Defining mechanisms used by the gut microbiome to control anti-tumor immunity may offer novel therapeutic modalities. Here, we demonstrate that Bacteroides rodentium and the closely related species Bacteroides uniformis induced anti-tumor immunity and limited melanoma development when colonized in germ-free (GF) mice, in mice with complex microbiome, or in WT mice. Enhanced CD8+ T cell infiltration seen in tumors of mice harboring B. rodentium coincided with increased expression of immune-stimulating pathways. Metabolomic analyses of cecal samples from GF mice colonized with ASF plus B. rodentium revealed lower tryptophan levels compared to ASF colonized controls. Feeding WT mice a tryptophan-deficient diet was sufficient to inhibit melanoma development. In silico genomic reconstruction of metabolic pathways revealed that B. rodentium harbors the tryptophanase A (TnaA) gene, which degrades tryptophan, the ortholog of which is found in B. uniformis. Administration of B. uniformis, which was mutated in TnaA, failed to inhibit melanoma growth in GF mice. Altogether, these findings highlight a novel mechanism of anti-tumor immunity and tumor growth inhibition dependent on tryptophan degradation by intestinal Bacteroides species. Ximena P. Diaz Olea, Kristin Beede, Gabriel Pereira, David Scott, Christopher Pettucci, Eric Martens, Dmitri Rodionov, Hyungsoo Kim, Ashok Kumar Sharma, Anthony Martin, Tongwu Zhang, Suzanne Devkota, Andrei Osterman, Amanda E. Ramer-Tait, Ze'ev A. Ronai. Tryptophan degradation by intestinal Bacteroides induces anti-tumor immunity and limits melanoma growth [abstract]. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr C062.

Defining mechanisms used by gut microbiota to control anti-tumor immunity may offer novel therapeutic modalities. Here, we demonstrate that Bacteroides rodentium and closely related Bacteroides uniformis species induce anti-tumor immunity and limit melanoma development when colonized in either germ-free (GF) mice, mice with a complex microbiome, or WT mice. Enhanced CD8 + T cell infiltration seen in tumors of mice harboring B. rodentium coincided with increased expression of immune-stimulating pathways and activation of bone marrow-derived dendritic cells that were co-cultured with the B. rodentium secretome. Metabolomic analyses of cecal samples from GF mice colonized with Altered Shedlar Flora (ASF) plus B. rodentium revealed lower tryptophan levels than in ASF-colonized controls, and WT mice fed a tryptophan-deficient diet exhibited inhibition of melanoma development. In silico genomic reconstruction of metabolic pathways revealed that both B. rodentium and B. uniformis harbor tryptophanase A ( TnaA ) and aromatic amino transferase ( ArAT ) genes, both of which function in tryptophan degradation. Administration of a B. uniformis harboring TnaA mutant failed to inhibit melanoma growth in gnotobiotic mice. Notably, administration of indoles, but not kynurenines, also effectively inhibited melanoma development, increasing immune cell infiltration into the tumors. Correspondingly, levels of bacterially encoded tryptophan-degrading enzymes were higher in cohorts of melanoma patients responding to immune checkpoint blockade. These findings highlight a novel mechanism of anti-tumor immunity and tumor growth inhibition dependent on the tryptophan degradation products, indoles, produced by intestinal Bacteroides species.

Study of gut microbiota control of anti-tumor immunity (ATI) identifies Bacteroides rodentium and the human-related Bacteroides uniformis species to be capable of inducing ATI and limiting melanoma development in germ-free (GF), complex microbiome, or wild-type (WT) mice. Enhanced CD8+ T cell infiltration within tumors of mice harboring B. rodentium coincides with increased expression of immune-stimulating pathways. Metabolomic analyses identify lower tryptophan levels in the cecal samples of GF mice harboring B. rodentium. In silico genomic reconstruction reveals that B. rodentium and B. uniformis harbor tryptophanase A (TnaA) and aromatic aminotransferase genes, which degrade tryptophan to indoles. Administration of B. uniformis harboring TnaA mutant fails to inhibit melanoma growth. Notably, administration of indoles effectively induces ATI and inhibits melanoma development. Correspondingly, the levels of bacterially encoded tryptophan-degrading enzymes are higher in cohorts of patients with melanoma responding to immunotherapy. These findings identify indoles as tryptophan breakdown products capable of inducing ATI resulting in melanoma inhibition.

IntroductionPersistent immune activation and inflammation remain significant barriers to managing comorbidities in people living with HIV (PLWH) on suppressive antiretroviral therapy (ART). While ART substantially reduces plasma viral loads to an undetectable level, it fails to fully restore gut microbial homeostasis and prevent microbial translocation, a critical pathogenic contributor to systemic persistent immune activation and inflammation. To evaluate the potential of human fecal microbiota transplantation (FMT) as an adjunctive therapy to restore gut health and attenuate inflammation and immune activation in PLWH on ART, we utilized a double humanized-BLT (dHu-BLT) mouse model, featuring a functional human immune system and a human-like microbiome.MethodsTwo groups of HIV-infected dHu-BLT mice were used in the study. One group received FMT in addition to ART, while the control group received ART alone. Using both a multi-omics approach (16S rRNA sequencing and RNA-seq) and an immune-based assay, we compared alterations in gut microbial composition, profiled transcriptomic changes in the intestinal tissue, and quantified markers of systemic immune activation and inflammation between the groups.ResultFMT supplementation in ART-treated mice increased the relative abundance of beneficial bacteria and modulated the transcriptomic profile of both human- and murine-related genes. Notably, genes associated with cellular structure and tissue maintenance, including Mcpt4, were upregulated, along with the extracellular matrix organization pathway predicted as the most strongly activated pathway in the FMT-supplemented group compared to ART alone. In contrast, genes and signaling pathways associated with inflammation were downregulated. Importantly, the FMT-supplemented group exhibited a significant reduction of plasma inflammatory markers, including CD62E, sCD14, sCD163, and FABP2, relative to the ART alone group.ConclusionThese results suggest that FMT may serve as a promising adjunctive strategy for mitigating systemic inflammation by improving gut health, thereby contributing to the reduction of comorbidities in PLWH on ART.

Targeting the gut microbiome through diet remains a priority in the treatment and management of inflammatory bowel diseases (IBD) for patients and clinicians alike. However, expert consensus is lacking, and cross-sectional studies indicate that patients with IBD often adopt low-fiber, high-sugar, and high-protein diets in an effort to control gastrointestinal symptoms and inflammation with negative implications for gut microbial functions and gastrointestinal inflammation. Although dietary carbohydrates are key modulators of gut microbial composition and function, substantial disagreement persists regarding how carbohydrate digestibility and structural properties influence IBD outcomes. This narrative review summarizes current knowledge on the role of dietary carbohydrates in IBD risk and progression and critically evaluates recent dietary interventions in which the gut microbiota was included among outcomes to elaborate a potential role of dietary carbohydrates in the clinical management of IBD. We conclude by outlining research priorities needed to affirm a role for dietary carbohydrate modification in the clinical management of IBD and to support the development of practical, evidence-based dietary guidance.

Myocarditis is an inflammatory disease of the myocardium caused by infection-induced autoimmunity toward heart-specific antigens. Although the gut microbiota has been linked to cardiovascular disease, its role in the pathogenesis of myocarditis is unclear. We therefore investigated whether the gut microbiota modulates disease severity in a mouse model of Experimental Autoimmune Myocarditis (EAM). Germ-free mice were colonized with one of three distinct microbiomes (M31B, W116 or MC608) prior to inducing EAM with adjuvanted myosin. Mice harboring MC608 had significantly higher inflammatory heart scores and myosin-specific autoimmune responses versus mice carrying M31B. Mice with severe signs of EAM also experienced alterations in composition of their microbiota, including a significant decrease in Lactobacillus. We further observed negative correlations between EAM severity and both L. reuteri and L. taiwanensis. To test for protective effects of these species, we inoculated Lactobacillus-deficient mice with both L. reuteri and L. taiwanensis prior to EAM induction. Treatment with both lactobacilli significantly decreased myosin-specific IFN-γ and IgG responses compared to controls. Altogether, these results show that the gut microbiota modulates EAM severity and autoreactive responses. Moreover, L. reuteri and L. taiwanensis provide protection against severe EAM, suggesting that live biotherapeutics may be a beneficial adjunct therapy for myocarditis patients. This project was supported by an NU Collaboration seed grant. Basic Autoimmunity (BA)

Competitive exclusion is conventionally believed to prevent the establishment of a secondary strain of the same bacterial species in the gut microbiome, raising concerns for the deployment of live bacterial therapeutics (LBTs), especially if the bacterial chassis is a strain native to the gut. In this study, we investigated factors influencing competition dynamics in the murine gut using isogenic native Escherichia coli strains. We found that competition outcomes are context-dependent, modulated by microbiome complexity, LBT transgene expression, intestinal inflammation, and host diet. Furthermore, we demonstrated that native LBTs can establish long-term engraftment in the gut alongside a parental strain, with transgene-associated fitness effects influencing competition. We identified various interventions, including strategic dosing and dietary modulation, that significantly enhanced LBT colonization levels by 2 to 3 orders of magnitude. These insights provide a framework for optimizing LBT engraftment and efficacy, supporting their potential translation for human therapeutic applications.

High-fat diet (HFD) is well known to impact various aspects of gut health and has been associated with many diseases and inflammation. However, the impact of HFD feeding on HIV-1 rectal transmission has not yet been well addressed. With an increasing threat of HIV-1 infection in men who have sex with men (MSM), where the rectal route is the primary mode of infection, it is imperative to understand the impact of HFD on gut microbiota and inflammation and consequently, its effect on HIV-1 rectal transmission. Here, we utilized our double humanized bone marrow, liver, thymus (dHu-BLT) mouse model to assess the impact of HFD feeding on the host's susceptibility to HIV-1 rectal transmission. We found that feeding an HFD successfully altered the gut microbial composition within 3 weeks in the dHu-BLT mouse model. In addition, levels of inflammatory mediators, specifically IL-12p70, IP-10, ICAM-1, and fecal calprotectin, were significantly higher in HFD-fed mice compared to control mice on a regular chow diet. We also observed that significantly different inflammatory markers (IL-12p70 and ICAM-1) were negatively correlated with the number of observed ASVs, Shannon diversity, and Faith's diversity in the HFD-fed group. Notably, when repeatedly challenged with a low dose of HIV-1 via a rectal route, mice receiving an HFD were significantly more susceptible to HIV-1 rectal infection than control mice. Together, these results underscore the impact of HFD feeding on the gut microbiota and inflammation and suggest the significance of diet-induced gut microbial dysbiosis and inflammation in promoting viral infection.IMPORTANCEHFD induces gut microbial dysbiosis and inflammation and has been associated with many infections and disease progression; however, its impact on HIV-1 rectal transmission is largely unknown. Given the increasing threat of HIV-1 incidence in men who have sex with men (MSM), it has become crucial to comprehend the impact of factors associated with gut health, like HFD consumption, on host susceptibility to HIV-1 rectal transmission. This is particularly important since anal intercourse remains the primary mode of HIV transmission within the MSM group. In this study, utilizing our unique mouse model, featuring both the human immune system and gut microbiota, we showed that HFD feeding led to gut microbial dysbiosis, induced inflammation, and increased HIV-1 rectal transmission. Collectively, our study highlights the significant impact of HFD on gut microbiota and inflammation and suggests an HFD consumption as a potential risk factor for promoting HIV-1 rectal susceptibility.

Abstract Growing evidence supports the importance of the gut microbiota in controlling tumor development, often via activation of anti-tumor immunity. Earlier studies from our laboratory identified 11 bacterial strains that were sufficient to inhibit melanoma development by inducing anti-tumor immunity in germ-free mice. Of these, three were predominant at the time of tumor collection. We thus set out to address whether fewer bacterial strains may be sufficient for melanoma growth inhibition. Here, we demonstrate that the administration of one bacterial strain, B. rodentium, was sufficient to attenuate melanoma growth in gnotobiotic mice. In all cases, gnotobiotic mice were also administered the altered Schadler flora (ASF), a mixture of 8 bacterial strains that are provided to establish minimal flora in these mice. RNAseq-based gene expression studies identified enrichment of immune-related genes, suggesting enhanced anti-tumor immunity. Metabolomic analysis identified reduced levels of tryptophane and isoleucine, which were implicated in control of immune system function. These findings point to a mechanism which may provide novel means to alter anti-tumor immunity and limit melanoma growth. Citation Format: Ximena Diaz Olea, Kristin Beede, Andrei Osterman, David Scott, Christopher Petucci, Daniel Kelly, Amanda Ramer-Tait, Ze’ev A. Ronai. Control of melanoma development by B. rodentium in germ free mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6685.

Humanized mice with human-like immune systems are commonly used to study immune responses to human-specific pathogens. However, one limitation of using humanized mice is their native murine gut microbiota, which significantly differs from that in humans. Given the importance of the gut microbiome to human health, these differences may profoundly impact the ability to translate results from humanized mouse studies to humans. Further, there is a critical need for improved pre-clinical models to study the complex in vivo relationships of the gut microbiome, immune system, and human disease. We previously created double humanized mice with a functional human immune system and a stable, human-like gut microbiome. Here, we characterized the engrafted human gut bacterial microbiome in our double humanized mouse model generated by transplanting fecal material from healthy human donors into the gut of humanized mice. Analysis of bacterial microbiomes in fecal samples from double humanized mice revealed they had unique 16S rRNA gene profiles consistent with those of the individual human donor samples. Importantly, transplanted human-like gut microbiomes were stable in mice for the duration of the study, extending up to 14.5 weeks post-transplant. Microbiomes of double humanized mice also harbored predicted functional capacities that more closely resembled those of the human donors than humanized mice. In conclusion, our study highlights the successful engraftment of human fecal microbiota in BLT humanized mice and underscores the stability of this model, offering a valuable platform for investigating the intricate interplay among the human gut microbiome, immune system, and various diseases in vivo .

Background Humanized mice featuring a functional human immune system are an important pre-clinical model for examining immune responses to human-specific pathogens. This model has been widely utilized to study human diseases that are otherwise impossible or difficult to investigate in humans or with other animal models. However, one limitation of using humanized mice is their native murine gut microbiome, which significantly differs from the one found in humans. These differences may be even greater for mice housed and bred in specific pathogen free conditions. Given the importance of the gut microbiome to human health and disease, these differences may profoundly impact the ability to translate the results from humanized mice studies to human disease. Further, there is a critical need for improved pre-clinical models to study the complex in vivo relationships of the gut microbiome, immune system, and human disease. We therefore created double humanized mice with both a functional human immune system and stable human-like gut microbiome.Results Surgery was performed on NOD. Cg-PrkdcscidII2rgtm1Wjl /SzJ (NSG) mice to create bone-marrow, liver, thymus (BLT) humanized mice. After immune reconstitution, mice were treated with broad spectrum antibiotics to deplete murine gut bacteria and then transplanted with fecal material from healthy human donors. Characterization of 173 fecal samples obtained from 45 humanized mice revealed that double humanized mice had unique 16S rRNA gene profiles consistent with those of the individual human donor samples. Importantly, transplanted human-like gut microbiomes were stable in mice for the duration of the study, up to 14.5 weeks post-transplant. Microbiomes of double humanized mice also harbored predicted functional capacities that more closely resembled those of the human donors compared to humanized mice.Conclusions Here, we describe successful engraftment of a stable human microbiome in BLT humanized mice to further improve this preclinical humanized mouse model. These double humanized mice represent a unique and tractable new model to study the complex relationships between the human gut microbiome, human immune system, and human disease in vivo .

The ecological relationships among antimicrobial producing, resistant, and sensitive strains have been proposed to follow rock-paper-scissors dynamics, but evidence is mainly based on Gram-negative bacteriocins in vitro. The ecological relevance of antimicrobials in vivo or in situ has not been systematically studied. This study therefore aimed to analyze binary and ternary competitions among reutericyclin-producing strain Limosilactobacillus reuteri TMW1.656, its reutericyclin-resistant, nonproducing isogenic derivative L. reuteri TMW1.656∆rtcN, and the reutericyclin-sensitive, nonproducing L. reuteri TMW1.656∆rtcN∆rtcT in vitro (liquid culture and static plate), in situ (sourdough fermentation), and in vivo (gut of germ-free mice). In liquid culture, L. reuteri TMW1.656 had a higher fitness than TMW1.656∆rtcN and TMW1.656∆rtcN∆rtcT. Limosilactobacillus reuteri TMW1.656∆rtcN∆rtcT had a higher fitness than TMW1.656∆rtcN. On agar plates, L. reuteri TMW1.656 had a higher fitness than TMW1.656∆rtcN∆rtcT. In situ, reutericyclin production and resistance had no influence on the fitness of the strains. In vivo, TMW1.656 had an advantage over TMW1.656∆rtcN and TMW1.656∆rtcN∆rtcT. Ternary competitions showed reutericyclin production was ecologically beneficial in all ecosystems. The findings support the ecological importance of reutericyclin in a variety of environments/niches, providing an explanation for the acquisition of the reutericyclin gene cluster in L. reuteri and its contribution to the ecological fitness of Streptococcus mutans.

The intestinal barrier orchestrates selective permeability to nutrients and metabolites while excluding noxious stimuli. Recent scientific advances establishing a causal role for the gut microbiota in human health outcomes have generated a resurgent interest toward intestinal permeability. Considering the well-established role of the gut barrier in protection against foreign antigens, there is mounting evidence for a causal link between gut permeability and the microbiome in regulating human health. However, an understanding of the dynamic host-microbiota interactions that govern intestinal barrier functions remains poorly defined. Furthermore, the system-level mechanisms by which microbiome-targeted therapies, such as probiotics and prebiotics, simultaneously promote intestinal barrier function and host health remain an area of active investigation. This review summarizes the recent advances in understanding the dynamics of intestinal permeability in human health and its integration with gut microbiota. We further summarize mechanisms by which probiotics/prebiotics influence the gut microbiota and intestinal barrier functions.

Enrichment of adherent-invasive Escherichia coli (AIEC) has been consistently detected in subsets of inflammatory bowel disease (IBD) patients. Although some AIEC strains cause colitis in animal models, these studies did not systematically compare AIEC with non-AIEC strains, and causal links between AIEC and disease are still disputed. Specifically, it remains unclear whether AIEC shows enhanced pathogenicity compared to that of commensal E. coli found in the same ecological microhabitat and if the in vitro phenotypes used to classify strains as AIEC are pathologically relevant. Here, we utilized in vitro phenotyping and a murine model of intestinal inflammation to systematically compare strains identified as AIEC with those identified as non-AIEC and relate AIEC phenotypes to pathogenicity. Strains identified as AIEC caused, on average, more severe intestinal inflammation. Intracellular survival/replication phenotypes routinely used to classify AIEC positively correlated with disease, while adherence to epithelial cells and tumor necrosis factor alpha production by macrophages did not. This knowledge was then applied to design and test a strategy to prevent inflammation by selecting E. coli strains that adhered to epithelial cells but poorly survived/replicated intracellularly. Two E. coli strains that ameliorated AIEC-mediated disease were subsequently identified. In summary, our results show a relationship between intracellular survival/replication in E. coli and pathology in murine colitis, suggesting that strains possessing these phenotypes might not only become enriched in human IBD but also contribute to disease. We provide new evidence that specific AIEC phenotypes are pathologically relevant and proof of principle that such mechanistic information can be therapeutically exploited to alleviate intestinal inflammation. IMPORTANCE Inflammatory bowel disease (IBD) is associated with an altered gut microbiota composition, including expansion of Proteobacteria. Many species in this phylum are thought to contribute to disease under certain conditions, including adherent-invasive Escherichia coli (AIEC) strains, which are enriched in some patients. However, whether this bloom contributes to disease or is just a response to IBD-associated physiological changes is unknown. Although assigning causality is challenging, appropriate animal models can test the hypothesis that AIEC strains have an enhanced ability to cause colitis in comparison to other gut commensal E. coli strains and to identify bacterial traits contributing to virulence. We observed that AIEC strains are generally more pathogenic than commensal E. coli and that bacterial intracellular survival/replication phenotypes contributed to disease. We also found that E. coli strains lacking primary virulence traits can prevent inflammation. Our findings provide critical information on E. coli pathogenicity that may inform development of IBD diagnostic tools and therapies.

any contemporary research challenges are associated with a level of complexity that requires solutions toward them to be developed by teams of scholars bringing relevant experience, innovative techniques, and interdisciplinary perspectives.As a result, over the last two decades, the scientific community has begun to pay considerable attention to team science as a method to enable advances not possible by a single laboratory or group. 1 Team science is now known to accelerate scientific and technological innovation and serve as an effective vehicle through which to translate research and scholarship into practice and policy. 2More recently, team science scholars documented that the outputs of diverse research teams tend to be more novel and highly cited. 3Because the fundamentals of team science and leadership are transferrable, and given the demonstrated affordances associated with inclusive team leadership, institutions are beginning to explore how they might best support faculty interested in obtaining the skills and experiences needed to lead large, collaborative, and diverse research teams.

Preclinical studies have shown that chronic alcohol abuse leads to alterations in the gastrointestinal microbiota that are associated with behavior changes, physiological alterations, and immunological effects. However, such studies have been limited in their ability to evaluate the direct effects of alcohol-associated dysbiosis. To address this, we developed a humanized alcohol-microbiota mouse model to systematically evaluate the immunological effects of chronic alcohol abuse mediated by intestinal dysbiosis. Germ-free mice were colonized with human fecal microbiota from individuals with high and low Alcohol Use Disorders Identification Test (AUDIT) scores and bred to produce human alcohol-associated microbiota or human control-microbiota F1 progenies. F1 offspring colonized with fecal microbiota from individuals with high AUDIT scores had increased susceptibility to Klebsiella pneumoniae and Streptococcus pneumoniae pneumonia, as determined by increased mortality rates, pulmonary bacterial burden, and post-infection lung damage. These findings highlight the importance of considering both the direct effects of alcohol and alcohol-induced dysbiosis when investigating the mechanisms behind alcohol-related disorders and treatment strategies.