High gene expression of inflammatory markers and IL-17A correlates with severity of injection site reactions of Atlantic salmon vaccinated with oil-adjuvanted vaccines

<p>Abstract</p> <p>Background</p> <p>Two decades after the introduction of oil-based vaccines in the control of bacterial and viral diseases in farmed salmonids, the mechanisms of induced side effects manifested as intra-abdominal granulomas remain unresolved. Side effects have been associated with generation of auto-antibodies and autoimmunity but the underlying profile of inflammatory and immune response has not been characterized. This study was undertaken with the aim to elucidate the inflammatory and immune mechanisms of granuloma formation at gene expression level associated with high and low side effect (granuloma) indices.</p> <p>Groups of Atlantic salmon parr were injected intraperitoneally with oil-adjuvanted vaccines containing either high or low concentrations of <it>Aeromonas salmonicida </it>or <it>Moritella viscosa </it>antigens in order to induce polarized (severe and mild) granulomatous reactions. The established granulomatous reactions were confirmed by gross and histological methods at 3 months post vaccination when responses were known to have matured. The corresponding gene expression patterns in the head kidneys were profiled using salmonid cDNA microarrays followed by validation by real-time quantitative PCR (qPCR). qPCR was also used to examine the expression of additional genes known to be important in the adaptive immune response.</p> <p>Results</p> <p>Granulomatous lesions were observed in all vaccinated fish. The presence of severe granulomas was associated with a profile of up-regulation of innate immunity-related genes such as complement factors C1q and C6, mannose binding protein, lysozyme C, C-type lectin receptor, CD209, Cathepsin D, CD63, LECT-2, CC chemokine and metallothionein. In addition, TGF-β (p = 0.001), IL-17A (p = 0.007) and its receptor (IL-17AR) (p = 0.009) representing T_H17 were significantly up-regulated in the group with severe granulomas as were arginase and IgM. None of the genes directly reflective of T_H1 T cell lineage (IFN-γ, CD4) or T_H2 (GATA-3) responses were differentially expressed.</p> <p>Conclusions</p> <p>Granulomatous reactions following vaccination with oil-based vaccines in Atlantic salmon have the profile of strong expression of genes related to innate immune responses. The expression of TGF-β, IL-17A and its receptor suggests an involvement of T_H17 T cell lineage and is in conformity with strong infiltration of neutrophils and macrophages into inflamed areas. Arginase upregulation shows that macrophages in these reactions are alternatively activated, indicating also a T_H2-profile. To what extent the expression of IL-17A and its receptor reflects an autoimmune vaccine-based reaction remains elusive but would be in conformity with previous observations of autoimmune reactions in salmon when vaccinated with oil-based vaccines.</p

Occurrence and antibiotic susceptibility of fish bacteria isolated from Oreochromis niloticus (Nile tilapia) and Clarias gariepinus (African catfish) in Uganda

Abstract The intention of this study was to identify the bacterial pathogens infecting Oreochromis niloticus (Nile tilapia) and Clarias gariepinus (African catfish), and to establish the antibiotic susceptibility of fish bacteria in Uganda. A total of 288 fish samples from 40 fish farms (ponds, cages, and tanks) and 8 wild water sites were aseptically collected and bacteria isolated from the head kidney, liver, brain and spleen. The isolates were identified by their morphological characteristics, conventional biochemical tests and Analytical Profile Index test kits. Antibiotic susceptibility of selected bacteria was determined by the Kirby-Bauer disc diffusion method. The following well-known fish pathogens were identified at a farm prevalence of; Aeromonas hydrophila (43.8%), Aeromonas sobria (20.8%), Edwardsiella tarda (8.3%), Flavobacterium spp. (4.2%) and Streptococcus spp. (6.3%). Other bacteria with varying significance as fish pathogens were also identified including Plesiomonas shigelloides (25.0%), Chryseobacterium indoligenes (12.5%), Pseudomonas fluorescens (10.4%), Pseudomonas aeruginosa (4.2%), Pseudomonas stutzeri (2.1%), Vibrio cholerae (10.4%), Proteus spp. (6.3%), Citrobacter spp. (4.2%), Klebsiella spp. (4.2%) Serratia marcescens (4.2%), Burkholderia cepacia (2.1%), Comamonas testosteroni (8.3%) and Ralstonia picketti (2.1%). Aeromonas spp., Edwardsiella tarda and Streptococcus spp. were commonly isolated from diseased fish. Aeromonas spp. (n = 82) and Plesiomonas shigelloides (n = 73) were evaluated for antibiotic susceptibility. All isolates tested were susceptible to at-least ten (10) of the fourteen antibiotics evaluated. High levels of resistance were however expressed by all isolates to penicillin, oxacillin and ampicillin. This observed resistance is most probably intrinsic to those bacteria, suggesting minimal levels of acquired antibiotic resistance in fish bacteria from the study area. To our knowledge, this is the first study to establish the occurrence of several bacteria species infecting fish; and to determine antibiotic susceptibility of fish bacteria in Uganda. The current study provides baseline information for future reference and fish disease management in the country

Alpha Interferon and Not Gamma Interferon Inhibits Salmonid Alphavirus Subtype 3 Replication In Vitro▿

Salmonid alphavirus (SAV) is an emerging virus in salmonid aquaculture, with SAV-3 being the only subtype found in Norway. Until now, there has been little focus on the alpha interferon (IFN-α)-induced antiviral responses during virus infection in vivo or in vitro in fish. The possible involvement of IFN-γ in the response to SAV-3 is also not known. In this study, the two IFNs were cloned and expressed as recombinant proteins (recombinant IFN-α [rIFN-α] and rIFN-γ) and used for in vitro studies. SAV-3 infection in a permissive salmon cell line (TO cells) results in IFN-α and IFN-stimulated gene (ISG) mRNA upregulation. Preinfection treatment (4 to 24 h prior to infection) with salmon rIFN-α induces an antiviral state that inhibits the replication of SAV-3 and protects the cells against virus-induced cytopathic effects (CPE). The antiviral state coincides with a strong expression of Mx and ISG15 mRNA and Mx protein expression. When rIFN-α is administered at the time of infection and up to 24 h postinfection, virus replication is not inhibited, and cells are not protected against virus-induced CPE. By 40 h postinfection, the alpha subunit of eukaryotic initiation factor 2 (eIF2α) is phosphorylated concomitant with the expression of the E2 protein as assessed by Western blotting. Postinfection treatment with rIFN-α results in a moderate reduction in E2 expression levels in accordance with a moderate downregulation of cellular protein synthesis, an approximately 65% reduction by 60 h postinfection. rIFN-γ has only a minor inhibitory effect on SAV-3 replication in vitro. SAV-3 is sensitive to the preinfection antiviral state induced by rIFN-α, while postinfection antiviral responses or postinfection treatment with rIFN-α is not able to limit viral replication

Molecular detection of tilapia lake virus (TiLV) genome in Nile tilapia (Oreochromis niloticus) from Lake Victoria

Tilapia lake virus (TiLV) is an emerging pathogen of Tilapiines associated with high mortalities of wild and farmed tilapia posing great threat to the fishery industry worldwide. The virus has been reported in Israel, Ecuador, Colombia, Thailand, Egypt, Taiwan, India and Malaysia. In this study, a reverse transcription polymerase chain reaction (RT-PCR) assay was developed and used to detect TiLV genome in Nile tilapia from Lake Victoria. Nile tilapia samples were collected from the Tanzanian (108 fish) and Ugandan (83 fish) parts of Lake Victoria in 2015 and 2016, respectively. Samples were screened for TiLV by using RT-PCR and the PCR products were sequenced. The findings show that out of the 191 fish examined, 28 had PCR products showing the presence of TiLV genome. The TiLV nucleic acids were detected in the spleen (10.99%, N=191), head kidney (7.69%, N=65), heart (3.45%, N=29) and liver (0.71%, N=140) samples while no PCR amplification was detected in the brain by the developed RT-PCR method. Generally, the findings show that the lymphoid organs, mainly comprising of the head kidney and spleen had the highest number of samples with positive nucleic acids for TiLV followed by heart samples. On the contrary, the liver and brain that have previously been shown to be target organs during acute infection either did not have or had the lowest level of TiLV nucleic acids detected in the present study. All the 28 sequences retrieved had an average length of 768 bp. A blast analysis on NCBI showed that all sequences obtained were homologous to TiLV segment-2 sequences obtained from previous outbreaks in Israel and Thailand. To our knowledge, this is the first detection of TiLV subclinical infections in Nile tilapia in Lake Victoria, a none-outbreak area.Keywords: Lake Victoria, Nile tilapia, PCR, phylogenetic, surveillance, tilapia lake viru

Molecular detection of tilapia lake virus (TiLV) genome in Nile tilapia (Oreochromis niloticus) from Lake Victoria

Proceedings of the 35 scientific conference of the Tanzania Veterinary Association held at AICC, Arusha, December 2017.ilapia lake virus (TiLV) is an emerging pathogen of Tilapiines associated with high mortalities of wild and farmed tilapia posing great threat to the fishery industry worldwide. The virus has been reported in Israel, Ecuador, Colombia, Thailand, Egypt, Taiwan, India and Malaysia. In this study, a reverse transcription polymerase chain reaction (RT-PCR) assay was developed and used to detect TiLV genome in Nile tilapia from Lake Victoria. Nile tilapia samples were collected from the Tanzanian (108 fish) and Ugandan (83 fish) parts of Lake Victoria in 2015 and 2016, respectively. Samples were screened for TiLV by using RT-PCR and the PCR products were sequenced. The findings show that out of the 191 fish examined, 28 had PCR products showing the presence of TiLV genome. The TiLV nucleic acids were detected in the spleen (10.99%, N=191), head kidney (7.69%, N=65), heart (3.45%, N=29) and liver (0.71%, N=140) samples while no PCR amplification was detected in the brain by the developed RT-PCR method. Generally, the findings show that the lymphoid organs, mainly comprising of the head kidney and spleen had the highest number of samples with positive nucleic acids for TiLV followed by heart samples. On the contrary, the liver and brain that have previously been shown to be target organs during acute infection either did not have or had the lowest level of TiLV nucleic acids detected in the present study. All the 28 sequences retrieved had an average length of 768 bp. A blast analysis on NCBI showed that all sequences obtained were homologous to TiLV segment-2 sequences obtained from previous outbreaks in Israel and Thailand. To our knowledge, this is the first detection of TiLV subclinical infections in Nile tilapia in Lake Victoria, a none-outbreak area

Molecular detection of tilapia lake virus (TiLV) genome in Nile tilapia (Oreochromis niloticus) from Lake Victoria

Proceedings of the 35 scientific conference of the Tanzania Veterinary Association held at AICC, Arusha, December 2017.ilapia lake virus (TiLV) is an emerging pathogen of Tilapiines associated with high mortalities of wild and farmed tilapia posing great threat to the fishery industry worldwide. The virus has been reported in Israel, Ecuador, Colombia, Thailand, Egypt, Taiwan, India and Malaysia. In this study, a reverse transcription polymerase chain reaction (RT-PCR) assay was developed and used to detect TiLV genome in Nile tilapia from Lake Victoria. Nile tilapia samples were collected from the Tanzanian (108 fish) and Ugandan (83 fish) parts of Lake Victoria in 2015 and 2016, respectively. Samples were screened for TiLV by using RT-PCR and the PCR products were sequenced. The findings show that out of the 191 fish examined, 28 had PCR products showing the presence of TiLV genome. The TiLV nucleic acids were detected in the spleen (10.99%, N=191), head kidney (7.69%, N=65), heart (3.45%, N=29) and liver (0.71%, N=140) samples while no PCR amplification was detected in the brain by the developed RT-PCR method. Generally, the findings show that the lymphoid organs, mainly comprising of the head kidney and spleen had the highest number of samples with positive nucleic acids for TiLV followed by heart samples. On the contrary, the liver and brain that have previously been shown to be target organs during acute infection either did not have or had the lowest level of TiLV nucleic acids detected in the present study. All the 28 sequences retrieved had an average length of 768 bp. A blast analysis on NCBI showed that all sequences obtained were homologous to TiLV segment-2 sequences obtained from previous outbreaks in Israel and Thailand. To our knowledge, this is the first detection of TiLV subclinical infections in Nile tilapia in Lake Victoria, a none-outbreak area