Quantitative trait loci (QTL) associated with resistance of rainbow trout Oncorhynchus mykiss against the parasitic ciliate Ichthyophthirius multifiliis.

The parasitic ciliate Ichthyophthirius multifiliis has a low host specificity eliciting white spot disease (WSD) in a wide range of freshwater fishes worldwide. The parasite multiplies rapidly whereby the infection may reach problematic levels in a host population within a few days. The parasite targets both wild and cultured fish but the huge economic impact of the protozoan is associated with mortality, morbidity and treatment in aquacultural enterprises. We have investigated the potential for genetic selection of WSD-resistant strains of rainbow trout. Applying the DNA typing system Affymetrix® and characterizing the genome of the individual fish by use of 57,501 single nucleotide polymorphisms (SNP) and their location on the rainbow trout chromosomes, we have genetically characterized rainbow trout with different levels of natural resistance towards WSD. Quantitative trait loci (QTL) used for the selection of breeders with specific markers for resistance are reported. We found a significant association between resistance towards I. multifiliis infection and SNP markers located on the two specific rainbow trout chromosomes Omy 16 and Omy 17. Comparing the expression of immune-related genes in fish-with and without clinical signs-we recorded no significant difference. However, trout surviving the infection showed high expression levels of genes encoding IgT, T-cell receptor TCRß, C3, cathelicidins 1 and 2 and SAA, suggesting these genes to be associated with protection.

Impact of Pseudomonas H6 surfactant on all external life cycle stages of the fish parasitic ciliate Ichthyophthirius multifiliis.

A bacterial biosurfactant isolated from Pseudomonas (strain H6) has previously been shown to have a lethal effect on the oomycete Saprolegnia diclina infecting fish eggs. The present work demonstrates that the same biosurfactant has a strong in vitro antiparasitic effect on the fish pathogenic ciliate Ichthyophthirius multifiliis. Three life cycle stages (the infective theront stage, the tomont and the tomocyst containing tomites) were all susceptible to the surfactant. Theronts were the most sensitive showing 100% mortality in as low concentrations as 10 and 13 µg/ml within 30 min. Tomonts were the most resistant but were killed in concentrations of 100 µg/ml. Tomocysts, which generally are considered resistant to chemical and medical treatment, due to the surrounding protective cyst wall, were also sensitive. The surfactant, in concentrations of 10 and 13 µg/ml, penetrated the cyst wall and killed the enclosed tomites within 60 min. Rainbow trout fingerlings exposed to the biosurfactant showed no adverse immediate or late signs following several hours incubation in concentrations effective for killing the parasite. This bacterial surfactant may be further developed for application as an antiparasitic control agent in aquaculture.

Effect of ES products from Anisakis (Nematoda: Anisakidae) on experimentally induced colitis in adult zebrafish.

Inflammatory bowel disease (IBD) in developed countries is linked with elevated hygienic standards. One of the several factors involved in this question may be reduced exposure to the immunomodulatory effects of parasitic helminths. Several investigations on treatment of mice and humans with helminth-derived substances have supported this notion, but underlying mechanisms remain unclear. This study therefore dissects to what extent a series of immune-related genes are modulated in zebrafish with experimentally induced colitis following exposure to excretory-secretory (ES) products isolated from larval Anisakis, a widely distributed fish nematode. Adult zebrafish intrarectally exposed to the colitis-inducing agent TNBS developed severe colitis leading to 80% severe morbidity, but if co-injected (ip) with Anisakis ES products, the morbidity rate was 50% at the end of the experiment (48 hours post-exposure). Gene expression studies of TNBS-treated zebrafish showed clear upregulation of a range of genes encoding inflammatory cytokines and effector molecules and some induction of genes related to the adaptive response. A distinct innate-driven immune response was seen in both TNBS and TNBS + ES groups, but expression values were significantly depressed for several important pro-inflammatory genes in the TNBS + ES group, indicating protective mechanisms of Anisakis ES compounds on intestinal immunopathology in zebrafish.

Rainbow trout (Oncorhynchus mykiss) immune response towards a recombinant vaccine targeting the parasitic ciliate Ichthyophthirius multifiliis.

The protective effect in rainbow trout (Oncorhynchus mykiss) of an experimental subunit vaccine targeting antigens in the parasite Ichthyophthirius multifiliis has been evaluated and compared to effects elicited by a classical parasite homogenate vaccine. Three recombinant parasite proteins (two produced in E. coli and one in insect cells) were combined and injected i.p., and subsequently, protection and antibody responses were analysed. Both the experimental and the benchmark vaccine induced partial but significant protection against I. multifiliis when compared to control fish. Specific antibody responses of vaccinated trout (subunit vaccine) were raised against one neurohypophysial n-terminal domain protein #10 of three recombinant proteins, whereas the benchmark vaccine group showed specific antibody production against all three recombinant proteins. The immunogenic parasite protein #10 may be a potential vaccine candidate supplementing the protective I-antigen in future vaccine trials.

Experimental evidence for direct in situ binding of IgM and IgT to early trophonts of Ichthyophthirius multifiliis (Fouquet) in the gills of rainbow trout, Oncorhynchus mykiss (Walbaum).

Freshwater fish are able to mount a protective immune response against the parasite Ichthyophthirius multifiliis (Ich) following a non-lethal exposure. Factors involved in immunity comprise cellular and humoral factors, but antibodies have been suggested to play a prominent role in protection. However, host antibodies have not yet been demonstrated to bind to the parasite in situ. By the use of immunohistochemical techniques, this study demonstrated that IgT and IgM bind to surface structures, including cilia, on the early feeding stage of the parasite in the gills of immune rainbow trout, Oncorhynchus mykiss, shortly (2 h) after invasion. No binding of IgT and no or only a weak binding of IgM was observed on the parasites in the gills of similarly exposed but naïve rainbow trout. This study indicates that antibodies play an important part in the protection of immune fish against Ich although additional humoral and cellular factors may contribute to this reaction.

Occurrence of gyrodactylids on wild Atlantic salmon, Salmo salar L., in Danish rivers.

Three hundred and one parr of wild salmon, Salmo salar L., from three river systems in Denmark were examined in December 2005 and August 2006 for gyrodactylid infections. The species determination was done by morphological analyses of the opisthaptoral hard parts and molecular analyses of the internal transcribed spacer (ITS) region using PCR restriction fragment length polymorphism (RFLP) and sequencing. Five species of the genus Gyrodactylus were found: Gyrodactylus derjavini, Gyrodactylus teuchis, Gyrodactylus truttae, Gyrodactylus arcuatus and Gyrodactylus salaris. The parasite species G. derjavini dominated over G. teuchis and G. truttae and only one specimen of G. arcuatus and one specimen of G. salaris were recovered. This specimen of G. salaris had the same mutation in the ITS-region as a previously described non-pathogenic Danish G. salaris variant. The prevalence of gyrodactylids on the salmon ranged from 0% to 100% but the mean abundance never exceeded 12.8.