Antemortem versus postmortem methods for detection of betanodavirus in Senegalese sole (Solea senegalensis)

The suitability of nested reverse transcription polymerase chain reaction (nRT-PCR) to detect betanodavirus in blood samples from naturally infected Senegalese sole (Solea senegalensis) was evaluated in comparison with other diagnostic methods. Results indicated that histologic examination of brain lesions could be regarded as the most consistent indicator of nodavirus infection in this species. The nRT-PCR showed low to moderate levels of detection; the best values were obtained in brain samples followed by blood samples. Inoculation of SSN-1 and SAF-1 cells with fish samples did not cause cytopathic effect, although virus was detected by reverse transcription polymerase chain reaction in approximately 25% of the SSN-1 inoculated wells. The efficiency of detection of the viral genome was dramatically increased by the use of nRTPCR, reaching 90.6% of positives in brain samples and 84.4% in blood samples. The sensitivity and the negative predictive value of nRT-PCR in blood samples were slightly lower than those obtained using brain samples. Nevertheless, it is suggested that the advantage of being able to perform diagnosis on live fish adequately counterbalances the slightly lower sensitivity of nRT-PCR on blood samples. This technique is proposed as a useful tool, not only for the selection of nodavirus-free breeders but also to check the fish status during ongrowing

Influência da estrutura populacional de Psectrogaster falcata (Perciformes: Curimatidae) no parasitismo de Excorallana berbicensis (Isopoda: Corallanidae) do reservatório da Usina Hidrelétrica Coaracy Nunes (UHECN), Rio Araguari, Ferreira Gomes, Amapá, Brasil.

Estudos sobre a fauna de ectoparasitos crustáceos em peixes de reservatórios hidrelétricos foram conduzidos em diferentes bacias hidrográficas do Brasil. Esses estudos registraram infestações por espécies de Isopoda, Copepoda e Branchiura. Em condições naturais, a presença e abundância de crustáceos ectoparasitos têm sido influenciadas tipicamente por uma variedade de fatores, tais como o ambiente, fisiologia, comportamento, imunologia e estado nutricional dos hospedeiros. O objetivo deste estudo foi avaliar a influência da estrutura populacional do hospedeiro P. falcata no parasitismo de E. berbicensis do reservatório da UHECN, Rio Araguari. Os espécimes de P. falcata foram capturados bimestralmente, de outubro de 2012 a agosto de 2013, em seis pontos distribuídos no reservatório da UHECN (00º54'11.8"N e 051º15'35.5"W), Rio Araguari. Para a pesca foram utilizadas redes de espera simples de diferentes malhas (20 a 60 mm entre nós opostos), com 12 horas de permanência e vistorias a cada duas horas. De cada peixe-hospedeiro foi mensurado o Comprimento Padrão (Cp), em centímetros. A sexagem foi realizada através de inspeção macroscópica das gônadas. A boca, câmara branquial, tegumento e nadadeiras de cada peixe foi examinada imediatamente após a captura, para verificar a presença de E. berbicensis. Para testar a proporção sexual de hospedeiros machos e fêmeas foi utilizado o teste qui-quadrado (X2), sendo que os valores de X2>3.84 foram considerados significativamente diferentes. O teste paramétrico Kolmogorov-Smirnov foi utilizado para comparar tanto a estrutura populacional em classes de tamanho entre os sexos, quanto a abundância de E. berbicensis por classe de tamanho de machos e fêmeas de P. falcata. O nível de significância estatística adotado foi p menor ou igual a 0.05. Um total de 65 espécimes de P. falcata foram capturados, sendo 28 machos (43,1%), com comprimento padrão variando entre 11,2 e 28,0 cm (16,5 ± 3,4 cm), e 37 fêmeas (56,9%), com comprimento padrão entre 11,2 e 27,8 cm (19,5 ± 3,9 cm). A proporção sexual de hospedeiros machos e fêmeas (1M:1,3F) não apresentou diferenças significativas (X2 = 1,904; p=0,168). A estrutura populacional para hospedeiros machos e fêmeas em comprimento foi construída por meio de distribuições de frequência de exemplares e o teste Kolmogorov-Smirnov (p>0,01) evidenciou fêmeas significativamente maiores que os machos. Após inspeção dos hospedeiros foram coletados 279 espécimes de E. berbicensis, sendo 82 presentes em hospedeiros machos (29,4%) e 197 em hospedeiros fêmeas (70,6%). Quanto a abundância de ectoparasitos crustáceos por classe de tamanho do hospedeiro, o teste Kolmogorov-Smirnov evidenciou diferenças significativas (p<0,01), onde a maioria dos parasitos coletados nos hospedeiros do sexo feminino (90,9%) estiveram no maior intervalo de classe (18 a 30 cm). Nos hospedeiros machos, esteve presente em sua maioria (73,0%) no intervalo de classe intermediário (12 a 24 cm). Essa diferença na abundância parasitária de E. berbicensis entre a estrutura populacional de machos e fêmeas de P. falcata pode estar relacionada, principalmente, a diferenças ecológicas e comportamentais entre os sexos dos hospedeiros. Além disso, hospedeiros maiores podem albergar uma maior quantidade parasitos que hospedeiros menores.Resumo 131-2

European sea bass brain DLB-1 cell line is susceptible to nodavirus: A transcriptomic study

Viral diseases are responsible for high rates of mortality and subsequent economic losses in modern aquaculture. The nervous necrosis virus (NNV) produces viral encephalopathy and retinopathy (VER), which affects the fish central nervous system. It is considered one of the most serious viral diseases in marine aquaculture, the European sea bass (Dicentrarchus labrax) being amongst the most susceptible. We have evaluated the European sea bass brain derived cell line (DLB-1) susceptibility to NNV genotypes and evaluated its transcriptomic profile. DLB-1 cells supported NNV gene transcription and replication since strains belonging to the four NNV genotypes produce cytopathic effects. Afterwards, DLB-1 cells were infected with an RGNNV strain, the one which showed the highest replication, for 12 and 72 h and an RNA-seq analysis was performed to identify potential genes involved in the host-NNV interactions. Differential expression analysis showed the up-regulation of many genes related to immunity, heat-shock proteins or apoptosis but not to proteasome or autophagy processes. These data suggest that the immune response, mainly the interferon (IFN) pathway, is not powerful enough to abrogate the infection, and cells finally suffer stress and die by apoptosis liberating infective particles. GO enrichment also revealed, for the first time, the down-regulation of terms related to brain/neuron biology indicating molecular mechanisms causing the pathogenic effect of NNV. This study opens the way to understand key elements in sea bass brain and NNV interactions.Versión del edito

Análisis de la relevancia y factibilidad de indicadores de calidad en las unidades de nutrición

Introducción: La evaluación de la calidad en las actividades sanitarias exige la elección de unos indicadores acordes con los resultados que queremos medir. De todos los posibles, debemos priorizar aquellos que nos permitan obtener la información más relevante sin sobrecargar el trabajo habitual de nuestras Unidades. Objetivo: Conocer la opinión de los socios de SENPE respecto a la relevancia y la viabilidad del uso de una selección de indicadores de calidad para su aplicación en nutrición clínica. Métodos: Encuesta remitida mediante correo electrónico a los socios de SENPE solicitando a los mismos su opinión sobre 12 indicadores de calidad, valorándose cada uno en cuanto a su relevancia y factibilidad de la aplicación en su medio. Resultados: Contestaron 40 encuestados de 40 centros diferentes de 12 comunidades autónomas. En general, los indicadores fueron considerados más relevantes que factibles. Los indicadores mejor puntuados fueron: “identificación en las bolsas de nutrición artificial”, “posición semi-incorporada del paciente con nutrición enteral por sonda nasogástrica” y “protocolos clínicos básicos”. Considerando los indicadores por grupos (de estructura, proceso o resultado) los mejor valorados fueron: “identificación del paciente en las bolsas de nutrición artificial” (estructura), “posición semi-incorporada” y “protocolos clínicos básicos” (proceso), y “cumplimiento del objetivo calórico” (resultado). Conclusión: Los resultados de la encuesta permiten seleccionar indicadores prioritarios para su aplicación en las Unidades de NutriciónIntroduction: The quality assessment in health activities requires the choice of indicators in line with the results we want to measure. Of all possible, we should prioritize those that allow us to obtain the most relevant information without overloading the regular work of our units. Objective: To determine the opinion of the members of SENPE regarding the relevance and feasibility of using a selection of quality indicators designed for use in clinical nutrition. Methods: E-mail survey sent to members of SENPE asking them their views on 12 quality indicators, evaluating each in terms of their relevance and feasibility of implementation in their environment. Results: 40 respondents answered from 40 centers in 12 different regions. In general, the indicators were considered more relevant than feasible. The indicators best rated were: “identification in artificial nutrition bags, “semi-recumbent position in patient with nasogastric tube feeding” and “basic clinical protocols”. Considering the type of indicator: “patient identification in the bags of artificial nutrition (structure),” a semi-incorporated “and” basic clinical protocols (process), and “fulfillment of the caloric goal” (result). Conclusion: The results of the survey can make a selection of indicators that could be considered for first-line introduction in a Nutrition Uni

Predicting high risk of exacerbations in bronchiectasis: the E-FACED score

BACKGROUND: Although the FACED score has demonstrated a great prognostic capacity in bronchiectasis, it does not include the number or severity of exacerbations as a separate variable, which is important in the natural history of these patients. OBJECTIVE: Construction and external validation of a new index, the E-FACED, to evaluate the predictive capacity of exacerbations and mortality. METHODS: The new score was constructed on the basis of the complete cohort for the construction of the original FACED score, while the external validation was undertaken with six cohorts from three countries (Brazil, Argentina, and Chile). The main outcome was the number of annual exacerbations/hospitalizations, with all-cause and respiratory-related deaths as the secondary outcomes. A statistical evaluation comprised the relative weight and ideal cut-off point for the number or severity of the exacerbations and was incorporated into the FACED score (E-FACED). The results obtained after the application of FACED and E-FACED were compared in both the cohorts. RESULTS: A total of 1,470 patients with bronchiectasis (819 from the construction cohorts and 651 from the external validation cohorts) were followed up for 5 years after diagnosis. The best cut-off point was at least two exacerbations in the previous year (two additional points), meaning that the E-FACED has nine points of growing severity. E-FACED presented an excellent prognostic capacity for exacerbations (areas under the receiver operating characteristic curve: 0.82 for at least two exacerbations in 1 year and 0.87 for at least one hospitalization in 1 year) that was statistically better than that of the FACED score (0.72 and 0.78, P<0.05, respectively). The predictive capacities for all-cause and respiratory mortality were 0.87 and 0.86, respectively, with both being similar to those of the FACED. CONCLUSION: E-FACED score significantly increases the FACED capacity to predict future yearly exacerbations while maintaining the score’s simplicity and prognostic capacity for death

Effectiveness and safety of obeticholic acid in a Southern European multicenter cohort of patients with primary biliary cholangitis and suboptimal response to ursodeoxycholic acid

Background Obeticholic acid (OCA) was recently approved as the only on-label alternative for patients with primary biliary cholangitis (PBC) with intolerance or suboptimal response to ursodeoxycholic acid (UDCA). However, few data are available outside clinical trials. Aim To assess the effectiveness and safety of OCA in a real-world cohort of patients with non-effective UDCA therapy. Methods Open-label, prospective, real-world, multicentre study, enrolling consecutive patients who did not meet Paris II criteria, from 18 institutions in Spain and Portugal. Effectiveness was assessed by the changes in GLOBE and UK-PBC scores from baseline. POISE and Paris II criteria were evaluated after 12 months of OCA . Liver fibrosis was evaluated by FIB-4 and AST to platelet ratio index (APRI). Results One hundred and twenty patients were eligible, median time since PBC diagnosis 9.3 (4.0-13.8) years, 21.7% had cirrhosis, and 26.7% received had previous or concomitant treatment with fibrates. Seventy-eight patients completed at least 1 year of OCA. The Globe-PBC score decreased to 0.17 (95% CI 0.05 to 0.28; P = 0.005) and the UK-PBC score decreased to 0.81 (95% CI -0.19 to 1.80; P = 0.11). There was a significant decrease in alkaline phosphatase of 81.3 U/L (95% CI 42.5 to 120; P < 0.001), ALT 22.1 U/L (95% CI 10.4 to 33.8; P < 0.001) and bilirubin 0.12 mg/dL (95% CI 0 to 0.24; P = 0.044). FIB-4 and APRI remained stable. According to the POISE criteria, 29.5% (23 out of 78) achieved response. The adverse events rate was 35%; 11.67% discontinued (8.3% due to pruritus). Conclusions This study supports data from phase III trials with significant improvement of PBC-Globe continuous prognostic marker score among OCA-treated patients with good tolerability

Nodavirus colonizes and replicates in the testis of gilthead seabream and European sea bass modulating its immune and reproductive functions

Viruses are threatening pathogens for fish aquaculture. Some of them are transmitted through gonad fluids or gametes as occurs with nervous necrosis virus (NNV). In order to be transmitted through the gonad, the virus should colonize and replicate inside some cell types of this tissue and avoid the subsequent immune response locally. However, whether NNV colonizes the gonad, the cell types that are infected, and how the immune response in the gonad is regulated has never been studied. We have demonstrated for the first time the presence and localization of NNV into the testis after an experimental infection in the European sea bass (Dicentrarchus labrax), and in the gilthead seabream (Sparus aurata), a very susceptible and an asymptomatic host fish species, respectively. Thus, we localized in the testis viral RNA in both species using in situ PCR and viral proteins in gilthead seabream by immunohistochemistry, suggesting that males might also transmit the virus. In addition, we were able to isolate infective particles from the testis of both species demonstrating that NNV colonizes and replicates into the testis of both species. Blood contamination of the tissues sampled was discarded by completely fish bleeding, furthermore the in situ PCR and immunocytochemistry techniques never showed staining in blood vessels or cells. Moreover, we also determined how the immune and reproductive functions are affected comparing the effects in the testis with those found in the brain, the main target tissue of the virus. Interestingly, NNV triggered the immune response in the European sea bass but not in the gilthead seabream testis. Regarding reproductive functions, NNV infection alters 17β-estradiol and 11-ketotestosterone production and the potential sensitivity of brain and testis to these hormones, whereas there is no disruption of testicular functions according to several reproductive parameters. Moreover, we have also studied the NNV infection of the testis in vitro to assess local responses. Our in vitro results show that the changes observed on the expression of immune and reproductive genes in the testis of both species are different to those observed upon in vivo infections in most of the casesMINECO and FEDER (AGL2010-20801-C02-01; AGL2010-20801-C02-02; AGL2013-43588-P); Fundación Séneca (04538/GERM/06)Versión del editor4,411

Longitudinal outcomes of obeticholic acid therapy in ursodiol-nonresponsive primary biliary cholangitis: Stratifying the impact of add-on fibrates in real-world practice

Background Suboptimal response to ursodeoxycholic acid occurs in 40% of primary biliary cholangitis (PBC) patients, affecting survival. Achieving a deep response (normalisation of alkaline phosphatase [ALP] and bilirubin ≤0.6 upper limit of normal) improves survival. Yet, the long-term effectiveness of second-line treatments remains uncertain. Aims To evaluate the long-term effectiveness of obeticholic acid (OCA) ± fibrates. Focusing on biochemical response (ALP ≤1.67 times the upper limit of normal, with a decrease of at least 15% from baseline and normal bilirubin levels), normalisation of ALP, deep response and biochemical remission (deep response plus aminotransferase normalisation). Methods We conducted a longitudinal, observational, multicentre study involving ursodeoxyccholic acid non-responsive PBC patients (Paris-II criteria) from Spain and Portugal who received OCA ± fibrates. Results Of 255 patients, median follow-up was 35.1 months (IQR: 20.2–53). The biochemical response in the whole cohort was 47.2%, 61.4% and 68.6% at 12, 24 and 36 months. GLOBE-PBC and 5-year UK-PBC scores improved (p < 0.001). Triple therapy (ursodeoxycholic acid plus OCA plus fibrates) had significantly higher response rates than dual therapy (p = 0.001), including ALP normalisation, deep response and biochemical remission (p < 0.001). In multivariate analysis, triple therapy remained independently associated with biochemical response (p = 0.024), alkaline phosphatase normalisation, deep response and biochemical remission (p < 0.001). Adverse effects occurred in 41.2% of cases, leading to 18.8% discontinuing OCA. Out of 55 patients with cirrhosis, 12 developed decompensation. All with baseline portal hypertension. Conclusion Triple therapy was superior in achieving therapeutic goals in UDCA-nonresponsive PBC. Decompensation was linked to pre-existing portal hypertension