Faculty Bibliography

Introduction/UNASSIGNED:Declining renal function results in the accumulation of solutes normally excreted by healthy kidneys. Data suggest that some of the protein-bound solutes mediate accelerated cardiovascular disease. Many of the poorly dialyzable protein-bound uremic retention solutes are products of gut bacterial metabolism. Methods/UNASSIGNED:We performed a blinded-randomized controlled trial comparing the changes in plasma concentrations of a panel of protein-bound solutes and microbiome structure in response to the once-weekly oral administration of 250 mg of vancomycin or placebo over a period of 12 weeks in a cohort of stable patients with end-stage kidney disease. We also examined the pattern of recovery of the solutes and gut microbiome over 12 weeks of placebo administration following vancomycin. Results/UNASSIGNED:. We demonstrated microbiome recovery after stopping vancomycin. However, recovery in the solutes was highly variable between subjects. Conclusions/UNASSIGNED:We demonstrated that microbiome suppression using vancomycin resulted in changes in multiple gut-derived uremic solutes. Future studies are needed to address whether reduction in those uremic solutes results in improvement of cardiovascular outcomes in ESKD patients.

Over-accumulation of oxalate in humans may lead to nephrolithiasis and nephrocalcinosis. Humans lack endogenous oxalate degradation pathways (ODP), but intestinal microbes can degrade oxalate using multiple ODPs and protect against its absorption. The exact oxalate-degrading taxa in the human microbiota and their ODP have not been described. We leverage multi-omics data (>3000 samples from >1000 subjects) to show that the human microbiota primarily uses the type II ODP, rather than type I. Further, among the diverse ODP-encoding microbes, an oxalate autotroph, Oxalobacter formigenes, dominates this function transcriptionally. Patients with Inflammatory Bowel Disease (IBD) frequently suffer from disrupted oxalate homeostasis and calcium oxalate nephrolithiasis. We show that the enteric oxalate level is elevated in IBD patients, with highest levels in Crohn's disease patients with both ileal and colonic involvement consistent with known nephrolithiasis risk. We show that the microbiota ODP expression is reduced in IBD patients, which may contribute to the disrupted oxalate homeostasis. The specific changes in ODP expression by several important taxa suggest that they play distinct roles in IBD-induced nephrolithiasis risk. Lastly, we colonize mice that are maintained in the gnotobiotic facility with O. formigenes, using either a laboratory isolate or an isolate we cultured from human stools, and observed a significant reduction in host fecal and urine oxalate levels, supporting our in silico prediction of the importance of the microbiome, particularly O. formigenes in host oxalate homeostasis.

BACKGROUND:Oxalobacter formigenes are bacteria that colonize the human gut and degrade oxalate, a component of most kidney stones. Clinical and epidemiological studies suggest that O. formigenes colonization reduces the risk for kidney stones. We sought to develop murine models to allow investigating O. formigenes in the context of its native human microbiome.For humanization, we transplanted pooled feces from healthy, non-colonized human donors supplemented with a human O. formigenes strain into recipient mice. We compared transplanting microbiota into mice that were either treated with broad-spectrum antibiotics to suppress their native microbiome, or were germ-free, or received humanization without pre-treatment or received a sham gavage (controls). RESULTS:All humanized mice were stably colonized with O. formigenes through 8 weeks post-gavage, whereas mice receiving sham gavage remained uncolonized (p<0.001). Humanization significantly changed the murine intestinal microbial community structure (p<0.001) with humanized germ-free and antibiotic-treated groups overlapping in β-diversity. Both the germ-free and antibiotic-treated mice had significantly increased numbers of human species compared to sham-gavaged mice (p<0.001). CONCLUSIONS:Transplanting mice with human feces and O. formigenes introduced new microbial populations resembling the human microbiome, with stable O. formigenes colonization; such models can define optimal O. formigenes strains to facilitate clinical trials.

PURPOSE OF REVIEW/OBJECTIVE:Enteric hyperoxaluria is commonly observed in malabsorptive conditions including Roux en Y gastric bypass (RYGB) and inflammatory bowel diseases (IBD). Its incidence is increasing secondary to an increased prevalence of both disorders. In this review, we summarize the evidence linking the gut microbiota to the risk of enteric hyperoxaluria. RECENT FINDINGS/RESULTS:In enteric hyperoxaluria, fat malabsorption leads to increased binding of calcium to free fatty acids resulting in more soluble oxalate in the intestinal lumen. Bile acids and free fatty acids in the lumen also cause increased gut permeability allowing more passive absorption of oxalate. In recent years, there is more interest in the role of the gut microbiota in modulating urinary oxalate excretion in enteric hyperoxaluria, stemming from our knowledge that microbiota in the intestines can degrade oxalate. Oxalobacter formigenes reduced urinary oxalate in animal models of RYGB. The contribution of other oxalate-degrading organisms and the microbiota community to the pathophysiology of enteric hyperoxaluria are also currently under investigation. SUMMARY/CONCLUSIONS:Gut microbiota might play a role in modulating the risk of enteric hyperoxaluria through oxalate degradation and bile acid metabolism. O. formigenes is a promising therapeutic target in this population; however, further studies in humans are needed to test its effectiveness.

There has been increasing interest in the human anaerobic colonic bacterium Oxalobacter formigenes because of its ability to metabolize oxalate, and its potential contribution to protection from calcium oxalate kidney stones. Prior studies examining the prevalence of this organism have focused on subjects in developed countries and on adults. Now using O. formigenes-specific PCR, we have compared the prevalence of these organisms among subjects in two remote areas in which modern medical practices have hardly been present with a USA group of mothers and their infants for the first three years of life. Among the Amerindians of the Yanomami-Sanema and Yekwana ethnic groups in Venezuela and the Hadza in Tanzania, O. formigenes was detected in 60-80% of the adult subjects, higher than found in adults from USA in this and prior studies. In young children, the prevalence was much lower in USA than in either tribal village. These data extend our understanding of the epidemiology of O. formigenes carriage, and are consistent with the hypothesis that the rising incidence of kidney stones is associated with the progressive loss of O. formigenes colonization in populations that have been highly impacted by modern medical practices.

Background: Cystinuria is a disease of impaired absorption of cystine and dibasic amino acids (DAA) from the intestine and renal tubule leading to formation of cystine kidney stones. However, the metabolic impact of reduced amino acid absorption and excessive loss in the urine is poorly understood. We measured endogenous, gut microbial, and xenobiotic metabolites, providing insight into consequences of the disease and its treatment.
Method(s): Urinary biochemicals were assayed using LC-MS in 293 urine specimens from patients with cystinuria or control urinary phenotypes. Multivariate statistical analyses were conducted to reveal statistically significant biochemical signatures of the disease and products of cysteine-binding thiol drugs (CBTDs). 16s rRNA gene sequencing was performed on fecal samples from 12 wildtype (WT) and 12 cystinuric (Slc3a1 knockout; KO) mice to evaluate their gut microbial composition.
Result(s): Cystinuric urine samples had elevated levels of cysteine-gamma-glutamyl cystine disulfide (glutathione precursor), indole-3-acetic acid (microbial tryptophan metabolism), and novel conjugated forms of putrescine (microbial DAA decomposition). Conversely, taurine (sulfur metabolism), indole-3-acetic acid-glucuronide, and novel urinary metabolite N-methyl pipecolic acid (lysine metabolism) were reduced in cystinuric urine. Where cysteine-bound CBTDs were observed, substantial amounts of "wasted" drug were also detected as CBTD homodimers, non-cysteine disulfides, and mixed drug disulfides. The differentiation of gut microbially-derived metabolites led us to evaluate the gut microbiome diversity and composition in a mouse model of cystinuria revealing clear beta diversity and taxa differentiation between WT and KO mice.
Conclusion(s): Cystinuria is associated with unique urinary metabolic profiles beyond hyperexcretion of cystine and DAA, indicating perturbed metabolic processes and potential gut microbial effects. Study of the gut microbiome of WT and KO mice provides the first evidence for them having distinct taxa, perhaps due to poorly absorbed DAA present in the intestinal lumen. Urinary profiles allow us to characterize the excretion profiles of CBTDs, providing insight which may be helpful to tailor treatment

Background: Kidney stones commonly affect US adults. In recent years, there has been increasing interest in the human anaerobic colonic bacterium Oxalobacter formigenes because of its ability to metabolize oxalate, and its potential to protect against calcium oxalate kidney stones. Currently, there are two known groups of O. formigenes (Group 1 and Group 2) with only a few isolates from each group characterized. In our experiments, we aimed to isolate O. formigenes from subjects with primary hyperoxaluria (PH), enteric hyperoxaluria (EH) and healthy controls (HC) to compare their metabolic activities. Understanding these differences will help expand our knowledge about this important organism and its effect on oxalate homeostasis in humans.
Method(s): We collected fecal samples from 37 patients via clinical trials at New York University Langone Medical Center and Mayo Clinic with PH, EH and HC. We cultured fecal samples in 25mM oxalate-rich selective media, then isolated O. formigenes by picking characteristic colonies from calcium oxalate agar. We identified and grouped isolates using PCR and Sanger sequencing of the oxc gene. We then tested their oxalate consumption via Oxalate Degradation Assay to compute mean oxalate degradation rates (ODR) for each group of isolates.
Result(s): We isolated 25 O. formigenes colonies from 14 subjects, with all isolates belonging to either HC (n=11) or PH (n= 14) patients, and none from EH patients. Based on oxc sequences, we identified Group 1 (n=17) and Group 2 (n=5) strains, and potentially a new taxonomic group Group 3 (n=3). We were able to regrow 13 (76%) of 17, 1 (20%) of 5, and 1 (33%) of 3 Group 1, 2, and 3 strains, respectively. All 14 PH patient colonies were identified as Group 1, while HC had a mix of all three groups. Mean ODR was significantly higher in Group 1 vs Group 2 isolates (8.5 +/- 3.3 vs 2.8 +/- 1.9 micromole/ hour, p=0.02). Group 3 isolates had intermediate ODR (5.7 +/- 3.1) values. As expected, the ODRs of our Group 1 isolates were similar to the control group 1 strain OXCC13 (11.1 +/- 1.2). Mean ODR between PH, EH and HC did not differ significantly.
Conclusion(s): We were able to isolate and characterize 25 colonies of O. formigenes, including a potential new group of O. formigenes. Group 1 strains appear to be most metabolically active in vitro, and were exclusively present in PH patients