The Microcystis-microbiome interactions: origins of the colonial lifestyle.

Species of the Microcystis genus are the most common bloom-forming toxic cyanobacteria worldwide. They belong to a clade of unicellular cyanobacteria whose ability to reach high biomasses during blooms is linked to the formation of colonies. Colonial lifestyle provides several advantages under stressing conditions of light intensity, ultraviolet light, toxic substances and grazing. The progression from a single-celled organism to multicellularity in Microcystis has usually been interpreted as individual phenotypic responses of the cyanobacterial cells to the environment. Here, we synthesize current knowledge about Microcystis colonial lifestyle and its role in the organism ecology. We then briefly review the available information on Microcystis microbiome and propose that changes leading from single cells to colonies are the consequence of specific and tightly regulated signals between the cyanobacterium and its microbiome through a biofilm-like mechanism. The resulting colony is a multi-specific community of interdependent microorganisms.

Lotka-Volterra model with Allee effect: equilibria, coexistence and size scaling of maximum and minimum abundance.

The Lotka-Volterra competition model (LVCM) is a fundamental tool for ecology, widely used to represent complex communities. The Allee effect (AE) is a phenomenon in which there is a positive correlation between population density and fitness, at low population densities. However, the interplay between the LVCM and AE has been seldom analyzed in multispecies models. Here, we analyze the mathematical properties of the LVCM [Formula: see text] AE, investigating the coexistence of species interacting through neutral diffuse competition, their equilibria and stable points. Minimum viable population density arises as the threshold below which species go extinct, characteristic of strong Allee effects. Then, by imposing relationships of main parameters to body size, i.e. allometric scaling, we derive a general solution to the size-scaling maximum and minimum expected density under plausible scenarios. The scaling of maximum population density is consistent with the literature, but we also provide novel predictions on the scaling of the lower limit to population density, a critical value for conservation science. The resulting framework is general and yields results that increase our current understanding of how complex demographic processes can be linked to ubiquitous ecological patterns.

Effect of hydrological modification on the potential toxicity of Microcystis aeruginosa complex in Salto Grande reservoir, Uruguay.

It is widely known that the environmental conditions caused by the construction of reservoirs favor the proliferation of toxic cyanobacteria and the formation of blooms due to the high residence time of the water, low turbidity, temperature regimes, among others. Microcystin-producing cyanobacteria such as those from the Microcystis aeruginosa complex (MAC) are the most frequently found organisms in reservoirs worldwide, being the role of the environment on microcystin production poorly understood. Here, we addressed the community dynamics and potential toxicity of MAC cyanobacteria in a subtropical reservoir (Salto Grande) located in the low Uruguay river. Samples were taken from five different sites (upstream, inside the reservoir and downstream) during contrasting seasons (summer and winter) to analyze: (i) the MAC community structure by amplicon sequencing of the phycocyanin gene spacer, (ii) the genotype diversity of microcystin-producing MAC by high resolution melting analysis of the mcyJ gene, and (iii) the abundance and mcy transcription activity of the microcystin-producing (toxic) fraction. We found that MAC diversity decreased from summer to winter but, despite the observed changes in MAC community structure, the abundance of toxic organisms and the transcription of mcy genes were always higher inside the reservoir, regardless of the season. Two different genotypes of toxic MAC were detected inside the reservoir, one associated with low water temperature (15 °C) and one thriving at high water temperature (31 °C). These findings indicate that the environmental conditions inside the reservoir reduce community diversity while promoting the proliferation of toxic genotypes that actively transcribe mcy genes, whose relative abundance will depend on the water temperature.

Rise of toxic cyanobacterial blooms is promoted by agricultural intensification in the basin of a large subtropical river of South America.

Toxic cyanobacterial blooms are globally increasing with negative effects on aquatic ecosystems, water use and human health. Blooms' main driving forces are eutrophication, dam construction, urban waste, replacement of natural vegetation with croplands and climate change and variability. The relative effects of each driver have not still been properly addressed, particularly in large river basins. Here, we performed a historical analysis of cyanobacterial abundance in a large and important ecosystem of South America (Uruguay river, ca 1900 km long, 365,000 km2 basin). We evaluated the interannual relationships between cyanobacterial abundance and land use change, river flow, urban sewage, temperature and precipitation from 1963 to the present. Our results indicated an exponential increase in cyanobacterial abundance during the last two decades, congruent with an increase in phosphorus concentration. A sharp shift in the cyanobacterial abundance rate of increase after the year 2000 was identified, resulting in abundance levels above public health alert since 2010. Path analyses showed a strong positive correlation between cyanobacteria and cropland area at the entire catchment level, while precipitation, temperature and water flow effects were negligible. Present results help to identify high nutrient input agricultural practices and nutrient enrichment as the main factors driving toxic bloom formation. These practices are already exerting severe effects on both aquatic ecosystems and human health and projections suggest these trends will be intensified in the future. To avoid further water degradation and health risk for future generations, a large-scale (transboundary) change in agricultural management towards agroecological practices will be required.

Las floraciones de cianobacterias tóxicas vienen aumentando drásticamente a nivel mundial con efectos negativos en los ecosistemas acuáticos, los usos del agua y la salud humana. Los principales mecanismos promotores de las floraciones son la eutrofización, la construcción de represas, la contaminación con residuos urbanos, la pérdida de vegetación natural y el cambio y la variabilidad climáticos. Los efectos relativos de cada determinante aún no se han abordado adecuadamente, particularmente en las grandes cuencas fluviales de América del Sur. En este trabajo, realizamos un análisis histórico de la abundancia de cianobacterias en un gran e importante ecosistema de América del Sur (el Río Uruguay, c.a. 1.900 km de largo, cuenca de 365.000 km2). Evaluamos las relaciones entre la abundancia de cianobacterias y el cambio en los usos del suelo, el caudal de los ríos, la contaminación urbana, la temperatura y la precipitación desde 1963 hasta el presente. Nuestros resultados evidencian un aumento exponencial en la abundancia de cianobacterias durante las últimas dos décadas, de forma congruente con el aumento en la concentración de fósforo en agua. Fue identificado además, un cambio brusco en la tasa de aumento de la abundancia de cianobacterias después del año 2000, lo que resultó en niveles de alerta por encima de riesgo para la salud pública desde 2010. Los análisis estadísticos indicaron una fuerte y positiva correlación entre las cianobacterias y el área de cultivo en la cuenca, mientras que la precipitación, la temperatura y el flujo de agua fueron insignificantes. Estos resultados contribuyen a identificar que las prácticas agrícolas con alto aporte de nutrientes y el enriquecimiento de nutrientes son los principales impulsores de la formación de floraciones tóxicas. Estas prácticas ya están teniendo graves efectos en los ecosistemas acuáticos y la salud humana y las proyecciones sugieren que se intensificarán en el futuro. Para evitar una mayor degradación de la calidad del agua y el incremento de los riesgos para la salud de las generaciones futuras, se requerirá un cambio a gran escala (transfronterizo) en la gestión agrícola hacia prácticas agroecológicas.

Initial clinical and treatment patterns of advanced differentiated thyroid cancer: ERUDIT study.

Background: Up to 30% of differentiated thyroid cancer (DTC) will develop advanced-stage disease (aDTC) with reduced overall survival (OS). Objective: The aim of this study is to characterize initial diagnosis of aDTC, its therapeutic management, and prognosis in Spain and Portugal. Methods: A multicentre, longitudinal, retrospective study of adult patients diagnosed with aDTC in the Iberian Peninsula was conducted between January 2007 and December 2012. Analyses of baseline characteristics and results of initial treatments, relapse- or progression-free survival ((RP)FS) from first DTC diagnosis, OS, and prognostic factors impacting the evolution of advanced disease were evaluated. Results: Two hundred and thirteen patients (median age: 63 years; 57% female) were eligible from 23 hospitals. Advanced disease presented at first diagnosis (de novo aDTC) included 54% of patients, while 46% had relapsed from early disease (recurrent/progressive eDTC). At initial stage, most patients received surgery (98%) and/or radioiodine (RAI) (89%), with no differences seen between median OS (95% CI) (10.4 (7.3-15.3) years) and median disease-specific-survival (95% CI) (11.1 (8.7-16.2) years; log-rank test P = 0.4737). Age at diagnosis being <55 years was associated with a lower risk of death (Wald chi-square (Wc-s) P < 0.0001), while a poor response to RAI to a higher risk of death ((Wc-s) P < 0.05). In the eDTC cohort, median (RP)FS (95% CI) was of 1.7 (1.0-2.0) years after RAI, with R0/R1 surgeries being the only common significant favourable factor for longer (RP)FS and time to aDTC ((Wc-s) P < 0.05). Conclusion: Identification of early treatment-dependent prognostic factors for an unfavourable course of advanced disease is possible. An intensified therapeutic attitude may reverse this trend and should be considered in poor-performing patients. Prospective studies are required to confirm these findings.

Peptide Receptor Radionuclide Therapy with [177Lu]Lu-DOTA-TATE in Patients with Advanced GEP NENS: Present and Future Directions.

This review article summarizes findings published in the last years on peptide receptor radionuclide therapy in GEP NENs, as well as potential future developments and directions. Unanswered questions remain, such as the following: Which is the correct dose and individual dosimetry? Which is the place for salvage PRRT-Lu? Whicht is the role of PRRT-Lu in the pediatric population? Which is the optimal sequencing of PRRT-Lu in advanced GEP NETs? Which is the place of PRRT-Lu in G3 NENs? These, and future developments such as inclusion new radiopharmaceuticals and combination therapy with different agents, such as radiosensitizers, will be discussed.

External Validity of Somatostatin Analogs Trials in Advanced Neuroendocrine Neoplasms: The GETNE-TRASGU Study.

INTRODUCTION: Somatostatin analogs (SSA) prolong progression-free survival (PFS) in patients with well-differentiated gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs). However, the eligibility criteria in randomized clinical trials (RCTs) have been restricted, which contrasts with the vast heterogeneity found in NENs. METHODS: We identified patients with well-differentiated (Ki-67% ≤20%), metastatic GEP-NENs treated in first line with SSA monotherapy from the Spanish R-GETNE registry. The therapeutic effect was evaluated using a Bayesian Cox model. The objective was to compare survival-based outcomes from real-world clinical practice versus RCTs. RESULTS: The dataset contained 535 patients with a median age of 62 years (range: 26-89). The median Ki-67% was 4 (range: 0-20). The most common primary tumor sites were as follows: midgut, 46%; pancreas, 34%; unknown primary, 10%; and colorectal, 10%. Half of the patients received octreotide LAR (n = 266) and half, lanreotide autogel (n = 269). The median PFS was 28.0 months (95% CI: 22.1-32.0) for octreotide versus 30.1 months (95% CI: 23.1-38.0) for lanreotide. The overall hazard ratio for lanreotide versus octreotide was 0.90 (95% credible interval: 0.71-1.12). The probability of effect sizes >30% with lanreotide versus octreotide was 2 and 6% for midgut and foregut NENs, respectively. CONCLUSION: Our study evaluated the external validity of RCTs examining SSAs in the real world, as well as the main effect-modifying factors (progression status, symptoms, tumor site, specific metastases, and analytical data). Our results indicate that both octreotide LAR and lanreotide autogel had a similar effect on PFS. Consequently, both represent valid alternatives in patients with well-differentiated, metastatic GEP-NENs.

Genotyping and Multivariate Regression Trees Reveal Ecological Diversification within the Microcystis aeruginosa Complex along a Wide Environmental Gradient.

Addressing the ecological and evolutionary processes underlying biodiversity patterns is essential to identify the mechanisms shaping community structure and function. In bacteria, the formation of new ecologically distinct populations (ecotypes) is proposed as one of the main drivers of diversification. New ecotypes arise when mutations in key functional genes or acquisition of new metabolic pathways by horizontal gene transfer allow the population to exploit new resources, permitting their coexistence with the parental population. We previously reported the presence of microcystin-producing organisms of the Microcystis aeruginosa complex (toxic MAC) through an 800-km environmental gradient ranging from freshwater to estuarine-marine waters in South America. We hypothesize that the success of toxic MAC in such a gradient is due to the existence of very closely related populations that are ecologically distinct (ecotypes), each specialized to a specific arrangement of environmental variables. Here, we analyzed toxic MAC genetic diversity through quantitative PCR (qPCR) and high-resolution melting analysis (HRMA) of a functional gene (mcyJ, microcystin synthetase cluster). We explored the variability of the mcyJ gene along the environmental gradient by multivariate classification and regression trees (mCART). Six groups of mcyJ genotypes were distinguished and associated with different combinations of water temperature, conductivity, and turbidity. We propose that each mcyJ variant associated with a defined environmental condition is an ecotype (or species) whose relative abundances vary according to their fitness in the local environment. This mechanism would explain the success of toxic MAC in such a wide array of environmental conditions. IMPORTANCE Organisms of the Microcystis aeruginosa complex form harmful algal blooms (HABs) in nutrient-rich water bodies worldwide. MAC HABs are difficult to manage owing to the production of potent toxins (microcystins) that resist water treatment. In addition, the role of microcystins in the ecology of MAC organisms is still elusive, meaning that the environmental conditions driving the toxicity of the bloom are not clear. Furthermore, the lack of coherence between morphology-based and genomic-based species classification makes it difficult to draw sound conclusions about when and where each member species of the MAC will dominate the bloom. Here, we propose that the diversification process and success of toxic MAC in a wide range of water bodies involves the generation of ecotypes, each specialized in a particular niche, whose relative abundance varies according to its fitness in the local environment. This knowledge can improve the generation of accurate prediction models of MAC growth and toxicity, helping to prevent human and animal intoxication.

Machine learning methods for imbalanced data set for prediction of faecal contamination in beach waters.

Predicting water contamination by statistical models is a useful tool to manage health risk in recreational beaches. Extreme contamination events, i.e. those exceeding normative are generally rare with respect to bathing conditions and thus the data is said to be imbalanced. Modeling and predicting those rare events present unique challenges. Here we introduce and evaluate several machine learning techniques and metrics to model imbalanced data and evaluate model performance. We do so by using a) simulated data-sets and b) a real data base with records of faecal coliform abundance monitored for 10 years in 21 recreational beaches in Uruguay (N ≈ 19000) using in situ and meteorological variables. We discuss advantages and disadvantages of the methods and provide a simple guide to perform models for a general audience. We also provide R codes to reproduce model fitting and testing. We found that most Machine Learning techniques are sensitive to imbalance and require specific data pre-treatment (e.g. upsampling) to improve performance. Accuracy (i.e. correctly classified cases over total cases) is not adequate to evaluate model performance on imbalanced data set. Instead, true positive rates (TPR) and false positive rates (FPR) are recommended. Among the 52 possible candidate algorithms tested, the stratified Random forest presented the better performance improving TPR in 50% with respect to baseline (0.4) and outperformed baseline in the evaluated metrics. Support vector machines combined with upsampling method or synthetic minority oversampling technique (SMOTE) performed well, similar to Adaboost with SMOTE. These results suggests that combining modeling strategies is necessary to improve our capacity to anticipate water contamination and avoid health risk.

Prevalence and Clinicopathological Characteristics of Moderate and High-Penetrance Genes in Non-BRCA1/2 Breast Cancer High-Risk Spanish Families.

(1) Background: Over the last decade, genetic counseling clinics have moved from single-gene sequencing to multigene panel sequencing. Multiple genes related to a moderate risk of breast cancer (BC) have emerged, although many questions remain regarding the risks and clinical features associated with these genes. (2) Methods: Ninety-six BC index cases (ICs) with high-risk features for hereditary breast and ovarian cancer (HBOC) and with a previous uninformative result for BRCA1/2 were tested with a panel of 41 genes associated with BC risk. The frequency of pathogenic variants (PVs) was related to the clinical characteristics of BC. (3) Results: We detected a PV rate of 13.5% (excluding two cases each of BRCA1 and MUTYH). Among the 95 assessed cases, 17 PVs were identified in 16 ICs, as follows: BRCA1 (n = 2), CHEK2 (n = 3), ATM (n = 5), MUTYH (n = 2), TP53 (n = 2), BRIP1 (n = 1), CASP8 (n = 1), and MSH2 (n = 1). We also identified a novel loss-of-function variant in CASP8, a candidate gene for increased BC risk. There was no evidence that the clinical characteristics of BC might be related to a higher chance of identifying a PV. (4) Conclusions: In our cohort, which was enriched with families with a high number of BC cases, a high proportion of mutations in ATM and CHEK2 were identified. The clinical characteristics of BC associated with moderate-risk genes were different from those related to BRCA1/2 genes.

Co-occurrence of germline pathogenic variants for different hereditary cancer syndromes in patients with Lynch syndrome.

BACKGROUND: Lynch syndrome (LS) is a hereditary condition characterized by a high risk of colorectal cancer, endometrial cancer, and other neoplasia associated with germline alterations in DNA mismatch repair genes. The classical genetic diagnostic strategy for LS consists of the Sanger sequencing of genes associated with the suspected syndrome. Next-generation sequencing (NGS) enables the simultaneous sequencing of a large number of hereditary cancer genes. Here, we aimed to study whether other germline pathogenic variants of hereditary cancer genes are present in patients with LS. METHODS: A cohort of 84 probands with a previous genetic diagnosis of LS by Sanger sequencing was reanalyzed using NGS via a commercial panel of 94 hereditary cancer genes by hybrid capture. The American College of Medical Genetics and Genomics criteria were used to classify the clinical significance of the variants. The findings of NGS were confirmed by Sanger sequencing. When possible, genetic analyses of the new findings in the proband's relatives were also performed by Sanger sequencing. RESULTS: We identified five families (6%), out of 84, with at least two germline pathogenic variants conferring to high or moderate risk in different dominant cancer-predisposing genes: [MLH1-BRCA2-NBN], [MLH1-BRCA1], [MSH2-ATM], [MSH6-NF1], and [MLH1-FANCA]. Interestingly, only one out of these five families exhibited a clinical phenotype associated with the new pathogenic variants. The family with three pathogenic variants of the [MLH1-BRCA2-NBN] genes showed a high aggregation of tumors associated with LS and breast and ovarian cancer syndrome. CONCLUSIONS: Our results showed that the co-occurrence of more than one pathogenic variant in cancer-predisposing genes was remarkable among cases of LS. In most cases, no clinicial manifestations were associated with the secondary pathogenic variants. Further studies are needed to confirm these findings and elucidate their clinical impact. Reanalysis of LS families should be considered only in families with mixed clinical phenotypes.

Physician-perceived utility of the EORTC QLQ-GINET21 questionnaire in the treatment of patients with gastrointestinal neuroendocrine tumours: a multicentre, cross-sectional survey (QUALINETS).

BACKGROUND AND OBJECTIVE: Patient-reported outcome measures can provide clinicians with valuable information to improve doctor-patient communication and inform clinical decision-making. The aim of this study was to evaluate the physician-perceived utility of the QLQ-GINET21 in routine clinical practice in patients with gastrointestinal neuroendocrine tumours (GI-NETs). Secondary aims were to explore the patient, clinician, and/or centre-related variables potentially associated with perceived clinical utility. METHODS: Non-interventional, cross-sectional, multicentre study conducted at 34 hospitals in Spain and Portugal (NCT02853422). Patients diagnosed with GI-NETs completed two health-related quality of life (HRQoL) questionnaires (QLQ-C30, QLQ-GINET21) during a single routine visit. Physicians completed a 14-item ad hoc survey to rate the clinical utility of QLQ-GINET21 on three dimensions: 1)therapeutic and clinical decision-making, 2)doctor-patient communication, 3)questionnaire characteristics. RESULTS: A total of 199 patients at 34 centres were enrolled by 36 participating clinicians. The highest rated dimension on the QLQ-GINET21 was questionnaire characteristics (86.9% of responses indicating "high utility"), followed by doctor-patient communication (74.4%), and therapeutic and clinical decision-making (65.8%). One physician-related variable (GI-NET patient volume > 30 patients/year) was associated with high clinical utility and two variables (older age/less experience treating GI-NETs) with low clinical utility. CONCLUSIONS: Clinician-perceived clinical utility of QLQ-GINET21 is high. Clinicians valued the instruments' capacity to provide a better understanding of patient perspectives and to identify the factors that had the largest influence on patient HRQoL.

Rapid freshwater discharge on the coastal ocean as a mean of long distance spreading of an unprecedented toxic cyanobacteria bloom.

Cyanobacterial toxic blooms are a worldwide problem. The Río de la Plata (RdlP) basin makes up about one fourth of South America areal surface, second only to the Amazonian. Intensive agro-industrial land use and the construction of dams have led to generalized eutrophication of main tributaries and increased the intensity and duration of cyanobacteria blooms. Here we analyse the evolution of an exceptional bloom at the low RdlP basin and Atlantic coast during the summer of 2019. A large array of biological, genetic, meteorological, oceanographic and satellite data is combined to discuss the driving mechanisms. The bloom covered the whole stripe of the RdlP estuary and the Uruguayan Atlantic coasts (around 500 km) for approximately 4 months. It was caused by the Microcystis aeruginosa complex (MAC), which produces hepatotoxins (microcystin). Extreme precipitation in the upstream regions of Uruguay and Negro rivers' basins caused high water flows and discharges. The evolution of meteorological and oceanographic conditions as well as the similarity of organisms' traits in the affected area suggest that the bloom originated in eutrophic reservoirs at the lower RdlP basin, Salto Grande in the Uruguay river, and Negro river reservoirs. High temperatures and weak Eastern winds prompted the rapid dispersion of the bloom over the freshwater plume along the RdlP northern and Atlantic coasts. The long-distance rapid drift allowed active MAC organisms to inoculate freshwater bodies from the Atlantic basin, impacting environments relevant for biodiversity conservation. Climate projections for the RdlP basin suggest an increase in precipitation and river water flux, which, in conjunction with agriculture intensification and dams' construction, might turn this extraordinary event into an ordinary situation.

Morphology captures toxicity in Microcystis aeruginosa complex: Evidence from a wide environmental gradient✰.

Blooms of the Microcystis aeruginosa complex (MAC) consist of mixtures of toxin-producing and non-toxin-producing populations, but the environmental conditions that determine their relative abundance and shift are not clear. Morphological traits reflect the responses of MAC organisms to environmental changes, thus they could be useful to improve the predictability of the abundance of both toxic and nontoxic populations. In this work, the response of MAC toxic populations to environmental conditions and their relationship with morphology (size of organisms) were investigated in different water bodies (reservoir, river, and estuary) covering wide salinity (0-33) and temperature (10-36 °C) gradients. Sub-surface water samples were collected and divided into 4 size classes (mesh size 〈20 µm, 20-60 µm, 60-150 µm and〉 150 µm) and three toxicity proxies were assessed (mcyE gene and transcripts copy numbers and microcystin concentration) for each size-class. For all the size-classes, the logarithm of the number of mcyE gene copies per sample was proportional to the logarithm of the corresponding biovolume fraction, showing that MAC biovolume is a good indicator of toxicity potential. When toxicity was analyzed through mcyE transcript abundance and microcystin concentration, the largest size fraction (>150 µm) showed the highest toxicity values of both proxies. Nevertheless, mcyE transcription and toxin production per cell were higher in the colonies retained in the 60 to 150 µm size fractions, followed by single cells (<20 µm). At the reservoir, where environmental variability is low, the total abundance of mcyE gene copies was significantly explained by MAC biovolume, regardless of the environmental conditions. However, when data from the reservoir to the estuary were modeled, biovolume and temperature (with a minor contribution of salinity and wind intensity) were selected in the best models. According to these results, the size distribution of MAC biovolume appears as a good predictor of active toxin production, being the colonies in the 60-150 µm size fraction good indicators of higher toxicity. These results can be used to predict MAC toxicity based on the size structure of the community.

Prediction of Progression-Free Survival in Patients With Advanced, Well-Differentiated, Neuroendocrine Tumors Being Treated With a Somatostatin Analog: The GETNE-TRASGU Study.

PURPOSE: Somatostatin analogs (SSAs) are recommended for the first-line treatment of most patients with well-differentiated, gastroenteropancreatic (GEP) neuroendocrine tumors; however, benefit from treatment is heterogeneous. The aim of the current study was to develop and validate a progression-free survival (PFS) prediction model in SSA-treated patients. PATIENTS AND METHODS: We extracted data from the Spanish Group of Neuroendocrine and Endocrine Tumors Registry (R-GETNE). Patient eligibility criteria included GEP primary, Ki-67 of 20% or less, and first-line SSA monotherapy for advanced disease. An accelerated failure time model was developed to predict PFS, which was represented as a nomogram and an online calculator. The nomogram was externally validated in an independent series of consecutive eligible patients (The Christie NHS Foundation Trust, Manchester, United Kingdom). RESULTS: We recruited 535 patients (R-GETNE, n = 438; Manchester, n = 97). Median PFS and overall survival in the derivation cohort were 28.7 (95% CI, 23.8 to 31.1) and 85.9 months (95% CI, 71.5 to 96.7 months), respectively. Nine covariates significantly associated with PFS were primary tumor location, Ki-67 percentage, neutrophil-to-lymphocyte ratio, alkaline phosphatase, extent of liver involvement, presence of bone and peritoneal metastases, documented progression status, and the presence of symptoms when initiating SSA. The GETNE-TRASGU (Treated With Analog of Somatostatin in Gastroenteropancreatic and Unknown Primary NETs) model demonstrated suitable calibration, as well as fair discrimination ability with a C-index value of 0.714 (95% CI, 0.680 to 0.747) and 0.732 (95% CI, 0.658 to 0.806) in the derivation and validation series, respectively. CONCLUSION: The GETNE-TRASGU evidence-based prognostic tool stratifies patients with GEP neuroendocrine tumors receiving SSA treatment according to their estimated PFS. This nomogram may be useful when stratifying patients with neuroendocrine tumors in future trials. Furthermore, it could be a valuable tool for making treatment decisions in daily clinical practice.

Implementation of massive sequencing in the genetic diagnosis of hereditary cancer syndromes: diagnostic performance in the Hereditary Cancer Programme of the Valencia Community (FamCan-NGS).

BACKGROUND: Approximately 5 to 10% of all cancers are caused by inherited germline mutations, many of which are associated with different Hereditary Cancer Syndromes (HCS). In the context of the Program of Hereditary Cancer of the Valencia Community, individuals belonging to specific HCS and their families receive genetic counselling and genetic testing according to internationally established guidelines. The current diagnostic approach is based on sequencing a few high-risk genes related to each HCS; however, this method is time-consuming, expensive and does not achieve a confirmatory genetic diagnosis in many cases. This study aims to test the level of improvement offered by a Next Generation Sequencing (NGS) gene-panel compared to the standard approach in a diagnostic reference laboratory setting. METHODS: A multi-gene NGS panel was used to test a total of 91 probands, previously classified as non-informative by analysing the high-risk genes defined in our guidelines. RESULTS: Nineteen deleterious mutations were detected in 16% of patients, some mutations were found in already-tested high-risk genes (BRCA1, BRCA2, MSH2) and others in non-prevalent genes (RAD51D, PALB2, ATM, TP53, MUTYH, BRIP1). CONCLUSIONS: Overall, our findings reclassify several index cases into different HCS, and change the mutational status of 14 cases from non-informative to gene mutation carriers. In conclusion, we highlight the necessity of incorporating validated multi-gene NGS panels into the HCSs diagnostic routine to increase the performance of genetic diagnosis.

Mouth gape determines the response of marine top predators to long-term fishery-induced changes in food web structure.

Here, we analyse changes throughout time in the isotopic niche of the Franciscana dolphin (Pontoporia blainvillei), the South American fur seal (Arctocephalus australis) and the South American sea lion (Otaria flavescens) from the Río de la Plata estuary and adjacent Atlantic Ocean to test the hypothesis that fishing may modify the diet of small-gape predators by reducing the average size of prey. The overall evidence, from stable isotope and stomach contents analyses, reveals major changes in resource partitioning between the three predators considered, mainly because of an increased access of Franciscana dolphins to juvenile demersal fishes. These results are consistent with the changes in the length distribution of demersal fish species resulting from fishing and suggest that Franciscana dolphin has been the most benefited species of the three marine mammal species considered because of its intermediate mouth gape. In conclusion, the impact of fishing on marine mammals goes beyond the simple reduction in prey biomass and is highly dependent on the mouth gape of the species involved.

Morphology-based differences in the thermal response of freshwater phytoplankton.

The thermal response of maximum growth rate in morphology-based functional groups (MBFG) of freshwater phytoplankton is analysed. Contrasting an exponential Boltzmann-Arrhenius with a unimodal model, three main features were evaluated: (i) the activation energy of the rise (Er), (ii) the presence of a break in the thermal response and (iii) the activation energy of the fall (Ef). The whole dataset (N = 563) showed an exponential increase (Er ∼ 0.5), a break around 24°C and no temperature dependence after the breakpoint (Ef = 0). Contrasting thermal responses among MBFG were found. All groups showed positive activation energy (Er > 0), four showed no evidence of decline in growth rate (temperature range = 0-35°C) and two presented a breakpoint followed by a sharp decrease in growth rate. Our results evidenced systematic differences between MBFG in the thermal response and a coherent response significantly related to morphological traits other than size (i.e. within MBFG). These results provide relevant information for water quality modelling and climate change predictions.

Occurrence of pesticide residues in fish from south American rainfed agroecosystems.

Environmental sustainability of South American rainfed agroecosystems is of current concern. In this work, we evaluate the occurrence of multiple pesticide residues in muscle tissue of wild fish species from two large rivers in South America (Uruguay and Negro Rivers). Two sampling campaigns (representing summer and winter crops) were performed during 2015 targeting a wide biodiversity of fish species used for human consumption (ranging from migratory to non-migratory and from detritivorous to top-predators). Three different localities associated to rainfed agriculture were assessed, two of them enclosed to a RAMSAR site (National Park "Esteros de Farrapos e Islas del Rio Uruguay"). Pesticide residues occurred in muscle tissue of 143 from 149 sampled fishes (96%). Thirty different pesticides were detected at concentrations from <1 to 194µgkg-1. Incidence of pesticides in fish were tightly related to: i) features of the contaminant: (Kow, environmental persistence and mobility) and ii) intensity of use of particular pesticides and land dedicated to rainfed agriculture. Trifloxystrobin, metolachlor and pyraclostrobin showed the highest rates of occurrence. Of great concern is that strobirulins have highest toxicity to fish from those detected compounds. From the pattern of pesticides occurring for non-migratory fish species it was possible to trend important spatial differences related to the intensity of rainfed agriculture. Results suggest a regular exposition of aquatic wild biota to sublethal concentrations of multiple semi-polar pesticides.

Neuroendocrine Tumor Heterogeneity Adds Uncertainty to the World Health Organization 2010 Classification: Real-World Data from the Spanish Tumor Registry (R-GETNE).

BACKGROUND: Gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs) are a complex family of tumors of widely variable clinical behavior. The World Health Organization (WHO) 2010 classification provided a valuable tool to stratify neuroendocrine neoplasms (NENs) in three prognostic subgroups based on the proliferation index. However, substantial heterogeneity remains within these subgroups, and simplicity sometimes entails an ambiguous and imprecise prognostic stratification. The purpose of our study was to evaluate the prognostic impact of histological differentiation within the WHO 2010 grade (G) 1/G2/G3 categories, and explore additional Ki-67 cutoff values in GEP-NENs. SUBJECTS, MATERIALS, AND METHODS: A total of 2,813 patients from the Spanish National Tumor Registry (RGETNE) were analyzed. Cases were classified by histological differentiation as NETs (neuroendocrine tumors [well differentiated]) or NECs (neuroendocrine carcinomas [poorly differentiated]), and by Ki-67 index as G1 (Ki-67 <2%), G2 (Ki-67 3%-20%), or G3 (Ki-67 >20%). Patients were stratified into five cohorts: NET-G1, NET-G2, NET-G3, NEC-G2, and NEC-G3. RESULTS: Five-year survival was 72%. Age, gender, tumor site, grade, differentiation, and stage were all independent prognostic factors for survival. Further subdivision of the WHO 2010 grading improved prognostic stratification, both within G2 (5-year survival: 81% [Ki-67 3%-5%], 72% [Ki-67 6%-10%], 52% [Ki-67 11%-20%]) and G3 NENs (5-year survival: 35% [Ki-67 21%-50%], 22% [Ki-67 51%-100%]). Five-year survival was significantly greater for NET-G2 versus NEC-G2 (75.5% vs. 58.2%) and NET-G3 versus NEC-G3 (43.7% vs. 25.4%). CONCLUSION: Substantial clinical heterogeneity is observed within G2 and G3 GEP-NENs. The WHO 2010 classification can be improved by including the additive effect of histological differentiation and the proliferation index. IMPLICATIONS FOR PRACTICE: Gastroenteropancreatic neuroendocrine neoplasms are tumors of widely variable clinical behavior, roughly stratified by the World Health Organization (WHO) 2010 classification into three subgroups based on proliferation index. Real-world data from 2,813 patients of the Spanish Registry RGETNE demonstrated substantial clinical heterogeneity within grade (G) 2 and G3 neuroendocrine neoplasms. Tumor morphology and further subdivision of grading substantially improves prognostic stratification of these patients and may help individualize therapy. This combined, additive effect shall be considered in future classifications of neuroendocrine tumors and incorporated for stratification purposes in clinical trials.