Sex-Biased Transcription Expression of Vitellogenins Reveals Fusion Gene and MicroRNA Regulation in the Sea Louse Caligus rogercresseyi.

The caligid ectoparasite, Caligus rogercresseyi, is one of the main concerns in the Chilean salmon industry. The molecular mechanisms displayed by the parasite during the reproductive process represent an opportunity for developing novel control strategies. Vitellogenin is a multifunctional protein recognized as a critical player in several crustaceans' biological processes, including reproduction, embryonic development, and immune response. This study aimed to characterize the C. rogercresseyi vitellogenins, including discovering novel transcripts and regulatory mechanisms associated with microRNAs. Herein, vitellogenin genes were identified by homology analysis using the reference sea louse genome, transcriptome database, and arthropods vitellogenin-protein database. The validation of expression transcripts was conducted by RNA nanopore sequencing technology. Moreover, fusion gene profiling, miRNA target analysis, and functional validation were performed using luciferase assay. Six putative vitellogenin genes were identified in the C. rogercresseyi genome with high homology with other copepods vitellogenins. Furthermore, miR-996 showed a putative role in regulating the Cr_Vitellogenin1 gene, which is highly expressed in females. Moreover, vitellogenin-fusion genes were identified in adult stages and highly regulated in males, demonstrating sex-related expression patterns. In females, the identified fusion genes merged with several non-vitellogenin genes involved in biological processes of ribosome assembly, BMP signaling pathway, and biosynthetic processes. This study reports the genome array of vitellogenins in C. rogercresseyi for the first time, revealing the putative role of fusion genes and miRNA regulation in sea lice biology.

Trypsin Genes Are Regulated through the miRNA Bantam and Associated with Drug Sensitivity in the Sea Louse Caligus rogercresseyi.

The role of trypsin genes in pharmacological sensitivity has been described in numerous arthropod species, including the sea louse Caligus rogercresseyi. This ectoparasite species is mainly controlled by xenobiotic drugs in Atlantic salmon farming. However, the post-transcriptional regulation of trypsin genes and the molecular components involved in drug response remain unclear. In particular, the miRNA bantam family has previously been associated with drug response in arthropods and is also found in C. rogercresseyi, showing a high diversity of isomiRs. This study aimed to uncover molecular interactions among trypsin genes and bantam miRNAs in the sea louse C. rogercresseyi in response to delousing drugs. Herein, putative mRNA/miRNA sequences were identified and localized in the C. rogercresseyi genome through genome mapping and blast analyses. Expression analyses were obtained from the mRNA transcriptome and small-RNA libraries from groups with differential sensitivity to three drugs used as anti-sea lice agents: azamethiphos, deltamethrin, and cypermethrin. The validation was conducted by qPCR analyses and luciferase assay of selected bantam and trypsin genes identified from in silico transcript prediction. A total of 60 trypsin genes were identified in the C. rogercresseyi genome, and 39 bantam miRNAs were differentially expressed in response to drug exposure. Notably, expression analyses and correlation among values obtained from trypsin and bantam revealed an opposite trend and potential binding sites with significant ΔG values. The luciferase assay showed a reduction of around 50% in the expression levels of the trypsin 2-like gene, which could imply that this gene is a potential target for bantam. The role of trypsin genes and bantam miRNAs in the pharmacological sensitivity of sea lice and the use of miRNAs as potential markers in these parasites are discussed in this study.

Functional diets modulate the acute phase protein response in Oncorhynchus mykiss subjected to chronic stress and challenged with Vibrio anguillarum.

The acute phase response to pathogens alters the production of proinflammatory cytokines that, in turn, activate the synthesis of acute phase proteins. These proteins neutralize, prevent, and indicate tissue damage, thereby influencing the specific immune response and allowing the organism to regain homeostasis. Functional diets based in pre- and probiotics are used in aquaculture to improve fish health and resistance to diseases, but there is an information gap on the mechanisms involved in these effects and if these diets are efficient when fish are raised under high stocking densities. This study aimed an evaluation of the acute phase response in Oncorhynchus mykiss fed functional diets supplemented with pre- and probiotics (i.e. mannan-oligosaccharides and Saccharomyces cerevisiae, respectively) and challenged by either Vibrio anguillarum or chronic stress via maintenance under high stocking densities. For this, the relative expression of acute phase response related genes in liver, and of inflammatory response related genes in head kidney was evaluated by RT-qPCR. The supplemented diets differentially modulated the acute phase protein response to the assessed challenge conditions, specifically evidencing an overexpression of the genes HAPT, SAA, LECT2, and IL-1ß under chronic stress and of HAPT, IL-1ß, IL8, and LECT2 at 24 h post-challenge with V. anguillarum. The observed early-stage regulation of acute phase proteins and of the immune response by the probiotic S. cerevisiae and by prebiotic mannan-oligosaccharides suggests that both supplements have high immunostimulatory potentials for fish farmed under high stocking densities.