Exoelectrogenic capacity of host microbiota predicts lymphocyte recruitment to the gut

Abstract

Electrotaxis, directional cell movement in response to an electric potential, has been demonstrated in a wide range of cell types including lymphocytes. Exoelectrogens, microorganisms capable of generating electrical currents, have been identified in microbial fuel cells. However, no studies have investigated exoelectrogenic microbes in fresh feces or the effects of an exoelectrogenic microbiota on the host organism. Here we show that commensal gut microbial populations differ in their capacity for electrical current production by exoelectrogens and that those differences are predictive of increased lymphocyte trafficking to the gut in vivo, despite the lack of increased production of canonical lymphocyte-specific chemokines. Additionally, we demonstrate that the difference in current production between mice purchased from different commercial sources correlates reproducibly with the presence or absence of segmented filamentous bacteria, and while our data do not support a direct role for segmented filamentous bacteria in ex vivo current production, an exoelectrogenic microbiota can be transferred in vivo via mucosa-associated bacteria present in the ileum. Moreover, we detect upregulation of microbial genes associated with extracellular electron transfer in feces of mice colonized with exoelectrogenic microbiota containing segmented filamentous bacteria. While still correlative, these results suggest a novel means by which the gut microbiota modulates the recruitment of cells of the immune system to the gut.

Keywords: electrotaxis; exoelectrogen; microbiota; segmented filamentous bacteria.

Fig. 1.

Current production by murine gut microbiota varies according to commercial source of mouse and is transferrable via feces. Schematic diagram (A) and photograph (B) of microbial electrolysis cells (MECs) used to measure current production. Ti, titanium; SS, stainless steel; BHI, brain heart infusion. Current production was measured in MECs inoculated with fecal samples collected from C57BL/6J and C57BL/6NHsd (C), A/J and A/JOlaHsd (D), BALB/cJ and BALB/cAnNHsd (E), and C3H/HeJ and C3H/HeNHsd (F) mice. Current (μA) is shown as mean ± SE from 4 identically treated MECs per group, and data from Jackson and Harlan mice are shown in green and orange, respectively. G: current production by feces from C57BL/6J recipients prior to gavage and 1 C57BL/6NHsd donor mouse. H: current production by feces from C57BL/6J recipient mice postgavage. I: transmission electron microscopic image of SFB adherent to ileal mucosa.

Fig. 2.

Fecal electrical current production is transferrable via material adherent to ileal mucosa and correlates with lymphocyte recruitment. Current production was measured in MECs inoculated with fecal samples (A, n = 4) or ileal mucosal scrapes (B, n = 1) collected from C57BL/6J or C57BL/6NHsd mice. Data from Jackson (Jax) and Harlan (HSD) mice are shown in green and orange, respectively. C: current production by fecal samples collected from 12 C57BL/6J recipients prior to gavage. D: current production in MECs inoculated with feces collected from recipients 48 h postgavage with ileal scrape material collected from C57BL/6NHsd (orange) or C57BL/6J (green) mice or following no gavage (blue). Current (μA) is shown as mean ± SE. E: immunohistochemical staining of ileal tissue for CD3 was performed in recipient mice 6 wk postgavage with ileal scrape material. Mean (± SE) number of CD3⁺ (F) and B220⁺ (G) cells per villus. Bars denote P ≤ 0.05 as determined via Mann-Whitney nonparametric analysis. H: immunohistochemical staining for CCL21 in the ileum of recipient mice 6 wk postgavage.

Fig. 3.

Gavage with exoelectrogenic material results in increased lymphocyte recruitment to the gut. Current production was measured in MECs inoculated with feces (A, n = 4) or ileal mucosal scrapes (B, n = 1) collected from C57BL/6J or C57BL/6NHsd mice. Current (μA) is shown as mean ± SE, and data from Jax and HSD mice are shown in green and orange, respectively. C: current production by Rag1^−/− recipients (n = 8) prior to gavage of material collected from ileal mucosal scrape. D: current production by recipients postgavage with ileal mucosal scrape. Data from Jax and HSD mice are shown in green and orange, respectively. E: photomicrographs of hematoxylin and eosin (H&E)-stained sections of ileal lymphoid follicles in Rag1^−/− recipients following adoptive transfer of eGFP⁺ lymphocytes. F: confocal microscopic images of the regions shown in E. G: mean number (± SE) of eGFP⁺ cells detected in Rag1^−/− recipients. Bar denotes P ≤ 0.05 as determined via t-test. H&E-stained (H) and confocal microscopic (I) images of spleens from recipient mice in E and F. Production of CCL19 (J), CCL21 (K), and CCL25 (L) in colonocytes collected from Rag1^−/− recipients 48 h postgavage and cultured ex vivo for 36 h. M: fold difference in expression of OmcA and MtrC, normalized to that of the housekeeping gene GyrB, in feces of Rag1^−/− recipients as determined via comparative C_T analysis.*Significant difference (P < 0.05).

Fig. 4.

The presence of segmented filamentous bacteria (SFB) in feces correlates with current production and the presence of microbial nanowires in MECs. A: relative abundance of operational taxonomic units (OTUs) in mucosal scrapes collected from C57BL/6NHsd mice; legend at right. B: Venn diagram depicting overlap between OTUs in feces of HSD mice (C57BL/6NHsd, A/JOlaHsd, and BALB/cAnNHsd mice, n = 8/strain); Jax mice (C57BL/6J, A/J, and BALB/cJ mice, n = 8/strain); and the ileal mucosal scrapes depicted in A. Unweighted (C) and weighted (D) principal component analysis of the fecal microbiota of C57BL/6, A/J, and BALB/c mice from Jackson (orange circles) and Harlan (red triangles) Labs (n = 8 mice/strain/vendor) and mucosal scrape material (blue squares) collected from C57BL/6NHsd mice (n = 4). E: scanning electron microscopic images of microbes present in MECs inoculated with feces from C57BL/6NHsd (HSD) or C57BL/6J (Jax) mice.