Changes in Lepeophtheirus salmonis gene expression during host switching and selection

Abstract

Lepeophtheirus salmonis is a copepod parasite of salmonid fish, commonly referred to as sea lice, and is particularly pathogenic to Atlantic salmon (Salmo salar). Although endemic on wild salmon, L. salmonis infection intensity rarely reaches clinical disease levels in wild fish. The stationary nature and high density stocking of Atlantic salmon in marine aquaculture allows the intensity of L. salmonis infection to reach clinical disease levels. In certain areas, during high levels of infection, salmon often develop gross lesions, most commonly on the head, dorsal and perianal regions. The lesions, coupled with the stress induced by infection, can result in decreased feeding, secondary infections and mortality of fish. Over the past 40 years a number of chemotherapeutants have been used to control L. salmonis infection in farmed fish, however, reliance on a relatively small number of treatments has driven the development of drug resistant populations of L. salmonis. Due to the high cost of developing new drugs, whose use may be limited by resistance development or environmental impacts, development of alternative treatment methods is becoming a necessary research avenue. Preliminary studies have shown that release of compounds derived from non-salmonid fish can reduce infection of salmon. The content of this thesis has the dual goal of trying to elucidate some of the behaviours of adult L. salmonis during host switching, and the development of an in vitro assay to assess the effects of the anti-attachment compound allyl isothiocyanate. Host switching behaviour was assessed using a cohabitation of L. salmonis infected and uninfected fish. It was found that male lice reached an equal distribution between the two populations of fish, while female lice did not switch hosts. This equal distribution of male lice is suggested to increase access of each lice to food and potential mates. Eight genes associated with various facets of lice survival underwent gene expression analysis (CYP18 A1-like gene, Trypsin-1, tissue plasminogen activator precursor-like (TPAP)-like gene, cyptochrome p450 isoform 1-like gene, peroxinectin-like gene, leukocyte receptor cluster member 9-like (LRCM9) gene, glycene receptor α-2-like gene, and a nicotinic acetylcholine receptor subunit-like (nAChR) gene) and five salmon genes associated with immune status were analysed (Interleukin (IL)-1β, IL-8, IL-12, IgT, and Matrix Metalloproteinase 9 (MMP9)) in an attempt to reveal patterns in gene expression during host colonization. Only one instance of significant difference occurred during the study, between MMP9 in the spleen of initially infected salmon and the spleen of initially uninfected salmon at 2 days post cohabitation. It was thought that the variability in gene expression of individual lice and fish that accompanied the use of outbred populations may have masked potential changes in gene expression. Pearson correlations were used to compare gene expression between individual lice and its respective host; expression of a peroxinectin-like gene had several instances of significant correlation with expression of host genes. This is thought to be caused either by peroxinectin-like having a role in modulation of host immune response, or by peroxinectin-like being expressed in response to the salmon reaction to infection. It appears that fish with low infection have more instances of significant correlation with several L. salmonis than fish with high infection levels. In vitro bioassays designed to assess lice response to allyl isothiocyanate revealed that adult and copepodid lice experienced increased mortality in higher treatment doses (>1ppm). Gene expression of 7 genes associated with different facets of survival were analyzed for dose dependent effects of allyl isothiocyanate on L. salmonis. The outcomes of this study suggests allyl isothiocyanate causes a significant amount of stress to L. salmonis, possibly causing the initiation of an immune response (increased then decreased LRCM9-like gene), reduced feeding (decreased Trypsin-1), and a nociceptive response (increased nAChR-like gene). Immobility was observed in lice at the high dose treatment, which is not necessarily one of the intended endpoints and may not be an achievable concentration in any management strategy, but provides evidence to suggest that exposure to allyl isothiocyanate may induce avoidance behaviour, reduce potential settlement on the host, and ultimately have a negative impact on lice survival