David Beltrán

Time course production of urolithins from ellagic acid by human gut microbiota.

Ellagic acid (EA) is converted to urolithins by gut microbiota. Urolithins have beneficial biological effects in humans, but differences in urolithin production capacity among individuals have been shown. Therefore, the identification of the urolithin production pathways and the microorganisms implicated is of high interest. EA was incubated with gut microbiota from two volunteers able to produce urolithins but with different in vivo urolithin profiles (urolithin A and isourolithin A producers). The metabolic capabilities observed in vivo were retained in vitro. Both individuals showed a much higher abundance of Clostridium leptum group of Firmicutes phylum than Bacteroides / Prevotella . EA was either dissolved in DMSO or suspended in water. DMSO increased EA solubility but decreased urolithin production rate due to a delay in growth of some microbial groups, principally, Clostridium coccoides . This allowed the detection of catabolic intermediates [urolithins M-5, M-6, M-7, C, and 2,3,8,10-tetrahydroxy urolithin (urolithin E)]. Bacteria from C. coccoides group (or genera co-occurring in vivo with this group) seem to be involved in production of different urolithins.

Effect of Ozone on the Inactivation of Yersinia enterocolitica and the Reduction of Natural Flora on Potatoes

Fresh vegetables contaminated with Yersinia enterocolitica have been implicated in foodborne disease outbreaks. Surfaces of vegetables can become contaminated with pathogenic microorganisms through contact with soil, irrigation water, fertilizers, equipment, humans, and animals. One approach to reduce this contamination is to treat fresh produce with sanitizers. In this study, the ability of ozone to inactivate Y. enterocolitica inoculated in water and on potato surfaces was evaluated. Furthermore, the efficacy of ozone in reducing natural flora on whole potato was determined. Total aerobic mesophilic and psychrotrophic bacteria, total coliforms, and Listeria monocytogenes were enumerated. Finally, several disinfection kinetic models were considered to predict Y. enterocolitica inactivation with ozone. Treatments with ozone (1.4 and 1.9 ppm) for 1 min decreased the Y. enterocolitica population in water by 4.6 and 6.2 log CFU ml(-1), respectively. Furthermore, ozonated water (5 ppm) for 1 min decreased Y. enterocolitica and L. monocytogenes from potato surfaces by 1.6 and 0.8 log CFU g(-1), respectively. Therefore, ozone can be an effective treatment for disinfection of wash water and for reduction of potato surface contamination.

Gordonibacter urolithinfaciens sp. nov., a urolithin-producing bacterium isolated from the human gut.

Urolithins are dibenzopyranone metabolites that exert anti-inflammatory activity in vivo and are produced by the gut microbiota from the dietary polyphenols ellagic acid (EA) and ellagitannins. However, the bacteria involved in this process remain unknown. We report here a novel bacterium, strain CEBAS 1/15P(T), capable of metabolizing EA to urolithins, that was isolated from healthy human faeces and characterized by determining phenotypic, biochemical and molecular methods. The strain was related to Gordonibacter pamelaeae 7-10-1-b(T), the type and only reported strain of the only species of the genus Gordonibacter, with about 97% 16S rRNA gene sequence similarity; they were both obligately anaerobic, non-spore-forming, Gram-stain-positive, short-rods/coccobacilli and metabolized only small numbers of carbon sources. L-Fucose, D-fructose, turanose, D-galacturonic acid and α-ketobutyric acid were metabolized by strain CEBAS 1/15P(T), while G. pamelaeae was negative for metabolism of these compounds. The whole-cell fatty acids consisted predominantly of saturated fatty acids (70%); strain CEBAS 1/15P(T) differed significantly from G. pamelaeae in the major fatty acid, which was C18 : 1ω9c, while anteiso-C15 : 0 was the major component for G. pamelaeae. The presence of a number of different fatty acid peaks, especially C19 : 0 cyclo and C18 : 1ω6c, was also indicative of distinct species. Six glycolipids (GL1-6) were recognized, while, in G. pamelaeae, only four glycolipids were described. On the basis of these data, the novel species Gordonibacter urolithinfaciens sp. nov. is described, with strain CEBAS 1/15P(T) ( = DSM 27213(T) = CCUG 64261(T)) as the type strain.

Isolation of Human Intestinal Bacteria Capable of Producing the Bioactive Metabolite Isourolithin A from Ellagic Acid

Urolithins are intestinal microbial metabolites produced from ellagitannin- and ellagic acid-containing foods such as walnuts, strawberries, and pomegranates. These metabolites, better absorbed than their precursors, can contribute significantly to the beneficial properties attributed to the polyphenols ellagitannins and ellagic acid (EA). However, both the ability of producing the final metabolites in this catabolism (urolithins A, B and isourolithin A) and the health benefits associated with ellagitannin consumption differ considerably among individuals depending on their gut microbiota composition. Three human urolithin metabotypes have been previously described, i.e., metabotype 0 (urolithin non-producers), metabotype A (production of urolithin A as unique final urolithin) and metabotype B (urolithin B and/or isourolithin A are produced besides urolithin A). Although production of some intermediary urolithins has been recently attributed to intestinal species from Eggerthellaceae family named Gordonibacter urolithinfaciens and Gordonibacter pamelaeae, the identification of the microorganisms responsible for the complete transformation of EA into the final urolithins, especially those related to metabotype B, are still unknown. In the present research we illustrate the isolation of urolithin-producing strains from human feces of a healthy adult and their ability to transform EA into different urolithin metabolites, including isourolithin A. The isolates belong to a new genus from Eggerthellaceae family. EA transformation and urolithin production arisen during the stationary phase of the growth of the bacteria under anaerobic conditions. The HPLC-DAD-MS analyses demonstrated the sequential appearance of 3,8,9,10-tetrahydroxy-urolithin (urolithin M6), 3,8,9-trihydroxy-urolithin (urolithin C) and 3,9-dihydroxy-urolithin (isourolithin A) while 3,8-dihydroxy-urolithin (urolithin A) and 3-hydroxy-urolithin (urolithin B) were not detected. For the first time isourolithin A production capacity of pure strains has been described. The biological activity attributed to urolithins A and B and isourolithin A (anti-inflammatory, anti-carcinogenic, cardioprotective, and neuroprotective properties) explains the relevance of identifying these urolithin-producing bacteria as potential novel probiotics with applications in the development of functional foods and nutraceuticals. Their human administration could improve the health benefits upon ellagitannin consumption, especially in metabotype 0 individuals. However, further research is necessary to probe well-established beneficial effects on the host and safety requirements before being considered among the next-generation probiotics.

Ellagibacter isourolithinifaciens gen. nov., sp. nov., a new member of the family Eggerthellaceae, isolated from human gut

Urolithins are gut microbial metabolites that exert health benefits in vivo and are generated from ellagic acid (EA) and ellagitannin-containing foods such as strawberries, pomegranates and walnuts. Gordonibacter species produce some intermediary urolithins but the micro-organisms responsible for the transformation of EA into the final and more bioactive urolithins, such as urolithin A and isourolithin A, are unknown. We report here a new bacterium, capable of metabolizing EA into isourolithin A, isolated from healthy human faeces and characterized by determining phenotypic, biochemical and molecular methods. Strain CEBAS 4A belongs to the Eggerthellaceae family and differed from other genera of this family, both phylogenetically and phenotypically. Based on 16S rRNA gene sequence similarity, the strain was related to Enterorhabdus musicola DSM 19490T (92.9 % similarity), Enterorhabdus caecimuris DSM 21839T (92.7 % similarity), Adlercreutzia equolifaciens DSM 19450T (92.5 % similarity), Asaccharobacter celatus DSM 18785T (92.5 % similarity) and Parvibacter caecicola DSM 22242T (91.2 % similarity). This strain was strictly anaerobic and Gram-stain-positive. The whole-cell fatty acids were saturated (98.3 %), a very high percentage that differs from the nearest genera ranging from 62 to 73 %. The major respiratory lipoquinone was menaquinone-7 and the diamino acid in the peptidoglycan was meso-diaminopimelic acid. Diphosphatidylglycerol and phosphatidylglycerol comprised the main polar lipid profile in addition to several phosphoglycolipids (PGL1-2), phospholipids (PL1-4), glycolipids (GL1-6) and lipids. Based on these data, a new genus, Ellagibacter gen. nov. is proposed with one species, Ellagibacter isourolithinifaciens sp. nov. The type strain of Ellagibacter isourolithinifaciens is CEBAS 4AT (=DSM 104140T=CCUG 70284T).