Lumpfish (Cyclopterus lumpus) Is Susceptible to Renibacterium salmoninarum Infection and Induces Cell-Mediated Immunity in the Chronic Stage

Abstract

Renibacterium salmoninarum is a Gram-positive, intracellular pathogen that causes Bacterial Kidney Disease (BKD) in several fish species in freshwater and seawater. Lumpfish (Cyclopterus lumpus) is utilized as a cleaner fish to biocontrol sea lice infestation in Atlantic salmon (Salmo salar) farms. Atlantic salmon is susceptible to R. salmoninarum, and it can transfer the infection to other fish species. Although BKD outbreaks have not been reported in lumpfish, its susceptibility and immune response to R. salmoninarum is unknown. In this study, we evaluated the susceptibility and immune response of lumpfish to R. salmoninarum infection. Groups of lumpfish were intraperitoneally (i.p.) injected with either R. salmoninarum (1×107, 1×108, or 1×109 cells dose-1) or PBS (control). R. salmoninarum infection kinetics and mortality were followed for 98 days post-infection (dpi). Transcript expression levels of 33 immune-relevant genes were measured in head kidney (n = 6) of fish infected with 1×109 cells/dose and compared to the control at 28 and 98 dpi. Infected lumpfish displayed characteristic clinical signs of BKD. Lumpfish infected with high, medium, and low doses had a survival rate of 65%, 93%, and 95%, respectively. Mortality in the high-dose infected group stabilized after 50 dpi, but R. salmoninarum persisted in the fish tissues until 98 dpi. Cytokines (il1β, il8a, il8b), pattern recognition receptors (tlr5a), interferon-induced effectors (rsad2, mxa, mxb, mxc), and iron regulation (hamp) and acute phase reactant (saa5) related genes were up-regulated at 28 dpi. In contrast, cell-mediated adaptive immunity-related genes (cd4a, cd4b, ly6g6f, cd8a, cd74) were down-regulated at 28 dpi, revealing the immune suppressive nature of R. salmoninarum. However, significant upregulation of cd74 at 98 dpi suggests induction of cell-mediated immune response. This study showed that R. salmoninarum infected lumpfish in a similar fashion to salmonid fish species and caused a chronic infection, enhancing cell-mediated adaptive immune response.

Inactivated Aeromonas salmonicida impairs adaptive immunity in lumpfish (Cyclopterus lumpus)

“…The sequences, amplicon sizes and amplification efficiencies for all primer pairs used in the RT‐qPCR analyses are presented in Table 1. Primer pairs for interleukin 1b ( i1b ), interleukin 8a ( il8a ), interleukin 8b ( il8b ), interleukin 10 ( il10 ), immunoglobulin heavy chain 1 ( igm‐h1 ), immunoglobulin heavy chain 2 ( igm‐h2 ), immunoglobulin mu heavy chain 1 ( igm‐mu‐1 ), HLA class II histocompatibility antigen ( mhc‐II ), cluster of differentiation 4 ( cd4‐1 and cd4‐2 ) and cluster of differentiation 8 ( cd8 ) and the endogenous control genes, 60S ribosomal protein L32 ( rpl32 ), elongation factor 1‐alpha ( ef1a ), eukaryotic translation initiation factor 3 subunit D ( etf3d ), polyadenylate‐binding protein 1a ( pabpc1a ) and polyadenylate‐binding protein 1b ( pabpc1b ), were designed and quality control (QC) was tested previously (Gnanagobal et al., 2021). Ct values of rpl32, ef1a , etf3d , pabpc1a and pabpc1b were analysed to select the best two endogenous controls.…”

“…pabpc1b and rpl32 were utilized as the two endogenous controls in the experimental RT‐qPCR analyses (St‐Pierre et al., 2017). RT‐qPCR reactions were done using PowerUp‐SYBR Master Mix (Applied BioSystems, Foster City, CA, USA) in a QuantStudio 3 (Applied BioSystems, ThermoFisher, Waltham, MA, USA) as described previously (Gnanagobal et al., 2021).…”

Assessment of listing and categorisation of animal diseases within the framework of the Animal Health Law (Regulation (EU) 2016/429): Bacterial kidney disease (BKD)

“…Clinical signs of BKD have been reported in lumpfish (Cyclopterus lumpus) following intraperitoneal inoculation of R. salmoninarum (Gnanagobal et al, 2021) without evidence of being susceptible.…”

First detection of Cyclopterus lumpus virus in England, following a mortality event in farmed cleaner fish

“…Whilst little is known about the full impact of CLuV on both farmed and wild lumpfish, less is known about whether lumpfish can act as vectors for salmon diseases of major concern, since some pathogens possess a broad host species range. For example, both viral haemorrhagic septicaemia virus (VHSV) and Renibacterium salmoninarum (the causative agent of bacterial kidney disease) have been previously reported affecting lumpfish (Guðmundsdóttir et al 2018;Gnanagobal et al 2021). Therefore, lumpfish could act as a vector for the transmission of these, and indeed other significant pathogens of Atlantic salmon.…”