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For the next #microtwjc session on Tuesday 3rd Feb 8pm GMT we will be discussing the following paper
Granulocytes Impose a Tight Bottleneck upon the Gut Luminal Pathogen Population during Salmonella Typhimurium Colitis
Published in December in PLoS Pathogens
The paper is available from the link below:
Sorry Im posting it so close to the session I forgot it was my turn!
Topological, chemical and immunological barriers are thought to limit infection by enteropathogenic bacteria. However, in many cases these barriers and their consequences for the infection process remain incompletely understood. Here, we employed a mouse model forSalmonella colitis and a mixed inoculum approach to identify barriers limiting the gut luminal pathogen population. Mice were infected via the oral route with wild type S. Typhimurium (S.Tm) and/or mixtures of phenotypically identical but differentially tagged S. Tm strains (“WITS”, wild-type isogenic tagged strains), which can be individually tracked by quantitative real-time PCR. WITS dilution experiments identified a substantial loss in tag/genetic diversity within the gut luminal S. Tm population by days 2–4 post infection. The diversity-loss was not attributable to overgrowth by S. Tm mutants, but required inflammation, Gr-1+ cells (mainly neutrophilic granulocytes) and most likely NADPH-oxidase-mediated defense, but not iNOS. Mathematical modelling indicated that inflammation inflicts a bottleneck transiently restricting the gut luminalS. Tm population to approximately 6000 cells and plating experiments verified a transient, inflammation- and Gr-1+ cell-dependent dip in the gut luminal S. Tm population at day 2 post infection. We conclude that granulocytes, an important clinical hallmark of S. Tm-induced inflammation, impose a drastic bottleneck upon the pathogen population. This extends the current view of inflammation-fuelled gut-luminal Salmonella growth by establishing the host response in the intestinal lumen as a double-edged sword, fostering and diminishing colonization in a dynamic equilibrium. Our work identifies a potent immune defense against gut infection and reveals a potential Achilles’ heel of the infection process which might be targeted for therapy.
Salmonella Typhimurium can colonize the human intestine and cause severe diarrhea. In recent years, it has become clear that this pathogen profits from inflammatory changes in the intestinal lumen, as the inflamed gut helps Salmonella to out-compete the resident microbiota. Granulocytes transmigrating into the gut lumen were found to “foster” luminal Salmonellagrowth by providing nutrients (used by Salmonella, not the microbiota) and by releasing growth inhibitors affecting the microbiota, but not the pathogen. In this study, we extend this “fostering” concept by showing that gut luminal Salmonella Typhimurium population is itself surprisingly vulnerable to the host’s inflammatory response. Indeed, inflammation reduces the size of the gut luminal Salmonella population by as much as 105-fold at day 2 post infection. Thus, triggering of mucosal inflammation is in fact a double-edged sword by providing S. Typhimurium with a relative growth advantage against the microbiota in the gut lumen and by killing 99.999% of the gut luminal pathogen population at day 2. However, the pathogen population can recover and grow up again during the subsequent days. This changes the current view: Inflammation is not simply “beneficial” for the pathogen in the gut lumen. Instead, pathogen growth in the inflamed gut must be considered as an equilibrium between inflammation-inflicted killing and fostering growth of the surviving bacteria.
1. Is the paper well written and concise?
2. Are the experiments well designed?
3. Do the results further our knowledge?
4. Anything you would have done differently?
If there is anything else you would like to discuss please use the comments box below.
See you all on Tues
Next week, on Tues 20th Jan at 8pm GMT we will be looking at this paper:
Insights into Vibrio cholerae Intestinal Colonization from Monitoring Fluorescently Labeled Bacteria
Yves A. Millet, David Alvarez, Simon Ringgaard, Ulrich H. von Andrian, Brigid M. Davis, Matthew K. Waldor
PLoS Pathog 10(10): e1004405. doi: 10.1371/journal.ppat.1004405
Vibrio cholerae, the agent of cholera, is a motile non-invasive pathogen that colonizes the small intestine (SI). Most of our knowledge of the processes required for V. cholerae intestinal colonization is derived from enumeration of wt and mutant V. cholerae recovered from orogastrically infected infant mice. There is limited knowledge of the distribution of V. choleraewithin the SI, particularly its localization along the villous axis, or of the bacterial and host factors that account for this distribution. Here, using confocal and intravital two-photon microscopy to monitor the localization of fluorescently tagged V. cholerae strains, we uncovered unexpected and previously unrecognized features of V. cholerae intestinal colonization. Direct visualization of the pathogen within the intestine revealed that the majority of V. choleraemicrocolonies attached to the intestinal epithelium arise from single cells, and that there are notable regiospecific aspects to V. cholerae localization and factors required for colonization. In the proximal SI, V. cholerae reside exclusively within the developing intestinal crypts, but they are not restricted to the crypts in the more distal SI. Unexpectedly, V. cholerae motility proved to be a regiospecific colonization factor that is critical for colonization of the proximal, but not the distal, SI. Furthermore, neither motility nor chemotaxis were required for proper V. choleraedistribution along the villous axis or in crypts, suggesting that yet undefined processes enable the pathogen to find its niches outside the intestinal lumen. Finally, our observations suggest that host mucins are a key factor limiting V. cholerae intestinal colonization, particularly in the proximal SI where there appears to be a more abundant mucus layer. Collectively, our findings demonstrate the potent capacity of direct pathogen visualization during infection to deepen our understanding of host pathogen interactions.
Vibrio cholerae is a highly motile bacterium that causes the diarrheal disease cholera. Despite our extensive knowledge of the genes and processes that enable this non-invasive pathogen to colonize the small intestine, there is limited knowledge of the pathogen’s fine localization within the intestine. Here, we used fluorescence microscopy-based techniques to directly monitor where and how fluorescent V. cholerae localize along intestinal villi in infected infant mice. This approach enabled us to uncover previously unappreciated features of V. cholerae intestinal colonization. We found that most V. cholerae microcolonies appear to arise from single cells attached to the epithelium. Unexpectedly, we observed considerable differences between V. cholerae fine localization in different parts of the small intestine and found that V. choleraemotility exerts a regiospecific influence on colonization. The abundance of intestinal mucins appears to be an important factor explaining at least some of the regiospecific aspects of V. cholerae intestinal localization. Overall, our findings suggest that direct observation of fluorescent pathogens during infection, coupled with genetic and/or pharmacologic manipulations of pathogen and host processes, adds a valuable depth to understanding of host-pathogen interactions.
- Was the paper well written? Clear? Easy to follow?
- Were the methods appropriate? Were there any methods/experiments that you thought were missing? Were the stats appropriate?
- Were the conclusions supported by the results? What impact do you think these results will have on the wider V cholera field?
- What experiments would you like to see done in the future to build upon this work?
- Anything else? Please comment in the box below
Looking forward to see you there (just search for #microtwjc on the night and the tweets should come up automatically). Please spread the word to your colleagues – the more the merrier 🙂