Contrasted survival under field or controlled conditions displays associations between mRNA levels of candidate genes and response to OsHV-1 infection in the Pacific oyster Crassostrea gigas

Abstract

Pacific oyster Crassostrea gigas suffers from chronic or sporadic mortality outbreaks worldwide, resulting from infectious diseases and/or physiological disorders triggered by environmental factors. Since 2008, ostreid herpesvirus OsHV-1 μVar has been identified as the main agent responsible for mass mortality of juvenile oysters in Europe. Previous studies of genome-wide expression profiling have provided candidate genes that potentially contribute to genetically-based resistance to summer mortality. To assess their value in determining resistance to the juvenile mass mortality that has occurred in France since 2008, we analyzed the expression of 17 candidate genes in an experimental infection by OsHV-1 μVar, and in an in vivo field experiment. Individual quantification of mRNA levels of 10 out of the 17 targeted genes revealed significant variation, of which 7 genes were showed differences between conditions that created significant differences in mortality, and 6 depended on the number of OsHV-1 genome copies individually quantified in mantle tissue. Complex SOD metalloenzymes known to be part of the antioxidant defense strategies may at least partly determine susceptibility or resistance to OsHV-1-associated mortality. Furthermore, inhibitor 2 of NF-κB, termed CgIκB2, exhibited highly significant variation of mRNA levels depending on OsHV-1 load in both experiments, suggesting its implication in the antiviral immune response of C. gigas. Our results suggest that CgIκB2 expression would make a good starting point for further functional research and that it could be used in marker-assisted selection.

Experimental infections of Pacific oyster Crassostrea gigas using the Australian ostreid herpesvirus-1 (OsHV-1) µVar strain

“…This result suggests that a sufficient initial dose of viral particles is needed to trigger high mortalities in Pacific oysters. Normand et al (2014) showed great differences in mortality between 2 groups injected with a virulent and a non-virulent inoculum at 2 × 10 9 and 4 × 10 6 copies of viral genomes µl −1 and prepared from dying and apparently healthy oysters, respectively. However, it is unclear whether the differences in mortality were only due to the virulence of the virus or whether the concentration of the inoculum might also have played a role.…”

“…Discrepancies occur in the literature when comparing the initial inoculum concentration and the mortalities obtained at 240 hpi after IM injection: Segarra et al (2014) observed a cumulative mortality of 63% at 90 hpi after the injection of 1.2 × 10 5 copies into 7 mo old oysters; Normand et al (2014) did not observe high mortalities (maximum cumulative mortality of 15% after 7 d of challenge) after the injection of 2 × 10 9 copies into 8 mo old oysters; and Schikorski et al (2011a) observed extreme mortality (> 90%) after the injection of 1.5 × 10 7 copies into 12 mo old oysters, which is in accordance with the results of the first 2 experiments presented in this paper. This variability in the short-term mortality kinetics after IM injection of a concentrated OsHV-1 inoculum highlights the potential issues and difficulties that may occur in the development of a reproducible and transferable infection model.…”

“…Mainly, these issues concern (1) the variation in the natural host due to the utilization of different oyster batches for each experiment; (2) the experimental parameters such as water renewal, density of infected host and nutrition; and (3) the variation in the viral inoculum itself (strain, virulence). Natural variation in the resistance to pathogens, particularly OsHV-1, is commonly observed in Pacific oyster populations and is mainly related to age, physiology, individual genetic basis and life history (Pernet et al 2012, Dégremont 2013, Normand et al 2014, Green et al 2014. Additionally, the density of the infected population within a tank may also greatly affect the kinetics of the disease.…”

“…Additionally, the density of the infected population within a tank may also greatly affect the kinetics of the disease. In the present study, a density of 8 to 10 oysters for 25 l of seawater (without food supply) was used, whereas Normand et al (2014) placed 40 infected oysters in 20 l tanks in a flow-through system with food, and Schikorski et al (2011a) used 100 naive oysters in cohabitation with 40 moribund animals in 25 l tanks. The water renewal regime was also different among studies.…”

“…Therefore some discrepancies between studies and/or infection trials seem quite inherent, and some precautions must be taken when comparing data from different studies. These discrepancies can also be explained by the variation in the viral inoculum itself (strain, virulence) as demonstrated by Normand et al (2014) as well as by its conservation and stability over time, as demonstrated by the results of Expt 3.…”