RNA-Seq analysis of ileocecal valve and peripheral blood from Holstein cattle infected with Mycobacterium avium subsp. paratuberculosis revealed dysregulation of the CXCL8/IL8 signaling pathway

Abstract

Paratuberculosis is chronic granulomatous enteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). Whole RNA-sequencing (RNA-Seq) is a promising source of novel biomarkers for early MAP infection and disease progression in cattle. Since the blood transcriptome is widely used as a source of biomarkers, we analyzed whether it recapitulates, at least in part, the transcriptome of the ileocecal valve (ICV), the primary site of MAP colonization. Total RNA was prepared from peripheral blood (PB) and ICV samples, and RNA-Seq was used to compare gene expression between animals with focal or diffuse histopathological lesions in gut tissues versus control animals with no detectable signs of infection. Our results demonstrated both shared, and PB and ICV-specific gene expression in response to a natural MAP infection. As expected, the number of differentially expressed (DE) genes was larger in the ICV than in the PB samples. Among the DE genes in the PB and ICV samples, there were some common genes irrespective of the type of lesion including the C-X-C motif chemokine ligand 8 (CXCL8/IL8), apolipoprotein L (APOLD1), and the interferon inducible protein 27 (IFI27). The biological processes (BP) enriched in the PB gene expression profiles from the cows with diffuse lesions included the killing of cells of other organism, defense response, immune response and the regulation of neutrophil chemotaxis. Two of these BP, the defense and immune response, were also enriched in the ICV from the cows with diffuse lesions. Metabolic analysis of the DE genes revealed that the N-glycan biosynthesis, bile secretion, one-carbon pool by folate and purine metabolism were significantly enriched in the ICV from the cows with focal lesions. In the ICV from cows with diffuse lesions; the valine, leucine and isoleucine degradation route, purine metabolism, vitamin digestion and absorption and the cholesterol routes were enriched. Some of the identified DE genes, BP and metabolic pathways will be studied further to develop novel diagnostic tools, vaccines and immunotherapeutics.

Identification of loci associated with susceptibility to bovine paratuberculosis and with the dysregulation of the MECOM, eEF1A2, and U1 spliceosomal RNA expression

“…RNA-Seq data and genotypes. Samples from the animals included in this study were tested by ELISA for the detection of MAP-specific antibodies, qPCR for MAP DNA detection, bacteriological culture, and histopathological analysis, as previously described (Table 1) 18 . All control animals (N = 4) were negative for all the diagnostic tests.…”

“…In contrast, the infected cows had detectable lesions in gut tissues with distinct severity; focal/ multifocal (N = 6) or diffuse (N = 4). RNA-extraction from PB and ICV samples of the fourteen cows, RNA-Seq libraries preparation, and sequencing using an Illumina NextSeq500 sequencer were performed as previously described 18 . The raw reads were filtered by their length (minimum size 75 bp long) and percentage of ambiguous base N less than 10% using Prinseq-lite 0.20.4 25 .…”

Detection of latent forms of Mycobacterium avium subsp. paratuberculosis infection using host biomarker-based ELISAs greatly improves paratuberculosis diagnostic sensitivity

“…SPARC, also known as osteonectin, is a matrix protein that binds collagen, and is required for the development of granuloma-like structures during chronic infections [ 57 ]. In our previous RNA-Seq study [ 39 ], the protein-protein interaction analysis revealed a COL1A2 centered network containing a COLI1A2-SPARC functional interaction. SPARC and COL1A2 were upregulated in the peripheral blood gene expression profiles from the cows with diffuse lesions which suggests that the expression of both proteins could lead to a bad prognosis in MAP-infected cows.…”

“…We previously identified several potential biomarkers for the diagnosis of MAP-infected animals [39]. Whole RNA-sequencing (RNA-Seq) was used to identify host genes differentially expressed in peripheral blood samples collected from animals with focal, or diffuse lesions in gut tissues vs. control animals without detected lesions nor bacterial load in gut tissues.…”

Epithelial processed Mycobacterium avium subsp. paratuberculosis induced prolonged Th17 response and suppression of phagocytic maturation in bovine peripheral blood mononuclear cells

“…Thanks to recent advances in sequencing technology, RNA-seq based transcriptomic analyses with MAP infection have been performed in vitro and in vivo environments [22][23][24][25] . However, in vivo studies could not elucidate the initial stage of infection related to persistence, such as the immune response to the formation of granuloma, and in vitro studies could not represent the actual course of infection because the infection models were mainly based on single cell types, such as monocyte-derived macrophages (MDMs).…”