Effect of antibiotic treatment on Oxalobacter formigenes colonization of the gut microbiome and urinary oxalate excretion.

The incidence of kidney stones is increasing in the US population. Oxalate, a major factor for stone formation, is degraded by gut bacteria reducing its intestinal absorption. Intestinal O. formigenes colonization has been associated with a lower risk for recurrent kidney stones in humans. In the current study, we used a clinical trial of the eradication of Helicobacter pylori to assess the effects of an antibiotic course on O. formigenes colonization, urine electrolytes, and the composition of the intestinal microbiome. Of 69 healthy adult subjects recruited, 19 received antibiotics for H. pylori eradication, while 46 were followed as controls. Serial fecal samples were examined for O. formigenes presence and microbiota characteristics. Urine, collected serially fasting and following a standard meal, was tested for oxalate and electrolyte concentrations. O. formigenes prevalence was 50%. Colonization was significantly and persistently suppressed in antibiotic-exposed subjects but remained stable in controls. Urinary pH increased after antibiotics, but urinary oxalate did not differ between the control and treatment groups. In subjects not on antibiotics, the O. formigenes-positive samples had higher alpha-diversity and significantly differed in Beta-diversity from the O. formigenes-negative samples. Specific taxa varied in abundance in relation to urinary oxalate levels. These studies identified significant antibiotic effects on O. formigenes colonization and urinary electrolytes and showed that overall microbiome structure differed in subjects according to O. formigenes presence. Identifying a consortium of bacterial taxa associated with urinary oxalate may provide clues for the primary prevention of kidney stones in healthy adults.

Development of a Humanized Murine Model for the Study of Oxalobacter formigenes Intestinal Colonization.

BACKGROUND: Oxalobacter formigenes are bacteria that colonize the human gut and degrade oxalate, a component of most kidney stones. Findings of 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. METHODS: For humanization, we transplanted pooled feces from healthy, noncolonized human donors supplemented with a human O. formigenes strain into recipient mice. We transplanted microbiota into mice that were treated with broad-spectrum antibiotics to suppress their native microbiome, were germ free, or received humanization without pretreatment or received sham gavage (controls). RESULTS: All humanized mice were stably colonized with O. formigenes through 8 weeks after gavage, whereas mice receiving sham gavage remained uncolonized (P < .001). Humanization significantly changed the murine intestinal microbial community structure (P < .001), with humanized germ-free and antibiotic-treated groups overlapping in ß-diversity. Both germ-free and antibiotic-treated mice had significantly increased numbers of human species compared with sham-gavaged mice (P < .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.

Probiotic properties of Oxalobacter formigenes: an in vitro examination.

Oxalobacter formigenes (O. formigenes) is a nonpathogenic, Gram-negative, obligate anaerobic bacterium that commonly inhabits the human gut and degrades oxalate as its major energy and carbon source. Results from a case-controlled study suggested that lack of O. formigenes colonization is a risk factor for recurrent calcium oxalate stone formation. Hence, O. formigenes colonization may prove to be an efficacious method for limiting calcium oxalate stone risk. However, challenges exist in the preparation of O. formigenes as a successful probiotic due to it being an anaerobe with fastidious growth requirements. Here we examine in vitro properties expected of a successful probiotic strain. The data show that the Group 1 O. formigenes strain OxCC13 is sensitive to pH < 5.0, persists in the absence of oxalate, is aerotolerant, and survives for long periods when freeze-dried or mixed with yogurt. These findings highlight the resilience of this O. formigenes strain to some processes and conditions associated with the manufacture, storage and distribution of probiotic strains.

Oxalobacter formigenes Colonization and Oxalate Dynamics in a Mouse Model.

Animal and human studies have provided compelling evidence that colonization of the intestine with Oxalobacter formigenes reduces urinary oxalate excretion and lowers the risk of forming calcium oxalate kidney stones. The mechanism providing protection appears to be related to the unique ability of O. formigenes to rely on oxalate as a major source of carbon and energy for growth. However, much is not known about the factors that influence colonization and host-bacterium interactions. We have colonized mice with O. formigenes OxCC13 and systematically investigated the impacts of diets with different levels of calcium and oxalate on O. formigenes intestinal densities and urinary and intestinal oxalate levels. Measurement of intestinal oxalate levels in mice colonized or not colonized with O. formigenes demonstrated the highly efficient degradation of soluble oxalate by O. formigenes relative to other microbiota. The ratio of calcium to oxalate in diets was important in determining colonization densities and conditions where urinary oxalate and fecal oxalate excretion were modified, and the results were consistent with those from studies we have performed with colonized and noncolonized humans. The use of low-oxalate purified diets showed that 80% of animals retained O. formigenes colonization after a 1-week dietary oxalate deprivation. Animals not colonized with O. formigenes excreted two times more oxalate in feces than they had ingested. This nondietary source of oxalate may play an important role in the survival of O. formigenes during periods of dietary oxalate deprivation. These studies suggest that the mouse will be a useful model to further characterize interactions between O. formigenes and the host and factors that impact colonization.

The role of Oxalobacter formigenes colonization in calcium oxalate stone disease.

About 75% of urinary stones contain oxalate. As Oxalobacter formigenes is a Gram-negative anaerobic bacterium that degrades oxalate in the intestinal tract, we assessed the role of O. formigenes in oxalate metabolism by evaluating its intestinal absorption, plasma concentration, and urinary excretion. Of 37 calcium oxalate stone formers, 26 tested negative for O. formigenes and were compared with the 11 patients who tested positive. Patients provided 24-h urine samples on both a self-selected and a standardized diet. Urinary oxalate excretion did not differ significantly on the self-selected diet, but was significantly lower in O. formigenes-positive than in O. formigenes-negative patients under controlled, standardized conditions. Intestinal oxalate absorption, measured using [(13)C2]oxalate, was similar in the patients with or without O. formigenes. Plasma oxalate concentrations were significantly higher in noncolonized (5.79 µmol/l) than in colonized stone formers (1.70 µmol/l). Colonization with O. formigenes was significantly inversely associated with the number of stone episodes. Our findings suggest that O. formigenes lowers the intestinal concentration of oxalate available for absorption at constant rates, resulting in decreased urinary oxalate excretion. Thus, dietary factors have an important role in urinary oxalate excretion. The data indicate that O. formigenes colonization may reduce the risk of stone recurrence.

Oral antibiotic treatment of Helicobacter pylori leads to persistently reduced intestinal colonization rates with Oxalobacter formigenes.

BACKGROUND AND PURPOSE: Oxalobacter formigenes (OF) may play a protective role in preventing calcium oxalate stones. This is the first prospective study to evaluate the effect of antibiotics on OF colonization. Intestinal colonization by OF is associated with reduced urinary oxalate excretion. Exposure to antibiotics may be an important factor determining rates of colonization. MATERIALS AND METHODS: The effect of antibiotics on OF colonization was compared in two groups: A group receiving antibiotics for gastric infection with Helicobacter pylori (HP) and a group without HP whose members were not receiving antibiotics. OF colonization in stool was detected by oxalate degradation at baseline and after 1 and 6 months. RESULTS: The prevalence at baseline of intestinal colonization with OF was 43.1% among all patients screened. Among the 12 patients who were positive for OF who did not receive antibiotics, 11 (92%) had OF on stool tests at 1 month and 6 months. Of the 19 participants who were positive for OF and who received antibiotics for HP, only 7 (36.8%) continued to be colonized by OF on follow-up stool testing at 1 and 6 months (P=0.003 by Fisher exact test). Amoxicillin and clarithromycin caused 62.5% of subjects to become negative for OF at 1 month; 56.2% remained negative for OF at 6 months. CONCLUSIONS: Antibiotics for HP infection effectively reduced colonization with OF, an effect present at 1 and 6 months after treatment. The lasting elimination of OF could be associated with hyperoxaluria and be a factor in recurrent kidney stone disease.

Oxalobacter colonization in the morbidly obese and correlation with urinary stone risk.

OBJECTIVE: To establish the baseline preoperative prevalence of Oxalobacter formigenes (OF) colonization in a cohort of obese patients scheduled for Roux-en-Y gastric bypass (RYGB) and determine the effect of OF colonization on urinary oxalate excretion. It has been proposed that loss of OF colonization after RYGB may contribute to the development of hyperoxaluria. METHODS: Adult patients scheduled to undergo RYGB were requested to provide a stool specimen and 24-hour urine collection before surgery. OF colonization status was determined by the calcium precipitation test. The 24-hour urine specimens were analyzed by the Litholink Corporation (Chicago, IL). RESULTS: Of the 51 patients submitting initial stool specimens, only 8 (16%) tested positive for OF, whereas 43 (84%) were negative. Patients colonized with OF were older than uncolonized subjects (52.9±6.8 vs 46.0±10.4 years, P=.03). Urinary oxalate was not significantly different between these groups (P=.14). CONCLUSION: OF colonization is uncommon in morbidly obese patients (16%) before surgery. Because hyperoxaluria develops in more than 50% of patients after RYGB, it is unlikely that loss of OF colonization is the primary cause.

Factors related to colonization with Oxalobacter formigenes in U.S. adults.

GOALS: To elucidate the determinants of Oxalobacter formigenes colonization in humans. BACKGROUND: O. formigenes is a gram-negative anaerobic bacterium that colonizes the colon of a substantial proportion of the normal population and metabolizes dietary and endogenous oxalate. The bacterium has been associated with a large reduction in the odds of recurrent calcium oxalate kidney stones. Subjects were 240 healthy individuals from Massachusetts and North Carolina. O. formigenes was detected by culture of fecal swabs. Information on factors of interest was obtained by telephone interviews and self-administered questionnaires. STUDY RESULTS: The overall prevalence of O. formigenes was 38%. Use of specific antibiotics previously thought to affect the bacterium was significantly related to colonization, with prevalences of 17%, 27%, and 36%, for those who had used these drugs <1, 1-5, and >5 years ago, compared with 55% in nonusers. There were no significant associations with demographic factors, nutrient intake, or medical history, although the prevalence appeared to increase somewhat with increasing oxalate consumption. CONCLUSIONS: Some antibiotics markedly affect colonization with O. formigenes. Although no other factor was identified as having a material influence on the prevalence of the bacterium, there is much to learn about how an individual acquires the organism and which factors affect persistence of colonization.

Enteric oxalate elimination is induced and oxalate is normalized in a mouse model of primary hyperoxaluria following intestinal colonization with Oxalobacter.

Oxalobacter colonization of rat intestine was previously shown to promote enteric oxalate secretion and elimination, leading to significant reductions in urinary oxalate excretion (Hatch et al. Kidney Int 69: 691-698, 2006). The main goal of the present study, using a mouse model of primary hyperoxaluria type 1 (PH1), was to test the hypothesis that colonization of the mouse gut by Oxalobacter formigenes could enhance enteric oxalate secretion and effectively reduce the hyperoxaluria associated with this genetic disease. Wild-type (WT) mice and mice deficient in liver alanine-glyoxylate aminotransferase (Agxt) exhibiting hyperoxalemia and hyperoxaluria were used in these studies. We compared the unidirectional and net fluxes of oxalate across isolated, short-circuited large intestine of artificially colonized and noncolonized mice. In addition, plasma and urinary oxalate was determined. Our results demonstrate that the cecum and distal colon contribute significantly to enteric oxalate excretion in Oxalobacter-colonized Agxt and WT mice. In colonized Agxt mice, urinary oxalate excretion was reduced 50% (to within the normal range observed for WT mice). Moreover, plasma oxalate concentrations in Agxt mice were also normalized (reduced 50%). Colonization of WT mice was also associated with marked (up to 95%) reductions in urinary oxalate excretion. We conclude that segment-specific effects of Oxalobacter on intestinal oxalate transport in the PH1 mouse model are associated with a normalization of plasma oxalate and urinary oxalate excretion in otherwise hyperoxalemic and hyperoxaluric animals.

Effect of antibiotics on Oxalobacter formigenes colonization of human gastrointestinal tract.

BACKGROUND AND PURPOSE: Oxalobacter formigenes is a bacterium residing in the human gastrointestinal tract that degrades oxalate and reduces its availability for absorption. This bacterium is assumed to be antibiotic sensitive, and repeated antibiotic therapies could eradicate it. The aim of the present study was to determine the differences in the colonization by O. formigenes of individuals who had been on antibiotics for at least 5 days at the time of sample collection and individuals who had not taken antibiotics for at least 3 months. PATIENTS AND METHODS: Stool samples were collected from 80 individuals without stone disease (35 with and 45 without antibiotic consumption) and 100 patients with stone disease (20 with and 80 without antibiotic consumption). Oxalobacter formigenes was detected by a polymerase chain reaction-based method, and the presence/absence of O. formigenes was correlated with urinary oxalate concentrations. RESULTS: Lower percentages of individuals without stone disease and with stone disease who were consuming antibiotics had O. formigenes colonization than individuals without antibiotic consumption. Urinary oxalate concentrations were higher in the individuals without O. formigenes than in colonized individuals. CONCLUSION: Our observations confirm a direct association between antibiotic consumption and absence of O. formigenes. Absence of intestinal O. formigenes could represent a pathogenic factor in calcium oxalate urolithiasis when antibiotics are prescribed generously.

Infrequency of colonization with Oxalobacter formigenes in inflammatory bowel disease: possible role in renal stone formation.

BACKGROUND AND AIM: Calcium oxalate renal stones (RS) and hyperoxaluria are common in patients with inflammatory bowel disease (IBD). The absence of intestinal oxalate degrading bacteria, Oxalobacter formigenes, may cause hyperoxaluria in IBD. The aim of the present study was to examine: (i) the colonization of O. formigenes in patients with IBD and controls and to correlate its presence with urinary oxalate excretion; and (ii) urinary analytes contributing to RS in IBD. METHODS: Stool samples were studied for O. formigenes using polymerase chain reaction and Southern blotting in patients with IBD (n = 48: ulcerative colitis, 37; Crohn's disease, 11), RS (n = 87) and healthy subjects that were used as controls (n = 48). Levels of urinary oxalate, citrate, calcium, magnesium, creatinine and uric acid were estimated spectrophotometrically in each patient and in 13 controls for 24 h. RESULTS: Five of the 48 (10.4%) patients with IBD had RS. Five of the 48 (10.4%) patients with IBD, 25 of the 87 (29%) with RS and 27 of the 48 (56%) controls were colonized with O. formigenes (P < 0.001 for RS vs controls and P = 0.01 for RS vs IBD). Patients without O. formigenes had higher urinary oxalate than those with it (IBD, median 0.48 [range 0.11-2.09]vs 0.43 [range 0.16-1.10] mmol/24 h, P = NS; RS, median 0.59 mmol/24 h, range 0.14-1.90 vs 0.44 mmol/24 h, range 0.23-0.97; P = 0.008, Mann-Whitney U-test). Median excretion of oxalate was higher in IBD and RS than in controls (0.47 [0.11-2.09], 0.56 [0.14-1.9] and 0.41 [0.21-0.62] mmol/24 h; P < 0.01), respectively. Median calcium was also higher in IBD and RS than in controls (6.50 [1.38-21.00], 6.78 [1.55-20.30] and 4.99 [1.47-9.60] mmol/24 h; P < 0.05, Kruskal-Wallis H-test), respectively. Median urinary magnesium was higher in IBD than in RS and controls (4.57 [1.50-12.30], 3.60 [0.90-6.35] and 2.49 [0.74-4.80]; P < 0.001, Kruskal-Wallis H-test), respectively. Urinary citrate excretion was comparable in IBD, RS and controls. CONCLUSIONS: Patients with IBD and RS rarely have O. formigenes in their stools as compared with controls; this may contribute to hyperoxaluria in IBD. Hyperoxaluria and hypercalciuria may contribute to RS in patients with IBD. Hypermagnesuria in patients with IBD may protect them from RS.

Colonization of the neonatal rat intestinal tract from environmental exposure to the anaerobic bacterium Oxalobacter formigenes.

Oxalobacter formigenes, an anaerobic bacterium that inhabits the mammalian gastrointestinal tract, has an important symbiotic relationship with its vertebrate hosts by regulating oxalic acid homeostasis. Epidemiological studies of O. formigenes colonization in man have shown that colonization occurs in young children, that every child can become colonized naturally, that >20% lose colonization during adolescence or as adults and that stable colonization can be disrupted by antibiotic use or changes in diet, greatly affecting subsequent health. As O. formigenes is a fastidious anaerobe that seldom re-colonizes adults, the question arises as to how initial colonization occurs. To investigate this question, non-colonized female laboratory rats were placed on diets high in oxalate and were colonized by oesophageal gavage with O. formigenes either before or after being impregnated. Faecal specimens from their offspring were tested for the presence of O. formigenes. Although the bacterium was first detected in a few neonates as early as 7 days post-partum, colonization of all the offspring did not occur until after weaning. In each case, the offspring were colonized with the bacterial strain carried by their mothers. To determine whether O. formigenes colonization occurs vertically or horizontally, newborn rats were placed with foster mothers that were either non-colonized or colonized with an O. formigenes strain different from that of their natural mothers. Colonization occurred temporally in a manner similar to natural colonization but all offspring became colonized only with the O. formigenes strain of the foster mothers. These data indicate that intestinal colonization occurs horizontally, but does not answer the question of how O. formigenes survives the aerobic environment in order to be transmitted.