Previously, our group has shown that the interbranchial lymphoid tissue (ILT) is a distinct structure largely consisting of T cells embedded in a meshwork of epithelial cells, with no direct resemblance to previously described lymphoid tissues. In this study, we aim to focus on the T cell population and the possibility of the ILT being a thymus analog. By characterizing structural responsiveness to Ag challenge, the presence of recombination activating genes, and different T cell-related transcripts, we attempt to further approach the immunological function of the ILT in salmonid gills. In addition to eight healthy individuals, a group of eight infectious salmon anemia virus-challenged fish were included to observe T cell responses related to infection. The results showed reduced size of ILT in the infected group, no expression of RAG-1 and -2, and a high degree of T cell diversity within the ILT. Taking into account that the ILT can be regarded as a strategically located T cell reservoir and possibly an evolutionary forerunner of mammalian MALTs right at the border to the external environment, the alteration in transcription observed may likely represent a shift in the T cell population to optimize local gill defense mechanisms.
Previously, it has been assumed that fish lack organized mucosa-associated lymphoid structures. Recently, an interbranchial lymphoid tissue (ILT) was described in salmonid gills at a site with substantial exposure to antigen. In this study, immune responses were examined in gills, mid-kidney and the laser-dissected ILT of Atlantic salmon (Salmo salar L.) infected with infectious salmon anaemia virus (ISAV). A strong innate response was observed in gills and mid-kidney and even in the laser-dissected ILT, despite the fact that no virus could be traced in this tissue. A small delayed increase in IgT transcripts, exclusively in the ILT, could indicate that this tissue has a role as a secondary lymphoid organ with clonal expansion of IgT expressing B-cells. Compared to the other examined tissues, gills displayed the earliest replication of the virus, further supporting this tissue as the main entry route for infection with ISAV.
Two strains of Atlantic salmon (Salmon salar) with different susceptibility to infectious salmon anaemia (ISA) were challenged with salmon pancreas disease virus (SPDV), the etiological agent of salmon pancreas disease (PD), by cohabitation. Serum and tissues were sampled at 0, 1, 3, 6 and 8 weeks post-challenge. Experimental challenge with SAV did not cause mortality, but virus loads and assessment of histopathology indicated that the fish more resistant to ISAV (ISAHi) also was more resistant to PD. Eight weeks post-challenge, the ISAHi strain had higher titres of SAV-neutralising antibodies than the less resistant strain (ISALo). Transcript levels of four adaptive and six innate immune parameters were analysed by real-time RT-PCR in heart, head kidney (HK) and gills of both strains. Secretory IgM (sIgM) and CD8 levels differed most between the two salmon strains. The ISAHi strain had significantly higher levels of sIgM in HK at all samplings, and significantly higher CD8 levels in gills at most samplings. In heart, both sIgM and CD8 levels increased significantly during the challenge, but the increase appeared earlier for the ISALo strain. By hierarchical clustering analysis of mRNA levels, a clear segregation was observed between the two strains prior to the virus challenge. As the viral infection developed, the clustering divide between fish strains disappeared, first for innate and later for adaptive parameters. At eight weeks post-challenge, the divide had however reformed for adaptive parameters. Possible pair-wise correlation between transcript levels of immune parameters was evaluated by a non-parametric statistical test. For innate parameters, the extent of correlation peaked at 3 wpc in all tissues; this came rapidly for ISALo and more gradual for ISAHi. The ISAHi strain tended to show higher correlation for innate parameters in heart and gill than ISALo at early sampling times. For adaptive immune parameters, little correlation was observed in general, except for ISAHi in heart at 6 wpc. Overall, the observed differences in immune parameters may provide important clues to the causes underlying the observed difference in susceptibility to PD.
The present study was performed to address putative links between the immune and pigmentary systems. A pigment-producing leukocyte-like cell-line (SHK-1 cells) of Atlantic salmon (Salmo salar L.) was exposed to different temperatures, poly I:C, bacterin or infected with virus (infectious pancreatic necrosis virus or infectious salmon anaemia virus). The effect of this stimulation regarding the transcription-pattern of the tyrosinase gene family (melanin genes) and the immune-related genes MHC class II and IFN-1 was analysed using real-time RT-qPCR. At 10°C cultivation, tyrosinase and dopachrome tautomerase remained unregulated. At 15°C, a moderate up-regulation was induced, while at 20°C, these genes were up-regulated in an exponential manner over time. Temperature did not affect the transcription of the immune-related genes. Virus infections, poly I:C or bacterin had no influence on the transcription of the melanogenesis-related genes, but triggered the immune-related genes. Our findings revealed no connections between the pigmentary and immune systems, but demonstrated a hereto undiscovered temperature-effect on the tyrosinase gene family.
Infectious salmon anemia (ISA) is an orthomyxoviral disease that has had devastating effects on farmed Atlantic salmon. ISA is still a disease resulting in continued loss of revenues and therefore development of effective vaccines is of great importance. Commercial vaccines against ISA are available, but the efficacy is poorly described. There is little information about vaccine-induced immune factors preventing ISA virus (ISAV) infection today. In this study we assessed the protective effects and immunogenicity of vaccines containing three different quantities of the inactivated ISAV antigen. Our findings indicated that immunization induced effective protection in Atlantic salmon with a relative percent survival (RPS) as high as 86. The level of protection was correlated to the amount of ISAV antigen in the vaccine, and fish immunized with high antigen amounts produced detectable ISAV-specific and neutralizing antibodies. While ISAV infection was detectable in non-vaccinated control fish challenged by cohabitation, no infection was detected in fish immunized with high antigen amounts. After challenge, transcriptional analysis of selected immune-related genes demonstrated activation of innate immune responses in ISAV-infected control fish, but not in vaccine protected fish. This study furthers the knowledge about vaccine efficacy and vaccine-induced immunity to ISAV challenge in Atlantic salmon.
The Prion protein (PRNP/Prp) plays a crucial role in transmissible spongiform encephalopathies (TSEs) like Creutzfeldt-Jakob disease (CJD), scrapie and mad cow disease. Notwithstanding the importance in human and animal disease, fundamental aspects of PRNP/Prp function and transmission remains unaccounted for.
The adhesion to inert solid surfaces was explored as a novel approach for the enrichment of previously uncultured bacteria from natural microbial communities. Enrichments on solid steel, glass and synthetic polymeric surfaces were established using samples from five freshwater lakes, a marine microbial mat and an alpine soil, and were subsequently analysed by molecular fingerprinting and sequencing of their 16S rRNA gene fragments. The majority of the enriched phylotypes grouped with the Alphaproteobacteria, Betaproteobacteria or Bacteroidetes and in several cases were related to typical biofilm-forming species and genera. Most enrichments were most closely related to previously uncultured phylotypes and none had previously been cultivated from the original environments even when applying improved high throughput liquid cultivation techniques. Of the 13 phylotypes enriched from freshwater samples, seven were previously unknown, three matched so-far uncultured environmental clones, and three were identical to previously cultivated bacteria. Of the 17 phylotypes recovered from soil, 12 were previously unknown with five of these phylotypes representing novel genera, whereas five phylotypes were identical to previously cultured soil bacteria. The feasibility of the biofilm-enrichment approach was exemplified by the successful isolation of a not-yet cultured Betaproteobacterium that constituted a discernible component of the alpine soil microbial community in situ and exhibited only 93% similarity to its closest cultured relative. Based on these results, cultivation on solid surfaces represents a promising approach to recover isolates that have so far escaped cultivation as suspended cultures in liquid media.
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