Without a pinch of salt: effect of low salinity on eggs and nauplii of the salmon louse (Lepeophtheirus salmonis).

The salmon louse is an economically important parasite on Atlantic salmon and poses a major threat to aquaculture. Several treatment methods have lost their effect due to resistance development in the lice. A rather new method for combatting sea lice is freshwater treatment where the various life stages of lice are differently affected by this treatment. In this study, we analyzed the effect of freshwater on the egg strings. A 3-h treatment with freshwater had a detrimental effect on the egg strings. First, the water penetrated the string, widening it, then entering the eggs and enlarging them. Finally, the ordered structure of the egg strings collapsed, and no alive animals hatched. Shorter treatments had a lower effectivity, and treatments with brackish water also showed milder effects. The egg strings were found to have a protective effect against low salinities, as hatched nauplii died rapidly under conditions that embryos survived. We also found that embryos react to low salinity on a molecular level by changing gene expression of several genes, when incubated in brackish water. Additionally, the hatching of embryos treated with brackish water was delayed in comparison to seawater controls.

The transcriptomic response of adult salmon lice (Lepeophtheirus salmonis) to reduced salinity.

Salmon lice (Lepeophtheirus salmonis) are marine parasitic copepods living on salmonids and are challenging for salmon aquaculture. One of several treatment methods is the application of freshwater to the fish which can lead to lice loss. However, lab experiments have shown that salmon lice, acclimated to seawater, are capable of surviving for several weeks in freshwater, when attached to a host. If not attached to a host, they die within a few hours in freshwater but can survive a longer time in brackish water. The molecular mechanisms involved in the adaptation to low salinity of the louse have not been identified yet. In this study we incubated salmon lice, being attached to a host, or detached, in seawater, brackish water and freshwater for 4 h and 1 d, sampled the animals and used RNA-Seq to identify genes involved in these mechanisms. Freshwater incubation led to a much stronger regulatory response than brackish water and a longer incubation time gave a stronger effect than a short incubation. Among the most interesting genes, upregulated in low salinity water are in addition to several transporters, several enzymes involved in amino acid metabolism and especially in the proline biosynthesis. A strong upregulation of these enzymes might lead to an accumulation of proline which is known to be used as an osmolyte in other species. While the RNA-Seq experiment was performed with female samples, qPCR showed that at least 10 genes regulated in females, were also regulated in males.

Host gill attachment causes blood-feeding by the salmon louse (Lepeophtheirus salmonis) chalimus larvae and alters parasite development and transcriptome.

BACKGROUND: Blood-feeding is a common strategy among parasitizing arthropods, including the ectoparasitic salmon louse (Lepeophtheirus salmonis), feeding off its salmon host's skin and blood. Blood is rich in nutrients, among these iron and heme. These are essential molecules for the louse, yet their oxidative properties render them toxic to cells if not handled appropriately. Blood-feeding might therefore alter parasite gene expression. METHODS: We infected Atlantic salmon with salmon louse copepodids and sampled the lice in two different experiments at day 10 and 18 post-infestation. Parasite development and presence of host blood in their intestines were determined. Lice of similar instar age sampled from body parts with differential access to blood, namely from gills versus lice from skin epidermis, were analysed for gene expression by RNA-sequencing in samples taken at day 10 for both experiments and at day 18 for one of the experiments. RESULTS: We found that lice started feeding on blood when becoming mobile preadults if sitting on the fish body; however, they may initiate blood-feeding at the chalimus I stage if attached to gills. Lice attached to gills develop at a slower rate. By differential expression analysis, we found 355 transcripts elevated in lice sampled from gills and 202 transcripts elevated in lice sampled from skin consistent in all samplings. Genes annotated with "peptidase activity" were among the ones elevated in lice sampled from gills, while in the other group genes annotated with "phosphorylation" and "phosphatase" were pervasive. Transcripts elevated in lice sampled from gills were often genes relatively highly expressed in the louse intestine compared with other tissues, while this was not the case for transcripts elevated in lice sampled from skin. In both groups, more than half of the transcripts were from genes more highly expressed after attachment. CONCLUSIONS: Gill settlement results in an alteration in gene expression and a premature onset of blood-feeding likely causes the parasite to develop at a slower pace.

Heavy and light chain homologs of ferritin are essential for blood-feeding and egg production of the ectoparasitic copepod Lepeophtheirus salmonis.

The salmon louse, Lepeophtheirus salmonis, is a hematophagous ectoparasite of salmonid fish. Due to its blood-feeding activity, the louse is exposed to great amounts of iron, which is an essential, yet potentially toxic mineral. The major known iron storage protein is ferritin, which the salmon louse encodes four genes of (LsFer1-4). Two of the ferritins are predicted to be secreted. These are one of the heavy chain homologs (LsFer1) and the light chain homolog (LsFer2). Here, we perform functional studies and characterize the two secreted ferritins. Our results show that knocking down LsFer1 and LsFer2 both negatively affect the parasite's physiology, as it is not able to properly feed and reproduce. In a starvation experiment, the transcript levels of both LsFer1 and LsFer2 decrease during the starvation period. Combined, these results demonstrate the importance of these genes for the normal parasite biology, and they could thus potentially be targets for pest management.

A scavenger receptor B (CD36)-like protein is a potential mediator of intestinal heme absorption in the hematophagous ectoparasite Lepeophtheirus salmonis.

Intestinal absorption of heme has remained enigmatic for years, even though heme provides the most bioavailable form of iron. The salmon louse, Lepeophtheirus salmonis, is a heme auxotrophic ectoparasite feeding on large quantities of blood from its host, the salmon. Here we show that a scavenging CD36-like receptor is a potential mediator of heme absorption in the intestine of the salmon louse. The receptor was characterized by a heme binding assay using recombinantly expressed protein, in situ hybridization and immunohistochemistry, as well as functional knockdown studies in the louse. A computational structural model of the receptor predicted a binding pocket for heme, as also supported by in silico docking. The mRNA and protein were expressed exclusively in the intestine of the louse. Further, knocking down the transcript resulted in lower heme levels in the adult female louse, production of shorter egg strings, and an overall lower hatching success of the eggs. Finally, starving the lice caused the transcript expression of the receptor to decrease. To our knowledge, this is the first time a CD36-like protein has been suggested to be an intestinal heme receptor.