Fecal Microbiota Transplant Restores Mucosal Integrity in a Murine Model of Burn Injury

Abstract

The gut microbiome is a community of commensal organisms that are known to play a role in nutrient production as well as gut homeostasis. The composition of the gut flora can be affected by many factors; however, the impact of burn injury on the microbiome is not fully known. Here, we hypothesized that burn-induced changes to the microbiome would impact overall colon health. After scald-burn injury, cecal samples were analyzed for aerobic and anaerobic colony forming units, bacterial community, and butyrate levels. In addition, colon and total intestinal permeabilities were determined. These parameters were further determined in a germ-reduced murine model. Following both burn injury and germ reduction, we observed decreases in aerobic and anaerobic bacteria, increased colon permeability and no change to small intestinal permeability. After burn injury, we further observed a significant decrease in the butyrate producing bacteria R. Gnavus, C. Eutactus, and Roseburia species as well as decreases in colonic butyrate. Finally, in mice that underwent burn followed by fecal microbiota transplant, bacteria levels and mucosal integrity were restored. Altogether our data demonstrate that burn injury can alter the microbiome leading to decreased butyrate levels and increased colon permeability. Of interest, fecal microbiota transplant treatment was able to ameliorate the burn-induced changes in colon permeability. Thus, fecal transplantation may represent a novel therapy in restoring colon health after burn injury.

Fig. 1. Burn injury decreases gut bacterial load and increases colon permeability

Mice were subjected to a 28% total body surface area dorsal scald-burn injury as described in the Methods. On post-burn day 6 (PBD6), cecal stool samples were cultured under (A) anaerobic or (B) aerobic conductions and CFU numbers determined. Sham or PBD6 mice underwent either (C) FITC dextran gavage to determine total intestinal permeability or (D) intestinal ligation proximal to the cecum and FITC dextran cecal injection to determine colon permeability. Sample size is 5 per group. *P<0.05, compared with sham as determined by Student t test.

Fig. 2. Antibiotic gavage decreases gut bacterial load and increases colon permeability

Mice underwent germ reduction by antibiotic gavage as described in the Methods. Isolated cecal stool samples from untreated or antibiotic-treated (ABX) mice were cultured under (A) anaerobic or (B) aerobic conductions and CFU numbers determined. Untreated or ABX mice underwent either (C) FITC dextran gavage to determine total intestinal permeability or (D) intestinal ligation proximal to the cecum and FITC dextran cecal injection to determine colon permeability. Sample size is 8 per group. *P<0.05, compared with saline as determined by Student t test.

Fig. 3. Burn injury and antibiotic gavage decrease gut butyrate levels

Mice were subjected to a 28% total body surface area dorsal scald-burn injury or germ reduction by antibiotic gavage as described in the Methods. Butyrate levels were determined from cecal stool samples isolated from (A) sham- or burn-injured mice (PBD6) or (B) untreated or antibiotic-treated mice. Sample size is 8 per group. *P<0.05 compared with sham as determined by Student t test.

Fig. 4. Burn injury selectively reduces gut bacterial phylum, class, family, and genus abundance

Mice were subjected to 28% total body surface area burn injury. On post-burn day 6, the mice were harvested and stool samples were collected from the cecum and sent for phylogenetic analysis using a sequence-specific hybridization assay of the entire 16S ribosomal RNA gene (V1–V3) to identify and measure relative abundance of >50,000 individual microbial taxa. The largest (A) phylum, (B) class, (C) family, and (D) genus from uninjured mice were compared with burn-injured mice. Sample size is 4 per group. *P<0.05 compared with sham as determined by Student t test.

Fig. 5. Burn injury selectively reduces butyrate producing bacterial strains Gnavus and Eutactus

Mice were subjected to 28% total body surface area burn injury. On post-burn day 6, the mice were harvested and stool samples were collected from the cecum and sent for phylogenetic analysis. The (A) Ruminococcus gnavus and (B) Coprococcus eutactus from uninjured mice were compared to burn-injured mice. Sample size is 4 per group. *P<0.05 compared with sham as determined by Student t test.

Fig. 6. FMT restores anaerobic bacterial counts and improves colon permeability after burn injury

Mice were subjected to a 28% total body surface area dorsal scald-burn injury as described in the Methods. On PBD2 and on PBD3, the mice underwent FMT as described. Sham or PBD6±FMT mice underwent intestinal ligation proximal to the cecum and FITC dextran cecal injection to determine colon permeability. Sample size is 8 per group. *P<0.05 compared with sham and PBD6 + FMT as determined by ANOVA followed by the Tukey post hoc comparison test. FMT indicates fecal microbiota transplant; PBD6, post-burn day 6.