Isolation and partial characterization of immunoglobulin from cod (Gadus morhua L.) (2023)

Cited by (71)

  • Development of a monoclonal antibody specific to burbot (Lota lota) IgM and optimization of an ELISA to measure anti-Aeromonas sp. antibody titers following pathogen challenge

    2023, Fish and Shellfish Immunology

    Burbot (Lota lota) are an ideal candidate for cool or cold-water aquaculture and are gaining interest because of their high economic value, low temperature requirements, and fast growth rate. Limited information exists on the innate and adaptive immune systems of this species. This is partly due to the lack of species-specific tools to determine antibody responses following disease or vaccination or to characterize the immune response in general. An anti-IgM monoclonal antibody (mAb 27C) was developed and characterized via enzyme-linked immunosorbent assay (ELISA) and Western blot for species specificity, affinity to the heavy chain of burbot IgM, and cross-reactivity to other reagents used in the analysis. The 27C monoclonal antibody was further utilized to develop an ELISA protocol to measure the specific antibody response of burbot following exposure to two pathogenic strains of Aeromonas sp. (A141 and IR004). This ELISA confirmed that vaccinated burbot that survived the challenge with either strain developed statistically higher titers of anti-Aeromonas antibodies specific for the relative strain when compared to fish that were not vaccinated or challenged. Western blot analysis further demonstrated that burbot surviving challenge had serum IgM that recognized distinct antigens specific to the strain they were challenged with, A141 bound to antigens in the 50-250Kda range and IR004 bound to a distinct 150Kda antigen. Western blots further indicated that each strain shared antigenic regions regardless of experimental Aeromonas strain exposure. Finally, immunofluorescent staining confirmed that mAb 27C binds to membrane-bound IgM (presumably B cells) on burbot head kidney cells. Taken together, results from this study demonstrate that mAb 27C specifically recognized burbot IgM and will be an important tool to further characterize the adaptive and cellular immune responses of this fish species.

  • Alternative adaptive immunity strategies: coelacanth, cod and shark immunity

    2016, Molecular Immunology

    The advent of high throughput sequencing has permitted to investigate the genome and the transcriptome of novel non-model species with unprecedented depth. This technological advance provided a better understanding of the evolution of adaptive immune genes in gnathostomes, revealing several unexpected features in different fish species which are of particular interest. In the present paper, we review the current understanding of the adaptive immune system of the coelacanth, the elephant shark and the Atlantic cod.

    The study of coelacanth, the only living extant of the long thought to be extinct Sarcopterygian lineage, is fundamental to bring new insights on the evolution of the immune system in higher vertebrates. Surprisingly, coelacanths are the only known jawed vertebrates to lack IgM, whereas two IgD/W loci are present. Cartilaginous fish are of great interest due to their basal position in the vertebrate tree of life; the genome of the elephant shark revealed the lack of several important immune genes related to T cell functions, which suggest the existence of a primordial set of TH1-like cells. Finally, the Atlantic cod lacks a functional major histocompatibility II complex, but balances this evolutionary loss with the expansion of specific gene families, including MHC I, Toll-like receptors and antimicrobial peptides.

    Overall, these data point out that several fish species present an unconventional adaptive immune system, but the loss of important immune genes is balanced by adaptive evolutionary strategies which still guarantee the establishment of an efficient immune response against the pathogens they have to fight during their life.

  • Francisella noatunensis in Atlantic cod (Gadus morhua L.); waterborne transmission and immune responses

    2011, Fish and Shellfish Immunology

    Citation Excerpt :

    Knowledge about the bacterium, the routes of infection and the immune responses in cod is of vital importance for better control of the disease and for development of vaccines. The immune system of Atlantic cod seems to differ from several other bony fish species as the antibody responses are shown to be absent or low after immunization with Vibrio salmonicida [4], Vibrio anguillarum [5] and hapten carriers [6,7]. In contrast to this, newer findings demonstrate an elevation of antibody levels against inactivated Aeromonas salmonicida, V. anguillarum and F. noatunensis [8].

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    This is the first report that confirms waterborne transmission of francisellosis in Atlantic cod. To investigate the transmission of disease, particle reduced water was transferred from a tank with intraperitoneally infected cod to a tank with healthy cod. Waterborne transmission of Francisella noatunensis was confirmed in the effluent group using immunohistochemistry and real-time quantitative PCR (RT-qPCR). The bacteria were located inside the accumulated macrophage-like cells. Specific and high antibody responses against live and inactivated bacteria were observed. Oil adjuvant had no effect on the antibody responses against inactivated F.noatunensis compared to saline formulation. The antigen epitope was a 20–25kDa component of F.noatunensis suggested to be lipopolysaccharide detected by Western blot, Sypro Ruby and Silver staining. Systemic immune reactions were investigated by measuring the expression of IFN-γ, IL-1β and IL-10 genes with RT-qPCR. After i.p. injection of live bacteria, a significant up-regulation of IFN-γ and IL-1β expression was observed from 15 to 60 days post infection in spleen and head kidney. In intestine, IFN-γ was significantly up-regulated after 30 days whereas rectum showed no significant differences in expression. Elevated expression of IL-10 was observed in all the organs tested but was only significantly up-regulated at 60 days post infection in intestine from i.p. infected fish. For the cohabitant group, IL-1β and IFN-γ was up-regulated in spleen whereas intestine and rectum showed a down-regulation after 60 days. IL-10 was up-regulated in intestine of cohabitant fish from day 30 to day 60. These results indicate that F.noatunensis infection provokes both specific antibody responses and long term inflammatory responses in cod. The present study provides new knowledge about infection routes and shows that both humoral and cellular defence mechanisms are triggered by F.noatunensis in cod.

  • Natural antibodies of cod (Gadus morhua L.): Specificity, activity and affinity

    2009, Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology

    Natural antibodies are present in the serum of vertebrates regardless of antigenic stimulation. Characteristic activity is commonly detected against haptenated proteins, single stranded DNA and thyroglobulin. Natural antibodies are believed to provide an instant protection against pathogens of a broad specificity and to participate in homeostasis. Cod is a poor antibody responder but shows a relatively high level of natural antibodies against haptenated proteins. In this project the specificity, activity and affinity of natural antibodies was studied in different groups of cod and the effects of age/size, environmental temperature, immunisation and infection examined. Antigen driven selection of natural antibodies was also studied in one group of cod. The results demonstrated a broad and yet characteristic specificity, primarily directed against haptenated proteins and possible food antigens. The antibody activity increased with increasing age and at higher temperature whereas immunostimulation by immunisation or infection resulted in variable response. The affinity index of natural antibodies of cod generally did not correlate with changes in the antibody activity but it was in the same range as the affinity index of acquired cod antibodies and that of some mammalian monoclonal acquired antibodies. Analysis of antigen driven antibody selection showed that the natural antibody repertoire of individual cod was heterogeneous with respect to its affinity for haptenated protein.

  • Comparison of antibody responses in Atlantic cod (Gadus morhua L.) to Vibrio anguillarum, Aeromonas salmonicida and Francisella sp.

    2009, Fish and Shellfish Immunology

    Citation Excerpt :

    Specific humoral antibody responses in cod immunised with V. anguillarum have been reported to be poor and primarily directed to LPS. Cod immune sera to serotype O2a and O2b were shown to distinguish between antigenic differences in the two sero-subtypes [21] and another study indicated that cod did not produce antibodies against a protein antigen [22,23]. Previously, we have shown that cod raised a weak but specific antibody response to V. anguillarum O2b [14], but only a few of the immunised individuals had raised antibodies with O-polysaccharide specificities that did not cross-react with either Va 1275 O2a or the serotype O2 isolates Va 1282 and Va 5022 on blot [16].

    Bacterial diseases such as vibriosis, atypical furunculosis and francisellosis, are registered as an increasing problem in cod farming in Norway. In order to develop efficient vaccines against diseases it is of interest to investigate if the cod immune system differentiates between various serotypes of Vibrio anguillarum and variants of Aeromonas salmonicida associated with the diseases by raising specific antibody responses. Cod of the same origin were shown to raise significant responses to V. anguillarum, A. salmonicida and the intracellular bacteria Francisella sp. Individual responses to V. anguillarum or A. salmonicida varied from none to high responses, while all individuals immunised with Francisella revealed a significant response. The cod immune system appeared in some degree to distinguish between V.anguillarum serotypes and A. salmonicida variants. Although all bacteria had induced significant antibody responses detectable in whole cell ELISA, only some had induced antibodies with specificity to linear O-polysaccharide epitopes on blot.

  • Characterization of IgM of Indian major carps and their cross-reactivity with anti-fish IgM antibodies

    2009, Fish and Shellfish Immunology

    Citation Excerpt :

    The heavy chain and light chain reported earlier for rohu are 85 and 23 kDa, respectively [5]. In general, the molecular weight of H and L chains varies from 60–77 to 23–26 kDa, respectively for most of the osteichthyan fishes but the higher molecular weight H chains, as found in Ig of IMCs, has also been observed in some other species such as cod (81 kDa) [16]; tilapia (90 kDa) [17] and barramundi (86 kDa) [18]. The bony fish Ig has been proved to be tetrameric [(H2L2)4] in nature by electron microscopic studies [19–21].

    Indian major carps (IMC), rohu (Labeo rohita), catla (Catla catla) and mrigal (Cirrhinus mrigala) were immunized with bovine serum albumin and the serum immunoglobulin M (IgM) was purified by affinity chromatography. The heavy and light chain of IgM of all the three species of IMC were about 88 and 26kDa, respectively. Anti-fish IgM antibody against all the three species were raised in mice and the reaction of anti-fish IgM antibodies with IgM of all the three species of IMC were studied by Western blot. The anti-fish IgM antibodies reacted strongly with the heavy chain of the same species against which it was raised while the reactions with the heavy chain of other species were milder indicating some degree of epitope sharing among the heavy chains of IgM of IMCs. However, there was no cross-reaction with the light chain of any of the IgM.

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