Ectopic thyroid adenoma diagnosed in a Senegal seabream (Diplodus bellottii, Steindachner 1882): Towards an accurate diagnosis.

Thyroid tissue in teleosts is located mainly in the pharyngeal region, usually reaching other adjacent anatomical locations. Herein, a nodular lesion located in the left operculum of a Senegal seabream (Diplodus bellottii) was surgically excised and sent for microscopical evaluation. Microscopically, the lesion presented irregular borders and consisted in columnar epithelial cells arranged in a tubulopapillary pattern, surrounding a central lumen filled with acellular, acidophilic and homogeneous, material ('colloid'). To determine the lesion's histogenesis, immunohistochemistry was performed employing antibodies for AE1/AE3, CK7, thyroglobulin and vimentin. The neoplastic cells presented low mitotic index and positive immunolabelling for CK7 and thyroglobulin. Therefore, a diagnosis of ectopic thyroid adenoma was made. Herein, the successful employment of antibodies classically used in mammals for accurate diagnosis of thyroid disorders is described. Proliferation of thyroid tissue in fish may reflect environmental and physiological imbalances, making the study and correct diagnosis of these tumours in this species important.

Assessment of male reproductive traits in endangered leuciscids from the Iberian Peninsula: first attempts to store gametes both at short- and long-term.

During the spring of 2022, several endangered leuciscid species (Anaecypris hispanica, Squalius aradensis, Anachondrostoma Occidentale, and Iberochondrostoma lusitanicum) were sampled both at the Vasco da Gama aquarium facilities and in some rivers of the Algarve region, Portugal. Sperm samples were extracted by gentle abdominal pressure and sperm motion parameters were assessed for the first time in four species, using a computerized analysis system. The results obtained showed that spermatozoa kinetic patterns were similar for all 4 species, with high motility and velocity values after the sperm activation time and with a marked decrease after 20. On the other hand, sperm longevity was highly variable between species, with short longevities (around 40 s) for A. hispanica and S. aradensis, and longer longevities (100-120 s) for A. occidentale and I. lusitanicum, which could indicate a latitudinal pattern in terms of sperm longevity. At the same time, morphometric analysis was carried out for the four target species, revealing that spermatozoa showed similar sizes and shapes to other external fertilizers belonging to Leuscididae, with small spherical heads, uniflagellate, and without acrosomes. In addition, a short-term gamete storage trail was performed by diluting sperm in 1:9 (sperm:extender) and storing them at 4ºC. Although the results obtained were uneven among the species studied, the dilution and extender used generated motilities above 40% up to day 4 of storage in S. aradensis and I. lusitanicum, and up to days 1-2 in A. hispanica and A. occidentale, respectively. Finally, gamete cryopreservation trials were also carried out on these threatened species. Although cryopreserved samples showed significantly lower motility than fresh samples, some protocols generate acceptable percentages of viability, DNA integrity, and sperm motility in some species such as I. lusitanicum and A. occidentale. The data revealed that the protocol based on 10% DMSO plus 7.5% egg yolk generated the best results.This study is the first to assess the reproductive traits of wild and captive populations of endangered leuciscids endemic from the Iberian Peninsula, describing the spermatozoa kinetics and developing protocols for managing male gametes both in short- and long-term storage. Outcomes will provide new and useful tools to complement the management and conservation of ex situ breeding programs that are being developed for these four endangered species.

Comparative analysis of transcriptomic responses to sub-lethal levels of six environmentally relevant pesticides in Saccharomyces cerevisiae.

Accidental spills and misuse of pesticides may lead to current and/or legacy environmental contamination and may pose concerns regarding possible risks towards non-target microbes and higher eukaryotes in ecosystems. The present study was aimed at comparing transcriptomic responses to effects of sub-lethal levels of six environmentally relevant pesticide active substances in the Saccharomyces cerevisiae eukaryotic model. The insecticide carbofuran, the fungicide pyrimethanil and the herbicides alachlor, S-metolachlor, diuron and methyl(4-chloro-2-methylphenoxy)acetate were studied. Some are currently used agricultural pesticides, while others are under restricted utilization or banned in Europe and/or North America albeit being used in other geographical locations. In the present work transcriptional profiles representing genome-wide responses in a standardized yeast population upon 2 h of exposure to concentrations of each compound exerting equivalent toxic effects, i.e., inhibition of growth by 20% relative to the untreated control cells, were examined. Hierarchical clustering and Venn analyses of the datasets of differentially expressed genes pointed out transcriptional patterns distinguishable between the six active substances. Functional enrichment analyses allowed predicting mechanisms of pesticide toxicity and response to pesticide stress in the yeast model. In general, variations in transcript numbers of selected genes assessed by Real-Time quantitative reverse transcription polymerase chain reaction confirmed microarray data and correlated well with growth inhibitory effects. A possible biological relevance of mechanistic predictions arising from these comparative transcriptomic analyses is discussed in the context of better understanding potential modes of action and adverse side-effects of pesticides.

The Saccharomyces cerevisiae response to stress caused by the herbicidal active substance alachlor requires the iron regulon transcription factor Aft1p.

In the Saccharomyces cerevisiae eukaryotic model, the induction of the iron regulon genes ARN1, FIT2 and CTH2 by growth-inhibitory concentrations of alachlor (ALA) was dependent on Aft1p expression. This transcription factor was found to be activated through its nuclear localization. The hypersensitivity of the aft1Δ mutant to ALA was abrogated by surplus exogenous iron, suggesting that the role of Aft1p in ALA tolerance may be associated with iron limitation under ALA stress. A transient decrease in the cellular iron content in the ALA-stressed cells supported this idea. In contrast to the upregulation of the nonreductive iron uptake genes ARN1 and FIT2 by ALA, the quantity of FET3 and FTR1 transcripts encoding the high-affinity iron uptake reductive pathway decreased. Yeast cells were apparently more sensitive to ALA when iron uptake occurred through the reductive pathway than when the nonreductive uptake of ferrichrome-bound ferric iron was dominant. On the other hand, the ALA hypersensitivity of the aft1Δ mutant was reversed by medium supplementation with glutathione or N-acetyl-L-cysteine. The results are compatible with possible links between ALA toxicity and perturbations in metal and antioxidant homeostasis, which may be relevant for environmental microbes and higher eukaryotes in situations of inadvertent herbicide contamination.

Validity and reliability of the Berlin questionnaire for the detection of moderate or severe obstructive sleep apnea in patients aged 40 years or older detected from primary care.

Background: The obstructive sleep apnea syndrome (OSA) is a highly prevalent condition. In Spain and other countries, only 5%-9% of patients with OSA have been diagnosed and treated. The lack of accessibility to diagnosis is considered the main cause of this situation through easy-to-use screening instruments, it is necessary to check their validity and reliability in the context where they are to be used. Objective: To validate the Spanish translation of the Berlin questionnaire for screening for moderate or severe OSA in patients aged 40 years or more detected in primary care. Methods: A descriptive observational study, with a first qualitative phase of transcultural adaptation to Spanish using the translation-back-translation method. Setting: primary care level of the Spanish National Health System. A total of 255 patients recruited from 7 healthcare centers completed the study. The Berlin questionnaire was administered to the recruited patients, and subsequently, a respiratory polygraphy was performed to confirm the diagnosis of OSA. The concurrent criterion validity of the questionnaire and its reliability in terms of internal consistency and reproducibility (intra-observer agreement) were analyzed. Results: The patients' mean age was 54.76 years (SD: 6.57; 95% CI: 53.53-54.99), and 54.12% were men (95% CI: 47.96-60.27). We found that 61.57% (95% CI: 55.57-67.57) presented OSA (apnea-hypopnea index-AHI >5), and 45.5% (95% CI: 17.05-57.92) presented moderate or severe (AHI >15) OSA. The Berlin questionnaire, with a cut-off point of 4.5, showed a sensitivity of 76.77% (95% CI: 67.94-85.59), a specificity of 74.49% (95% CI: 65.35-83.63), a positive predictive value of 75.25% (95% CI: 66.34-84.16), a negative predictive value of 76.04% (95% CI: 66.98-85.10), and an area under the curve of 0.786 (95% CI: 0.721-0.851). Cronbach's alpha coefficient was 0.730 (95% CI: 0.668-0.784), and the Kappa index was 0.739 (95% CI, 0.384-1.000). Conclusion: The Spanish adaptation of the Berlin questionnaire has good validity and reliability as a test for the diagnostic screening of moderate or severe OSA in patients aged 40 years or older. The findings of our study confirm that primary care physicians should use such screening tools to predict OSA.

Aeromonas spp. Prevalence, Virulence, and Antimicrobial Resistance in an Ex Situ Program for Threatened Freshwater Fish-A Pilot Study with Protective Measures.

Ex situ breeding programs are important conservation tools for endangered freshwater fish. However, developing husbandry techniques that decrease the likelihood of disease, antimicrobial resistance, and virulence determinants acquisition during this process is challenging. In this pilot study, we conducted a captivity experiment with Portuguese nase (Iberochondrostoma lusitanicum), a critically endangered leuciscid species, to investigate the influence of simple protective measures (i.e., material disinfection protocols and animal handling with gloves) on the dynamics of a potential pathogenic genus, Aeromonas, as well as its virulence profiles and antimicrobial resistance signatures. Our findings show that antimicrobial resistance in Aeromonas spp. collected from I. lusitanicum significantly increased during the extent of the assay (5 weeks), with all isolates collected at the end of the study classified as multidrug-resistant. Additionally, humans handling fishes without protective measures were colonized by Aeromonas spp. The use of protective measures suggested a decreasing trend in Aeromonas spp. prevalence in I. lusitanicum, while bacterial isolates displayed significantly lower virulence index values when virulence phenotypical expression was tested at 22 °C. Despite this study representing an initial trial, which needs support from further research, protective measures tested are considered a simple tool to be applied in ex situ breeding programs for aquatic animals worldwide. Furthermore, current results raise concern regarding antimicrobial resistance amplification and zoonotic transmission of Aeromonas spp. in aquatic ex situ programs.

Gadolinium ecotoxicity is enhanced in a warmer and acidified changing ocean as shown by the surf clam Spisula solida through a multibiomarker approach.

Humans have exhaustively combusted fossil fuels, and released pollutants into the environment, at continuously faster rates resulting in global average temperature increase and seawater pH decrease. Climate change is forecasted to exacerbate the effects of pollutants such as the emergent rare earth elements. Therefore, the objective of this study was to assess the combined effects of rising temperature (Δ = + 4 °C) and decreasing pH (Δ = - 0.4 pH units) on the bioaccumulation and elimination of gadolinium (Gd) in the bioindicator bivalve species Spisula solida (Surf clam). We exposed surf clams to 10 µg L-1 of GdCl3 for seven days, under warming, acidification, and their combination, followed by a depuration phase lasting for another 7 days and investigated the Gd bioaccumulation and oxidative stress-related responses after 1, 3 and 7 days of exposure and the elimination phase. Gadolinium accumulated after just one day with values reaching the highest after 7 days. Gadolinium was not eliminated after 7 days, and elimination is further hampered under climate change scenarios. Warming and acidification, and their interaction did not significantly impact Gd concentration. However, there was a significant interaction on clam's biochemical response. The augmented total antioxidant capacity and lipid peroxidation values show that the significant impacts of Gd on the oxidative stress response are enhanced under warming while the increased superoxide dismutase and catalase values demonstrate the combined impact of Gd, warming & acidification. Ultimately, lipid damage was greater in clams exposed to warming & Gd, which emphasizes the enhanced toxic effects of Gd in a changing ocean.

Single and combined ecotoxicological effects of ocean warming, acidification and lanthanum exposure on the surf clam (Spisula solida).

Lanthanum (La) is one of the most abundant emergent rare earth elements. Its release into the environment is enhanced by its use in various industrial applications. In the aquatic environment, emerging contaminants are one of the stressors with the ability to compromise the fitness of its inhabitants. Warming and acidification can also affect their resilience and are another consequence of the growing human footprint on the planet. However, from information gathered in the literature, a study on the effects of ocean warming, acidification, and their interaction with La was never carried out. To diminish this gap of knowledge, we explored the effects, combined and as single stressors, of ocean warming, acidification, and La (15 µg L-1) accumulation and elimination on the surf clam (Spisula solida). Specimens were exposed for 7 days and depurated for an additional 7-day period. Furthermore, a robust set of membrane-associated, protein, and antioxidant enzymes and non-enzymatic biomarkers (LPO, HSP, Ub, SOD, CAT, GPx, GST, TAC) were quantified. Lanthanum was bioaccumulated after just one day of exposure, in both control and climate change scenarios. A 7-day depuration phase was insufficient to achieve control values and in a warming scenario, La elimination was more efficient. Biochemical response was triggered, as highlighted by enhanced SOD, CAT, GST, and TAC levels, however as lipoperoxidation was observed it was insufficient to detoxify La and avoid damage. The HSP was largely inhibited in La treatments combined with warming and acidification. Concomitantly, lipoperoxidation was highest in clams exposed to La, warming, and acidification combined. The results highlight the toxic effects of La on this bivalve species and its enhanced potential in a changing world.

The usefulness of the Spanish version of the STOP-Bang questionnaire for screening for moderate or severe sleep apnea syndrome in primary care.

Rationale: Sleep apnea-hypopnea syndrome (OSA) is a highly prevalent disease and has been related to cardiovascular diseases and occupational and traffic accidents. Currently, it is estimated that there is a significant underdiagnosis of OSA, mainly due to the difficulty accessing the tests for that purpose. Objective: To determine the usefulness of the Spanish version of the STOP-Bang questionnaire (SBQ) for screening for moderate or severe OSA in the adult population attending primary care. Methods: A descriptive observational multicenter study was conducted. Through an opportunistic search, (patients over 18 years old), were recruited in seven primary care centers. The SBQ was applied to them and home respiratory polygraphy (HRP) was subsequently performed to confirm the diagnosis of OSA. The criterion validity of the SBQ was analyzed, comparing the score obtained by the SBQ with the apnea-hypopnea index (AHI) obtained by RP, establishing the diagnosis of OSA for an AHI>5. The reliability of the questionnaire was evaluated. Results: A total of 255 subjects, 54.1% men, with a mean age of 54.76 ± 10 years, were recruited in the study. The results showed that 61.57% (95% Confidence Interval: 55.57-67.57) of the subjects presented OSA, presenting 22.75% (17.57-57.92) a mild OSA (530) (11.54-20.62). The Kuder and Richardson coefficient was 0.623 (0.335-0.788) and Cohen's Kappa coefficient was 0.871 (0.520-1.00; p < 0.001). For moderate/severe OSA screening (AHI>15) the SBQ obtained an ROC curve of 0.769 (0.704-0.833) that with an optimal cutoff of 3, achieved a sensitivity of 84.85% (77.28-92.42) and a specificity of 55.10% (44.74-65.46). Conclusions: The SBQ is very effective for detecting moderate/severe OSA. Its psychometric properties are similar to those obtained in studies on other populations. Because of its ease of use, the SBQ is a very useful tool for primary health care professionals.

Suitability of a Saccharomyces cerevisiae-based assay to assess the toxicity of pyrimethanil sprayed soils via surface runoff: comparison with standard aquatic and soil toxicity assays.

The present study is aimed at evaluating whether a gene expression assay with the microbial eukaryotic model Saccharomyces cerevisiae could be used as a suitable warning tool for the rapid preliminary screening of potential toxic effects on organisms due to scenarios of soil and water contamination with pyrimethanil. The assay consisted of measuring changes in the expression of the selected pyrimethanil-responsive genes ARG3 and ARG5,6 in a standardized yeast population. Evaluation was held by assessing the toxicity of surface runoff, a major route of pesticide exposure in aquatic systems due to non-point-source pollution, which was simulated with a pyrimethanil formulation at a semifield scale mimicking worst-case scenarios of soil contamination (e.g. accident or improper disposal). Yeast cells 2-h exposure to the runoff samples led to a significant 2-fold increase in the expression of both indicator genes. These results were compared with those from assays with organisms relevant for the aquatic and soil compartments, namely the nematode Caenorhabditis elegans (reproduction), the freshwater cladoceran Daphnia magna (survival and reproduction), the benthic midge Chironomus riparius (growth), and the soil invertebrates Folsomia candida and Enchytraeus crypticus (survival and reproduction). Under the experimental conditions used to simulate accidental discharges into soil, runoff waters were highly toxic to the standard test organisms, except for C. elegans. Overall, results point out the usefulness of the yeast assay to provide a rapid preview of the toxicity level in preliminary screenings of environmental samples in situations of inadvertent high pesticide contamination. Advantages and limitations of this novel method are discussed.

Potential mechanisms underlying response to effects of the fungicide pyrimethanil from gene expression profiling in Saccharomyces cerevisiae.

Pyrimethanil is a fungicide mostly applied in vineyards. When misused, residue levels detected in grape must or in the environment may be of concern. The present work aimed to analyze mechanisms underlying response to deleterious effects of pyrimethanil in the eukaryotic model Saccharomyces cerevisiae. Pyrimethanil concentration-dependent effects at phenotypic (inhibition of growth) and transcriptomic levels were examined. For transcriptional profiling, analysis focused on two sublethal exposure conditions that inhibited yeast growth by 20% or 50% compared with control cells not exposed to the fungicide. Gene expression modifications increased with the magnitude of growth inhibition, in numbers and fold-change of differentially expressed genes and in diversity of over-represented functional categories. These included mostly biosynthesis of arginine and sulfur amino acids metabolism, as well as energy conservation, antioxidant response, and multidrug transport. Several pyrimethanil-responsive genes encoded proteins sharing significant homology with proteins from phytopathogenic fungi and ecologically relevant higher eukaryotes.

Transcriptional profiling in Saccharomyces cerevisiae relevant for predicting alachlor mechanisms of toxicity.

Alachlor has been a commonly applied herbicide and is a substance of ecotoxicological concern. The present study aims to identify molecular biomarkers in the eukaryotic model Saccharomyces cerevisiae that can be used to predict potential cytotoxic effects of alachlor, while providing new mechanistic clues with possible relevance for experimentally less accessible eukaryotes. It focuses on genome-wide expression profiling in a yeast population in response to two exposure scenarios exerting effects from slight to moderate magnitude at phenotypic level. In particular, 100 and 264 genes, respectively, were found as differentially expressed on a 2-h exposure of yeast cells to the lowest observed effect concentration (110 mg/L) and the 20% inhibitory concentration (200 mg/L) of alachlor, in comparison with cells not exposed to the herbicide. The datasets of alachlor-responsive genes showed functional enrichment in diverse metabolic, transmembrane transport, cell defense, and detoxification categories. In general, the modifications in transcript levels of selected candidate biomarkers, assessed by quantitative reverse transcriptase polymerase chain reaction, confirmed the microarray data and varied consistently with the growth inhibitory effects of alachlor. Approximately 16% of the proteins encoded by alachlor-differentially expressed genes were found to share significant homology with proteins from ecologically relevant eukaryotic species. The biological relevance of these results is discussed in relation to new insights into the potential adverse effects of alachlor in health of organisms from ecosystems, particularly in worst-case situations such as accidental spills or careless storage, usage, and disposal.

Contribuiçäo ao conhecimento de plantas medicinais brasileiras: Gromphrena officinalis, Mart.: Amaranthaceae / Knowledge of Brazilian medicial plants: Gromphrena officinalis, Mart: Amaranthaceae

Sendo a Gomphrena officinalis Martius - Amaranthaceae uma controvertida planta usada como panacéia em Medicina popular. O material de ensaio determinado por Calos Rizzini foi coletado e preservado também herborizado; na Reserva biológica I.B.G.E. em Brasília D.F. foi feito estudo botânico como meio de auxiliar a Farmacognósia, no estudo anatômico abordam-se a sede morfogenética, ou seja a folha, limbo, mesófilo, bordo, ápice, pecíolo, bem como nervuras, cutícula e particularmente tricomas. No estudo farmacoquímico foram procurados: heterosídios antociânicos, saponinas, taninos, antraquinonas, flavonóides, osídios redutores, alcalóides e bases orgânicas. Encontrou-se grande quantidade de saponinas e discreta de açúcares, antacianas, tanóides e flavonóides. Nos ensaios biológicos näo se achou açäo antiviral nem antibiótica, somente pequena açäo hemolítica