Study on Immune Response of Organs of Epinephelus coioides and Carassius auratus After Immersion Vaccination With Inactivated Vibrio harveyi Vaccine

Abstract

Immersion vaccination relies on the response of fish mucosa-associated lymphoid tissues, the Crucian carp (Carassius auratus) and Grouper (Epinephelus coioides) were researched in this paper to examine local mucosal immune responses and associated humoral system responses following immersion vaccination. We administered 1.5 × 10⁷ CFU/ml formalin-inactivated Vibrio harveyi cells and measured mucus and serum antibody titers as well as IgM, MHC II mRNA levels in immune organs. The mucosal antibody response preceded the serum response indicating a role for local mucosal immunity in immersion vaccination. IgM and MHC II mRNA levels were relatively greater for the spleen and head kidney indicating the importance and central position of systemic immunity. Expression levels were also high for the gills while skin levels were the lowest. IgM and MHC II mRNA levels were altered over time following vaccination and the hindgut, liver and spleen were similar indicating a close relationship, so the absolute value of r is used to analyze the correlation among different organs immunized. It can be inferred the existence of an internal immune molecular mechanism for Immune synergy hindgut-liver-spleen, from the peak time (14^th day), the relative ratio of genes expression in the same tissues between the immunized grouper and the control group (26 times), and Pearson correlation coefficient (0.8<|r|<1). Injection challenges with live V. harveyi indicated that the relative protection rates for the crucian carp and Grouper was basically the same at 44.4% and 47.4%, respectively. It is believe that crucian carp may be used as a substitute for the valuable grouper in immunity experiment, just from aspect of the relative percent survival (RPS) and how it changes with time. But they were not consistent about the IgM mRNA expression between that of crucian carp and grouper after immersion the Vibrio vaccine.

Keywords: IgM; MHC II; Pearson correlation coefficient(r); immersion vaccination; immune mechanism; immune synergy of hindgut-liver-spleen; mucosal immunity.

Figure 1

Time-and antibody titers of Carassius auratus (left) and Epinephelus coioides (right).

Figure 2

Relative quantification of crucian carp IgM gene mRNA in different organs. The relative abundance for IgM mRNA from the tissues was ranked as spleen > HK> gill >liver > hindgut>skin. The alterations in mRNA levels over time were similar for the gills, skin, hindgut and liver with increases at day 4 post-inoculation. And there were clear change difference between spleen and HK at this experiment, although both organs are central immune organs.

Figure 3

Relative quantification of grouper IgM gene mRNA in different organs. The relative abundance for IgM mRNA from the tissues was ranked as spleen > HK> liver>gill> hindgut>skin. The alterations in mRNA levels over time were similar for the hindgut, liver, and spleen with increases at day 14 post-inoculation. Organ IgM mRNA expression levels changed with time and the hindgut, liver and spleen followed the same temporal trend.

Figure 4

Relative quantification of grouper MHC II gene mRNA in different organs. The relative abundance for MHC II mRNA from the tissues was ranked as spleen > HK> liver>gill> hindgut>skin. The alterations in mRNA levels over time were similar for the hindgut, liver, and spleen with increases at day 14 post-inoculation. Organ MHC II mRNA expression levels changed with time and the hindgut, liver and spleen followed the same temporal trend.

Figure 5

Correlation analysis of grouper organs after immersion with inactivated V. harveyi vaccine. The relation of gene expression up-regulation was analyzed by the absolute value of r between the quantitative changes of gene expression in different organs after immunization; while the relation of gene expression up-regulation multiple was analyzed by the absolute r value of between the relative ratio of gene expression in different organs after immunization is analyzed, and this relative ratio is the ratio of gene expression in the same tissues between the immunized grouper and the control group.