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. 2022 Jun 8;30(6):836-847.e6.
doi: 10.1016/j.chom.2022.04.012. Epub 2022 May 13.

"V体育官网" Host cells subdivide nutrient niches into discrete biogeographical microhabitats for gut microbes

Megan J Liou  1 , Brittany M Miller  1 , Yael Litvak  2 , Henry Nguyen  1 , Dean E Natwick  3 , Hannah P Savage  1 , Jordan A Rixon  4 , Scott P Mahan  1 , Hirotaka Hiyoshi  5 , Andrew W L Rogers  1 , Eric M Velazquez  1 , Brian P Butler  6 , Sean R Collins  3 , Stephen J McSorley  4 , Rasika M Harshey  7 , Mariana X Byndloss  8 , Scott I Simon  9 , Andreas J Bäumler  10
Affiliations

"V体育官网" Host cells subdivide nutrient niches into discrete biogeographical microhabitats for gut microbes

Megan J Liou et al. Cell Host Microbe. .

Abstract

Changes in the microbiota composition are associated with many human diseases, but factors that govern strain abundance remain poorly defined. We show that a commensal Escherichia coli strain and a pathogenic Salmonella enterica serovar Typhimurium isolate both utilize nitrate for intestinal growth, but each accesses this resource in a distinct biogeographical niche VSports手机版. Commensal E. coli utilizes epithelial-derived nitrate, whereas nitrate in the niche occupied by S. Typhimurium is derived from phagocytic infiltrates. Surprisingly, avirulent S. Typhimurium was shown to be unable to utilize epithelial-derived nitrate because its chemotaxis receptors McpB and McpC exclude the pathogen from the niche occupied by E. coli. In contrast, E. coli invades the niche constructed by S. Typhimurium virulence factors and confers colonization resistance by competing for nitrate. Thus, nutrient niches are not defined solely by critical resources, but they can be further subdivided biogeographically within the host into distinct microhabitats, thereby generating new niche opportunities for distinct bacterial species. .

Keywords: Enterobacterales; Escherichia coli; Salmonella; biogeography; chemotaxis; gut microbiota; nitrate; nutrient niches. V体育安卓版.

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Conflict of interest statement (VSports最新版本)

Declaration of interests The authors declare no competing interests.

VSports手机版 - Figures

Figure 1:
Figure 1:. Visualization of microhabitats occupied by E. coli and S. Typhimurium in the colon of streptomycin pre-treated mice.
(A-C) Groups of streptomycin pre-treated lys-EGFP-ki mice (N = 4) were infected with the S. Typhimurium wild type (STm wt) (A), an avirulent invA spiB mutant (STm invA spiB) (B), or mock-infected (Mock) (C). Sections of the cecum were stained for EGFP (green fluorescence), the O12-antigen of S. Typhimurium lipopolysaccharide (LPS) (white fluorescence), actin (red fluorescence), and DAPI nuclear stain (blue fluorescence). (A) The red rectangle in the top panel indicates an area that is shown at higher magnification in the three bottom panels, which show staining for S. Typhimurium LPS (anti-STm LPS; left panel), EGFP synthesis in phagocytes (EGFP; middle panel) and an overlay (right panel). (D-G) Groups of streptomycin pre-treated C57BL/6 mice (N is indicated in panel G) were mock infected (Mock) or infected with an avirulent S. Typhimurium invA spiB mutant (STm invA spiB), a non-motile avirulent S. Typhimurium invA spiB fliC fljB mutant (STm invA spiB fliC fljB), E. coli Nissle 1917 (EcN wt), or a non-motile E. coli Nissle 1917 fliC mutant (EcN fliC). (D) Sections of the colon were stained for actin (red fluorescence), DAPI nuclear stain (blue fluorescence), and either the O12-antigen of S. Typhimurium LPS (white fluorescence, top panels), or the O6-antigen of E. coli Nissle 1917 LPS (white fluorescence, bottom panels). (E-G) Quantification of bacterial distance from epithelium. (E) Example of quantitative analysis of an immunofluorescence image (top left panel) by segmenting the actin brush border (actin boundary) and bacterial colonization zones (top right panel), removing background outside the masked areas (bottom left panel) and generating seed pixels within colonization zones to represent bacterial positions (bottom right panel). (F) Lines connecting each seed pixel with the closest pixel within the actin boundary (left panel) are colored to indicate their lengths according to the color scheme shown below. The graph on the right plots the number of lines (values) against their lengths for a section from a mouse infected with E. coli Nissle 1917 (EcN wt). (G) The graph plots the number of lines (values) against their lengths for a section from a mouse infected with an avirulent S. Typhimurium invA spiB mutant (STm invA spiB). (H) The graph shows the average interquartile range of distances from the actin boundary for the indicated bacterial strains (bars). Each symbol (dots/squares) represents data from one microscopic image. *, P < 0.05; ***, P < 0.001; ns, P > 0.05.
Figure 2:
Figure 2:. E. coli and S. Typhimurium use motility and energy taxis to access nitrate in distinct microhabitats.
(A-B) Groups of streptomycin pre-treated C57BL/6 mice (N is indicated by the number of dots) were infected with the indicated mixtures of E. coli Nissle 1917 (EcN) or S. Typhimurium (STm) strains and colon contents collected 6 days after infection (A) or 4 days after infection (B) to determine the competitive index. Bars represent geometric means ± geometric error. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, P ≥ 0.05.
Figure 3:
Figure 3:. Phagocyte-derived nitrate constructs a nutrient-niche for S. Typhimurium that can also be accessed by commensal E. coli.
(A-D) Groups of streptomycin pre-treated C57BL/6J mice were mock infected (Mock) or infected with E. coli Nissle 1917 (EcN wt), avirulent S. Typhimurium (STm invA spiB), or virulent S. Typhimurium (STm wt) and organs collected 4 days after infection. (A) Histological sections of the cecum were blinded and scored for lesions by a veterinary pathologist. Each bar represents data from one animal. (B) Transcript levels of the indicated pro-inflammatory genes were determined by quantitative real-time PCR in RNA isolated from the cecal mucosa. (C) Nitrate concentrations in cecal mucus scrapings determined by a modified Griess assay. (D) Levels of myeloperoxidase in cecal contents was determined by ELISA. (C and D) Each dot represents data from one animal. (E and F) Lethally irradiated C57BL6/J-Ly5.1 mice received a bone marrow transplant from wild-type (WT) C57BL/6 donor mice or Nos2-deficient donor mice. The resulting groups of bone-marrow chimera mice along with groups of C57BL/6 and Nos2-deficient donor mice were pre-treated with streptomycin and then infected with the indicated mixtures of S. Typhimurium strains. (E) Competitive index of S. Typhimurium strains recovered from colon contents. Bars represent geometric means ± geometric error. (F) Representative images showing flow cytometric analysis of intestinal cells from bone marrow chimera mice to distinguish hematopoietic cells of the recipient (CD45.1+) from hematopoietic cells of the C57BL/6 (top panel) or Nos2-deficient (bottom panel) donor (CD45.2+). (G) Groups of streptomycin pre-treated C57BL/6 mice (N is indicated by the number of dots) were infected with the indicated S. Typhimurium (STm) strains, followed by inoculation with E. coli (EcN) strain mixtures. Bars represent geometric means ± geometric error. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, P > 0.05.
Figure 4:
Figure 4:. Epithelial-derived nitrate constructs a nutrient-niche for E. coli that cannot be accessed by avirulent S. Typhimurium.
(A and B) The colonic epithelium (A) or cecal mucus (B) were collected from groups of mice (N is indicated by the number of dots) lacking expression of Pparg in the intestinal epithelium (Ppargfl/fl Villincre/−) or littermate controls (Ppargfl/fl Villin−/−). (A) Nos2 transcript levels were determined by quantitative real-time PCR in RNA isolated from preparations of the colonic epithelium. (B) Nitrate concentrations in mucus scrapings were determined by a modified Griess assay. (A and B) The boxes in the whisker blot represent the first to third quartiles, and the line indicates the median value. Each dot represents data from one animal. (C and D) Groups of mice lacking expression of Pparg in the intestinal epithelium (Ppargfl/fl Villincre/−) or littermate controls (Ppargfl/fl Villin−/−) were infected with the indicated S. Typhimurium (STm) or E. coli (EcN) strain mixtures. (C) The competitive index in colon contents was determined 4 days after infection. Each dot represents data from one animal. Bars represent geometric means ± geometric error. (D) Histological sections of the cecum were blinded and scored for lesions by a veterinary pathologist. Each bar represents data from one animal. *, P < 0.05; **, P < 0.01; ns, P > 0.05.
Figure 5:
Figure 5:. McpB and McpC exclude S. Typhimurium from a nutrient-niche containing nitrate.
(A) Genetic repertoire of chemotaxis receptor genes (blue and red arrows) in E. coli Nissle 1917 (left panel) and S. Typhimurium strain ATCC14028 (right panel). (B) Groups of streptomycin pre-treated C57BL/6J mice were infected with the indicated mixtures of avirulent S. Typhimurium (STm) strains and cecal contents were collected 6 days after infection to determine the competitive index. Bars represent geometric means ± geometric error. (C) Groups (N = 8) of streptomycin treated mice were infected with one of the indicated S. Typhimurium strains and the cecum was collected for sectioning two days later. The boxes in the whisker blot represent the first to third quartiles, and the line indicates the median value of the interquartile range of distances from the actin boundary. Each symbol (dots/squares) represents data from one microscopic image. ***, P < 0.001.
Figure 6:
Figure 6:. E. coli uses nitrate respiration to confer colonization resistance against Salmonella.
(A and B) Groups of streptomycin pre-treated C57BL/6J mice were inoculated with sterile LB broth (Mock) or with the indicated E. coli Nissle 1917 (EcN) strains. Two days after inoculation with E. coli, mice were challenged with a 1:1 mixture of S. Typhimurium (STm) wild type (wt) and a nitrate respiration-deficient mutant (narZ narG napA). (A) Total numbers of S. Typhimurium recovered from cecal contents. (B) Competitive index of the indicated S. Typhimurium strains. (C) Groups of streptomycin pre-treated C57BL/6J mice were inoculated with the indicated mixtures of S. Typhimurium and E. coli strains. The graph shows the competitive index comparing fitness of the S. Typhimurium strain with the E. coli strain. (A-C) Bars represent geometric means ± geometric error. Each symbol (circles or squares) represents data from one animal. *, P < 0.05; **, P < 0.01; ***, P < 0.001. (D) Model for the nutrient-niche occupied by commensal E. coli (left panel), virulent S. Typhimurium (middle panel) and the competition between both species for nitrate (right panel). Created with BioRender.com. NO, nitric oxide; O2−, superoxide radical; NO3−, nitrate.
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