Screening of Streptococcus Suis serotype 2 resistance genes with GWAS and transcriptomic microarray analysis

Abstract

Background: Swine streptococcosis has caused great economic loss in the swine industry, and the major pathogen responsible for this disease is Streptococcus Suis serotype 2 (SS2). Disease resistance breeding is a fundamental way of resolving this problem. With the development of GWAS and transcriptomic microarray technology, we now have powerful research tools to identify SS2 resistance genes.

Results: In this research, we generated an F₂ generation of SS2 resistant C57BL/6 and SS2 susceptive A/J mice. With the F₂ generation of these two mice strains and GWAS analysis, we identified 286 significant mouse genome SNPs sites associated with the SS2 resistance trait. Gene expression profiles for C57BL/6 and A/J were analyzed under SS2 infection pressure by microarray. In total, 251 differentially expressed genes were identified between these two mouse strains during SS2 infection. After combining the GWAS and gene expression profile data, we located two genes that were significantly associated with SS2 resistance, which were the UBA domain containing 1 gene (Ubac1) and Epsin 1 gene (Epn 1). GO classification and over-representation analysis revealed nine up-regulated related to immune function, which could potentially be involved in the C57BL/6 SS2 resistance trait.

Conclusion: This is the first study to use both SNP chip and gene express profile chip for SS2 resistance gene identification in mouse, and these results will contribute to swine SS2 resistance breeding.

Keywords: Genome-wide association study; Resistance genes; Streptococcus suis serotype 2; Transcriptome analysis.

Fig. 1

Survival curves of A/J and C57BL/6 mice after infection with S. suis serotype 2. a Mice were injected i.p. with 1 mL 5 × 10⁷ CFU/mL SS2. b Mice were injected i.p. with 1 mL 5 × 10⁸ CFU/mL. Mortality of each group was recorded daily for 15 days. A/J mice showed significantly higher susceptibility to SS2 compared with C57BL/6 mice. (P<0.05 with Log-rank Test)

Fig. 2

Bacterial loads and antibody titers of F₂ progeny mice. a SS2 can be isolated from brains of all susceptible F₂ mice. Bacterial loads are calculated as CFU from 0.05 g brain tissue. b ELISA results show that all sera of SS2 resistant F₂ mice are SS2 antibody positive. Negative control sera were isolated from uninfected F₂ mice. ELISA results were read at an optical density (OD) 450 nm

Fig. 3

Manhattan plot of F₂ susceptible and F₂ resistant samples. Significant association signals were observed on the qB, qC1.1 cytobands of chromosome 2. Compared to other chromosomes, SNPs on qA1, qE3 cytobands of chromosome 7 and qF2 cytobands of chromosome 12 are also significant. Thus, these SNPs may be related to mouse SS2 susceptibility

Fig. 4

Schematic of different expression genes between C57BL/6 and A/J following infection with SS2. There are 105 up-regulated and 146 down-regulated genes in C57BL/6 relative to A/J mice

Fig. 5

GO classification (biological process) of 56 up-regulated genes (a) and 73 down-regulated genes (b) with known Entrez ID. The x-axis indicates the number of genes for each GO term

Fig. 6

GO over-representation analysis. a Up-regulated immunity related GO terms. Larger than expected numbers of up-regulated genes associated with the GO term biological processes are indicated by P-values. b Connections for the immunity related GO terms and their genes

Fig. 7

Enlarged Manhattan plots of interest SNP and related DEGs. Enlarged Manhattan plots demonstrate positions of significant SNPs and DEGs alongside cytobands. The red triangle indicates the significant SNP. X-axis shows the scale of chromosome. Strings below X-axis denote genes and their locations. Genes in the green fragment are SNP related DEGs