Detection of different Betanodavirus genotypes in wild fish from Spanish Atlantic coastal waters (Galicia, northwestern Spain).

OBJECTIVE: The nervous necrosis virus (NNV; genus Betanodavirus) is an aquatic pathogen that is responsible for a neurological disease affecting marine fish. Despite its almost worldwide distribution, global warming could favor the spread of NNV to new areas, highlighting the importance of conducting epidemiological surveys on both wild and farmed marine fish species. In this study, we assessed NNV prevalence in wild fish caught along the Galician Atlantic coast. METHODS: In total, 1277 fish were analyzed by reverse transcription real-time polymerase chain reaction. RESULT: Twenty two (1.72%) of those fish tested positive for NNV, including two species in which the pathogen had not yet been reported. CONCLUSION: The reassortant RGNNV/SJNNV (red-spotted grouper NNV/striped jack NNV) was detected in 55% of NNV-positive individuals, while the remaining 45% harbored the SJNNV-type genome. Moreover, from European Pilchard Sardina pilchardus and Atlantic Mackerel Scomber scombrus, we isolated four reassortant strains that carried amino acid mutations at key sites related to NNV-host interaction.

Nervous necrosis virus viability modulation by water salinity and temperature.

Nervous necrosis virus (NNV) is a hazardous aquatic pathogen, distributed worldwide and in a wide range of temperatures. Viral persistence in water has been demonstrated to be affected by different factors, such as temperature, UV, or biological load. In this study, we have investigated the viability of NNV strains in low- and high-salinity seawater (LS and HS, respectively) both in laboratory and aquarium conditions, at different storage temperatures, and for comparative purposes, in culture medium. Our results showed the highest NNV viability in seawater at 15°C and as temperature increased, a drop in viral persistence was observed. Additionally, survival at 15 and 30°C was strongly affected by increasing salt content, while no differences were observed between LS and HS groups at 20 and 25°C. The results of the incubation under aquarium conditions indicated that the effect of UV light and oxygen exposure accelerate the inactivation of infective particles. According to previous studies, NNV persistence in cell culture medium was higher than in seawater, and as observed in the latter, increasing incubation temperatures led to a decrease in viral survival.

Effect of rearing density on nervous necrosis virus infection in Senegalese sole (Solea senegalensis).

Intensive fish farming at high densities results in a wide range of adverse consequences on fish welfare, including pathogen spreading, stress and increased mortality rates. In this work, we have assessed whether the survival of Senegalese sole infected with the nervous necrosis virus (NNV), a pathogen responsible for severe disease outbreaks, is affected by rearing density. Based on the different fish ratios per surface area (g cm-2 ) and water volume (g L-1 ), our research showed an earlier mortality onset in the tanks containing NNV-infected fish reared at medium density (MD: 0.071 g cm-2 /5 g L-1 ) and high density (HD: 0.142 g cm-2 /10 g L-1 ), as well as higher cumulative mortality values. However, transcription analysis of hsp70, gr1 and pepck genes, well-known stress biomarkers, seems to indicate that none of the challenged fish were under high stress conditions. NNV load was slightly higher both in dead and in sampled fish from MD and HD groups, and especially in the rearing water from these groups, where peaks in mortality seemed to correlate with increasing NNV load in the water. In conclusion, our results suggest that rearing NNV-infected Senegalese sole at high densities resulted in an earlier mortality onset and higher cumulative values and viral load.

Amino acidic substitutions in the polymerase N-terminal region of a reassortant betanodavirus strain causing poor adaptation to temperature increase.

Nervous necrosis virus (NNV), Genus Betanodavirus, is the causative agent of viral encephalopathy and retinopathy (VER), a neuropathological disease that causes fish mortalities worldwide. The NNV genome is composed of two single-stranded RNA molecules, RNA1 and RNA2, encoding the RNA polymerase and the coat protein, respectively. Betanodaviruses are classified into four genotypes: red-spotted grouper nervous necrosis virus (RGNNV), striped jack nervous necrosis virus (SJNNV), barfin flounder nervous necrosis virus (BFNNV) and tiger puffer nervous necrosis virus (TPNNV). In Southern Europe the presence of RGNNV, SJNNV and their natural reassortants (in both RNA1/RNA2 forms: RGNNV/SJNNV and SJNNV/RGNNV) has been reported. Pathology caused by these genotypes is closely linked to water temperature and the RNA1 segment encoding amino acids 1-445 has been postulated to regulate viral adaptation to temperature. Reassortants isolated from sole (RGNNV/SJNNV) show 6 substitutions in this region when compared with the RGNNV genotype (positions 41, 48, 218, 223, 238 and 289). We have demonstrated that change of these positions to those present in the RGNNV genotype cause low and delayed replication in vitro when compared with that of the wild type strain at 25 and 30 °C. The experimental infections confirmed the impact of the mutations on viral replication because at 25 °C the viral load and the mortality were significantly lower in fish infected with the mutant than in those challenged with the non-mutated virus. It was not possible to challenge fish at 30 °C because of the scarce tolerance of sole to this temperature.

Betanodavirus infection in primary neuron cultures from sole.

Nervous necrosis virus (NNV), G. Betanodavirus, is the causative agent of viral encephalopathy and retinopathy, a disease that causes mass mortalities in a wide range of fish species. Betanodaviruses are neurotropic viruses and their replication in the susceptible fish species seems to be almost entirely restricted to nerve tissue. However, none of the cell lines used for NNV propagation has a nervous origin. In this study, first we established a protocol for the primary culture of neurons from Senegalese sole, which made it possible to further study virus-host cell interactions. Then, we compared the replication of three NNV strains with different genotypes (SJNNV, RGNNV and a RGNNV/SJNNV reassortant strain) in sole neuron primary cultures and E-11 cells. In addition, to study how two amino acid substitutions at the c-terminal of the capsid protein (positions 247 and 270) affect the binding to cell receptors, a recombinant strain was also tested. The results show that sole neural cells enabled replication of all the tested NNV strains. However, the recombinant strain shows a clearly delayed replication when compared with the wt strain. This delay was not observed in virus replicating in E-11 cells, suggesting a viral interaction with different cell receptors. The establishment of a sole primary neuronal culture protocol provides an important tool for research into betanodavirus infection in sole.

Brain Recovery after a Plane Crash: Treatment with Growth Hormone (GH) and Neurorehabilitation: A Case Report.

UNLABELLED: The aim of this study is to describe the results obtained after growth hormone (GH) treatment and neurorehabilitation in a young man that suffered a very grave traumatic brain injury (TBI) after a plane crash. METHODS: Fifteen months after the accident, the patient was treated with GH, 1 mg/day, at three-month intervals, followed by one-month resting, together with daily neurorehabilitation. Blood analysis at admission showed that no pituitary deficits existed. At admission, the patient presented: spastic tetraplegia, dysarthria, dysphagia, very severe cognitive deficits and joint deformities. Computerized tomography scanners (CT-Scans) revealed the practical loss of the right brain hemisphere and important injuries in the left one. Clinical and blood analysis assessments were performed every three months for three years. Feet surgery was needed because of irreducible equinovarus. RESULTS: Clinical and kinesitherapy assessments revealed a prompt improvement in cognitive functions, dysarthria and dysphagia disappeared and three years later the patient was able to live a practically normal life, walking alone and coming back to his studies. No adverse effects were observed during and after GH administration. CONCLUSIONS: These results, together with previous results from our group, indicate that GH treatment is safe and effective for helping neurorehabilitation in TBI patients, once the acute phase is resolved, regardless of whether or not they have GH-deficiency (GHD).

A Potential Nervous Necrosis Virus (NNV) Live Vaccine for Sole Obtained by Genomic Modification.

Viral Encephalopathy and Retinopathy (VER) is a neurological infectious fish disease that causes vacuolization and necrosis in the central nervous system, which lead to swimming abnormalities and, generally, host death in the early stages of development. VER is caused by the Nervous Necrosis Virus (NNV), a non-enveloped virus with a bisegmented and positive-stranded (+) RNA genome. The largest segment (RNA1) codes for viral polymerase while capsid protein is encoded by RNA2. The aim of this study was to explore the potential of a reverse-engineered RGNNV/SJNNV strain that harbors mutations in both 3'NCRs (position 3073 of RNA1 and 1408 and 1412 of RNA2) as an attenuated live vaccine for sole. The attenuation of this strain was confirmed through experimental infections in sole at 22 °C. Vaccination trials were performed by bath, intramuscular, and intraperitoneal injection, at two temperatures (18 and 22 °C). Our results indicate the improved survival of vaccinated fish and delayed and poorer viral replication, as well as an overexpression of immune response genes linked to T cell markers (cd4 and cd8), to an early inflammatory response (tlr7 and tnfα), and to antiviral activity (rtp3 and mx). In conclusion, our study indicates that the attenuated strain is a good vaccine candidate as it favors sole survival upon infection with the wt strain while inducing a significant immune response.

Role of Rotifers in Betanodavirus Transmission to European Sea Bass Larvae.

Marine invertebrates such as rotifers or Artemia, frequently used for fish larvae feeding, can be a potential source of pathogens. It has been demonstrated that Artemia can act as a nervous necrosis virus (NNV)-vector to Senegalese sole larvae. Therefore, in this study, we aimed to clarify the role of rotifers in NNV transmission to sea bass larvae following an oral challenge. Our results showed that sea bass larvae fed on a single dose of rotifers retaining NNV displayed clinical signs, mortality, and viral replication similar to the immersion challenge, although the course of the infection was slightly different between the two infection routes. Furthermore, we also demonstrated that rotifers can internalize NNV particles due to their filtering nature and maintain virus viability since viral particles were detected by immunohistochemistry, immunofluorescence, and cell culture within the rotifer body. However, viral quantification data suggested that rotifers are not permissive to NNV replication. In conclusion, this research demonstrated NNV horizontal transmission through rotifers to sea bass larvae, highlighting the importance of establishing strict routine controls on live food to prevent the introduction of potential pathogens to hatcheries.

Effect of risk of malnutrition on 30-day mortality among older patients with acute heart failure in Emergency Departments.

BACKGROUND: Little is known about the prevalence and impact of risk of malnutrition on short-term mortality among seniors presenting with acute heart failure (AHF) in emergency setting. The objective was to determine the impact of risk of malnutrition on 30-day mortality risk among older patients who attended in Emergency Departments (EDs) for AHF. MATERIAL AND METHODS: We performed a secondary analysis of the OAK-3 Registry including all consecutive patients ≥65 years attending in 16 Spanish EDs for AHF. Risk of malnutrition was defined by the Mini Nutritional Assessment Short Form (MNA-SF) < 12 points. Unadjusted and adjusted logistic regression models were used to assess the association between risk of malnutrition and 30-day mortality. RESULTS: We included 749 patients (mean age: 85 (SD 6); 55.8% females). Risk of malnutrition was observed in 594 (79.3%) patients. The rate of 30-day mortality was 8.8%. After adjusting for MEESSI-AHF risk score clinical categories (model 1) and after adding all variables showing a significantly different distribution among groups (model 2), the risk of malnutrition was an independent factor associated with 30-day mortality (adjusted OR by model 1 = 3.4; 95%CI 1.2-9.7; p = .020 and adjusted OR by model 2 = 3.1; 95%CI 1.1-9.0; p = .033) compared to normal nutritional status. CONCLUSIONS: The risk of malnutrition assessed by the MNA-SF is associated with 30-day mortality in older patients with AHF who were attended in EDs. Routine screening of risk of malnutrition may help emergency physicians in decision-making and establishing a care plan.

Parálisis Cerebral: situación alimentaria en pacientes con soporte nutricional / Cerebral Palsy: nutritional status in patients with nutritional support

Introducción: la parálisis cerebral (PC) es la afección neuromotriz más común de la primera infancia siendo la principal causa de discapacidad física en niños. Esta patología afecta el estado nutricional de quienes la padecen y, a su vez, la malnutrición exacerba las alteraciones neurológicas preexistentes. Objetivos: determinar la distribución de frecuencias del tipo de alimentación y el tipo de fórmula o leche seleccionada en los pacientes con diagnóstico de PC con soporte enteral exclusivamente. Determinar el aporte calórico, proteico, de fibra, líquidos, calcio y hierro. Determinar la concordancia entre el estado nutricional de los pacientes y el percibido por sus cuidadores. Materiales y Método: estudio observacional, transversal, descriptivo y analítico. Muestreo consecutivo, por conveniencia. Resultados: la muestra quedó conformada por 30 pacientes y sus cuidadores. El 70% de la muestra utilizaba fórmula y/o leche, el 3% alimentos modificados y el 27% realizaba una alimentación mixta; el tipo de fórmula más utilizado fue la infantil (33%). El 60% de la muestra fue normopeso, 33% bajo peso y el 7% tuvo exceso de peso. El aporte calórico promedio fue de 11,1 kcal/cm (DS + 6,5 kcal/cm) en los niños con bajo peso, 11,6 kcal/cm (DS + 6,4 kcal/cm) en aquellos con normopeso y 9,8 kcal/cm (+ 2,4 kcal/cm) en los niños con exceso de peso; no se observaron diferencias según estado nutricional (p=0,13). El 90% cubrió el requerimiento de proteínas, el 80% el de hierro, 73% el hídrico, el 70% el de calcio y un 3% el de fibra. Se halló concordancia moderada entre el estado nutricional real del niño y el percibido por su cuidador. (kappa 0,44; p=0,0013; IC95 0,22-0,62%). Conclusión: Los pacientes con PC requieren el acompañamiento de Licenciados en nutrición para adecuar el aporte a sus requerimientos nutricionales considerando las características de la PC y su tratamiento(AU).

Introduction: cerebral palsy (CP) is the most common neuromotor condition during early childhood, being the main cause of physical disability in children. This pathology affects the nutritional status of those affected, and in turn, malnutrition exacerbates pre-existing neurological disorders. Objectives: to determine the distributional frequency of feeding type and milk or formula type selected within patients diagnosed with PC and enteral support exclusively. To determine the caloric, protein, fiber, fluids, calcium and iron intake. To determine concordance between the nutritional status of patients and that perceived by their caregivers. Materials and Methods: an observational, transversal, prospective, descriptive, analytical study with a consecutive sampling was conducted. Results: the study included 30 patients and their caregivers. 70% of the sample used formula and/or milk, 3% modified foods and 27% a mixed diet; infant formula was the one mostly used (33%). 60% of the sample was normal weight, 33% low weight and 7% overweight. The average caloric intake was 11.1 kcal/cm (SD +6.5 kcal/cm) in children with low weight, 11.6 kcal/cm (SD +6.4 kcal/cm) in those with normal weight and 9.8 kcal/cm (SD +2.4 kcal/cm) in children with overweight; no differences were observed following nutritional status (p=0.13). 90% covered protein requirement, 80% iron, 73% water, 70% calcium and 3% fiber. Moderate agreement was found between the real nutritional status of the child and that perceived by the caregiver (kappa 0.44, p=0.0013, IC95 0.22-0.62%). Conclusion: patients with CP require the accompaniment of graduates in nutrition to adapt nutritional requirements, taking into consideration the characteristics of the CP and its treatment(AU).

Trastornos hipertensivos del embarazo en sala de labor y parto del Hospital Escuela / Hypertensive Disorders of Pregnancy in the Labor and Delivery Ward of the Hospital Escuela

Los Trastornos Hipertensivos del Embarazo (THE) se caracterizan por presión arterial sistólica mayor o iguala 140 mm Hg o diastólica mayor o igual a 90 mm Hg. Representan la segunda causa de mortalidad materna en Honduras y provocan elevada morbimortalidad perinatal. Objetivo: Describir los datos clínico-epidemiológicos, evaluación, manejo y complicaciones materno-fetales de los trastornos hipertensivos del embarazo en el Hospital Escuela. Métodos:Estudio transversal, descriptivo, realizado del 16 de marzo al 26 de julio de 2010, se revisaron los expedientes clínicos de mujeres con diagnóstico de trastorno hipertensivo del embarazo, excluyendo los casos de hipertensión gestacional. Resultados: La prevalencia de los trastornos hipertensivos del embarazo es 7.8%, de estos 47% presentaron preeclampsia leve y 38% preeclampsia severa; se les realizó inducción y cesárea al 40% respectivamente. Se practicó cardiotocografía en reposo al 10.4% y prueba de provocación con oxitocina al 9%; se presentó crisis hipertensiva en el 35% de los casos. La principal complicación materna fue el Síndrome de HELLP y la principal complicación neonatal el Síndrome de Distrés Respiratorio (SDR), aproximadamente el 50% de los recién nacidos fueron pequeños para la edad gestacional. Se presentaron 8 muertes fetales, 11 muertes neonatales y una muerte materna. Conclusión: las mujeres con THE tienen el doble de riesgo de terminar su embarazo en cesárea comparado con las mujeres sin este trastorno. La mortalidad materna se asocia con eclampsia y Síndrome deHELLP (por sus siglas en inglés: H de hemolysis, EL de elevated liver enzymes y LP de low platelet count); la mortalidad perinatal con preeclampsia severa, APGAR menor a 7 al primer minuto, prematurez y restricción del crecimiento intrauterino...

Agências regulatórias na experiência brasileira: um panorama do atual desenho institucional / Texto para Discussão (TD) 941: Agências regulatórias na experiência brasileira: um panorama do atual desenho institucional

Examina o projeto de reforma do Estado empreendido pelo governo Fernando Henrique, representado pela criação de agências regulatórias após a privatização de serviços públicos. Constata-se a existência de dois tipos distintos de agências regulatórias: o primeiro, representado pelas agências de governo, que executam diretrizes de governo, e o segundo, equivalentes ao modelo anglo-saxão, que podem ser denominados agências de Estado e que regulam a oferta de serviços públicos por meio de aplicação de legislação própria específica. Apresenta propostas de aperfeiçoamento institucional do modelo.