Longitudinal Evaluation of Gut Bacteriomes and Viromes after Fecal Microbiota Transplantation for Eradication of Carbapenem-Resistant Enterobacteriaceae

Abstract

Understanding the role of fecal microbiota transplantation (FMT) in the decolonization of multidrug-resistant organisms (MDRO) is critical. Specifically, little is known about virome changes in MDRO-infected subjects treated with FMT. Using shotgun metagenomic sequencing, we characterized longitudinal dynamics of the gut virome and bacteriome in three recipients who successfully decolonized carbapenem-resistant Enterobacteriaceae (CRE), including Klebsiella spp. and Escherichia coli, after FMT. We observed large shifts of the fecal bacterial microbiota resembling a donor-like community after transfer of a fecal microbiota dominated by the genus Ruminococcus. We found a substantial expansion of Klebsiella phages after FMT with a concordant decrease of Klebsiella spp. and striking increase of Escherichia phages in CRE E. coli carriers after FMT. We also observed the CRE elimination and similar evolution of Klebsiella phage in mice, which may play a role in the collapse of the Klebsiella population after FMT. In summary, our pilot study documented bacteriome and virome alterations after FMT which mediate many of the effects of FMT on the gut microbiome community. IMPORTANCE Fecal microbiota transplantation (FMT) is an effective treatment for multidrug-resistant organisms; however, introducing a complex mixture of microbes also has unknown consequences for landscape features of gut microbiome. We sought to understand bacteriome and virome alterations in patients undergoing FMT to treat infection with carbapenem-resistant Enterobacteriaceae. This finding indicates that transkingdom interactions between the virome and bacteriome communities may have evolved in part to support efficient FMT for treating CRE.

Keywords: CRE; FMT; bacteriome; virome.

FIG 1

FMT to treat CRE. Timeline of sample collection for donors and CRE recipients. Results for CRE were based on rectal swabs from the recipients.

FIG 2

Analysis of bacterial composition of FMT donors and recipients. (A) Alpha diversity (Shannon diversity) and richness (Chao1 index) of fecal bacterial composition of donors, recipients before FMT, and recipients after FMT at different time points. (B) Relative abundance of Klebsiella spp. detected in all recipients substantially decreased after FMT. (C) PCoA based on Bray-Curtis distance for recipients and donors. PCoA plot showing separation of relative abundance of gut microbiota populations before and after FMT. (D) Fecal bacterial composition profile at the order level in donors and in CRE-colonized FMT recipients pre-FMT and post-FMT. Bacterial community composition changed toward that of donors, with increased Firmicutes and decreased Actinobacteria in recipients 2 weeks after FMT. The dominance of Firmicutes in stool of donors and CRE recipients post-FMT is denoted by the blue color. (E) Relative abundance of fecal bacterial taxa (at species level) of the donors and recipients.

FIG 3

Functionality alterations following FMT in donor and recipients. (A) Metagenomic functional predictions for donor, pre-FMT, and post-FMT samples. Mean relative gene pathway abundances for pre- and post-FMT samples were not significantly different (Mann-Whitney test; P > 0.05). Gene pathway abundances were calculated using the HUMAnN pipeline and grouped by major functional categories. (B) Abundance distribution of bacteria function pathways in donors and in recipients following FMT.

FIG 4

FMT influences gut microbiome interactions in patients with CRE. The correlation networks of the gut bacteriomes and gut viromes of patients with CRE before and after FMT (last collection sample from each recipient) and donors. Vertices indicate omics variables, and lines indicate a significant Pearson correlation coefficient at a |ρ| value of >0.6 and a P value of <0.05.

FIG 5

Analysis of viral composition of donors and CRE recipients, and alterations of bacteriophages following FMT. (A) Relative abundance of gut virome at the order level of donor and recipient before and after FMT. (B) Alpha diversity (Shannon’s diversity) of viromes in stools of donors and recipients at different time points after FMT. (C) Relative abundance of Klebsiella species in recipients (samples from recipient 1 prior to starting amoxicillin-clavulanic acid [Augmentin]). (D) Alterations of Klebsiella phages relative abundance from stool VLP metagenomes before and after FMT. (E) Alteration of Escherichia phages stool VLP metagenomes from three recipients before and after FMT. (F) Density plots of log₁₀ (read count per sample) distribution of Klebsiella phages.

FIG 6

FMT decolonizes carbapenem-resistant Klebsiella pneumoniae and reconstitutes the microbiota in mice. (A) Experimental scheme for Klebsiella pneumoniae challenge and treatment with FMT and with FVT alone. (B) PCoA of gut microbiota composition of mice stool identifies separation between control (gray), FMT (yellow), and FVT (blue) samples. (C to E) Fecal microbiota composition at the genus level of treated mice (5 mice per group). (F and G) Diversity (Shannon index) and richness of mouse gut bacteriomes.

FIG 7

Alteration of mice gut virome and impact of FMT in CRE-challenged mice. (A) Abundance distribution of virus in mouse stool following FMT. (B) The relative abundance of 10 predominant Klebsiella phages increased after FMT.