Potential biological control agents for the salmon louse Lepeophtheirus salmonis (Kroyer, 1837)

Abstract

The salmon louse, Lepeophtheirus salmonis (Krøyer), is an obligate ectoparasitic copepod that lives on the external surface of salmonid fish. It is the most common parasite on cage reared Atlantic salmon (Salmo salar L.), costing the aquaculture industry in Scotland millions of pounds each year to control. Traditional methods used to control sea lice have centred on the use of chemical pesticide treatments, which are expensive, hazardous to handle, potentially deleterious to the marine environment, and are sometimes ineffective. Furthermore, the misuse use of two previously efficacious chemotherapeutants has led to a build up of resistance in sea lice populations. The aims of this study were to investigate a potential alternative control strategy, concentrating on the isolation of the naturally occurring enemies of sea lice, and evaluating their potential for use as biological control agents. A screening protocol was undertaken to examine sea lice removed from harvest size farmed fish and wild Atlantic salmon, to look for the presence of epibiotic and hyperparasitic organisms on the external surfaces of the sea lice. A screening protocol was also undertaken to look for invasive microorganisms, such as fungal pathogens, internal symbionts, and other internal hyperparasites. Sea lice were examined microscopically for the presence of external epibionts and obvious signs of internal invasion by microorganisms and parasites. Surface sterilised sea lice were incubated on growth media to screen for the presence of fungal pathogens. Fresh tissue squashes were performed on lice showing clinical signs of infection and screening / diagnostic PCRs were used to detect and identify endosymbionts and invasive pathogens and parasites. Wax histology, TEM and SEM were used to further investigate host parasite interactions in order to evaluate pathogenicity where appropriate. In vitro and in vivo challenge trials were performed with an isolated hyperparasite to effect transmission and to determine pathogenicity. The stalked suctorian ciliates Ephelota gemmipara and Ephelota gigantea, and the monogenean worm Udonella sp. were frequently found utilising L. salmonis as a substrate. High densities of Udonella were observed on the cephalothoracic shield and genital segment of adult lice, and very high densities of Ephelota spp. were found on the genital segment, abdomen, and egg strings. The prevalence and seasonal pattern of occurrence of these epibionts were assessed. Fourteen fungal isolates were obtained, but failed to sporulate in vitro culture and were hence not identifiable. No endosymbionts were detected in either the body cavity or the egg strings by using the screening and diagnostic PCRs. An hyperparasitic microsporidian was found heavily parasitising adult sea lice. The microsporidian infection arose in the epidermal cells lying beneath the cuticle, infection was not observed in other tissues. Horizontal transmission was assumed as developing eggs and egg strings were not seen to be infected. The ultrastructure and complete developmental cycle of this hyperparasite in the salmon louse were described. Specific PCR primers were designed for use as a diagnostic tool and a molecular phylogeny was constructed using rRNA gene sequences. Taking into account its taxonomic positioning, its morphology and its unique characteristics, it appears that this parasite represents both a new genus and a new species. The microsporidian was not found at all farm sites visited and was not detected in sea lice from wild caught Atlantic salmon. The microsporidian infected up to 10% of lice sampled when most prevalent, but was sometimes absent, and showed no clear seasonal pattern in its occurrence. Transmission of the microsporidian hyperparasite to uninfected sea lice was not achieved and evidence for an alternative / intermediate host for the microsporidian is presented. This is the first report of a microsporidian from sea lice, and indeed from marine copepods; it is also the first report of a hyperparasitic microsporidian in crustacea. The potential for the epibionts and endobionts found in association with sea lice to be used as biological control agents are presented.