Transcriptomic Analysis of Yersinia enterocolitica Biovar 1B Infecting Murine Macrophages Reveals New Mechanisms of Extracellular and Intracellular Survival

Abstract

c Yersinia enterocolitica is typically considered an extracellular pathogen; however, during the course of an infection, a significant number of bacteria are stably maintained within host cell vacuoles. Little is known about this population and the role it plays during an infection. To address this question and to elucidate the spatially and temporally dynamic gene expression patterns of Y. enterocolitica biovar 1B through the course of an in vitro infection, transcriptome sequencing and differential gene expression analysis of bacteria infecting murine macrophage cells were performed under four distinct conditions. Bacteria were first grown in a nutrient-rich medium at 26°C to establish a baseline of gene expression that is unrelated to infection. The transcriptomes of these bacteria were then compared to bacteria grown in a conditioned cell culture medium at 37°C to identify genes that were differentially expressed in response to the increased temperature and medium but not in response to host cells. Infections were then performed, and the transcriptomes of bacteria found on the extracellular surface and intracellular compartments were analyzed individually. The upregulated genes revealed potential roles for a variety of systems in promoting intracellular virulence, including the Ysa type III secretion system, the Yts2 type II secretion system, and the Tad pilus. It was further determined that mutants of each of these systems had decreased virulence while infecting macrophages. Overall, these results reveal the complete set of genes expressed by Y. enterocolitica in response to infection and provide the groundwork for future virulence studies.T he genus Yersinia includes three species that are pathogenic to humans: Y. pestis, the causative agent of the plague, as well as Y. enterocolitica and Y. pseudotuberculosis, both of which cause gastrointestinal diseases. Of the three organisms, Y. enterocolitica is the species most frequently isolated from humans (1). Y. enterocolitica infections are typically acquired through ingestion of the bacteria in contaminated food or water, especially raw or undercooked pork products (2). After ingestion, the bacteria travel through the gastrointestinal tract to the terminal ileum, where they are able to penetrate the M cells of the Peyer's patches and infect the mesenteric lymph nodes (1-3). This leads to a self-limiting gastroenteritis and mesenteric lymphadenitis in otherwise healthy patients (3). In immunocompromised patients or young children with developing immune systems, the bacteria can spread from the lymph nodes to systemic sites, leading to potentially fatal septicemia (1). Over half of all reported Y. enterocolitica infections occur in children under the age of five, the group that is most predisposed to developing the systemic form of the infection (4).Pathogenic Y. enterocolitica isolates can be divided into six distinct biovars based on several biochemical and physiological attributes (2, 5). Of the six biovars, the North American isolate, biovar 1B, is the most...

Type II Secretion Is Necessary for Optimal Association of the Legionella-Containing Vacuole with Macrophage Rab1B but Enhances Intracellular Replication Mainly by Rab1B-Independent Mechanisms

“…Thus, our study represents the first documentation of a role for T2S in the association of the LCV with Rab1B. Although T2S was recently shown to be important for the intracellular survival or growth of Burkholderia cenocepacia, Chlamydia trachomatis, and Yersinia enterocolitica (113)(114)(115)(116), the current study is also the first demonstration of a link between T2S and Rab1 proteins associated with vesicular trafficking. The recruitment of the Rab1B GTPase to the LCV and the role that T4S effectors play in that process have been the focus of an extraordinary amount of research, revealing a remarkable array of enzymatic reactions.…”

Comparison ofYersinia enterocoliticaDNA methylation at ambient and host temperatures

“…Therefore, there is precedence for LTTR expression correlating with promoter methylation. Transcription of YE0914 in the RNA-seq transcriptomic analysis by Bent et al (Bent et al , 2015) does not show temperature-dependent regulation between Y. enterocolitica cells grown in LB at 26°C compared to conditioned RPMI at 37°C (Bent et al , 2015). However, the p-value in this study for YE0914 at these two temperatures is very high (0.65-0.93), suggesting significant heterogeneity in expression within sampled populations.…”

“…al. (Bent et al , 2015). This study is a comprehensive RNA-seq analysis of Y. enterocolitica 8081v transcripts prepared from cultures grown at 25°C in LB broth and from 37°C cultures grown in (i) conditioned RPMI, (ii) in surface contact with mouse macrophages, or (iii) internalized by mouse macrophages.…”

The YsrS Paralog DygS Has the Capacity To Activate Expression of the Yersinia enterocolitica Ysa Type III Secretion System

“…During mouse infections, activation of ysa expression was evident in intestinal and lymphatic tissue by 48 h postinfection (18). Upregulation of ysa genes was also detected from intracellular Y. enterocolitica during mouse macrophage tissue culture infection (19). Thus, the Ysa system may promote survival of Y. enterocolitica strains that fail to subvert phagocytosis.…”