MicroRNA responses associated with Salmonella enterica serovar typhimurium challenge in peripheral blood: effects of miR-146a and IFN-γ in regulation of fecal bacteria shedding counts in pig

Abstract

Background: MicroRNAs are involved in a broad range of biological processes and are known to be differentially expressed in response to bacterial pathogens.

Results: The present study identified microRNA responses in porcine peripheral blood after inoculation with the human foodborne pathogen Salmonella enterica serovar Typhimurium strain LT2. We compared the microRNA transcriptomes of the whole blood of pigs (Duroc × Landrace × Yorkshire) at 2-days post inoculation and before Salmonella infection. The analysis identified a total of 29 differentially expressed microRNAs, most of which are implicated in Salmonella infection and immunology signaling pathways. Joint analysis of the microRNA and mRNA transcriptomes identified 24 microRNAs with binding sites that were significantly enriched in 3' UTR of differentially expressed mRNAs. Of these microRNAs, three were differentially expressed after Salmonella challenge in peripheral blood (ssc-miR-146a-5p, ssc-miR-125a, and ssc-miR-129a-5p). Expression of 23 targets of top-ranked microRNA, ssc-miR-146a-5p, was validated by real-time PCR. The effects of miR-146a, IFN-γ, and IL-6 on the regulation of fecal bacteria shedding counts in pigs were investigated by in vivo study with a Salmonella challenge model.

Conclusions: The results indicated that induction of miR-146a in peripheral blood could significantly increase the fecal bacterial load, whereas IFN-γ had the reverse effect. These microRNAs can be used to identify targets for controlling porcine salmonellosis.

Keywords: IFN-γ; Immune response; Salmonella; Swine; miR-146a; microRNA.

Fig. 1

The miR-146a regulation network. The microRNA is plotted at the center of the graph (red). The gene regulators regulated by the microRNA are plotted around the microRNA (blue) with edges weighted by the number of target genes. The target genes are plotted as black nodes and labeled by the official gene symbol

Fig. 2

Hierarchical clustering of the gene expression data in PBMCs measured by real-time PCR. Cells were treated in vitro with three different doses of LPS (1 ng/ml, 10 ng/ml, and 100 ng/ml) and miR-146 overexpression or knockdown. Color codes of yellow, black, and blue represent expression levels of high, average, and low, respectively, across the treatments shown

Fig. 3

Effects of miR-146a, IFN-γ and IL-6 on Salmonella shedding counts in pigs. Salmonella shedding counts of a three (0 μg: black square, 10 μg: red cycle, and 50 μg: blue triangle) by three (Control, IFN-γ and IL-6) factor, completely randomized design with five animals in each group