TRANSCRIPTOME ANALYSIS OF RED BLOOD CELLS IN RESPONSE TO THE RED SPOTTED GROUPER NERVOUS NECROSIS VIRUS (RGNNV)

Abstract

Red spotted grouper nervous necrosis virus (RGNNV) is the most widespread betanodavirus strain affecting numerous fish species, with European seabass (Dicentrarchus labrax) being highly susceptible. RGNNV causes up to 100% mortality in larvae and juveniles and significant losses in adult fish by targeting central nervous tissue, particularly the retina and brain.

In order to develop effective vaccines, the host immune response to the pathogen should be fully understood. However, in the infection produced by RGNNV in European sea bass, the role of RBCs is unknown. Nucleated RBCs are known to act as phagocytes, antigen-presenting cells, and modulate leukocyte activity, producing cytokines and other signaling molecules in response to viral infections (1).  The aim of this work was to study the participation of European sea bass RBCs in the immune response to RGNNV infection.

European sea bass purified RBCs were exposed ex vivo to RGNNV, and the expression of genes involved in the antiviral immune response was evaluated. On the other hand, in vivo assays were performed in which juvenile individuals were infected with RGNNV and at 1- and 3-days post infection brain and purified RBCs from peripheral blood samples were used to study gene expression by RT-qPCR. In addition, RBCs transcriptome was also evaluated by means of RNA-sequencing.

The ex vivo results showed that RGNNV was able to stimulate the expression of some genes related to the antiviral immune response in RBCs exposed to RGNNV, such as isg15 (an interferon-induced protein that has been implicated as a central player in the host antiviral response). The in vivo results showed that RGNNV was able to stimulate the expression of mx (a GTPase that is part of the antiviral response and is known to inhibit viral replication) in RBCs, both at 1- and 3-days post-infection. It is noteworthy that mx was also highly expressed in the brain, the target organ of RGNNV infection. The RNA-sequencing results showed over-representation of antiviral immune response pathways in RBCs from RGNNV infected European sea bass, depicting a clear response to the virus, mediated by cytokines and antiviral proteins.

References:

1. Nombela I, Requena-Platek R, Morales-Lange B, Chico V, Puente-Marin S, Ciordia S, Mena MC, Coll J, Perez L, Mercado L, Ortega-Villaizan M. Rainbow Trout Red Blood Cells Exposed to Viral Hemorrhagic Septicemia Virus Up-Regulate Antigen-Processing Mechanisms and MHC I&II, CD86, and CD83 Antigen-presenting Cell Markers. Cells. 2019 8(5):386.

Acknowledgements:

This study was funded by ThinkInAzul programme, GVA-THINKINAZUL/2021/020, supported by MCIN with funding from European Union NextGenerationEU (PRTR-C17.1) and by Generalitat Valenciana.