Introduction
The rapid growth of aquaculture and the intensive culture conditions are leading to chronic stress situations favoring infections including those caused by viruses (Kibenge et al., 2019). Betanodavirus (NNV) is one of the main viral threatens in the Mediterranean coast. In larvae and juvenile stages, NNV infection reaches mortality rates up to 100%, but adults are asymptomatic carriers of infection. NNV can be both horizontal and vertically transmitted (Bandin & Souto, 2020). Focusing on vertical transmission, we must consider the ability of NNV to reach the gonad and establish a latent infection and the transmission of NNV through gametes to their progeny, which is highly susceptible (Valero et al., 2018a). Up to date, vaccines are the preventive method more explored. Although, maternal transfer of immune factors, defined as the transfer of innate or adaptive immune factors by an immunocompetent female to an immunologically naïve neonate has been described in fish (Swain & Nayak, 2009). This process could not explain why female vaccination improve the development of the immune system of the progeny in terms of gen expression up-regulation without promoting the transfer of maternal mRNAs, as recently demonstrated (Valero et al., 2023). Thus, maternal immune priming has been proposed to influence the immune development of the progeny in fish (Valero et la., 2023). However, nothing is known about the molecular pathways that orchestrated this issue. The objective of this study is to identified expression pathways modulated by females’ vaccination in both ovary and fertilized eggs using massive transcriptomics.
Material and methods
The females’ specimens of a mature broodstock, but not the males, were vaccinated against NNV with and effective vaccine (pBAD), produced and characterized previously (Gonzalez-Silvera et al., 2019) and a similar group but sham-vaccinated was established as control. Serum and ovary samples from vaccinated or sham-vaccinated females were obtained 21 days from the vaccination. Several pools of 0.08g of early fertilized eggs, 0 days post-hatching (dph), were obtained from vaccinated and sham-vaccinated broodstocks one year after females’ vaccination. Also, larvae from 4 and 9 dph were sampled. Anti-NNV IgM levels were analyzed in females’ serum samples (Valero et al., 2018b). Ovary and egg fertilized samples were submitted to mRNA-seq (García-Beltrán et al., 2024). Also, ovary, egg and larvae samples were used to analyze beta-defensins by qPCR (García-Beltrán et al., 2024).
Results
The efficiency of vaccination was confirmed by the production of anti-NNV IgM antibodies in vaccinated females, but not in the sham-vaccinated group. Transcriptomic analysis revealed the following differentially expressed genes (DEGs): 245 up-regulated and 3179 down-regulated genes in ovary while 3143 up-regulated genes and 4773 down-regulated genes in fertilized eggs, comparing in all cases between control and vaccinated groups. In ovary, there were six KEGG pathways up-regulated: ribosome, oxidative phosphorylation, spliceosome, cell cycle, oocyte meiosis and proteasome. In egg fertilized, there were ten KEGG pathways up-regulated including spliceosome, RNA degradation and mRNA surveillance pathway. Bdef expression was up-regulated at 4 and 9 dph in eggs fertilized from vaccinated females compared to controls.
Discussion and conclusions
None of the canonical immune pathways resulted altered. This issue together with the transcriptional study of Bdef revealed that their expression increased in larvae from vaccinated-females without a detectable maternal transfer of mRNAs as previously described (Valero et al., 2023). However, we detected some up-regulated pathways that could be related to immunity and viral infection resolution such as oxidative phosphorylation (Li et al., 2019) or spliceosome (Chang & Zhang, 2017). Moreover, other up-regulate pathways such as RNA degradation and mRNA surveillance might be orchestrating the previously observed effect of higher levels of expression of immune-related genes in the progeny from vaccinated-females. In fact, RNA degradation and mRNA surveillance pathway are considered to exert antiviral roles (Balistreri et al., 2017). In conclusion and although further studies are mandatory to ascertain all the molecular mechanism involved, our data suggest that general molecular regulation pathways of the cells might be the clue for understanding the maternal priming of immunity in the fish progeny.
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