hoffmannii, C. petroselineus, C. sertipes, C. subturibulosus, C. urdaibaiensis and C. vernus). To this end we have performed a combined study of morphological and molecular data (rDNA ITS sequences). The morphological analysis was carried out on 114 collections and the molecular analysis involved 31 of the 114 collections, including 11 type collections (types for C. casimiri and C. fraternus were not available). In addition, a study of spores under field emission scanning electron microscopy (FESEM) was conducted. The results of the combined analysis allowed us to asign the studied material to five species (C. casimiri s.l., C. decipiens s.l., C. gallurae, C. subturibulosus s.l. and C. vernus s.l.). Thus, all collections from more continental areas, which were originally identified as six different taxa (C. atrocoeruleus, C. contrarius, C. decipiens, C. fraternus, C. sertipes, C. flexipes fo. sertipes) corresponded to C. decipiens sensu lato, a widely distributed, genetically and morphologically variable species. Cortinarius casimiri is also found in such habitats, but it is confirmed as distinct taxon. Collections from Mediterranean sclerophyllous communities correspond to C. gallurae, C. vernus sensu lato and C. subturibulosus sensu lato. Due to close phylogenetic relationships we propose the new combinations C. casimiri var. hoffmannii (=C. decipiens var. hoffmannii non C. hoffmannii) and C. subturibulosus var. bombycinus (=C. bombycinus), and the new variety C. vernus var. nevadavernus (=C. vernus H. Lindstr. & Melot sensu auct.).

Paraoxonase-1 (PON1) is a serum enzyme which catalyzes the hydrolysis of organophosphate pesticides. In this study we conducted a cross-sectional study and reported on the distribution of three common genetic polymorphisms of the PON1 gene in a population of floriculture workers from Mexico as well as the association between those polymorphisms and other predictors with serum PON1 activity on paraoxon, diazoxon and phenylacetate. The genotype frequencies at position PON1(55) were 89% (LL), 10% (LM) and 0.6% (MM), at position PON1(192) they were 16% (QQ), 47% (QR) and 37% (RR), and 26% (TT), 42% (TC) and 32% (CC) at position PON1(-108). Thus, the frequencies of alleles L, Q and T were 0.94, 0.40 and 0.47, respectively. The PON1(55) polymorphism had no significant effect on serum PON1 activity on any substrate. We found a significant association between the PON1(192) polymorphism and PON1 activity towards paraoxon and diazoxon, which increased in genotypes as follows: 192RR>192QR>192QQ for paraoxonase activity and, inversely, 192QQ>192QR>192RR for diazoxonase activity. The PON1(-108) polymorphism also had a significant effect on PON1 activity level towards paraoxon in the following order among the genotype groups: -108CC>-108TC>-108TT. Serum PON1 activity towards diazoxon was not associated with the PON1(-108) polymorphism but it was influenced by the intensity exposure to pesticides at the floriculture industry and the years of the occupational exposure to pesticides. No polymorphism significantly influenced serum PON1 activity on phenylacetate.

In the course of a histopathological survey performed to discover the cause of mass mortality of the striped clam Chamelea gallina in the Balearic Islands (Spain, Mediterranean Sea), we detected a Marteilia-like parasite in 3 clams. Molecular methods were applied to identify the parasite. DNA extracted from a paraffin block was used to carry out a PCR assay for Marteilia refringens detection based on a rDNA sequence of the parasite (the intergenic spacer of ribosomal genes, IGS). The nucleotide sequence of the IGS amplified fragment and the positive signal obtained by in situ hybridisation analysis with a M. refringens-specific probe allowed us to confirm the presence of this parasite in the digestive gland tissue of C. gallina.

Here, a new satellite-DNA family is isolated and characterized from wedge sole, Dicologoglossa cuneata Moreau, 1881 (Pleuronectiformes), a fish having a small genome. This satellite-DNA family of sequences was isolated by conventional cloning after digestion of genomic DNA with the DraI restriction enzyme. Repeat units are 171 bp in length with a high AT content (63%). Several runs of consecutive adenines and thymines were found, and concomitantly computer analyses revealed that these regions are prone to acquire stable sequence-directed curvature. Especially remarkable is that the DraI sequences are composed almost entirely of the repetition of up to fourteen 9-bp motifs (T/C)GTC(A/C)AAAA similar to other vertebrate centromeric satellite-DNA sequences. In fact, we demonstrate the origin of this satellite through duplication of this motif plus the addition of a stretch of cytosines. The centromeric location and the presence in this satellite-DNA sequence of not only different vertebrate motifs (CENP-B box, pJα) but also others such as the CDEIII motif of Saccharomyces cerevisiae reveal a possible role in centromere function. All these characteristics provide important information on the origin, function, and the evolution of the centromeric satellite DNAs in wedge sole.

8 pages, 4 figures, The life cycle of the species forming the genus Marteilia (Phylum Paramyxea), which are marine protozoan parasites of molluscs, is still poorly known, although there is evidence of the need for intermediate hosts. To date, only the calanoid copepod Acartia (Paracartia) grani has been demonstrated to be involved in the life cycle of the parasite infecting flat oysters growing in ponds with a low biodiversity. However, no complex natural environments, such as bays or estuaries, have been studied for a similar purpose. Therefore, a survey for the presence of the protozoan Marteilia was conducted by PCR on the zooplankton community of a natural ecosystem, the Alfacs and Fangar bays in the Delta de l'Ebre (NW Mediterranean). Identification of zooplankton species found infected allowed two new Marteilia hosts to be proposed: the cyclopoida Oithona sp. and an indeterminate harpaticoida species. This is the first report of Marteilia in zooplankton species sampled from a natural enzootic area. © 2007 Elsevier B.V. All rights reserved, This study was supported by funds from an INIA predoctoral grand provided by the Spanish Minister of Science, the MCYT Project CTM-02575/MAR to M.A., the EU Community Reference Laboratory for Mollusc Diseases and Technology, and the Marie Curie Host Fellowships (n°405865H/P297815F) provided by the European Commission

Background: Childrens exposure to neurotoxic compounds poses a major problem to public health because of their actively developing brain that makes them highly vulnerable. However, limited information is available on neuropsychological effects in children associated with pre- and postnatal exposures to pesticides. Objective: To evaluate the association between current and pre- and postnatal exposures to pesticides and their effects on neurodevelopment in children aged 6–11 years living in agricultural communities from South-Eastern Spain. Methods: An ambispective study was conducted on 305 children aged 6–11 years randomly selected from public schools of the study area. Current exposure to organophosphate pesticides was assessed measuring children's urinary levels of dialkylphosphates (DAPs). Both prenatal and postnatal residential exposure to pesticides was estimated by developing a geographical information system (GIS) technology-based index that integrated distance-weighted measure of agricultural surface, time-series of crop areas per municipality and year, and land-use maps. Neuropsychological performance was evaluated with the Wechsler Intelligence Scale for Children-Fourth Edition (WISC-IV). The association of pre- and postnatal and current pesticide exposure with WISC-IV scale scores was assessed using multivariate linear regression models and generalized estimating equation (GEE) models, respectively. Results: Greater urinary DAP levels were associated with a poorer performance on intelligence quotient and verbal comprehension domain, with effects being more prominent in boys than in girls. The influence of an increase in 10 ha per year in crop surface around the child's residence during the postnatal period was associated with decreased intelligence quotient, processing speed and verbal comprehension scores. As regards prenatal exposure to pesticides, a poor processing speed performance was observed. These effects were also more prominent in boys than in girls. Conclusions: Our results suggest that postnatal exposure to pesticides can negatively affect children's neuropsychological performance. Prenatal exposure was weakly associated to neurodevelopment impairment. Keywords: Pesticides, Neuropsychological development, Neurotoxicity, Children, Environmental

Blood pH controls the activity of important regulatory enzymes in the metabolism. Serine dehydratase (SerDH) transforms l-serine into pyruvate and ammonium and is involved in the regulation of gluconeogenesis from serine in the rat liver. In this work, we investigate the effect of chronic metabolic acidosis on the kinetics, specific protein level, tissue location, and mRNA levels of rat liver SerDH. Experimental acidosis was induced in rats by ingestion of 0.28 M ammonium chloride solution for 10 days. Acidosis significantly ( P < 0.05) decreased SerDH activity at all substrate concentrations assayed. Moreover, the Vmaxvalue was 38.50 ± 3.51 mU/mg ( n = 7) of mitochondrial protein in the acidotic rats and 92.49 ± 6.79 mU/mg ( n = 7) in the control rats. Western blot analysis revealed a significant reduction (14%) in the level of SerDH protein content in the rat liver during acidosis. Immunohistochemical analysis showed that SerDH location did not change in response to chronic metabolic acidosis and confirmed previous results on SerDH protein levels. Moreover, the SerDH mRNA level, estimated by RT-PCR, was also significantly 33.8% lower than in control. These results suggest that during experimental acidosis a specific repression of rat-liver SerDH gene transcription could result, lowering the amount and activity of this enzyme. The changes found in SerDH expression are part of an overall metabolic response of liver to maintain acid-base homeostasis during acidosis.

Marteilia refringens is a paramyxean parasite which infects the flat oyster Ostrea edulis and mussels (Mytilus galloprovincialis), where it has been attributed to a separate species, Marteilia maurini, by several authors. Doubts persist though as to the existence or not of two species of Marteilia in Europe. We have devised a molecular method for the diagnosis of M. refringens based on 358 bp nested-PCR of the rDNA intergene spacer (rDNA IGS) which is capable of detecting 0.5 fg of M. refringens DNA. Molecular characterization of this spacer indicates that the Marteilia parasites which infect oysters and mussels are two different strains of the same species.

Premise of the study: We characterize 10 microsatellite loci in the endangered fern Vandenboschia speciosa (Hymenophyllaceae), enabling studies on the genetic population structure of this Macaronesian-European species using DNA hypervariable markers. Methods and Results: Ten primer sets were developed and tested on 47 individuals in a total of two Iberian populations of V. speciosa. The primers amplified di- and hexanucelotide repeats. The number of alleles ranged from two to eight, and the expected heterozygosity ranged from 0.107 to 0.807 among the populations analyzed. Conclusions: The 10 microsatellite markers developed will be useful in characterizing the genetic diversity of V. speciosa and understanding its population structure (including the possible structure between sporophyte and gametophyte phases) and biogeographic history, and will provide important genetic data for the conservation of this species., This study was supported by the Regional Andalusian Government (project P10-RNM-6198). C.G.L. was funded by a postdoctoral grant from the Regional Andalusian Government. S.B.S. and L.T.C. were funded by a Beca-Iniciación a la Investigación grant from the Universidad de Granada during 2012–2013 and 2014–2015, respectively. I.M.A. was funded by a Beca-Colaboración grant (Spanish Government).

In a previous work, we characterized the HinfI satellite DNA family in the subtribe Centaureinae (Cardueae) demonstrating that a “library” of eight HinfI subfamilies would exist in the common ancestor of all Centaureinae, which were differentially amplified in different lineages. Now, we extend our study by analyzing a total of 219 additional repeats from fifteen species belonging to Carlininae, Echinopsinae and Carduinae, and comparing them to those of Centaureinae. Most HinfI sequences belonged to the subfamily II, although a few sequences of other subfamilies were detected in some species. Additionally, a new subfamily characteristic of several Carduinae species was discovered. Although phylogenetic trees grouped sequences by subfamily affinity instead of species provenance, when comparing repeats of the same subfamily, the degree of divergence between any pair of sequences was related to the evolutionary distance between the species compared in most cases. Exceptions were in comparisons between sequences of some Centaureinae species, and between sequences of some Carduinae species and those of Centaureinae. Our results demonstrate that: (1) At least nine HinfI subfamilies would exist in the common ancestor of Cardueae, each one differentially amplified in different lineages; (2) After differential spreading, sequences of each subfamily evolved concertedly through molecular drive, resulting in the gradual divergence of repeats between different species; (3) The rate to which concerted evolution occurred was different between lineages according to the evolutionary history of each one.

We report on the isolation and characterization of eight microsatellite markers from enriched libraries for the critically endangered Atropa baetica. These are the first microsatellite loci reported for Atropa species. The total number of alleles found was 18, the expected heterozygosity ranged from 0.198 to 0.505. These markers will be useful to establish the real census of individuals and the genetic diversity both within and among the different populations of A. baetica.

We report on the isolation and characterization of eight microsatellite markers from enriched libraries for Arenaria nevadensis, one of the most critically endangered plant species in the Iberian Peninsula. These are the first microsatellite loci reported for Arenaria species. The number of alleles ranged from two to eight, and the expected heterozygosity from 0.067 to 0.873. These markers will be useful for characterizing the genetic diversity in A. nevadensis and understanding its population structure, and will provide important genetic data for the conservation and recovery of this species.

BACKGROUND AND AIMS Subtribe Centaureinae appears to be an excellent model group in which to analyse satellite DNA and assess the influence that the biology and/or the evolution of different lineages have had on the evolution of this class of repetitive DNA. Phylogenetic analyses of Centaureinae support two main phases of radiation, leading to two major groups of genera of different ages. Furthermore, different modes of evolution are observed in different lineages, reflected by morphology and DNA sequences. METHODS The sequences of 502 repeat units of the HinfI satellite DNA family from 38 species belonging to ten genera of Centaureinae were isolated and compared. A phylogenetic reconstruction was carried out by maximum likelihood and Bayesian inference. KEY RESULTS Up to eight different HinfI subfamilies were found, based on the presence of a set of diagnostic positions given by a specific mutation shared by all the sequences of one group. Subfamilies V-VIII were mostly found in older genera (first phase of radiation in the subtribe, late Oligocene-Miocene), although some copies of these types of repeats were also found in some species of the derived genera. Subfamilies I-IV spread mostly in species of the derived clade (second phase of radiation, Pliocene to Pleistocene), although repeats of these subfamilies exist in older species. Phylogenetic trees did not group the repeats by taxonomic affinity, but sequences were grouped by subfamily provenance. Concerted evolution was observed in HinfI subfamilies spread in older genera, whereas no genetic differentiation was found between species, and several subfamilies even coexist within the same species, in recently radiated groups or in groups with a history of recurrent hybridization of lineages. CONCLUSIONS The results suggest that the eight HinfI subfamilies were present in the common ancestor of Centaureinae and that each spread differentially in different genera during the two main phases of radiation following the library model of satellite DNA evolution. Additionally, differential speciation pathways gave rise to differential patterns of sequence evolution in different lineages. Thus, the evolutionary history of each group of Centaureinae is reflected in HinfI satellite DNA evolution. The data reinforce the value of satellite DNA sequences as markers of evolutionary processes.

Eukaryotic genomes are composed of both unique and repetitive DNA sequences. These latter form families of different classes that may be organized in tandem or may be dispersed within genomes with a moderate to high degree of repetitiveness. The repetitive DNA fraction may represent a high proportion of a particular genome due to correlation between genome size and abundance of repetitive sequences, which would explain the differences in genomic DNA contents of different species. In this review, we analyze repetitive DNA diversity and abundance as well as its impact on genome structure, function, and evolution.

Background and Aims: p,p´-dichlorodiphenyldichloroethene (p,p´-DDE) acts as an androgen receptor antagonist, (Kelce et al 1995), however the data regarding its hormonal effects in men are limited. ...

SarkOne is a genus-specific satellite-DNA family, isolated from the genomes of the species of the genus Sarcocapnos. This satellite DNA is composed of repeats with a consensus length of 855 bp and a mean G+C content of 52.5%. We have sequenced a total of 189 SarkOne monomeric repeats belonging to a total of seven species of the genus Sarcocapnos. The comparative analysis of these sequences both at the intraspecific and the interspecific levels have revealed divergence patterns between species are proportional to between-species divergence according to the phylogeny of the genus. Our study demonstrates that the molecular drive leading to the concerted-evolution pattern of this satellite DNA is a time-dependent process by which new mutations are spreading through genomes and populations at a gradual pace. However, time is a limiting factor in the observation of concerted evolution in some pairwise comparisons. Thus, pairwise comparisons of species sharing a recent common ancestor did not reveal nucleotide sites in transitional stages higher than stage III according to the Strachan’s model. By contrast, there was a gradation in the percentage of upper transition stages (IV, V, VI) the more phylogenetically distant the species were. In addition, closely related species shared a high number of polymorphic sites, but these types of sites were not common when comparing more distant species. All these data are discussed in the light of current life-cycle models of satellite-DNA evolution.

Organophosphate pesticides are widely used in agricultural purposes. Recently, a few studies have demonstrated the ability of these chemicals to alter the function of the thyroid gland in human. Moreover, the paraoxonase-1 enzyme (PON1) plays an important role in the toxicity of some organophosphate pesticides, with low PON1 activity being associated with higher pesticide sensitivity. This study evaluates the interaction between exposure to organophosphate compounds and PON1 enzyme activity on serum levels of TSH and thyroid hormones in a population of workers occupationally exposed to pesticides. A longitudinal study was conducted on a population of floriculture workers from Mexico, during two periods of high and low-intensity levels of pesticide application. A structured questionnaire was completed by workers containing questions on sociodemographic characteristics and other variables of interest. Urine and blood samples were taken, and biomarkers of exposure (dialkylphosphates), susceptibility (PON1 polymorphisms and activity) and effect (thyroid hormone levels) were determined. Interaction between dialkylphosphates and PON1 polymorphisms or PON1 activity on hormone levels was evaluated by generalized estimating equation (GEE) models. A significant interaction was found between serum diazoxonase activity and total dialkylphosphates ({Sigma}DAP) on TSH levels. Thus, when PON1 activity was increased we observed a decrease in the percentage ofmore » variation of TSH level for each increment in one logarithmic unit of the {Sigma}DAP levels. This interaction was also observed with the PON1{sub 192}RR genotype. These results suggest a stronger association between organophosphate pesticides and thyroid function in individuals with lower PON1 activity.« less

When genomic DNA from the European flat oyster Ostrea edulis L. was digested by BclI enzyme, a band of about 150 bp was observed in agarose gel. After cloning and sequencing this band and analysing their molecular characteristics and genomic organization by means of Southern blot, in situ hybridisation, and polymerase chain reaction (PCR) protocols, we concluded that this band is an interspersed highly repeated DNA element, which is related in sequence to the flanking regions of (CT)-microsatellite loci of the species O. edulis and Crassostrea gigas. Furthermore, we determined that this element forms part of a longer repetitive unit of 268 bp in length that, at least in some loci, is present in more than one copy. By Southern blot hybridisation and PCR amplifications-using primers designed for conserved regions of the 150-bp BclI clones of O. edulis-we determined that this repetitive DNA family is conserved in five other oyster species (O. stentina, C. angulata, C. gigas, C. ariakensis, and C. sikamea) while it is apparently absent in C. gasar. Finally, based on the analysis of the repetitive units in these oyster species, we discuss the slow degree of concerted evolution in this interspersed repetitive DNA family and its use for phylogenetic analysis.

The ability of organophosphate pesticides to disturb thyroid gland function has been demonstrated by experimental studies on animal, but evidence of such effects on human remains scarce. The aim of this study was to assess the association between exposure to organophosphate compounds and serum levels of thyroid hormones in floriculture workers. A longitudinal study was conducted on 136 male subjects from the State of Mexico and Morelos, Mexico, occupationally exposed to organophosphate pesticides, during agricultural periods of high (rainy season) and low (dry season) levels of pesticide application. Using a structured questionnaire, a survey was carried out on sociodemographic characteristics, anthropometry, clinical history, alcohol and tobacco consumption, residential chemical exposure, and occupational history. Urine and blood samples were taken the day after pesticide application to determine urine dialkylphosphate (DAP) levels, serum levels of TSH, total T(3), total T(4), serum PON1 activity, and serum p,p'-DEE levels. The analysis of the association between DAP levels and thyroid hormonal profile was carried out using multivariate generalized estimating equation (GEE) models. Our results showed an increase in both TSH and T(4) hormones in serum associated with a increase in total dimethylphosphate levels (SigmaDMP) in urine (p-trend

10 pages, 4 figures, Since the first description of Marteilia refringens (Paramyxea) in flat oysters Ostrea edulis in 1968 in the Aber Wrach, Brittany (France), the life-cycle of this parasite has remained unknown. However, recent studies, conducted in the ‘ claire ’ system, have proposed the planktonic copepod Acartia grani as a potential intermediate host for the parasite. Nevertheless, experimental transmission of the parasite through the copepod has failed. Recent studies in this field have reported the presence of the parasite in zooplankton from the bays of the Delta de l’Ebre, a more complex and natural estuarine environment than that of the claire. As a result, 2 new Marteilia host species were proposed: the copepods Oithona sp. (Cyclopoida) and an indeterminate Harpaticoida. Consequently, the objective of the present work was to study the dynamics of Marteilia in the zooplankton community from one of the bays, Alfacs Bay, as well as the dynamics of the parasite in cultivated mussels during 1 complete year. Six different zooplankton taxa appeared to be parasitized by M. refringens, including copepods (3 Calanoida, Acartia discaudata, A. clausi and A. italica ; 1 Cyclopoida, Oithona sp.; and 1 Harpacticoida, Euterpina acutifrons), and larval stages of decapod crustaceans (zoea larvae of Brachyura, probably Portumnus sp.). These taxa are thus proposed as new subjects for study, since they could be intermediate hosts in the infection process of mussels by Marteilia, This work was carried out with funding from an INIA predoctoral grant from the Spanish Government and a Marie Curie training site UE grant to N.C., and INIA project RTA04-023 co-ordinated by Dr M. Ramón. Also an IFAPA post-doctoral grant (Junta de Andalucia) to I.L. and MCYT grants CTM2004-02575/MAR and REN2002-04151-C02-01/MAR to M.A.

Marteilia refringens is a protozoan parasite recognized as a significant pathogen of the European flat oyster Ostrea edulis. It is believed to have a complex life-cycle involving several hosts. In this study, we applied molecular approaches to identify this parasite in samples of the razor clam Solen marginatus from the south west coast of Spain. We used a PCR assay to amplify a fragment of the IGS rDNA region. PCR products were sequenced and the phylogenetic affinity of the sequences was determined. In situ hybridization analysis showed tissue distribution and presence of different developmental stages of the parasite in the digestive diverticula epithelium, which suggested a true parasitism in these individuals. This is the first report of the occurrence of M. refringens in the razor clam S. marginatus in the south Atlantic. The methodology described herein may be useful for accurate identification of the parasite strain in different hosts and thus provide valuable information for marteiliosis control programmes.

Serine dehydratase (SerDH) is a gluconeogenic enzyme involved in the catabolism of serine, which is regulated by the composition of their diet and their hormonal status in rats. This study examines how chronic injury caused to the liver of rats by the ingestion of thioacetamide (TAA) affects SerDH protein, mRNA levels, enzyme kinetics and its tissue location. After 97 days’ oral intake of TAA, the activity of SerDH at all substrate concentrations assayed was about 60% lower than in controls. No significant differences in Km values were found between the treated group and controls. Immunoblotting and immunohistochemistry revealed a significant reduction in the level of SerDH protein in the livers of the treated rats. SerDH was detected specifically in the periportal zone of the hepatic acinus and this location did not change in response to TAA treatment. The level of SerDH mRNA, quantified by reverse transcription and polymerase chain reaction, was significantly lower in treated rats than in the controls. The present findings suggest that the SerDH expression is rendered to be down regulatory during chronic liver injury induced by TAA. These results enhance our understanding about the biochemical mechanisms implied in the control and integration of serine catabolism during liver injury in rat. (Mol Cell Biochem 268: 33–43, 2005)

Se ofrece en este trabajo el estudio de una tumba visigoda localizada en la Fase III de la Intervención Arqueológica en la Plaza de España de Écija (Sevilla), en la que confluye la información arqueológica con la antropológica y epigráfica. In this paper is offered the study of a visigothic tomb excavated in the 3rd phase of the archaeological excavations carried out in the Plaza de España of Ecija (Seville), in which the archaeological evidence meets with anthropological and epigraphic documentation.

We have analysed a centromeric satellite DNA family that is conserved in several commercial and non-commercial oyster species (Ostrea edulis, O. stentina, Crassostrea angulata, C. gigas, C. gasar, C. ariakensis, C. virginica and C. sikamea). This satellite DNA family is composed of AT-rich repeat sequences of 166+/-2 bp and presents a 9-bp motif similar to the mammalian CENP-B box. The homology of oyster HindIII satellite DNA with satellite DNAs from other bivalves and its relation to a part of a mobile element suggest the existence of an ancient transposable element as a generating unit of satellite DNA in bivalve molluscs. Taking advantage of its degree of conservation in oyster species, we have used this element as a taxonomic marker. This marker clearly supports a high degree of differentiation between O. edulis and O. stentina, and, conversely, upholds the contention that C. gigas and C. angulata are the same species. Finally, we have used HindIII satellite DNA as a phylogenetic marker between these species, revealing two clades, one formed by Asiatic species (C. angulata, C. gigas and C. ariakensis) and another by the European, American and African species (O. edulis, C. virginica and C. gasar, respectively).