Desarrollo de una vacuna DNA para el control de la enfermedad de linfocistis

Leiva, R., Borrego, J.J., Castro, D. y Labella, A.M. 2018. Desarrollo de una vacuna DNA para el control de la enfermedad de linfocistis. En: La Acuicultura en el Litoral Suratlántico. IX Jornadas de Acuicultura en el Litroral Suratlántico, p 96. Instituto de Investigación y Formación Agraria y Pesquera (IFAPA), Consejería de Agricultura, Pesca y Desarrollo Rural, Junta de Andalucía.El virus de la enfermedad de linfocistis (LCDV) es el agente etiológico de la enfermedad de linfocistis, que afecta tanto a peces marinos como dulceacuícolas. Esta enfermedad supone un grave problema para el sector de la acuicultura, provocando pérdidas económicas debido a su alta morbilidad. En el presente estudio se ha desarrollado un plásmido recombinante con objeto de ser utilizado como vacuna de DNA para limitar la incidencia de dicha enfermedad en el cultivo de dorada (Sparus aurata). El gen codificante de la proteína principal de la cápside (MCP) del LCDV de dorada (LCDV-Sa) se clonó en primer lugar en el vector de expresión pGEX-6P-3, con el fin de expresar la MCP viral como proteína de fusión con GST (glutathione S-transferase) en Escherichia coli BL-21. La expresión de la proteína de fusión MCP-GST en células bacterianas se comprobó mediante Western blot e inmunoensayo dot-blot con anticuerpos específicos dirigidos contra la MCP y la GST. Esta MCP recombinante se empleará como antígeno de captura para la detección mediante ELISA de anticuerpos anti-LCDV en peces vacunados. Por otra parte, el gen codificante de la MCP se subclonó en el vector de expresión pcDNA3.1/NT-GFP-TOPO, obteniéndose así el plásmido vacunal. Tras su amplificación en E. coli TOP 10, el plásmido se purificó y se utilizó para transfectar la línea celular de dorada SAF-1, evaluándose dos métodos de transfección: Nucleofector Kit y lipofectamina. La expresión de la MCP viral en células SAF-1 transfectadas con el plásmido vacunal se ha demostrado mediante detección de la GFP (green fluorescent protein) por microscopía de epifluorescencia, y por inmunodetección de la MCP viral utilizando anticuerpos específicos. El plásmido vacunal obtenido se inyectará intramuscularmente en ejemplares de dorada para determinar la distribución y expresión temporal de la vacuna, así como para detectar anticuerpos anti-LCDV en los peces vacunados.Proyecto de Excelencia de la Junta de Andalucía (P12-RNM-2261). Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Análisis de la expresión de inmunogenes en doradas vacunadas en respuesta a la infección por LCDV-Sa

El objetivo del presente estudio ha sido evaluar la respuesta inmune frente a la infección por Lymphocystis Disease Virus 3 (LCDV-Sa) en ejemplares de dorada (Sparus aurata) vacunados con un plásmido que codifica la proteína principal de la cápside del virus, con el fin de identificar inmunogenes relacionados con la protección inducida por la vacuna. Se utilizaron juveniles de dorada inyectados intramuscularmente con la vacuna (grupo vacunado) o el plásmido vacío (grupo placebo), así como con PBS (dos grupos). A los 30 d post-vacunación, los peces se inocularon con un aislado de LCDV-Sa (106 TCID50/pez), excepto uno de los grupos originalmente inyectado con PBS que se mantuvo como control no-infectado. Se tomaron muestras de riñón cefálico a los 1 y 3 d post-infección (dpi) (6 peces por tiempo) y se realizó un análisis de la expresión relativa de 49 inmunogenes de dorada utilizando la plataforma OpenArray®, basada en qPCR con sondas TaqMan, usando las muestras del control no-infectado como calibrador. Los resultados mostraron una mayor expresión diferencial de inmunogenes en los peces vacunados en comparación a los peces que recibieron el plásmido vacío o a los no vacunados. En todos los grupos experimentales se observó una sub-regulación génica a 1 dpi, que cambió a sobre-regulación a los 3 dpi. Además, en los peces vacunados se observó la estimulación temprana de la expresión del gen activador de recombinación (rag1) y una sobre-regulación tardía de mx1 y mx2.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Evaluación de la respuesta inmune en lenguado senegalés conferida por una vacuna inactivada frente a Betanodavirus

La necrosis nerviosa viral es una enfermedad que afecta a peces cultivados en todo el mundo. Su agente etiológico es el virus de la necrosis nerviosa, género Betanodavirus, familia Nodaviridae, que presenta un genoma compuesto por dos segmentos de RNA monocatenario. Los betanodavirus se clasifican en cuatro especies, Striped Jack-, Tiger puffer-, Redspotted grouper- y Barfin flonder nervous necrosis virus (SJNNV, TPNNV, RGNNV y BFNNV, respectivamente). En el sur de Europa se han descrito recombinantes de los segmentos genómicos de las especies SJNNV y RGNNV como agentes causantes de epizootías en lenguado senegalés y dorada. El control de esta enfermedad es de gran importancia para la acuicultura europea y la vacunación es una de las estrategias más prometedoras. Sin embargo, solo existen vacunas comercializadas contra la especie RGNNV las cuales no protegen frente a los aislados recombinantes, por lo que se ha desarrollado una vacuna inactivada utilizando el aislado recombinante SpSsIAusc160.03 que produce una moderada protección frente a la infección vírica en lenguado (Valero et al., 2021). El objetivo de este estudio es evaluar la capacidad de dicha vacuna de inducir una respuesta inmune eficaz en lenguado (Solea senegalensis). Se tomaron muestras de cerebro y riñón cefálico de lenguados vacunados y sin vacunar a 2, 3 y 7 días post-vacunación (dpv), analizándose la expresión de 106 inmunogenes mediante la plataforma OpenArray® (Gémez et al., 2020). Se detectó una respuesta inmune temprana en muestras de riñón, expresándose diferencialmente 39 y 29 genes a 2 y 3 dpv, respectivamente. Esta modulación fue significativamente menor a 7 dpv, con solo 3 genes expresados diferencialmente (DEG). En muestras de cerebro, tejido diana de la infección, se observó una menor modulación génica, detectándose expresión diferencial exclusivamente a 2 y 3 dpv (5 y 12 DEG, respectivamente). Financiación: Proyecto RTI2018-094687-B-C21/C22 del MICIU cofinanciado por FEDER.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Myofascial trigger points in cluster headache patients: a case series

Active myofascial trigger points (MTrPs) have been found to contribute to chronic tension-type headache and migraine. The purpose of this case series was to examine if active trigger points (TrPs) provoking cluster-type referred pain could be found in cluster headache patients and, if so, to evaluate the effectiveness of active TrPs anaesthetic injections both in the acute and preventive headache's treatment. Twelve patients, 4 experiencing episodic and 8 chronic cluster headache, were studied. TrPs were found in all of them. Abortive infiltrations could be done in 2 episodic and 4 chronic patients, and preemptive infiltrations could be done in 2 episodic and 5 chronic patients, both kind of interventions being successful in 5 (83.3%) and in 6 (85.7%) of the cases respectively. When combined with prophylactic drug therapy, injections were associated with significant improvement in 7 of the 8 chronic cluster patients. Our data suggest that peripheral sensitization may play a role in cluster headache pathophysiology and that first neuron afferent blockade can be useful in cluster headache management

Informe COTEC: situación y evolución de la economía circular en España

Tercera entrega de este informe bienal analiza la evolución y la situación actual de la economía circular en España respecto a Europa.La economía circular es una de las grandes transiciones en las que Cotec está inmersa y una de sus líneas prioritarias en las que está centrando sus esfuerzos. Con este tercer informe, Cotec quiere mantener su contribución al análisis de la situación de la economía circular en España y analizar las políticas puestas en marcha desde los distintos niveles administrativos. Esta entrega además incorpora un estudio detallado de las competencias y capacidades disponibles a nivel nacional para impulsar la transición circular, así como de las barreras y los elementos facilitadores para dicha transición. La economía circular ofrece una alternativa al actual modelo de producción, basado en una cadena de valor lineal que genera residuos en todas las etapas, desde la extracción de materias primas hasta la generación de residuos, pasando por las fases de fabricación, distribución y consumo. La alternativa consiste en prolongar la vida económica útil de los materiales y los recursos tanto como sea posible, reduciendo al mínimo la generación de residuos.Peer ReviewedPostprint (published version

Why Are Outcomes Different for Registry Patients Enrolled Prospectively and Retrospectively? Insights from the Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF).

Background: Retrospective and prospective observational studies are designed to reflect real-world evidence on clinical practice, but can yield conflicting results. The GARFIELD-AF Registry includes both methods of enrolment and allows analysis of differences in patient characteristics and outcomes that may result. Methods and Results: Patients with atrial fibrillation (AF) and ≥1 risk factor for stroke at diagnosis of AF were recruited either retrospectively (n = 5069) or prospectively (n = 5501) from 19 countries and then followed prospectively. The retrospectively enrolled cohort comprised patients with established AF (for a least 6, and up to 24 months before enrolment), who were identified retrospectively (and baseline and partial follow-up data were collected from the emedical records) and then followed prospectively between 0-18 months (such that the total time of follow-up was 24 months; data collection Dec-2009 and Oct-2010). In the prospectively enrolled cohort, patients with newly diagnosed AF (≤6 weeks after diagnosis) were recruited between Mar-2010 and Oct-2011 and were followed for 24 months after enrolment. Differences between the cohorts were observed in clinical characteristics, including type of AF, stroke prevention strategies, and event rates. More patients in the retrospectively identified cohort received vitamin K antagonists (62.1% vs. 53.2%) and fewer received non-vitamin K oral anticoagulants (1.8% vs . 4.2%). All-cause mortality rates per 100 person-years during the prospective follow-up (starting the first study visit up to 1 year) were significantly lower in the retrospective than prospectively identified cohort (3.04 [95% CI 2.51 to 3.67] vs . 4.05 [95% CI 3.53 to 4.63]; p = 0.016). Conclusions: Interpretations of data from registries that aim to evaluate the characteristics and outcomes of patients with AF must take account of differences in registry design and the impact of recall bias and survivorship bias that is incurred with retrospective enrolment. Clinical Trial Registration: - URL: http://www.clinicaltrials.gov . Unique identifier for GARFIELD-AF (NCT01090362)

REQUITE: A prospective multicentre cohort study of patients undergoing radiotherapy for breast, lung or prostate cancer

Purpose: REQUITE aimed to establish a resource for multi-national validation of models and biomarkers that predict risk of late toxicity following radiotherapy. The purpose of this article is to provide summary descriptive data. Methods: An international, prospective cohort study recruited cancer patients in 26 hospitals in eight countries between April 2014 and March 2017. Target recruitment was 5300 patients. Eligible patients had breast, prostate or lung cancer and planned potentially curable radiotherapy. Radiotherapy was prescribed according to local regimens, but centres used standardised data collection forms. Pre-treatment blood samples were collected. Patients were followed for a minimum of 12 (lung) or 24 (breast/prostate) months and summary descriptive statistics were generated. Results: The study recruited 2069 breast (99% of target), 1808 prostate (86%) and 561 lung (51%) cancer patients. The centralised, accessible database includes: physician-(47,025 forms) and patient-(54,901) reported outcomes; 11,563 breast photos; 17,107 DICOMs and 12,684 DVHs. Imputed genotype data are available for 4223 patients with European ancestry (1948 breast, 1728 prostate, 547 lung). Radiation-induced lymphocyte apoptosis (RILA) assay data are available for 1319 patients. DNA (n = 4409) and PAXgene tubes (n = 3039) are stored in the centralised biobank. Example prevalences of 2-year (1-year for lung) grade >= 2 CTCAE toxicities are 13% atrophy (breast), 3% rectal bleeding (prostate) and 27% dyspnoea (lung). Conclusion: The comprehensive centralised database and linked biobank is a valuable resource for the radiotherapy community for validating predictive models and biomarkers. Patient summary: Up to half of cancer patients undergo radiation therapy and irradiation of surrounding healthy tissue is unavoidable. Damage to healthy tissue can affect short-and long-term quality-of-life. Not all patients are equally sensitive to radiation "damage" but it is not possible at the moment to identify those who are. REQUITE was established with the aim of trying to understand more about how we could predict radiation sensitivity. The purpose of this paper is to provide an overview and summary of the data and material available. In the REQUITE study 4400 breast, prostate and lung cancer patients filled out questionnaires and donated blood. A large amount of data was collected in the same way. With all these data and samples a database and biobank were created that showed it is possible to collect this kind of information in a standardised way across countries. In the future, our database and linked biobank will be a resource for research and validation of clinical predictors and models of radiation sensitivity. REQUITE will also enable a better understanding of how many people suffer with radiotherapy toxicity

Risk profiles and one-year outcomes of patients with newly diagnosed atrial fibrillation in India: Insights from the GARFIELD-AF Registry.

BACKGROUND: The Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF) is an ongoing prospective noninterventional registry, which is providing important information on the baseline characteristics, treatment patterns, and 1-year outcomes in patients with newly diagnosed non-valvular atrial fibrillation (NVAF). This report describes data from Indian patients recruited in this registry. METHODS AND RESULTS: A total of 52,014 patients with newly diagnosed AF were enrolled globally; of these, 1388 patients were recruited from 26 sites within India (2012-2016). In India, the mean age was 65.8 years at diagnosis of NVAF. Hypertension was the most prevalent risk factor for AF, present in 68.5% of patients from India and in 76.3% of patients globally (P < 0.001). Diabetes and coronary artery disease (CAD) were prevalent in 36.2% and 28.1% of patients as compared with global prevalence of 22.2% and 21.6%, respectively (P < 0.001 for both). Antiplatelet therapy was the most common antithrombotic treatment in India. With increasing stroke risk, however, patients were more likely to receive oral anticoagulant therapy [mainly vitamin K antagonist (VKA)], but average international normalized ratio (INR) was lower among Indian patients [median INR value 1.6 (interquartile range {IQR}: 1.3-2.3) versus 2.3 (IQR 1.8-2.8) (P < 0.001)]. Compared with other countries, patients from India had markedly higher rates of all-cause mortality [7.68 per 100 person-years (95% confidence interval 6.32-9.35) vs 4.34 (4.16-4.53), P < 0.0001], while rates of stroke/systemic embolism and major bleeding were lower after 1 year of follow-up. CONCLUSION: Compared to previously published registries from India, the GARFIELD-AF registry describes clinical profiles and outcomes in Indian patients with AF of a different etiology. The registry data show that compared to the rest of the world, Indian AF patients are younger in age and have more diabetes and CAD. Patients with a higher stroke risk are more likely to receive anticoagulation therapy with VKA but are underdosed compared with the global average in the GARFIELD-AF. CLINICAL TRIAL REGISTRATION-URL: http://www.clinicaltrials.gov. Unique identifier: NCT01090362

Global Retinoblastoma Presentation and Analysis by National Income Level.

Importance: Early diagnosis of retinoblastoma, the most common intraocular cancer, can save both a child's life and vision. However, anecdotal evidence suggests that many children across the world are diagnosed late. To our knowledge, the clinical presentation of retinoblastoma has never been assessed on a global scale. Objectives: To report the retinoblastoma stage at diagnosis in patients across the world during a single year, to investigate associations between clinical variables and national income level, and to investigate risk factors for advanced disease at diagnosis. Design, Setting, and Participants: A total of 278 retinoblastoma treatment centers were recruited from June 2017 through December 2018 to participate in a cross-sectional analysis of treatment-naive patients with retinoblastoma who were diagnosed in 2017. Main Outcomes and Measures: Age at presentation, proportion of familial history of retinoblastoma, and tumor stage and metastasis. Results: The cohort included 4351 new patients from 153 countries; the median age at diagnosis was 30.5 (interquartile range, 18.3-45.9) months, and 1976 patients (45.4%) were female. Most patients (n = 3685 [84.7%]) were from low- and middle-income countries (LMICs). Globally, the most common indication for referral was leukocoria (n = 2638 [62.8%]), followed by strabismus (n = 429 [10.2%]) and proptosis (n = 309 [7.4%]). Patients from high-income countries (HICs) were diagnosed at a median age of 14.1 months, with 656 of 666 (98.5%) patients having intraocular retinoblastoma and 2 (0.3%) having metastasis. Patients from low-income countries were diagnosed at a median age of 30.5 months, with 256 of 521 (49.1%) having extraocular retinoblastoma and 94 of 498 (18.9%) having metastasis. Lower national income level was associated with older presentation age, higher proportion of locally advanced disease and distant metastasis, and smaller proportion of familial history of retinoblastoma. Advanced disease at diagnosis was more common in LMICs even after adjusting for age (odds ratio for low-income countries vs upper-middle-income countries and HICs, 17.92 [95% CI, 12.94-24.80], and for lower-middle-income countries vs upper-middle-income countries and HICs, 5.74 [95% CI, 4.30-7.68]). Conclusions and Relevance: This study is estimated to have included more than half of all new retinoblastoma cases worldwide in 2017. Children from LMICs, where the main global retinoblastoma burden lies, presented at an older age with more advanced disease and demonstrated a smaller proportion of familial history of retinoblastoma, likely because many do not reach a childbearing age. Given that retinoblastoma is curable, these data are concerning and mandate intervention at national and international levels. Further studies are needed to investigate factors, other than age at presentation, that may be associated with advanced disease in LMICs

CARB-ES-19 Multicenter Study of Carbapenemase-Producing Klebsiella pneumoniae and Escherichia coli From All Spanish Provinces Reveals Interregional Spread of High-Risk Clones Such as ST307/OXA-48 and ST512/KPC-3

ObjectivesCARB-ES-19 is a comprehensive, multicenter, nationwide study integrating whole-genome sequencing (WGS) in the surveillance of carbapenemase-producing K. pneumoniae (CP-Kpn) and E. coli (CP-Eco) to determine their incidence, geographical distribution, phylogeny, and resistance mechanisms in Spain.MethodsIn total, 71 hospitals, representing all 50 Spanish provinces, collected the first 10 isolates per hospital (February to May 2019); CPE isolates were first identified according to EUCAST (meropenem MIC &gt; 0.12 mg/L with immunochromatography, colorimetric tests, carbapenem inactivation, or carbapenem hydrolysis with MALDI-TOF). Prevalence and incidence were calculated according to population denominators. Antibiotic susceptibility testing was performed using the microdilution method (EUCAST). All 403 isolates collected were sequenced for high-resolution single-nucleotide polymorphism (SNP) typing, core genome multilocus sequence typing (cgMLST), and resistome analysis.ResultsIn total, 377 (93.5%) CP-Kpn and 26 (6.5%) CP-Eco isolates were collected from 62 (87.3%) hospitals in 46 (92%) provinces. CP-Kpn was more prevalent in the blood (5.8%, 50/853) than in the urine (1.4%, 201/14,464). The cumulative incidence for both CP-Kpn and CP-Eco was 0.05 per 100 admitted patients. The main carbapenemase genes identified in CP-Kpn were blaOXA–48 (263/377), blaKPC–3 (62/377), blaVIM–1 (28/377), and blaNDM–1 (12/377). All isolates were susceptible to at least two antibiotics. Interregional dissemination of eight high-risk CP-Kpn clones was detected, mainly ST307/OXA-48 (16.4%), ST11/OXA-48 (16.4%), and ST512-ST258/KPC (13.8%). ST512/KPC and ST15/OXA-48 were the most frequent bacteremia-causative clones. The average number of acquired resistance genes was higher in CP-Kpn (7.9) than in CP-Eco (5.5).ConclusionThis study serves as a first step toward WGS integration in the surveillance of carbapenemase-producing Enterobacterales in Spain. We detected important epidemiological changes, including increased CP-Kpn and CP-Eco prevalence and incidence compared to previous studies, wide interregional dissemination, and increased dissemination of high-risk clones, such as ST307/OXA-48 and ST512/KPC-3