Oral bacteria relative abundance in faeces increases due to gut microbiota depletion and is linked with patient outcomes.

The detection of oral bacteria in faecal samples has been associated with inflammation and intestinal diseases. The increased relative abundance of oral bacteria in faeces has two competing explanations: either oral bacteria invade the gut ecosystem and expand (the 'expansion' hypothesis), or oral bacteria transit through the gut and their relative increase marks the depletion of other gut bacteria (the 'marker' hypothesis). Here we collected oral and faecal samples from mouse models of gut dysbiosis (antibiotic treatment and DSS-induced colitis) and used 16S ribosomal RNA sequencing to determine the abundance dynamics of oral bacteria. We found that the relative, but not absolute, abundance of oral bacteria increases, reflecting the 'marker' hypothesis. Faecal microbiome datasets from diverse patient cohorts, including healthy individuals and patients with allogeneic haematopoietic cell transplantation or inflammatory bowel disease, consistently support the 'marker' hypothesis and explain associations between oral bacterial abundance and patient outcomes consistent with depleted gut microbiota. By distinguishing between the two hypotheses, our study guides the interpretation of microbiome compositional data and could potentially identify cases where therapies are needed to rebuild the resident microbiome rather than protect against invading oral bacteria.

Altered microbial bile acid metabolism exacerbates T cell-driven inflammation during graft-versus-host disease.

Microbial transformation of bile acids affects intestinal immune homoeostasis but its impact on inflammatory pathologies remains largely unknown. Using a mouse model of graft-versus-host disease (GVHD), we found that T cell-driven inflammation decreased the abundance of microbiome-encoded bile salt hydrolase (BSH) genes and reduced the levels of unconjugated and microbe-derived bile acids. Several microbe-derived bile acids attenuated farnesoid X receptor (FXR) activation, suggesting that loss of these metabolites during inflammation may increase FXR activity and exacerbate the course of disease. Indeed, mortality increased with pharmacological activation of FXR and decreased with its genetic ablation in donor T cells during mouse GVHD. Furthermore, patients with GVHD after allogeneic hematopoietic cell transplantation showed similar loss of BSH and the associated reduction in unconjugated and microbe-derived bile acids. In addition, the FXR antagonist ursodeoxycholic acid reduced the proliferation of human T cells and was associated with a lower risk of GVHD-related mortality in patients. We propose that dysbiosis and loss of microbe-derived bile acids during inflammation may be an important mechanism to amplify T cell-mediated diseases.

Immune-Related Colitis Is Associated with Fecal Microbial Dysbiosis and Can Be Mitigated by Fecal Microbiota Transplantation.

Colitis induced by treatment with immune-checkpoint inhibitors (ICI), termed irColitis, is a substantial cause of morbidity complicating cancer treatment. We hypothesized that abnormal fecal microbiome features would be present at the time of irColitis onset and that restoring the microbiome with fecal transplant from a healthy donor would mitigate disease severity. Herein, we present fecal microbiota profiles from 18 patients with irColitis from a single center, 5 of whom were treated with healthy-donor fecal microbial transplantation (FMT). Although fecal samples collected at onset of irColitis had comparable α-diversity to that of comparator groups with gastrointestinal symptoms, irColitis was characterized by fecal microbial dysbiosis. Abundances of Proteobacteria were associated with irColitis in multivariable analyses. Five patients with irColitis refractory to steroids and biologic anti-inflammatory agents received healthy-donor FMT, with initial clinical improvement in irColitis symptoms observed in four of five patients. Two subsequently exhibited recurrence of irColitis symptoms following courses of antibiotics. Both received a second "salvage" FMT that was, again, followed by clinical improvement of irColitis. In summary, we observed distinct microbial community changes that were present at the time of irColitis onset. FMT was followed by clinical improvements in several cases of steroid- and biologic-agent-refractory irColitis. Strategies to restore or prevent microbiome dysbiosis in the context of immunotherapy toxicities should be further explored in prospective clinical trials.

Modulating microbiome-immune axis in the deployment-related chronic diseases of Veterans: report of an expert meeting.

The present report summarizes the United States Department of Veterans Affairs (VA) field-based meeting titled "Modulating microbiome-immune axis in the deployment-related chronic diseases of Veterans." Our Veteran patient population experiences a high incidence of service-related chronic physical and mental health problems, such as infection, irritable bowel syndrome (IBS), inflammatory bowel disease (IBD), various forms of hematological and non-hematological malignancies, neurologic conditions, end-stage organ failure, requiring transplantation, and posttraumatic stress disorder (PTSD). We report the views of a group of scientists who focus on the current state of scientific knowledge elucidating the mechanisms underlying the aforementioned disorders, novel therapeutic targets, and development of new approaches for clinical intervention. In conclusion, we dovetailed on four research areas of interest: 1) microbiome interaction with immune cells after hematopoietic cell and/or solid organ transplantation, graft-versus-host disease (GVHD) and graft rejection, 2) intestinal inflammation and its modification in IBD and cancer, 3) microbiome-neuron-immunity interplay in mental and physical health, and 4) microbiome-micronutrient-immune interactions during homeostasis and infectious diseases. At this VA field-based meeting, we proposed to explore a multi-disciplinary, multi-institutional, collaborative strategy to initiate a roadmap, specifically focusing on host microbiome-immune interactions among those with service-related chronic diseases to potentially identify novel and translatable therapeutic targets.

Tissue-specific features of the T cell repertoire after allogeneic hematopoietic cell transplantation in human and mouse.

T cells are the central drivers of many inflammatory diseases, but the repertoire of tissue-resident T cells at sites of pathology in human organs remains poorly understood. We examined the site-specificity of T cell receptor (TCR) repertoires across tissues (5 to 18 tissues per patient) in prospectively collected autopsies of patients with and without graft-versus-host disease (GVHD), a potentially lethal tissue-targeting complication of allogeneic hematopoietic cell transplantation, and in mouse models of GVHD. Anatomic similarity between tissues was a key determinant of TCR repertoire composition within patients, independent of disease or transplant status. The T cells recovered from peripheral blood and spleens in patients and mice captured a limited portion of the TCR repertoire detected in tissues. Whereas few T cell clones were shared across patients, motif-based clustering revealed shared repertoire signatures across patients in a tissue-specific fashion. T cells at disease sites had a tissue-resident phenotype and were of donor origin based on single-cell chimerism analysis. These data demonstrate the complex composition of T cell populations that persist in human tissues at the end stage of an inflammatory disorder after lymphocyte-directed therapy. These findings also underscore the importance of studying T cell in tissues rather than blood for tissue-based pathologies and suggest the tissue-specific nature of both the endogenous and posttransplant T cell landscape.

Circulating microbial cell-free DNA is increased during neutropenia after hematopoietic stem cell transplantation.

We used a next-generation sequencing platform to characterize microbial cell-free DNA (mcfDNA) in plasma samples from patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HCT). In this observational study, we sought to characterize plasma mcfDNA in order to explore its potential association with the immunologic complications of transplantation. We compared serially collected patient samples with plasma collected from healthy control subjects. We observed changes in total mcfDNA burden in the plasma after transplantation, which was most striking during the early posttransplant neutropenic phase. This elevation could be attributed to a number of specific bacterial taxa, including Veillonella, Bacteroides, and Prevotella (genus level). For an additional cohort of patients, we compared the data of mcfDNA from plasma with 16s-ribosomal RNA sequencing data from stool samples collected at matched time points. In a number of patients, we confirmed that mcfDNA derived from specific microbial taxa (eg, Enterococcus) could also be observed in the matched stool sample. Quantification of mcfDNA may generate novel insights into mechanisms by which the intestinal microbiome influences systemic cell populations and, thus, has been associated with outcomes for patients with cancer.

High-resolution analyses of associations between medications, microbiome, and mortality in cancer patients.

Discerning the effect of pharmacological exposures on intestinal bacterial communities in cancer patients is challenging. Here, we deconvoluted the relationship between drug exposures and changes in microbial composition by developing and applying a new computational method, PARADIGM (parameters associated with dynamics of gut microbiota), to a large set of longitudinal fecal microbiome profiles with detailed medication-administration records from patients undergoing allogeneic hematopoietic cell transplantation. We observed that several non-antibiotic drugs, including laxatives, antiemetics, and opioids, are associated with increased Enterococcus relative abundance and decreased alpha diversity. Shotgun metagenomic sequencing further demonstrated subspecies competition, leading to increased dominant-strain genetic convergence during allo-HCT that is significantly associated with antibiotic exposures. We integrated drug-microbiome associations to predict clinical outcomes in two validation cohorts on the basis of drug exposures alone, suggesting that this approach can generate biologically and clinically relevant insights into how pharmacological exposures can perturb or preserve microbiota composition. The application of a computational method called PARADIGM to a large dataset of cancer patients' longitudinal fecal specimens and detailed daily medication records reveals associations between drug exposures and the intestinal microbiota that recapitulate in vitro findings and are also predictive of clinical outcomes.

The TaxUMAP atlas: Efficient display of large clinical microbiome data reveals ecological competition in protection against bacteremia.

Longitudinal microbiome data provide valuable insight into disease states and clinical responses, but they are challenging to mine and view collectively. To address these limitations, we present TaxUMAP, a taxonomically informed visualization for displaying microbiome states in large clinical microbiome datasets. We used TaxUMAP to chart a microbiome atlas of 1,870 patients with cancer during therapy-induced perturbations. Bacterial density and diversity were positively associated, but the trend was reversed in liquid stool. Low-diversity states (dominations) remained stable after antibiotic treatment, and diverse communities had a broader range of antimicrobial resistance genes than dominations. When examining microbiome states associated with risk for bacteremia, TaxUMAP revealed that certain Klebsiella species were associated with lower risk for bacteremia localize in a region of the atlas that is depleted in high-risk enterobacteria. This indicated a competitive interaction that was validated experimentally. Thus, TaxUMAP can chart comprehensive longitudinal microbiome datasets, enabling insights into microbiome effects on human health.

Enhanced Feature Selection for Microbiome Data using FLORAL: Scalable Log-ratio Lasso Regression.

Identifying predictive biomarkers of patient outcomes from high-throughput microbiome data is of high interest, while existing computational methods do not satisfactorily account for complex survival endpoints, longitudinal samples, and taxa-specific sequencing biases. We present FLORAL (https://vdblab.github.io/FLORAL/), an open-source computational tool to perform scalable log-ratio lasso regression and microbial feature selection for continuous, binary, time-to-event, and competing risk outcomes, with compatibility of longitudinal microbiome data as time-dependent covariates. The proposed method adapts the augmented Lagrangian algorithm for a zero-sum constraint optimization problem while enabling a two-stage screening process for extended false-positive control. In extensive simulation and real-data analyses, FLORAL achieved consistently better false-positive control compared to other lasso-based approaches, and better sensitivity over popular differential abundance testing methods for datasets with smaller sample size. In a survival analysis in allogeneic hematopoietic-cell transplant, we further demonstrated considerable improvement by FLORAL in microbial feature selection by utilizing longitudinal microbiome data over only using baseline microbiome data.

A phase 2 study of interleukin-22 and systemic corticosteroids as initial treatment for acute GVHD of the lower GI tract.

Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality following allogeneic hematopoietic transplantation. In experimental models, interleukin-22 promotes epithelial regeneration and induces innate antimicrobial molecules. We conducted a multicenter single-arm phase 2 study evaluating the safety and efficacy of a novel recombinant human interleukin-22 dimer, F-652, used in combination with systemic corticosteroids for treatment of newly diagnosed lower gastrointestinal acute GVHD. The most common adverse events were cytopenias and electrolyte abnormalities, and there were no dose-limiting toxicities. Out of 27 patients, 19 (70%; 80% confidence interval, 56%-79%) achieved a day-28 treatment response, meeting the prespecified primary endpoint. Responders exhibited a distinct fecal microbiota composition characterized by expansion of commensal anaerobes, which correlated with increased overall microbial α-diversity, suggesting improvement of GVHD-associated dysbiosis. This work demonstrates a potential approach for combining immunosuppression with tissue-supportive strategies to enhance recovery of damaged mucosa and promote microbial health in patients with gastrointestinal GVHD. This trial was registered at www.clinicaltrials.gov as NCT02406651.

Profiling the Fungal Microbiome after Fecal Microbiota Transplantation for Graft-versus-Host Disease: Insights from a Phase 1 Interventional Study.

Disruption of the intestinal bacterial microbiota is frequently observed in the context of allogeneic hematopoietic cell transplantation (HCT) and is particularly pronounced in patients who develop graft-versus-host disease (GVHD). Donor fecal microbiota transplantation (FMT) restores gut microbial diversity and reduces GVHD in HCT recipients. The composition of the intestinal fungal community in patients with GVHD, and whether fungal taxa are transferred during FMT are currently unknown. We performed a secondary analysis of our clinical trial of FMT in patients with steroid-refractory GVHD with a focus on the mycobiota. We characterized the fecal mycobiota of 17 patients and healthy FMT donors using internal transcribed spacer amplicon sequencing. The donor who provided the majority of FMT material in our study represents an n-of-one study of the intestinal flora over time. In this donor, mycobiota composition fluctuated over time while the bacterial microbiota remained stable over 16 months. Fungal DNA was detected more frequently in baseline stool samples from patients with steroid-refractory GVHD than in patients with steroid-dependent GVHD. We could detect fungal taxa in the majority of samples but did not see evidence of mycobiota transfer from donor to recipient. Our study demonstrates the feasibility of profiling the mycobiota alongside the more traditional bacterial microbiota, establishes the methodology, and provides a first insight into the mycobiota composition of patients with GVHD.

Conditioning Regimens are Associated with Distinct Patterns of Microbiota Injury in Allogeneic Hematopoietic Cell Transplantation.

PURPOSE: The gut microbiota is subject to multiple insults in allogeneic hematopoietic cell transplantation (allo-HCT) recipients. We hypothesized that preparative conditioning regimens contribute to microbiota perturbation in allo-HCT. EXPERIMENTAL DESIGN: This was a retrospective study that evaluated the relationship between conditioning regimens exposure in 1,188 allo-HCT recipients and the gut microbiome. Stool samples collected from 20 days before transplantation up to 30 days after were profiled using 16S rRNA sequencing. Microbiota injury was quantified by changes in α-diversity. RESULTS: We identified distinct patterns of microbiota injury that varied by conditioning regimen. Diversity loss was graded into three levels of conditioning-associated microbiota injury (CMBI) in a multivariable model that included antibiotic exposures. High-intensity regimens, such as total body irradiation (TBI)-thiotepa-cyclophosphamide, were associated with the greatest injury (CMBI III). In contrast, the nonmyeloablative regimen fludarabine-cyclophosphamide with low-dose TBI (Flu/Cy/TBI200) had a low-grade injury (CMBI I). The risk of acute GVHD correlated with CMBI degree. Pretransplant microbial compositions were best preserved with Flu/Cy/TBI200, whereas other regimens were associated with loss of commensal bacteria and expansion of Enterococcus. CONCLUSIONS: Our findings support an interaction between conditioning at the regimen level and the extent of microbiota injury.

Diet-derived metabolites and mucus link the gut microbiome to fever after cytotoxic cancer treatment.

Not all patients with cancer and severe neutropenia develop fever, and the fecal microbiome may play a role. In a single-center study of patients undergoing hematopoietic cell transplant (n = 119), the fecal microbiome was characterized at onset of severe neutropenia. A total of 63 patients (53%) developed a subsequent fever, and their fecal microbiome displayed increased relative abundances of Akkermansia muciniphila, a species of mucin-degrading bacteria (P = 0.006, corrected for multiple comparisons). Two therapies that induce neutropenia, irradiation and melphalan, similarly expanded A. muciniphila and additionally thinned the colonic mucus layer in mice. Caloric restriction of unirradiated mice also expanded A. muciniphila and thinned the colonic mucus layer. Antibiotic treatment to eradicate A. muciniphila before caloric restriction preserved colonic mucus, whereas A. muciniphila reintroduction restored mucus thinning. Caloric restriction of unirradiated mice raised colonic luminal pH and reduced acetate, propionate, and butyrate. Culturing A. muciniphila in vitro with propionate reduced utilization of mucin as well as of fucose. Treating irradiated mice with an antibiotic targeting A. muciniphila or propionate preserved the mucus layer, suppressed translocation of flagellin, reduced inflammatory cytokines in the colon, and improved thermoregulation. These results suggest that diet, metabolites, and colonic mucus link the microbiome to neutropenic fever and may guide future microbiome-based preventive strategies.

Xylose versus Bacteroides in graft-versus-host.

In a recent Cell paper, Hayase et al. define a mechanistic link between a carbapenem antibiotic and graft-versus-host disease (GVHD) following hematopoietic cell transplantation. Bacteroidesthetaiotaomicron was observed to induce degradation of colonic mucus, an effect that was reversed by treatment with xylose, which served as a prebiotic to reduce mortality.

Pilot Trial of Homebound Hematopoietic Cell Transplantation.

For eligible patients with multiple myeloma (MM) and amyloid light chain (AL) amyloidosis, high-dose chemotherapy and autologous hematopoietic cell transplantation (HCT) is a standard and widely used consolidation therapy. Autologous HCT requires specialized care at a transplantation center and investment from patients and caregivers. We studied the safety and feasibility of delivering transplantation care in a homebound setting to decrease the burden of therapy and increase access to autologous HCT. Patients with MM and AL amyloidosis undergoing autologous HCT were eligible if they resided in designated ZIP codes and had a full-time caregiver, Wi-Fi connection, HCT Comorbidity Index ≤3, and Karnofsky Performance Status score ≥80. High-dose melphalan (on day -2) and hematopoietic cell reinfusion (day 0) were administered in the outpatient clinic. Protocol-specific home care was provided from day +1 through engraftment. Patients were assessed and blood was drawn daily by advanced practice providers. Interventions were delivered by registered nurses. Attending physicians communicated daily through telemedicine. Quality of life, patient and caregiver satisfaction, and fecal microbiota profiling data were collected. Fifteen patients were enrolled and received transplantation care at home starting on day +1 following hematopoietic cell infusion. Patients remained in the program for an average of 12 days and required an average of 2 outpatient visits while receiving home care. Seven of 15 patients were admitted for a median of 4 days (range, 3 to 10 days); admission occurred on day +7 in 5 patients, on day +8 in 1 patient, and on day +12 in 1 patient for neutropenic fever in 2 patients, fever attributed to engraftment syndrome in 2 patients, diarrhea in 2 patients, and dehydration in 1 patient. Only 1 patient had a documented infection (Clostridioides difficile). One patient admitted with neutropenic fever required intensive care unit admission for a gastrointestinal bleed. Forty-seven percent of the patients experienced a grade ≥3 nonhematologic toxicity. There were no deaths on the study. Patients and caregivers reported high satisfaction with care. Microbiota diversity patterns were similar to those of autologous HCT recipients who did not receive post-HCT care at home, although a subset of the cohort maintained microbiota diversity throughout. Homebound HCT in an urban setting is safe and feasible, with less than one-half of patients requiring inpatient admission. Despite increased patient and caregiver responsibility in the homebound setting compared with an inpatient setting, patient and caregiver satisfaction was high. These results support expansion of homebound transplantation care programs.