Understanding host-microbiome interactions and influence on STEC colonization in cattle using integrated omics

• Author / Creator

  Pan, Zhe

• Shiga toxin producing Escherichia coli (STEC) is the major foodborne pathogen in humans with Shiga toxin 1 (stx1) and 2 (stx2) being the main virulence factors. Cattle are the major reservoir of STEC with those shedding >104 CFU/g STEC being defined as super shedders (SS). The rectal anal junction (RAJ) is the primary colonization site of STEC and previous studies have revealed that both fecal and rectal mucosal microbiota impact STEC colonization at the RAJ. To date, the extent to which stx in STEC affects host-microbial interactions remains unidenfined. This thesis aimed to identify how host and fecal/mucosal microbiota respond to stxs and STEC colonization. Study 1 (Chapter 2) consisted of an epidemiological survey to reveal the abundance (DNA) and expression (RNA) of stx1 and stx2 in STEC as it was affected by sampling type (fecal vs. rectal mucosa) and breed (Angus, Charolais, Kinsella Composite Expression of stx2 was influenced by the expressions of host immune genes previously reported to be downregulated in SS including MS4A1, CCL21, CD19, and LTB. The random forest model and Boruta method further revealed that MS4A1 was the most predictive of stx2 expression, a response that appeared to be linked to host immunity. Study 2 (Chapter 3) performed amplicon sequencing to characterize differences in rectal digesta microbial profiles and interactions using samples collected from steers in which stx2 was not expressed (Stx2- group) and those with stx2 expressed (Stx2+ group). Although microbial diversities and similarities did not differ between the two groups, microbial networks were remarkably different, with group-specific microbes being the most connected taxa within the network. These results suggested that the expression of stx2 in bacteria altered microbial community structures even when their diversity and composition were comparable to the Stx2- group. Study 3 (Chapter 4) identified the variation in host transcriptomes in veal calves challenged with STEC O157 that lacked stx2a (WT group) or possessed stx2a (RE group) using rectal mucosa samples collected at pre- (T1), 7 days (T2), and 26 days post-challenge (T5). The stx2a is a subtype of stx2 and is critical for STEC pathogenicity. A total of 214 downregulated differentially expressed genes (DEGs) were identified in WT-T2 compared to RE-T2. No upregulated DEGs were identified in WT and RE at T2. At T5, a total of 152 upregulated DEGs and 45 GO terms were shared between WT and RE, while no downregulated DEGs were identified for WT and RE. Functional analysis revealed that WT inhibited responses at extracellular regions and impaired tissue barrier integrity at T2, while those responses were enhanced at T5. For RE, no functional variations were found at T2, with the aforementioned functions enhanced at T5. In study 4 (Chapter 5), cDNA amplicon sequencing was performed to characterize the activity of the rectal mucosa microbial profiles, interactions, and assembly as well as host-microbial interactions related to the expression of stx2a gene in RAJ mucosa colonized STEC O157. The rectal mucosa microbial diversities were not affected by the presence of stx2a in STEC. Instead, the dynamics of microbial interactions and assembly patterns differed in response to strain-specific STEC O157 colonization. The relative abundance of Paeniclostridium and Gallibacterium were identified as connectors in microbial networks and specialists in microbial assembly. Host immune responses varied after challenge with B-cell and T-cell signaling receptor pathways, antigen processing and presentations being upregulated regardless of stx2a production. The beneficial microbes (e.g. Prevotella) dominated interactions with host immune genes, while the opportunistic pathogen Paeniclostridium dominated interactions with expressions of host immune genes post challenge and such relationship depended on the production of stx2a. In summary, this thesis provides knowledge of host-microbial interactions in response to stx gene expression and STEC colonization. Our findings suggest that STEC colonization and stx gene expression could be systematically attributed to differences in genetic variations, host responses, and fecal/mucosal microbiota.

• Subjects / Keywords

  • Omics
  • Beef
  • Shiga toxin producing E.coli
  • RNA-seq
  • Amplicon seq

• Graduation date

  Fall 2023

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-jy8q-4g14

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.