In vitro Selection of Probiotics for Microbiota Modulation in Normal-Weight and Severely Obese Individuals: Focus on Gas Production and Interaction With Intestinal Epithelial Cells

The intestinal microbiota plays important roles in the maintenance of health. Strategies aiming at its modulation, such as probiotics, have received a deal of attention. Several strains have been studied in different in vitro models; however, the correlation of results obtained with the in vivo data has been limited. This questions the usefulness of such in vitro selection models, traditionally relying on over-simplified tests, not considering the influence of the accompanying microbiota or focusing on microbiota composition without considering functional traits. Here we assess the potential of six Bifidobacterium, Lactobacillus and Lacticaseibacillus strains in an in vitro model to determine their impact on the microbiota not just in terms of composition but also of functionality. Moreover, we compared the responses obtained in two different population groups: normal-weight and severely obese subjects. Fecal cultures were conducted to evaluate the impact of the strains on specific intestinal microbial groups, on the production of short-chain fatty acids, and on two functional responses: the production of gas and the interaction with human intestinal epithelial cells. The response to the different probiotics differed between both human groups. The addition of the probiotic strains did not induce major changes on the microbiota composition, with significant increases detected almost exclusively for the species added. Higher levels of gas production were observed in cultures from normal-weight subjects than in the obese population, with some strains being able to significantly reduce gas production in the latter group. Moreover, in obese subjects all the Bifidobacterium strains tested and Lacticaseibacillus rhamnosus GG were able to modify the response of the intestinal cells, restoring values similar to those obtained with the microbiotas of normal-weight subjects. Our results underline the need for the screening and selection of probiotics in a target-population specific manner by using appropriate in vitro models before enrolling in clinical intervention trials.The present work was mainly financed by the project AGL2013-43770-R (receiving funds from the Ministry of Economy and Competitiveness—MINECO and the European Union FEDER funds), and partly by projects AGL2017-83653-R (AEI/FEDER, UE), and RTI2018-098288-B-I00 (MCIU/AEI/FEDER, UE) and by a contract with the company Takanashi Milk Products (Yokohama, Japan). AN was the recipient of a predoctoral contract granted by MINECO (reference BES-2014-068796). SA is currently financed by a postdoctoral Juan de la Cierva contract (Ministry of Science, Innovation and Universities, Spain, ref. IJCI-2017-32156) and NS is currently the recipient of a postdoctoral contract awarded by the Fundación para la Investigación y la Innovación Biosanitaria del Principado de Asturias (FINBA)

Impact of extreme obesity and diet-induced weight loss on the fecal metabolome and gut microbiota

[Scope]: A limited number of human studies have characterized fecal microbiota and metabolome in extreme obesity and after diet-induced weight loss.[Methods and results]: Fecal samples from normal-weight and extremely obese adults and from obese participants before and after moderate diet-induced weight loss are evaluated for their interaction with the intestinal adenocarcinoma cell line HT29 using an impedance-based in vitro model, which reveals variations in the interaction between the gut microbiota and host linked to obesity status. Microbiota composition, short chain fatty acids, and other intestinal metabolites are further analyzed to assess the interplay among diet, gut microbiota, and host in extreme obesity. Microbiota profiles are distinct between normal-weight and obese participants and are accompanied by fecal signatures in the metabolism of biliary compounds and catecholamines. Moderate diet-induced weight loss promotes shifts in the gut microbiota, and the primary fecal metabolomics features are associated with diet and the gut–liver and gut–brain axes.[Conclusions]: Analyses of the fecal microbiota and metabolome enable assessment of the impact of diet on gut microbiota composition and activity, supporting the potential use of certain fecal metabolites or members of the gut microbiota as biomarkers for the efficacy of weight loss in extreme obesity.The present work was mainly financed by the project AGL2013-43770-R (receiving funds from the Ministry of Economy and Competitiveness—MINECO and the European Union FEDER funds), and partly by projects AGL2017-83653-R (AEI/FEDER, UE), and RTI2018-098288-B-I00 (MCIU/AEI/FEDER, UE). A.M.N. was the recipient of a predoctoral contract granted by MINECO (reference BES-2014-068796) and N.S. received a postdoctoral Juan de la Cierva Contract (Ministry of Science, Innovation and Universities, Spain, reference IJCI-2015-19885) and currently she has a postdoctoral contract awarded by the Biosanitary Research Foundation in Asturias (FINBA, Spain).Peer reviewe

Probiotic-Induced Modulation of Microbiota Composition and Antibiotic Resistance Genes Load, an In Vitro Assessment

The imbalance of the gut microbiota (GM) is known as dysbiosis and is associated with disorders such as obesity. The increasing prevalence of microorganisms harboring antibiotic resistance genes (ARG) in the GM has been reported as a potential risk for spreading multi-drug-resistant pathogens. The objective of this work was the evaluation, in a fecal culture model, of different probiotics for their ability to modulate GM composition and ARG levels on two population groups, extremely obese (OB) and normal-weight (NW) subjects. Clear differences in the basal microbiota composition were observed between NW and OB donors. The microbial profile assessed by metataxonomics revealed the broader impact of probiotics on the OB microbiota composition. Also, supplementation with probiotics promoted significant reductions in the absolute levels of tetM and tetO genes. Regarding the blaTEM gene, a minor but significant decrease in both donor groups was detected after probiotic addition. A negative association between the abundance of Bifidobacteriaceae and the tetM gene was observed. Our results show the ability of some of the tested strains to modulate GM. Moreover, the results suggest the potential application of probiotics for reducing the levels of ARG, which constitutes an interesting target for the future development of probiotics.The present work was funded by project AGL2017-83653-R (AEI/FEDER, UE), by the project AYUD/2021/ 50981 from the Principality of Asturias, to support the activity of their research groups. The authors declare that this study has also received funding from Takanashi Milk Products (Yokohama, Japan). The funder had the following involvement with the study: metataxonomic analyses and review of the manuscript. S.A. was financed by a postdoctoral Juan de la Cierva contract (Ministry of Science, Innovation and Universities, Spain, ref. IJCI-2017-32156). A.M.N., S.S. and S.A. are currently the recipient of contracts awarded by the Fundación para la Investigación Biosanitaria de Asturias (FINBA).Peer reviewe

Nutrición parenteral domiciliaria en España, 2019: informe del Grupo de Nutrición Artificial Domiciliaria y Ambulatoria NADYA

RESUMEN Objetivo: comunicar los datos de nutrición parenteral domiciliaria (NPD) obtenidos del registro del grupo NADYA-SENPE (www.nadyasenpe.com) del año 2019. Material y métodos: análisis descriptivo de los datos recogidos de pacientes adultos y pediátricos con NPD en el registro NADYA-SENPE desde el 1 de enero al 31 de diciembre de 2019. Resultados: se registraron 283 pacientes (51,9 %, mujeres), 31 niños y 252 adultos procedentes de 47 hospitales españoles, lo que representa una tasa de prevalencia de 6,01 pacientes/millón de habitantes/año 2019. El diagnóstico más frecuente en los adultos fue “oncológico paliativo” y “otros” (21,0 %). En los niños fue la enfermedad de Hirschsprung junto a la enterocolitis necrotizante, las alteraciones de la motilidad intestinal y la pseudoobstrucción intestinal crónica, con 4 casos cada uno (12,9 %). El primer motivo de indicación fue el síndrome del intestino corto tanto en los niños (51,6 %) como en los adultos (37,3 %). El tipo de catéter más utilizado fue el tunelizado tanto en los niños (75,9 %) como en los adultos (40,8 %). Finalizaron 68 episodios, todos en adultos: la causa más frecuente fue el fallecimiento (54,4 %). Pasaron a la vía oral el 38,2 %. Conclusiones: el número de centros y profesionales colaboradores con el registro NADYA va incrementándose. Se mantienen estables las principales indicaciones y los motivos de finalización de la NPD

Microbiota y perfil fecal de aminoácidos y metabolitos derivados según el grado de obesidad

Trabajo presentado al XIII Workshop Sociedad Española de Microbiota, Probióticos y Prebióticos, celebrado en Valencia (España), del 7 la 9 de junio de 2022

A fecal-culture model, monitoring gas production, for assessing prebiotics' fermentability in normal-weight and obese subjects

Trabajo presentado en el 10th Workshop on Probiotics and Prebiotics (SEPyP 2019), celebrado en Las Palmas de Gran Canaria (España) del 6 al 8 de febrero de 2019[Background/Aims] The gut microbiota is altered in different conditions, which leads to the interest in prebiotics to restore it and to identify the best suited compounds. The use of fecal culture models provides an interesting strategy, allowing to identify prebiotics with suitable fermentation profiles, and appropriate microbiota-modulation properties, for each population. Here we aimed at comparing the fermentability and microbiota-modulation ability of different prebiotics, including the novel prebiotic 1-Kestose, in normal-weight and obese adults. [Methods] Fresh fecal samples from 9 normal-weight (BMI 40) volunteers were collected, transported to the laboratory (under anaerobic conditions), homogenized, diluted (10% v/v) into a carbohydrate-free basal medium and stabilized at 37°C in an anaerobiosis cabinet. The prebiotics tested (Actilight, Synergy, P95, Inulin, GOS and 1-kestose) were added (0.3%, w/v) to the stabilized fecal cultures and incubated at 37°C for 24 hours. Gas production along incubation was monitored in real-time by using the ANKOM RF System. Samples were taken at 0 and 24 hours of incubation for pH measurements, determination of short-chain-fatty-acids by gas chromatography and gut microbiota analyses by qPCR. [Results] In both volunteers’ groups kestose resulted the more fermentable prebiotic as indicated by the largest accumulation of gas and the lowest pH after incubation. On the contrary, inulin was the less fermentable compound. Interestingly, intestinal microbiota from obese individuals tended to show a lower ability to produce gas than that from normal-weight volunteers. The different prebiotics were able to induce changes in microbiota composition in both volunteer groups, although showing differences among them. [Conclusions] The fecal culture model used, with real-time monitoring of gas production, constitutes a fast and easy method for assessing the fermentability of prebiotics in different population groups. 1-kestose showed good characteristics suggesting its applicability as a readily fermentable prebiotic substrate for intestinal microbiota modulation

Selección de prebióticos dirigidos a individuos con obesidad mórbida y normopeso a través de un modelo funcional in vitro

Trabajo presentado en la 13ª Reunión de la Red Española de Bacterias Lácticas (RedBAL), celebrada en Madrid (España) del 17 al 18 de junio de 2019La microbiota intestinal constituye un ecosistema complejo y dinámico, cuyo desequilibrio conduce a un estado de disbiosis que se ha relacionado con diversas enfermedades, entre ellas la obesidad. Por esta razón, existe gran interés en la modulación de la microbiota a través estrategias nutricionales, como la administración de prebióticos. Existen numerosos modelos para estudiar los efectos de los prebióticos, pero aún es necesario el desarrollo de modelos in vitro sencillos y económicamente asequibles, que permitan evaluar la fermentación de los mismos considerando el complejo contexto de la microbiota intestinal permitiendo una mejor predicción de su funcionalidad en humanos. En este trabajo se estudió la respuesta in vitro de la microbiota intestinal a diferentes prebióticos, utilizando la producción de gas como medida en tiempo real de la funcionalidad de los compuestos. Se llevaron a cabo cultivos fecales a partir de muestras obtenidas de población adulta normopeso (NP, n=9), y voluntarios diagnosticados con obesidad mórbida (OB, n=9). Tras la estabilización de la microbiota se añadieron prebióticos de tipo fructano (Actilight, Synergy1, P95, 1-kestosa, e inulina), y de tipo galactano (GOS: Bimuno) a una concentración final de 0,3% p/v acompañados de un control negativo de fermentación y un control positivo no-prebiótico (glucosa como fuente de carbono). Se monitorizó la acumulación de gas utilizando la tecnología ANKOM RF, tomando además muestras para el estudio de la composición microbiana mediante qPCR y de ácidos grasos de cadena corta (AGCC) por cromatografía de gases (tiempo 0 y tras 24 horas de incubación). Los resultados de la producción de gas así como descenso del pH muestran una alta fermentabilidad del nuevo prebiótico 1-kestosa mientras que la inulina fue el substrato con fermentación más lenta, obteniéndose la misma tendencia para ambas poblaciones, si bien se observaron algunas diferencias entre ambas. La adición de prebióticos 1- kestosa y GOS produjo profundos cambios en la microbiota de OB, aumentando durante la incubación los niveles de los siguientes grupos microbianos: Bifidobacterium, grupo Bacteroides, Faecalibacterium así como niveles totales de bacterias. En cambio, estos mismos sustratos afectaron de forma diferente a la microbiota de NP, en la que tan solo se detectó un aumento de los niveles de los géneros Faecalibacterium y Bifidobacterium. Respecto a los AGCC, los prebióticos que más incrementaron su producción por la microbiota fueron la 1-kestosa en OB, y GOS en NP. La comparación de los resultados muestra además una mayor actividad fermentativa en cuanto a niveles de gas acumulado y metabolitos producidos por la microbiota de la población NP respecto a la OB. La aplicación del modelo in vitro utilizado nos permitió comparar la fermentabilidad de diversos prebióticos. 1-kestosa y GOS presentaron mayor capacidad moduladora que el resto de los substratos ensayados, pero con diferente efecto según la población analizada. La microbiota de NP presentó una actividad metabólica mayor, mientras que en OB los cambios en la composición microbiana fueron mayores que en NP. Los resultados obtenidos señalan la utilidad del modelo in vitro para el estudio de la fermentabilidad de diferentes substratos y su seguimiento en tiempo real, además su aplicación permite la selección del compuesto con mayor potencial para cada población diana

Genetic diagnosis of familial hypercholesterolemia using a DNA-array based platform

The aim of this study was to validate the Lipochip genetic diagnostic platform by assessing effectiveness, sensitivity, specificity and costs for the identification of patients with familial hypercholesterolemia (FH) in Spain. This platform includes the use of a DNA micro array, the detection of large gene rearrangements and the complete resequencing of the low-density lipoprotein receptor gene. DNA samples of patients with clinically diagnosed FH were analyzed for mutations by application of the Lipochip platform. Results obtained were confirmed by DNA sequencing and MLPA analysis by two other, independent laboratories. Of 808 patients tested, Lipochip detected a mutation in 66% of the cases and of these 78% were detected by the micro array. A specificity of 99.5% at a sensitivity of 99.8% was reached. A positive test result could be reported within 22 days after start of analysis. The total average screening costs of $350 per case were significantly lower compared to other existing screening programs. Lipochip provides a reliable, fast and cheap alternative for the genetic testing of patients with clinically diagnosed F

Aplicación de inteligencia artificial en el estudio de la microbiota intestinal asociada con obesidad mórbida

Resumen del trabajo presentado en el XI Workshop de la Sociedad Española de Microbiota, Probióticos y Prebióticos, celebrado en Granada (España), del 12 al 14 de febrero de 2020Introducción/Objetivo. La obesidad representa una patología compleja y multifactorial con una elevada prevalencia a nivel mundial. Numerosos estudios han tratado de caracterizar la composición de la microbiota intestinal asociada a la obesidad. Sin embargo, los resultados no son concluyentes ni muestran información discriminatoria para los diferentes grados de obesidad. Sujetos y métodos. Se estudió una muestra de 184 sujetos, clasificada en 5 categorías de IMC según la Sociedad Española para el Estudio de la Obesidad. La composición de la microbiota intestinal se determinó por qPCR y la información dietética a partir de un cuestionario de frecuencias de consumo validado. La influencia de las variables microbiológicas sobre la obesidad se calculó mediante técnicas de aprendizaje matemático. Tras el entrenamiento de diferentes métodos se seleccionó el árbol de decisión C4.5 tanto por su capacidad discriminatoria como por la interpretabilidad de los datos obtenidos en el contexto de la obesidad. Resultados. A través de árboles de decisión (con un porcentaje de acierto 95%, precisión 92% y cobertura 1), ha sido posible identificar al grupo Bacteroides como el grupo microbiano con mayor valor predictivo para obesidad mórbida en la muestra analizada. Niveles del grupo Bacteroides por encima de 10,17 log nºcel/g clasifican a los sujetos con un IMC> 40 kg/m2. Dichos sujetos presentaron también una mayor ingesta dietética de carnes y derivados (170,7 vs. 123,7 g/d) así como de huevos (24,4 vs. 17,6 g/d). Por el contrario, el consumo de ácidos grasos poliinsaturados fue menor que los sujetos pertenecientes al resto de categorías de IMC y no se observaron diferencias significativas en la ingesta energética. Conclusiones. Hemos desarrollado la inteligencia artificial para la identificación de un nivel de referencia de microrganismos fecales, Bacteroides > 10,17 log nºcel/g característico de los individuos con obesidad mórbida frente a individuos con menor valor de IMC

Teduglutide: a review of its use in short bowel syndrome

Introduction: Long-term Parenteral Support (PS) can be associated with serious complications, with a significant deterioration in the quality of life of patients with short bowel syndrome (SBS). Teduglutide is a recombinant analogue of glucagon-like peptide-2; it belongs to a novel therapeutic family and represents the first non-symptomatic approach against SBS.Objectives: To review the non-clinical and clinical data on efficacy and safety of teduglutide.Results: Teduglutide approval was based on results from a pivotal Phase III, 24-week, double-blind, placebo-controlled study (STEPS). SBS patients dependent on PS >= 3 times/week for >= 12 months received 0.05 mg/kg teduglutide (n = 43) or placebo (n = 43) 1 time/day. At week 24 there were significantly more responders in the teduglutide group vs. placebo (63 vs. 30%; p = 0.002). The overall mean reduction vs. PS baseline volume at week 24 was significantly higher with teduglutide vs. placebo (4.4 vs. 2.3 l/ week, p = 3 times/week for >= 12 months received 0.05 mg/kg teduglutide (n = 43) or placebo (n = 43) 1 time/day. At week 24 there were significantly more responders in the teduglutide group vs. placebo (63 vs. 30%; p = 0.002). The overall mean reduction vs. PS baseline volume at week 24 was significantly higher with teduglutide vs. placebo (4.4 vs. 2.3 l/ week, p = 12 months received 0.05 mg/kg teduglutide (n = 43) or placebo (n = 43) 1 time/day. At week 24 there were significantly more responders in the teduglutide group vs. placebo (63 vs. 30%; p = 0.002). The overall mean reduction vs. PS baseline volume at week 24 was significantly higher with teduglutide vs. placebo (4.4 vs. 2.3 l/ week,