Grandparental immune priming in the pipefish Syngnathus typhle

Abstract

Background: Phenotypic changes in response to environmental influences can persist from one generation into the next. In many systems parental parasite experience influences offspring immune responses, known as transgenerational immune priming (TGIP). TGIP in vertebrates is mainly maternal and short-term, supporting the adaptive immune system of the offspring during its maturation. However, if fathers and offspring have a close physical connection, evolution of additional paternal immune priming can be adaptive. Biparental TGIP may result in maximized immunological protection. Here, we investigate multigenerational biparental TGIP in the sex-role reversed pipefish Syngnathus typhle by exposing grandparents to an immune challenge with heat-killed bacteria and assessing gene expression (44 target genes) of the F2-generation.

Results: Grandparental immune challenge induced gene expression of immune genes in one-week-old grandoffspring. Similarly, genes mediating epigenetic regulation including DNA-methylation and histone modifications were involved in grandparental immune priming. While grand-maternal impact was strong on genes of the complement component system, grand-paternal exposure changed expression patterns of genes mediating innate immune defense.

Conclusion: In a system with male pregnancy, grandparents influenced the immune system of their grandoffspring in a sex-specific manner, demonstrating multigenerational biparental TGIP. The involvement of epigenetic effects suggests that TGIP via the paternal line may not be limited to the pipefish system that displays male pregnancy. While the benefits and costs of grandparental TGIP depend on the temporal heterogeneity of environmental conditions, multigenerational TGIP may affect host-parasite coevolution by dampening the amplitude of Red Queen Dynamics.

Keywords: Epigenetic inheritance; Gene expression; Grandparental effects; Host-parasite interaction; Immune defense; Immune priming.

Fig. 1

Experimental design. The grandparental generation (F0) was vaccinated using a combination of heat-killed immunological novel Vibrio spp. and Tenacibaculum maritimum (F0-bacteria), or were left naïve (F0-N) as control. Immune-challenged mature pipefish were used in following mating design: 1. Control: [♀F0-naïve x ♂F0-naïve]; 2. Paternal: [♀F0-naïve x ♂F0-bacteria]; 3. Maternal: [♀F0-bacteria x ♂F0-naïve] and 4. Biparental: [♀F0-bacteria x ♂F0-bacteria] and kept according to their mating pairs (families) in separate 36 × 80 L semi-flow through aquaria (16 family replicates per parental bacteria treatment and eight per control group; 56 families). F1-individuals were crossed within former parental treatment groups but left immunologically naïve (out of each of the four grandparental treatment groups five families were chosen to do F1-crosses resulting in 20 F1-families). In spring 2014, F2-juveniles were exposed one-week post birth to the same heat-killed Vibrio (F2-V+) and Tenacibaculum (F2-T+) bacteria used for the F0-generation or left naïve (F2-N) (per F1-crossing four families produced F2-offspring resulting in 16 F1-families). Out of each family 12 individuals were chosen for the direct immune challenge. Per F2-offspring treatment (F2-V+, F2-T+, F2-N) four individual replicates were used; resulting in a total of 192 samples

Fig. 2

Principle Component Analysis (PCA) depicting the grandparental bacteria treatment effect on gene expression of one-week-old F2-juveniles. PCA to visualize gene categories revealing a significant different gene expression profiles per grandparental control (F0-control), grand-paternal (F0-paternal), grand-maternal (F0-maternal) and grand-biparental (F0-biparental) bacteria treatment groups (Panels a-f) on relative gene expression data (−∆Ct-values) using an Euclidean distance matrix (N = 192). Panel a all immune genes (29 genes-total), Panel b genes of the innate immune system (13 genes), Panel c genes of the innate & adaptive immune system (5 genes); Panel d complement component genes (3 genes); Panel e epigenetic regulation genes (15 genes-total) and Panel f histone acetylation genes (2 genes). The variance in percentage (%) explained by the respective principle coordinates (PCs) is indicated below (for PC1) and besides (for PC2) the corresponding axis. The size (cm) of the grid is indicated by `d´ for dimension in the upper right corner

Fig. 3

Factor maps to demonstrate the contribution of variance retained by each principal component for immune genes (29 genes-total) and epigenetic regulation genes (15 genes-total) of one-week-old F2-juveniles. The response variables (genes) are symbolized by arrows whereby the length of the arrow is directional proportional with the contribution of variance of each gene to the total variability. The colour gradient in the left corner highlights the most important genes in explaining the variation (contribution %) retained by the principle components calculated according to [97]