Taxonomic and functional characterization of the microbial community involved in obtaining biogas and other compounds of biotechnological interest from citrus peel waste

A recuperação de energia a partir de biomassa lignocelulósica, como os resíduos sólidos de citros (RSC), vem sendo estudada como alternativa para seu reaproveitamento. O objetivo desta pesquisa foi avaliar a utilização de RSC como substrato para a obtenção de H2 e demais bioprodutos em reatores em batelada, bem como caracterizar a comunidade microbiana envolvida neste processo. Foi possível obter um consórcio autóctone por meio da autofermentação do RSC, além de isolar uma cepa similar à Enterococcus casseliflavus (99%), e obter os parâmetros cinéticos de seu crescimento em glicose, como crescimento específico (μ = 0,35 h), tempo de geração celular (Tg = 1,98 h-1), potencial máximo de concentração de H2 (P = 9,1 mmol H2.L-1), velocidade específica máxima de obtenção de H2 (Rm = 1,99 mmol H2.L-1) e tempo de início da obtenção de H2 (λ = 4,08 h). Na etapa seguinte, foram avaliados dois diferentes pré-tratamentos, sendo que a maior concentração de H2 ocorreu nos reatores contendo RSC in natura (13,31 mmol H2.L-1), quando comparado à hidrotermólise (P = 8,19 mmol H2.L-1) e deslignificação alcalina (P = 7,27 mmol H2.L-1). Por meio do método de Plackett & Burman, foram avaliadas as variáveis pH (5,5, 7,0 e 8,5), temperatura (30, 37 e 44 °C), concentração de inóculo alóctone (1, 2 e 3 gSTV.L-1) e de RSC (5, 10 e 15 g.L-1), volume do headspace (40, 50 e 60%) e componentes do meio de cultura, como extrato de levedura (0, 0,5 e 1 g.L-1), CaCO3 (0, 2,5 e 5 g.L-1), NaCl (0, 2,5 e 5 g.L-1) e peptona (0, 2,5 e 5 g.L-1). O pH, concentração de inóculo e substrato (RSC) foram as variáveis significativas. A condição ótima estabelecida por meio de Delineamento do Composto Central Rotacional foi de 4 gSTV.L-1 de inóculo (lodo granular aplicado ao processamento termofílico de vinhaça de cana-de-açúcar), 29,8 g.L-1 de RSC e pH 8,98, com P = 48,47 mmol H2.L-1. A partir do sequenciamento shotgun, foi possível confirmar a partir da expressão gênica diferencial que as vias de degradação da celulose pela via acetogênica foram superexpressas nos reatores otimizados, sendo Clostridium e Paraclostridium os gêneros mais abundantes.Energy recovery from lignocellulosic biomass, such as citrus peel waste (CPW), has been studied as an alternative to its reuse. This research aimed at evaluating the use of CPW as substrate to obtain H2 and other by-products in batch reactors, as well as characterizing the microbial community involved in this process. It was possible to obtain an autochthonous consortium through the self-fermentation of CPW, in addition to isolating a strain similar to Enterococcus casseliflavus (99%), as well as obtaining its growth kinetic parameters in glucose, namely specific growth (μ = 0.35 h), generation time (Tg = 1.98 h-1), maximum H2 concentration potential (P = 9.1 mmol H2.L-1), maximum H2 production rate (Rm = 1.99 mmol H2.h-1) and starting time of H2 production (λ = 4.08 h). The effect of hydrothermolysis (180 °C; 15 minutes) and alkaline delignification (NaOH 5 mol.L-1; 1% H2O2; 24 hours) pretreatments in CPW were evaluated, where it was observed that the H2 obtainment was higher in reactors containing fresh CPW (13.31 mmol H2.L-1) when compared to hydrothermolysis (P = 8.19 mmol H2.L-1) and alkaline delignification (P = 7.27 mmol H2.L-1). By means of applying the Plackett & Burman statistical method, the variables pH (5.5,7.0 and 8.5), temperature (30, 37 and 44 °C), inoculum (1, 2 and 3 gSTV.L-1) and CPW (5,10 and 15 g.L-1) concentrations, headspace volume (40, 50 and 60%) and components of the culture medium were evaluated, namely yeast extract (0, 0.5 and 1 g.L-1), CaCO3, NaCl and peptone (0, 2.5 and 5 g.L-1) concentrations, with pH, inoculum and CPW concentrationthe most significant variables. The optimal condition established by the Rotational Central Composite Design was 4 gSTV.L-1 of inoculum (sludge of sugarcane vinasse anaerobic treatment), 29.8 g.L-1 of CPW and pH 8.98, with maximum production of 48.47 mmol H2.L-1. After the shotgun sequencing, it was possible to confirm, from differential gene expression, that the cellulose degradation acetogenic pathways were overexpressed in the optimized reactor, with Clostridium and Paraclostridium the most abundant genera

A Microbiologia no caderno do aluno e em livros didáticos: análise documental

Este trabalho tem o objetivo de avaliar em que medida as propostas didáticas de livros da área contribuem, ou não, para a superação de obstáculos no processo de ensino-aprendizagem de Microbiologia, por meio de uma análise comparativa dos temas relacionados à Microbiologia nos materiais didáticos analisados, a saber: coleção de livros didáticos e apostilas destinadas aos anos finais do Ensino Fundamental (Caderno do Aluno). A pesquisa é de natureza qualitativa e tem como método a análise documental. Concluiu-se que os livros didáticos analisados contemplaram os tópicos relacionados à Microbiologia de forma mais apropriada quando comparados às apostilas, entretanto, ambos os materiais são deficientes, e seu uso como única fonte de pesquisa ou como cronograma de aulas não é aconselhável

Revista iberoamericana de educación

Resumen basado en el de la publicaciónTítulo, resumen y palabras clave en portugués e inglésNúmero especial que acompaña al monográfico "Neurodidáctica en el aula: transformando la educación"Se busca evaluar en qué medida las propuestas didácticas de libros del área contribuyen, o no, a la superación de obstáculos en el proceso de enseñanza-aprendizaje de Microbiología, a través de un análisis comparativo de los temas relacionados a la Micro-biología en los materiales didácticos analizados, a saber: colección de libros didácticos y apostillas destinadas a los años finales de la Enseñanza Fundamental (Cuaderno del Alumno). La investigación es de naturaleza cualitativa y tiene como método el análisis documental. Se concluyó que los libros didácticos analizados contemplaron los tópicos relacionados con la Microbiología de forma más apropiada cuando comparados a las apostillas, sin embargo, ambos materiales son deficientes, y su uso como única fuente de investigación o como cronograma de clases no es aconsejable.ES

Screening design of nutritional and physicochemical parameters on bio-hydrogen and volatile fatty acids production from Citrus Peel Waste in batch reactors

International audienceDifferent nutritional and physicochemical conditions to obtain H2 from Citrus Peel Waste (CPW) were evaluated. For this, a screening design was carried out using Plackett and Burman design, in order to verify the main significant conditions of this process. The variables studied were pH (5.5–8.5), temperature (30–44 °C), autochthonous inoculum (0.75–2.25 gTVS.L−1), allochthonous inoculum (1–3 gTVS.L−1) and substrate (5–15 g.L−1), headspace (40–60%) and nutritional medium components, such as yeast extract (0–1 g.L−1), CaCO3, NaCl and peptone (0–5 g.L−1). The most significant operational variables were pH (8.5), allochthonous inoculum (3 gTVS.L−1) and substrate concentration (15 g CPW.L−1), conditions that favored the highest H2 (13.29 mmol.L−1) and acetic acid productions (1340 mg.L−1). Escherichia (34.5%) and Clostridium (29.93%) were the main genera identified under these conditions

Microbial and functional characterization of granulated sludge from full-scale UASB thermophilic reactor applied to sugarcane vinasse treatment

International audienceConsidering the scarcity of data in the literature regarding phylogenetic and metabolic composition of different inocula, especially those from thermophilic conditions, this research aimed at characterizing the microbial community and preferable metabolic pathways of an UASB reactor sludge applied to the thermophilic treatment (55°C) of sugarcane vinasse, by means of shotgun metagenomics. After its metabolic potential was depicted, it was possible to observe several genes encoding enzymes that are of great importance to anaerobic digestion processes with different wastes as substrate, especially regarding the biodegradation of carbohydrates and ligninolytic compounds, glycerolypids, volatile fatty acids and alcohols metabolism and biogas (H2 and CH4) production. The genera identified in higher relative abundances for Bacteria domain were Sulfirimonas (37.52 ± 1.8%), possibly related to the sludge endogenic activity due to its strong relation with a peptidoglycan lyase enzymes family, followed by Fluviicola (5.01 ± 1.0%), Defluviitoga (4.36 ± 0.2%), Coprothermobacter (4.32 ± 0.5%), Fervidobacterium (2.93 ± 0.3%), Marinospirillum (2.75 ± 0.2%), Pseudomonas (2.14 ± 0.2%) and Flavobacterium (1.78 ± 0.1%), mostly related with carbohydrates fermentations and/or H2 production. For Archaea domain, Methanosarcina (0.61 ± 0.1%), Methanothermobacter (0.38 ± 0.0%), Methanoculleus (0.30 ± 0.1%), Thermococcus (0.03 ± 0.0%), Methanolobus (0.02 ± 1.8%), Methanobacterium (0.013 ± 0.0%), Aciduliprofundum and Pyrococcus (0.01 ± 0.0%) were the most dominant ones, being Methanosarcina the most related with methanogenesis. It was concluded that the robust inoculum description performed in this study may subside future biotechnological researches by using similar inocula (UASB sludges), focusing on the obtainment of value-added by-products by means of anaerobic digestion, such as volatile fatty acids, alcohols and biogas (H2 and CH4), by using several types of waste as substrate

Microbial and functional characterization of an allochthonous consortium applied to hydrogen production from Citrus Peel Waste in batch reactor in optimized conditions

International audienceEnergy recovery from lignocellulosic waste has been studied as an alternative to the problem of inappropriate waste disposal. The present study aimed at characterizing the microbial community and the functional activity of reactors applied to H2 production through lignocellulosic waste fermentation in optimized conditions. The latter were identified by means of Rotational Central Composite Design (RCCD), applied to optimize allochthonous inoculum concentration (2.32–5.68 gTVS/L of granular anaerobic sludge), pH (4.32–7.68) and Citrus Peel Waste (CPW) concentration (1.55–28.45 g/L). After validation, the conditions identified for optimal H2 production were 4 gSTV/L of allochthonous inoculum, 29.8 g/L of CPW (substrate) and initial pH of 8.98. In these conditions, 48.47 mmol/L of H2 was obtained, which is 3.64 times higher than the concentration in unoptimized conditions (13.31 mmol H2/L using 15 g/L of CPW, 2 gTVS/L of allochthonous inoculum, pH 7.0). Acetogenesis was the predominant pathway, and maximal concentrations of 3,731 mg/L of butyric acid and 3,516 mg/L of acetic acid were observed. Regarding the metataxonomic profile, Clostridium genus was dramatically favored in the optimized condition (79.78%) when compared to the allochthonous inoculum (0.43%). It was possible to identify several genes related to H2 (i.e dehydrogenases) and volatile fatty acids (VFA) production and withcellulose degradation, especially some CAZymes from the classes Auxiliary Activities, Glycoside Hydrolases and Glycosyl Transferase. By means of differential gene expression it was observed that cellulose degradation and acetic acid production pathways were overabundant in samples from the optimized reactors, highlighting endoβ-1,4-glucanase/cellulose, endo-β-1,4-xylanase, β-glucosidase, β-mannosidase, cellulose β-1,4-cellobiosidase, cellobiohydrolase, and others, as main the functions

Microbial community dynamics during the co-digestion of manure and the organic fraction of household waste

International audienceIntroduction: Organic waste co-digestion is an attractive option for territories to valorize different types of locally produced waste and optimize biogas production in digestion plants. We studied the dry batch co-digestion of reconstituted racehorse manure (MA) and the organic fraction of residual household waste (HW), in pilot reactors. We identified optimal proportions of co-substrates and evaluated the effects of their initial proportions on microbial community dynamics.Methodology: Batch pilots for dry anaerobic digestion (60 L each) were filled with HW:MA in various proportions (w:w), ranging from 0:100 to 92:8, for a total of 14 incubations. The co-substrates were fully immersed in digested mixed urban wastewater sludge serving both as inoculum and liquid input. The pilots were operated at ~36°C during up to 5 weeks. The conversion dynamics was characterized by monitoring relevant physico-chemical parameters: biogas production and composition, pH, volatile fatty acids (VFA) and total alkalinity content. The composition of archaeal and bacterial communities was determined by 16S rDNA metabarcoding targeting the V4-V5 region of archaea and bacteria, using an Ion Torrent PGM sequencer. Principal component analysis was performed with R packages such as FactoMineR.Results: When co-digesting HW, a transient accumulation of VFA was observed. Maximal VFA concentrations were all the higher as the initial proportion of HW was high and reached up to ~17 g/L. Moreover, a partial inhibition of methane production occurred in the presence of HW, mostly during the 8-15 first days of incubation, especially for proportions 79:21 and 92:8. The total cumulated productions of methane were similar for HW proportions up to 79%, suggesting the adaptation of microbial communities to high VFA conditions. Consistently, clear changes in the microbial composition occurred according to the HW proportion. In the absence of HW, bacteria were dominated by classes Bacteroidia and Spirochaetia; for high HW proportions, Spirochaetia proportion decreased at the benefit of Clostridia and Bacilli. For archaea, Bathyarchaeia members were the most abundant in the absence of HW, while Methanobrevibacter and Methanosarcina genera were selected at the highest HW proportions. Moreover, the proportions of archaea were significantly lower as soon as HW was employed.Conclusions: The adaptation of microbial communities over time to high VFA concentrations suggests the possibility to employ a pre-adapted inoculum, to avoid the transient inhibition phase while digesting significant proportions of HW. Moreover, it opens the perspective of identifying inhibition biomarkers

Microbial community dynamics during the co-digestion of manure and the organic fraction of household waste

International audienceIntroduction: Organic waste co-digestion is an attractive option for territories to valorize different types of locally produced waste and optimize biogas production in digestion plants. We studied the dry batch co-digestion of reconstituted racehorse manure (MA) and the organic fraction of residual household waste (HW), in pilot reactors. We identified optimal proportions of co-substrates and evaluated the effects of their initial proportions on microbial community dynamics.Methodology: Batch pilots for dry anaerobic digestion (60 L each) were filled with HW:MA in various proportions (w:w), ranging from 0:100 to 92:8, for a total of 14 incubations. The co-substrates were fully immersed in digested mixed urban wastewater sludge serving both as inoculum and liquid input. The pilots were operated at ~36°C during up to 5 weeks. The conversion dynamics was characterized by monitoring relevant physico-chemical parameters: biogas production and composition, pH, volatile fatty acids (VFA) and total alkalinity content. The composition of archaeal and bacterial communities was determined by 16S rDNA metabarcoding targeting the V4-V5 region of archaea and bacteria, using an Ion Torrent PGM sequencer. Principal component analysis was performed with R packages such as FactoMineR.Results: When co-digesting HW, a transient accumulation of VFA was observed. Maximal VFA concentrations were all the higher as the initial proportion of HW was high and reached up to ~17 g/L. Moreover, a partial inhibition of methane production occurred in the presence of HW, mostly during the 8-15 first days of incubation, especially for proportions 79:21 and 92:8. The total cumulated productions of methane were similar for HW proportions up to 79%, suggesting the adaptation of microbial communities to high VFA conditions. Consistently, clear changes in the microbial composition occurred according to the HW proportion. In the absence of HW, bacteria were dominated by classes Bacteroidia and Spirochaetia; for high HW proportions, Spirochaetia proportion decreased at the benefit of Clostridia and Bacilli. For archaea, Bathyarchaeia members were the most abundant in the absence of HW, while Methanobrevibacter and Methanosarcina genera were selected at the highest HW proportions. Moreover, the proportions of archaea were significantly lower as soon as HW was employed.Conclusions: The adaptation of microbial communities over time to high VFA concentrations suggests the possibility to employ a pre-adapted inoculum, to avoid the transient inhibition phase while digesting significant proportions of HW. Moreover, it opens the perspective of identifying inhibition biomarkers

Co-digestion anaérobie de boues urbaines : effets du fumier et de la fraction organique des déchets ménagers employés comme cosubstrats, sur la composition des communautés microbienne

International audienceLa co-digestion anaérobie ou méthanisation des déchets organiques est une option intéressante pour valoriser différents types de déchets organiques produits localement car elle permet notamment d’envisager une optimisation de la production de biogaz. Nous avons étudié la co-digestion de boues urbaines associées à du fumier de cheval (F) et à la fraction organique des ordures ménagères (FOR). Nous nous sommes intéressés aux effets de différents mélanges de ces 3 co-substrats sur la production de biogaz et la dynamique des communautés microbiennes catalysant les bioconversions au sein de ce procédé. I. PROTOCOLE EXPERIMENTALDes pilotes de digestion anaérobie en voie sèche (60 L chacun) ont été alimentés avec des mélanges de F:FOR dans des proportions (masse:masse) variant de 0:100 à 92:8 (Figure 1). Les co-substrats ont été entièrement immergés dans des boues mixtes d'eaux usées urbaines digérées (digestat), servant à la fois d'inoculum et d'apport liquide. Les digestions ont été menées à ~36°C pendant une période pouvant aller jusqu'à 5 semaines. La production et la composition du biogaz, le pH, la concentration en acides gras volatils (AGV) et l'alcalinité totale ont été mesurées au cours du temps. La composition des communautés microbiennes (archées et bactéries) a été déterminée par métabarcoding de l'ADN codant pour l’ARN ribosomique 16S et les résultats ont été soumis à des analyses multivariées. II. RESULTATS EXPERIMENTAUXLors de la co-digestion en présence de déchets ménagers, une accumulation transitoire d’acides gras volatils a été observée et leur concentration totale pouvait atteindre ~17 g/L lorsque la proportion initiale de déchets ménagers était la plus élevée. Une inhibition partielle de la production de méthane a été observée en présence de déchets ménagers, principalement pendant les 8-15 premiers jours d'incubation, et surtout pour les proportions les plus élevées. En fin d’incubation, les productions totales cumulées de méthane étaient cependant similaires pour des proportions initiales de déchets ménagers allant jusqu'à 79%, soulignant l'adaptation des communautés microbiennes à des concentrations d'acides gras volatils élevées. De manière cohérente, des changements clairs de composition des communautés microbiennes se sont produits en fonction de la proportion de déchets ménagers. En l'absence de déchets ménagers, les bactéries étaient dominées par les classes Bacteroidia et Spirochaetia ; pour des proportions élevées de déchets ménagers, la proportion de Spirochaetia a diminué au profit de Clostridia et Bacilli. De plus, les proportions d'archées étaient significativement plus faibles dès lors que des déchets ménagers était ajoutés. Pour conclure, l'adaptation des communautés microbiennes au fil du temps à des concentrations élevées d'acides gras volatiles suggère la possibilité d'utiliser un inoculum pré-adapté, afin d'éviter la phase d'inhibition transitoire lors de la co-digestion avec des proportions importantes de déchets ménagers

Co-digestion anaérobie de boues urbaines : effets du fumier et de la fraction organique des déchets ménagers employés comme cosubstrats, sur la composition des communautés microbienne

International audienceLa co-digestion anaérobie ou méthanisation des déchets organiques est une option intéressante pour valoriser différents types de déchets organiques produits localement car elle permet notamment d’envisager une optimisation de la production de biogaz. Nous avons étudié la co-digestion de boues urbaines associées à du fumier de cheval (F) et à la fraction organique des ordures ménagères (FOR). Nous nous sommes intéressés aux effets de différents mélanges de ces 3 co-substrats sur la production de biogaz et la dynamique des communautés microbiennes catalysant les bioconversions au sein de ce procédé. I. PROTOCOLE EXPERIMENTALDes pilotes de digestion anaérobie en voie sèche (60 L chacun) ont été alimentés avec des mélanges de F:FOR dans des proportions (masse:masse) variant de 0:100 à 92:8 (Figure 1). Les co-substrats ont été entièrement immergés dans des boues mixtes d'eaux usées urbaines digérées (digestat), servant à la fois d'inoculum et d'apport liquide. Les digestions ont été menées à ~36°C pendant une période pouvant aller jusqu'à 5 semaines. La production et la composition du biogaz, le pH, la concentration en acides gras volatils (AGV) et l'alcalinité totale ont été mesurées au cours du temps. La composition des communautés microbiennes (archées et bactéries) a été déterminée par métabarcoding de l'ADN codant pour l’ARN ribosomique 16S et les résultats ont été soumis à des analyses multivariées. II. RESULTATS EXPERIMENTAUXLors de la co-digestion en présence de déchets ménagers, une accumulation transitoire d’acides gras volatils a été observée et leur concentration totale pouvait atteindre ~17 g/L lorsque la proportion initiale de déchets ménagers était la plus élevée. Une inhibition partielle de la production de méthane a été observée en présence de déchets ménagers, principalement pendant les 8-15 premiers jours d'incubation, et surtout pour les proportions les plus élevées. En fin d’incubation, les productions totales cumulées de méthane étaient cependant similaires pour des proportions initiales de déchets ménagers allant jusqu'à 79%, soulignant l'adaptation des communautés microbiennes à des concentrations d'acides gras volatils élevées. De manière cohérente, des changements clairs de composition des communautés microbiennes se sont produits en fonction de la proportion de déchets ménagers. En l'absence de déchets ménagers, les bactéries étaient dominées par les classes Bacteroidia et Spirochaetia ; pour des proportions élevées de déchets ménagers, la proportion de Spirochaetia a diminué au profit de Clostridia et Bacilli. De plus, les proportions d'archées étaient significativement plus faibles dès lors que des déchets ménagers était ajoutés. Pour conclure, l'adaptation des communautés microbiennes au fil du temps à des concentrations élevées d'acides gras volatiles suggère la possibilité d'utiliser un inoculum pré-adapté, afin d'éviter la phase d'inhibition transitoire lors de la co-digestion avec des proportions importantes de déchets ménagers