Evaluation of gilthead seabream (Sparus aurata) immune response after LCDV DNA vaccination

A DNA vaccine against Lymphocystis Disease Virus (LCDV) was developed and its protective efficacy in gilthead seabream (Sparus aurata) has been established. The aim of the present study is the evaluation of immune-related gene expression after vaccination to identify which genes could be relevant to control the viral infection. The vaccine was administered intramuscularly to gilthead seabream specimens (100 g weight) at 10 µg/fish dose. In addition, two control groups, injected with the empty plasmid at the same dose or PBS, were established to evaluate non-specific immune response and basal response of fish, respectively. In this study 23 genes related to the immune response (tlr5, tlr9, ifnI, irf1, irf3, irf9, pkr, mx1, mx2, mx3, isg15, tnfα, casp1, il1β, il6, il10, ck3, ck10, c3, nccrp1, mhcII, tcrβ, and ighm) and 2 reference genes (ef1α and actβ) were analysed using real-time PCR (RT-qPCR) in samples of head kidney and intestine at 1, 3, and 8 d post-vaccination. DNA-vaccination of gilthead seabream induced the differential expression of 9 genes in head kidney and 15 genes in intestine samples. Through the course of the experiment, 9 of those genes reached high level of up-regulation comparing to control groups. These genes were related to IFN type I pathway (irf9 and mx3, in head kidney), inflammation (il1β, il6, tnfα, ck10, c3 and nccrp-1, in both organs analysed), and adaptive immune response (mhcII, in intestine). Conclusion: The results obtained allow us to understand which genes could be responsible for the protection against LCDV infection conferred by the DNA vaccine in gilthead seabream. Inflammation is the biological process mainly triggered as a systemic response in vaccinated fish. Different gene expression profiles have been observed in each organ, which may indicate specialized roles relative to immune defensive mechanisms.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Immune response of gilthead seabream (Sparus aurata) after experimental infection with lymphocystis disease virus (LCDV-Sa)

Lymphocystis disease (LCD) is caused by the lymphocystis disease virus (LCDV) (family Iridoviridae), affecting more than 150 fish species from both marine and freshwater environments. A few studies have been focused on the immune defensive mechanisms of fish against LCDV, but only one was conducted during a natural LCD outbreak in gilthead seabream, which is one of the most important cultured fish species in the Mediterranean and the European Atlantic coasts. The aim of this study was the analysis of 23 genes related to the immune response in gilthead seabream specimens after experimental infection with LCDV-Sa using real-time PCR (qRT-PCR) in samples of head kidney and intestine at 1, 3, and 8 dpi. To study the progression of LCDV-Sa infection in gilthead seabreams, the number of viral DNA copies and the expression of mcp were determined in samples of caudal fin, head kidney and intestine. LCDV-Sa was detected by qPCR in all the samples from inoculated fish analysed, whereas no amplification was obtained in samples from the control group. Regarding the gene expression following LCDV-Sa infection, a total of 22 of the 23 genes studied were differentially expressed in head kidney or intestine samples at some time points analysed. Different gene expression profiles were obtained between the organs studied, detecting 18 differentially expressed genes (DEGs) in head kidney samples, four of them exclusively up- or down-regulated (nccrp1, il10, mhcII, and tnfα genes), and 5 genes with a significant change in the expression tendency from 1 to 8 dpi (irf3, isg15, il10, ck10, and c3). In the intestine, 18 DEGs were also detected (14 shared with head kidney), being mx1, casp1, ck3 and tlr9 genes exclusively detected in these samples, and mx1, mx3, irf9 and ighm differentially regulated over time. The results obtained allow us to understand which genes are essential for host-pathogen interactions and could be used as molecular markers for vaccine efficacy evaluation.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech Proyecto de Excelencia Junta de Andalucía Ref. P12-RNM-226

Evaluation of immune response after LCDV-Sa infection in DNA-vaccinated gilthead seabream

The immune-related gene expression in vaccinated gilthead seabream after Lymphocystis Disease Virus 3 (LCDV-Sa) infection was analysed by using an OpenArray based on TaqMan qPCR. The DNA vaccine used in this study encodes the viral major capsid protein and confers protection against LCDV-Sa infection in juvenile gilthead seabream. Gilthead seabream juveniles were distributed into four experimental groups and intramuscularly injected with the vaccine (vaccinated group), the empty-plasmid (mock-vaccinated group), or PBS (control groups). Thirty days after vaccination, vaccinated and mock-vaccinated fish, as well as one of the control groups, were injected intraperitoneally with LCDV-Sa (106 TCID50/fish). Samples of head-kidney (HK) from 6 fish were individually collected 1 and 3 days post-infection (dpi). The relative expression levels of 49 genes related to the immune response and 4 reference genes were analysed using an OpenArray. Samples from the non-infected control group were used as calibrator. The number of genes differentially expressed (DEG) in HK at 1 dpi was higher in vaccinated fish compared with both mock-vaccinated and non-vaccinated animals. At 3 dpi, most DEG were upregulated, and the differences in their number among groups were minimized. The recombination-activating gene 1 (rag1), a mediator of development of B and T lymphocytes, was the only gene upregulated in HK samples at 1 dpi. This gene was also upregulated in non-vaccinated animals but at 3 dpi. In contrast, early mx induction was observed in non-vaccinated animals (upregulation of mx2 at 1 dpi) in comparison to vaccinated seabreams (upregulation of mx1 and mx2 at 3 dpi). The results that will be discussed could evidence the role of the DNA vaccine as regulator of the primary lymphoid tissues (HK) in gilthead seabream against LCDV-Sa infection, through downregulation of inflammation related-genes, early upregulation of rag1, and a later expression of interferon stimulated genes.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Immune response of vaccinated juvenile gilthead seabream (Sparus aurata) after LCDV-Sa infection

Lymphocystis disease is one of the main viral pathologies affecting cultured gilthead seabream in the Mediterranean area. In our group, a DNA-vaccine has been developed based on the major capsid protein (MCP) of the Lymphocystis Disease Virus 3 (LCDV-Sa). The aim of the present study is the evaluation of immune-related gene expression in vaccinated fish after viral infection to identify immunogenes involved in the vaccine-induced protection. To fulfil this objective an OpenArray® platform has been developed to study 49 genes related to the immune response. Reference and viral genes were also evaluated. Gilthead seabream specimens (5 g mean weight) were distributed into 3 experimental groups, inoculated with the vaccine at 0.1 µg/g fish dose, the empty plasmid at the same dose or PBS. Thirty days post-vaccination, fish were intramuscularly injected with the virus at 106 TCID50/fish dose. Samples of head-kidney, spleen, intestine and caudal fin from 6 fish were individually collected at 1, 2 and 3-days post-injection in all groups. The quantification of viral DNA in fins of fish challenged with LCDV-Sa were carried out by a qPCR assay targeting a viral structural gene (putative myristoylated membrane protein, MMP) alternative to the mcp gene contained in the vaccine. The results obtained showed an increase of genes deregulated within the haematopoietic organs between vaccinated and non-vaccinated fish. However, in the intestine and fin, the results showed the opposite trend. The global effect of fish vaccination was a diminished immune response compared to non-vaccinated fish, being 83 and 99 genes differentially expressed through the experiment, respectively. Moreover, viral replication decreased in groups of fish previously vaccinated. The modulation of the immune response provoked by the vaccination trial seems to control the progression of the disease.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

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

Immune gene expression in gilthead seabream after nervous necrosis virus (NNV) challenge

Viral nervous necrosis (VNN) is a disease that affects farmed fish worldwide. Its etiologic agent is the nervous necrosis virus (NNV), genus Betanodavirus, family Nodaviridae. NNV are small and non-enveloped viruses with a genome consisting of two molecules of positive-sense single-stranded RNA, RNA1 and RNA2, which encode the RNA-dependent RNA polymerase and the capsid protein, respectively. The betanodaviruses have been classified into four species: Striped jack nervous necrosis virus (SJNNV), Tiger puffer nervous necrosis virus (TPNNV), Red-spotted grouper nervous necrosis virus (RGNNV), and Barfin flounder nervous necrosis virus (BFNNV). In Southern Europe, natural reassortants between RGNNV and SJNNV have been isolated from Senegalese sole (Solea senegalensis), gilthead seabream (Sparus aurata) and European seabass (Dicentrarchus labrax) associated to VNN outbreaks. Immune response against betanodavirus infections has been poorly studied in gilthead seabream. In this study, fish were challenged by intramuscular (im) injection or by immersion, using a reassortant strain containing RGNNV-type RNA1 and SJNNV-type RNA2 segments. Head kidney and brain samples were collected at 24, 48 and 72 h post-challenge (pc) for the injection experiment, while in the bath challenge sampling was performed at 48 and 72 h pc. The immunogen expression analysis was carried out using the platform OpenArray®. In the im-injected fish, 21 differentially expressed genes (DEGs) were identified in head kidney samples at 24 h pc, whereas a lower immune response was detected at 48 and 72 h pc (11 and 9 DEGs, respectively). In brain samples, a delayed response was observed, with 32 DEGs recorded at 72 h pc. Regarding the bath-challenged fish, fewer immunogenes were differentially expressed although all of them were up-regulated. This research was funded by the Ministerio de Ciencia, Innovación y Universidades (MCIUI) and FEDER under Grant RTI2018-094687-B.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

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

CD4陽性T細胞に発現するA20(tnfaip3) はTh2型アレルギー性気道炎症を抑制する

研究科: 千葉大学大学院医学薬学府（先端医学薬学専攻）学位記番号: 千大院医薬博甲第医1412号博士（医学）千葉大学 = Chiba Universit

RICORS2040 : The need for collaborative research in chronic kidney disease

Chronic kidney disease (CKD) is a silent and poorly known killer. The current concept of CKD is relatively young and uptake by the public, physicians and health authorities is not widespread. Physicians still confuse CKD with chronic kidney insufficiency or failure. For the wider public and health authorities, CKD evokes kidney replacement therapy (KRT). In Spain, the prevalence of KRT is 0.13%. Thus health authorities may consider CKD a non-issue: very few persons eventually need KRT and, for those in whom kidneys fail, the problem is 'solved' by dialysis or kidney transplantation. However, KRT is the tip of the iceberg in the burden of CKD. The main burden of CKD is accelerated ageing and premature death. The cut-off points for kidney function and kidney damage indexes that define CKD also mark an increased risk for all-cause premature death. CKD is the most prevalent risk factor for lethal coronavirus disease 2019 (COVID-19) and the factor that most increases the risk of death in COVID-19, after old age. Men and women undergoing KRT still have an annual mortality that is 10- to 100-fold higher than similar-age peers, and life expectancy is shortened by ~40 years for young persons on dialysis and by 15 years for young persons with a functioning kidney graft. CKD is expected to become the fifth greatest global cause of death by 2040 and the second greatest cause of death in Spain before the end of the century, a time when one in four Spaniards will have CKD. However, by 2022, CKD will become the only top-15 global predicted cause of death that is not supported by a dedicated well-funded Centres for Biomedical Research (CIBER) network structure in Spain. Realizing the underestimation of the CKD burden of disease by health authorities, the Decade of the Kidney initiative for 2020-2030 was launched by the American Association of Kidney Patients and the European Kidney Health Alliance. Leading Spanish kidney researchers grouped in the kidney collaborative research network Red de Investigación Renal have now applied for the Redes de Investigación Cooperativa Orientadas a Resultados en Salud (RICORS) call for collaborative research in Spain with the support of the Spanish Society of Nephrology, Federación Nacional de Asociaciones para la Lucha Contra las Enfermedades del Riñón and ONT: RICORS2040 aims to prevent the dire predictions for the global 2040 burden of CKD from becoming true

Study of the B−→Λc+Λˉc−K−B^{-} \to \Lambda_{c}^{+} \bar{\Lambda}_{c}^{-} K^{-} decay

The decay $B^{-} \to \Lambda_{c}^{+} \bar{\Lambda}_{c}^{-} K^{-}$ is studied in proton-proton collisions at a center-of-mass energy of $\sqrt{s}=13$ TeV using data corresponding to an integrated luminosity of 5 $\mathrm{fb}^{-1}$ collected by the LHCb experiment. In the $\Lambda_{c}^+ K^{-}$ system, the $\Xi_{c}(2930)^{0}$ state observed at the BaBar and Belle experiments is resolved into two narrower states, $\Xi_{c}(2923)^{0}$ and $\Xi_{c}(2939)^{0}$, whose masses and widths are measured to be $$m(\Xi_{c}(2923)^{0}) = 2924.5 \pm 0.4 \pm 1.1 \,\mathrm{MeV}, \\ m(\Xi_{c}(2939)^{0}) = 2938.5 \pm 0.9 \pm 2.3 \,\mathrm{MeV}, \\ \Gamma(\Xi_{c}(2923)^{0}) = \phantom{000}4.8 \pm 0.9 \pm 1.5 \,\mathrm{MeV},\\ \Gamma(\Xi_{c}(2939)^{0}) = \phantom{00}11.0 \pm 1.9 \pm 7.5 \,\mathrm{MeV},$$ where the first uncertainties are statistical and the second systematic. The results are consistent with a previous LHCb measurement using a prompt $\Lambda_{c}^{+} K^{-}$ sample. Evidence of a new $\Xi_{c}(2880)^{0}$ state is found with a local significance of $3.8\,\sigma$, whose mass and width are measured to be $2881.8 \pm 3.1 \pm 8.5\,\mathrm{MeV}$ and $12.4 \pm 5.3 \pm 5.8 \,\mathrm{MeV}$, respectively. In addition, evidence of a new decay mode $\Xi_{c}(2790)^{0} \to \Lambda_{c}^{+} K^{-}$ is found with a significance of $3.7\,\sigma$. The relative branching fraction of $B^{-} \to \Lambda_{c}^{+} \bar{\Lambda}_{c}^{-} K^{-}$ with respect to the $B^{-} \to D^{+} D^{-} K^{-}$ decay is measured to be $2.36 \pm 0.11 \pm 0.22 \pm 0.25$, where the first uncertainty is statistical, the second systematic and the third originates from the branching fractions of charm hadron decays.Comment: All figures and tables, along with any supplementary material and additional information, are available at https://cern.ch/lhcbproject/Publications/p/LHCb-PAPER-2022-028.html (LHCb public pages