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      Efficacy of heat‐killed and formalin‐killed vaccines against Tilapia tilapinevirus in juvenile Nile tilapia ( Oreochromis niloticus)

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          Abstract

          Tilapia tilapinevirus (also known as tilapia lake virus, TiLV) is considered to be a new threat to the global tilapia industry. The objective of this study was to develop simple cell culture‐based heat‐killed (HKV) and formalin‐killed (FKV) vaccines for the prevention of disease caused by TiLV. The fish were immunized with 100 µl of either HKV or FKV by intraperitoneal injection with each vaccine containing 1.8 × 10 6 TCID 50‐inactivated virus. A booster vaccination was carried out at 21‐day post‐vaccination (dpv) using the same protocol. The fish were then challenged with a lethal dose of TiLV at 28 dpv. The expression of five immune genes ( IgM, IgD, IgT, CD4 and CD8) in the head kidney and spleen of experimental fish was assessed at 14 and 21 dpv and again after the booster vaccination at 28 dpv. TiLV‐specific IgM responses were measured by ELISA at the same time points. The results showed that both vaccines conferred significant protection, with relative percentage survival of 71.3% and 79.6% for HKV and FKV, respectively. Significant up‐regulation of IgM and IgT was observed in the head kidney of fish vaccinated with HKV at 21 dpv, while IgM, IgD and CD4 expression increased in the head kidney of fish receiving FKV at the same time point. After booster vaccination, IgT and CD8 transcripts were significantly increased in the spleen of fish vaccinated with the HKV, but not with FKV. Both vaccines induced a specific IgM response in both serum and mucus. In summary, this study showed that both HKV and FKV are promising injectable vaccines for the prevention of disease caused by TiLV in Nile tilapia.

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          Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.

          The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA).
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            Ontogeny of the immune system of fish.

            Information on the ontogeny of the fish immune system is largely restricted to a few species of teleosts (e.g., rainbow trout, catfish, zebrafish, sea bass) and has previously focused on morphological features. However, basic questions including the identification of the first lympho-hematopoietic sites, the origin of T- and B-lymphocytes and the acquisition of full immunological capacities remain to be resolved. We review these three main topics with special emphasis on recent results obtained from the zebrafish, a new experimental model particularly suitable for study of the ontogeny of the immune system because of its rapid development and easy manipulation. This species also provides an easy way of creating mutations that can be detected by various types of screens. In some teleosts (i.e., angelfish) the first blood cells are formed in the yolk sac. In others, such as zebrafish, the first hematopoietic site is an intraembryonic locus, the intermediate cell mass (ICM), whereas in both killifish and rainbow trout the first blood cells appear for a short time in the yolk sac but later the ICM becomes the main hematopoietic area. Erythrocytes and macrophages are the first blood cells to be identified in zebrafish embryos. They occur in the ICM, the duct of Cuvier and the peripheral circulation. Between 24 and 30 hour post-fertilization (hpf) at a temperature of 28 degrees C a few myeloblasts and myelocytes appear between the yolk sac and the body walls, and the ventral region of the tail of 1-2 day-old zebrafish also contains developing blood cells. The thymus, kidney and spleen are the major lymphoid organs of teleosts. The thymus is the first organ to become lymphoid, although earlier the kidney can contain hematopoietic precursors but not lymphocytes. In freshwater, but not in marine, teleosts the spleen is the last organ to acquire that condition. We and other authors have demonstrated an early expression of Rag-1 in the zebrafish thymus that correlates well with the morphological identification of lymphoid cells. On the other hand, the origins and time of appearance of B lymphocytes in teleosts are a matter of discussion and recent results are summarized here. The functioning rather than the mere morphological evidence of lymphocytes determines when the full immunocompetence in fish is attained. Information on the histogenesis of fish lymphoid organs can also be obtained by analysing zebrafish mutants with defects in the development of immune progenitors and/or in the maturation of non-lymphoid stromal elements of the lymphoid organs. The main characteristics of some of these mutants will also be described.
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              Identification of a novel RNA virus lethal to tilapia.

              Tilapines are important for the sustainability of ecological systems and serve as the second most important group of farmed fish worldwide. Significant mortality of wild and cultured tilapia has been observed recently in Israel. The etiological agent of this disease, a novel RNA virus, is described here, and procedures allowing its isolation and detection are revealed. The virus, denominated tilapia lake virus (TiLV), was propagated in primary tilapia brain cells or in an E-11 cell line, and it induced a cytopathic effect at 5 to 10 days postinfection. Electron microscopy revealed enveloped icosahedral particles of 55 to 75 nm. Low-passage TiLV, injected intraperitoneally in tilapia, induced a disease resembling the natural disease, which typically presents with lethargy, ocular alterations, and skin erosions, with >80% mortality. Histological changes included congestion of the internal organs (kidneys and brain) with foci of gliosis and perivascular cuffing of lymphocytes in the brain cortex; ocular inflammation included endophthalmitis and cataractous changes of the lens. The cohabitation of healthy and diseased fish demonstrated that the disease is contagious and that mortalities (80 to 100%) occur within a few days. Fish surviving the initial mortality were immune to further TiLV infections, suggesting the mounting of a protective immune response. Screening cDNA libraries identified a TiLV-specific sequence, allowing the design of a PCR-based diagnostic test. This test enables the specific identification of TiLV in tilapines and should help control the spread of this virus worldwide.
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                Author and article information

                Contributors
                Channarong.R@chula.ac.th
                htdong@ait.ac.th
                Journal
                J Fish Dis
                J Fish Dis
                10.1111/(ISSN)1365-2761
                JFD
                Journal of Fish Diseases
                John Wiley and Sons Inc. (Hoboken )
                0140-7775
                1365-2761
                03 September 2021
                December 2021
                : 44
                : 12 ( doiID: 10.1111/jfd.v44.12 )
                : 2097-2109
                Affiliations
                [ 1 ] Center of Excellence in Fish Infectious Diseases (CE FID), Department of Veterinary Microbiology Faculty of Veterinary Science Chulalongkorn University Bangkok Thailand
                [ 2 ] The International Graduate Program of Veterinary Science and Technology (VST) Faculty of Veterinary Science Chulalongkorn University Bangkok Thailand
                [ 3 ] Division of Aquacultural Biotechnology Biotechnology Center of Ho Chi Minh City Ho Chi Minh Vietnam
                [ 4 ] Center of Excellence in Aquatic Animal Health Management Faculty of Fisheries Kasetsart University Bangkok Thailand
                [ 5 ] Fish Health Platform Center of Excellence for Shrimp Molecular Biology and Biotechnology (Centex Shrimp) Faculty of Science Mahidol University Bangkok Thailand
                [ 6 ] National Center for Genetic Engineering and Biotechnology (BIOTEC) National Science and Technology Development Agency (NSTDA) Pathum Thani Thailand
                [ 7 ] Aquaculture Research Group Moredun Research Institute Edinburgh UK
                [ 8 ] Infection and Immunity Division Roslin Institute Edinburgh UK
                [ 9 ] Faculty of Science and Technology Suan Sunandha Rajabhat University Bangkok Thailand
                [ 10 ] Department of Food, Agriculture and Bioresources School of Environment, Resources and Development Asian Institute of Technology Pathum Thani Thailand
                Author notes
                [*] [* ] Correspondence

                Ha Thanh Dong, School of Environment, Resources and Development, Asian Institute of Technology, Pathum Thani, Thailand.

                Email: htdong@ 123456ait.ac.th

                Channarong Rodkhum, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand.

                Email: Channarong.R@ 123456chula.ac.th

                Author information
                https://orcid.org/0000-0003-1953-1103
                https://orcid.org/0000-0002-6187-2772
                https://orcid.org/0000-0002-4834-8693
                https://orcid.org/0000-0002-5599-4343
                https://orcid.org/0000-0002-2243-0146
                https://orcid.org/0000-0002-9582-1228
                https://orcid.org/0000-0003-3550-4842
                https://orcid.org/0000-0002-5711-9294
                https://orcid.org/0000-0002-7190-757X
                Article
                JFD13523
                10.1111/jfd.13523
                9291230
                34477227
                24d0c5ea-e0ee-41a9-81ae-fca1e62570c5
                © 2021 The Authors. Journal of Fish Diseases published by John Wiley & Sons Ltd.

                This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.

                History
                : 03 August 2021
                : 05 June 2021
                : 03 August 2021
                Page count
                Figures: 3, Tables: 3, Pages: 0, Words: 8374
                Funding
                Funded by: International Veterinary Vaccinology Network
                Categories
                Research Article
                Research Articles
                Custom metadata
                2.0
                December 2021
                Converter:WILEY_ML3GV2_TO_JATSPMC version:6.1.7 mode:remove_FC converted:18.07.2022

                immune responses,inactivated vaccine,nile tilapia,protection,tilapia lake virus

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