Comparative susceptibilities and immune-related gene expressions of brown trout strains and their hybrids infected with Lactococcus garvieae and Yersinia ruckeri.

Brown trout are polymorphic salmonid species, and it is of importance to investigate whether hybridization affects disease resistance. In this study, susceptibility of brown trout (Salmo trutta Abant, Anatolian, Black Sea, and Caspius) strains and their hybrids to Lactococcus garvieae and Yersinia ruckeri as well as their immune-related gene expression profiles were studied. Results indicated that reciprocal hybridization did not affect disease resistance in brown trout strains. Purebred Black Sea strain of brown trout was the most resistant group against Y. ruckeri, followed by other Black Sea strain hybrids. On the other hand, purebred Anatolian strain was the most resistant group to L. garvieae, followed by other Anatolian strain hybrids. Expression pattern of target genes differed in families, but the overall gene expression was comparatively high in Y. ruckeri infected families. Upregulations were mainly significant at 7 and 28 d post infection while marginal regulations were observed 8 h after infection. Disease resistance status of strains was supported by high expression of immune-related genes such as major histocompatibility complex class I (MHC-I), immunoglobulin light chain (IgL), and antioxidant- and hemoglobin-related gene expression. Therefore, our findings suggest that Black Sea and Anatolian strains could be used to develop fish stock that are resistant for yersiniosis and lactocaccosis, respectively.

Genetic and molecular evidence that brown trout Salmo trutta belonging to the Danubian lineage are a single biological species.

We tested for reproductive isolation between Salmo trutta abanticus, S.t. labrax, S.t. caspius and S.t. fario by conducting crosses to produce F1 and F2 offspring. We also estimated the extent of genetic divergence between all three entities by examining sequence variation across the coI, d-loop and cytb mitochondrial genes. All of the F1 cross-types were successfully produced. After 2 years of culturing, F2 generation were produced as well. Fertilization, hatching and survival rates and hatching performance of F1 and F2 generations were evaluated. F2 generation had similar performance to their parent. Fertilization, hatching, larval survival rate and hatchery performance of F1 and F2 generation were similar except pure bred F2 S.t. abanticus. Purebred F1 individuals shared similar coloration patterns and spots with their parents but direction of the hybridization appeared to be decisive on morphology of hybrids. Some of the hybrids exhibit different morphological characters than their parents. Based on partial alignments of the three genes, phylogenetic analysis showed that these S. trutta are gathering within the same clade and appeared as monophyletic group. We found that there were some morphologic and genetic variation among S. trutta subspecies but the degree of variation does not warrant species level recognition. These findings indicate that the four subspecies constitute a single biological entity, corresponding to different morphs of the Danubian lineage. We therefore recommend that S. trutta belonging to Danubian lineage in Turkey be referred to as Salmo trutta and that strains be named according to location, such as Abant, Caspian, Black Sea and Anatolian.

Protection of rainbow trout against yersiniosis by lpxD mutant Yersinia ruckeri.

Yersinia ruckeri is a Gram negative bacteria causing yersiniosis in freshwater and marine fish. Lipid A, important for pathogenesis of Gram negative bacteria, biosynthesis pathway requires nine enzyme catalyzed steps. Although there are nine genes encoding lipid A biosynthesis in bacteria, biosynthesis of lipopolysaccharides relies on lpxD gene that encodes the third pathway enzyme. The roles of LpxD in Y. ruckeri virulence have not been studied. In the present study, in-frameshift deletion of lpxD gene and their role in Y. ruckeri virulence in rainbow trout were determined. For this purpose, 92% of the Y. ruckeri lpxD genes were deleted by homologous recombination. After running in SDS-PAGE and staining with silver stain, no LPS was detectable in the Y. ruckeri ΔlpxD mutant. Virulence and immunogenicity of the Y. ruckeri ΔlpxD mutant (YrΔlpxD) were determined in rainbow trout. Rainbow trout immunized with YrΔlpxD with immersion, or intraperitoneal injection method displayed superior protection (relative percentage survival ≥ 84%) after exposure to wild type Y. ruckeri. In conclusion, our results indicated that deletion of the lpxD gene causes significant attenuation of Y. ruckeri in rainbow trout, and LPS deficient YrΔlpxD could be used as a live attenuated vaccine against Y. ruckeri in rainbow trout. This vaccine can protect fish and it can be applied to fish with different methods such as immersion or injection.