A prospective study in severely injured patients reveals an altered gut microbiome is associated with transfusion volume

Abstract

Background: Traumatic injury can lead to a compromised intestinal epithelial barrier and inflammation. While alterations in the gut microbiome of critically injured patients may influence clinical outcomes, the impact of trauma on gut microbial composition is unknown. Our objective was to determine if the gut microbiome is altered in severely injured patients and begin to characterize changes in the gut microbiome due to time and therapeutic intervention.

Methods: We conducted a prospective, observational study in adult patients (n = 72) sustaining severe injury admitted to a Level I Trauma Center. Healthy volunteers (n = 13) were also examined. Fecal specimens were collected on admission to the emergency department and at 3, 7, 10, and 13 days (±2 days) following injury. Microbial DNA was isolated for 16s rRNA sequencing, and α and β diversities were estimated, according to taxonomic classification against the Greengenes database.

Results: The gut microbiome of trauma patients was altered on admission (i.e., within 30 minutes following injury) compared to healthy volunteers. Patients with an unchanged gut microbiome on admission were transfused more RBCs than those with an altered gut microbiome (p < 0.001). Although the gut microbiome started to return to a β-diversity profile similar to that of healthy volunteers over time, it remained different from healthy controls. Alternatively, α diversity initially increased postinjury, but subsequently decreased during the hospitalization. Injured patients on admission had a decreased abundance of traditionally beneficial microbial phyla (e.g., Firmicutes) with a concomitant decrease in opportunistic phyla (e.g., Proteobacteria) compared to healthy controls (p < 0.05). Large amounts of blood products and RBCs were both associated with higher α diversity (p < 0.001) and a β diversity clustering closer to healthy controls.

Conclusion: The human gut microbiome changes early after trauma and may be aided by early massive transfusion. Ultimately, the gut microbiome of trauma patients may provide valuable diagnostic and therapeutic insight for the improvement of outcomes postinjury.

Level of evidence: Prognostic and Epidemiological, level III.

Figure 1:. Alpha (α) and Beta (β) diversity for all injured patients over time and healthy controls.

A. α-diversity, as measured by observed OTUs, was significantly increased on admission (day 0) compared to healthy controls and subsequently decreased at day 5 (p<0.05). β-diversity is represented by principle components analysis plots for all injured patients over time and healthy controls for the following indices: B. Weighted UniFrac (Variability accounted for PC1=19.6%, PC2=13.4%, PC3=10.3%); C. Jaccard UniFrac (Variability accounted for PC1=3.5%, PC2=2.3%, PC3=1.8%); D. Bray-Curtis (Variability accounted for PC1=8.3%, PC2=3.6%, PC3=3.5%). B-diversity was significantly different between injured patients and healthy controls at all time points and compared to day 0 (p<0.05). Specifically, admission samples (red dots) displayed a massive spatial shift compared to healthy controls (yellow dots) indicating substantial dissimilarity in gut flora. While there is a general shift for the gut microbiome of trauma patients to resemble healthy controls as early as days 1-4 (orange dots), there are still significant differences on days 5-8 (green dots), days 9-12 (purple dots) and days 13+ (blue dots).

Figure 2:. Alpha (α) and Beta (β) diversity for all injured patients for total blood products, RBCs, and injury severity score (ISS).

Top row illustrates β-diversity represented by the Weighted UniFrac principle components analysis plots, while the bottom row shows α-diversity (i.e., observed OTUs) for total blood products (A), RBCs (B), and ISS score (C). The weighted UniFrac Index was significantly different according to amount of total blood products infused (Variability accounted for PC1=11.2%, PC2=7.2%, PC3=5.5%), wherein patients receiving none (orange dots) or low (red dots) amounts of total blood products transfused had significantly different microbiome than those getting large (purple dots) or extreme (green dots) of blood products. The same can be said RBCs (Variability accounted for PC1=23.0%, PC2=16.6%, PC3=8.8%), wherein patients receiving no RBCS (blue dots), or a low amount (orange dots) were significantly different than those receiving large amounts of RBCs (green dots). Additionally, β-diversity was different for those with an ISS score above 15 (red dots) versus those with an ISS under 15 (orange dots) (Variability accounted for PC1=23.0%, PC2=16.6%, PC3=8.8%). α-diversity also differed according to blood products and RBCs infused, but not by ISS score.

Figure 3:. Gut microbial composition following injury over time and healthy controls characterized by phyla.

The dominant phyla in both healthy controls and the post-injury groups at all time points were Firmicutes and Bacteroidetes. Relative abundance of the phylum Firmicutes was significantly decreased at days 0, days 1-4, days 5-8, days 9-12, and >13 days (*-p<0.05, ****-P<0.0001) compared to healthy controls. The relative abundance in the phylum of Proteobacteria was increased at day 0 following injury and at each time point compared to admission samples (^@@@@-p<0.0001).