Orientador: Jorge de Lucas Junior / Resumo: Foram avaliadas as características quantitativas, qualitativas e o potencial e distribuição da produção de biogás, da cama de frangos de corte.Os estudos foram realizados em galpões experimentais, laboratórios e biodigestores. O experimento foi analisado como inteiramente casualisado, adotando-se esquema fatorial 4x3x2 considerando quatro quantidades de cama (0,4, 0,5, 0,6 e 0,7 kg MS.ave-1), três materiais de cobertura dos galpões AMC (telha de fibrocimento), AMB ( fibrocimento com pintura) e ALU (alumínio ondulada) e dois lotes de criação de aves em camas com uma utilização e reutilização em Dourados - MS. O ambiente dos galpões, proporcionado pelos diferentes tipos de cobertura foi caracterizado a partir de índices de temperatura do bulbo seco, umidade relativa e velocidade do vento. Para avaliar o desempenho animal foram observados: o peso vivo, consumo de ração e mortalidade e para a quantificação da cama, foram estudados peso da cama produzida, teor de matéria seca, coeficiente de resíduo, umidade, proteína bruta, potencial de emissão de amônia, temperatura da superfície e do interior da cama de frango e produção de biogás...(Resumo completo, clicar acesso eletrônico abaixo) / Abstract:Quantitative, qualitative characteristics and potential and biogas distribution from poultry litter were evaluated.. The experiment was analyzed as a completely randomized, using a 4x3x2 factorial scheme established by four quantities of litter (0.4; 0.5; 0.6 and 0.7 kgMS.bird-1), three covering materials of aviaries: AMC (asbestos cement tile), AMB (asbestos cement tile with painting) and ALU (wavy aluminum tile) and two lots of breeding in Dourados - MS. Aviary environment, which was promoted by different kinds of covering, was characterized descriptively considering dried bulb temperature index, humidity index and wind velocity. In order to evaluate broiler performance, it was observed: live weight, ration consumption, mortality and for poultry litter quantification: produced litter weight, dry matter content, residue coefficient, dampness, crude protein, ammonia emission potential, surface, and inside temperature of poultry litter and biogas production were studied...(Complete abstract, click electronic access below) / Doutor

A vinhaça, água residuária do processo, apresenta em sua composição DQO de 20 a 100 g DQO.L-1 e um ótimo potencial de produção de biogás por meio da digestão anaeróbia. Para avaliar a capacidade de produção de biogás, foram realizados experimentos sob o efeito gradual da carga orgânica volumétrica (COV) e da relação A/M, avaliando a etapa metanogênica. Como no processo de fabricação do etanol é adicionado ácido sulfúrico para evitar a contaminação bacteriana, foi avaliado a interferência da adição de diferentes concentrações de sulfetos no processo em valore de pH 7,0 e 7,5. O fermentador foi alimentado com Na2S.9H2O, variando a concentração do íon S2- de 0 a 1000 mg.L-1. Para obter uma boa eficiência na produção de metano, a etapa acidogênica também foi avaliada. Para isso, se manteve as mesmas concentrações de biomassa e substrato, porém, ajustando os valores de pH em 5,0, 5,5, 6,0 e 6,5. Na etapa metanogênica uma satisfatória redução da elevada carga orgânica presente na vinhaça foi alcançada, com eficiência de remoção entre 82,0% e 90,3% em processo mesofílico. Um ajuste polinomial foi realizado para avaliar a produção específica de metano, que variou de 379 mL CH4.h-1 a 872 mL CH4.h-1 e atividade metanogênica de 0,33 mmol CH4.gSVT-1 h-1 a 0,77 mmol CH4.gSVT-1 h-1. Para a etapa da sulfetogênese, o pH afetou consideravelmente o desempenho das arqueas metanogênicas, ocorrendo inibição mais acentuada para o pH 7,5. Na concentração mais baixa testada, de 50 mg S2-.L-1, foi observada inibição de 31,85% para pH 7,0 e de 67% para pH 7,5 e para a concentração mais elevada de 1000 mg S2-.L-1 a inibição foi de 59,75% e de 94,07% respectivamente. Na última etapa da acidogênese, maiores concentrações de ácido propiônico e acético foram alcançadas em pH 5,0 e 5,5, com 1374,66 e 1477,23 mg C3H6O2.L-1 e 993,05 e 767,80 mg CH3COOH.L-1. Maiores taxas de produção de AGV ocorreram em pH 6,5, alcançando para o ácido propiônico 8,82 mmol.d-1 gSVT-1 e 7,99 mmol.d-1 gSVT-1 para ácido acético. A produção acumulada de metano nas primeiras 60 horas atingiu 2210 mL, 5300 mL, 7210 mL e 7620 mL CH4, respectivamente para pH 5,0, 5,5, 6,0 e / Centro de Pesquisa e Desenvolvimento Leopoldo Américo Miguêz de Mello, CENPES. / The vinasse, wastewater of the process, presents in its composition a COD of 20 to 100 g COD.L-1 and an excellent biogas production potential through anaerobic digestion. To evaluate the biogas production capacity, experiments were carried out under the gradual effect of organic volumetric load (OVL) and A/M ratio, evaluating the methanogenic step. Considering that sulfuric acid is added to the ethanol production process to prevent bacterial contamination, the interference of the addition of different sulfide concentrations in the process at pH 7.0 and 7.5 was evaluated. The fermenter was fed with Na2S.9H2O, varying the concentration of the S2- ion from 0 to 1000 mg.L-1. In order to obtain good efficiency in the production of methane, the acidogenic step was also evaluated. For this, the same concentrations of biomass and substrate were maintained, however, the pH to was adjusted 5.0, 5.5, 6.0 and 6.5. In the methanogenic stage a satisfactory reduction of the organic load present in the vinasse was achieved, with removal efficiency in the range of 82.0% and 90.3% for mesophilic process. A polynomial fit was performed to evaluate the specific production of methane, ranging from 379 mL CH4.h-1 to 872 mL CH4.h-1 and a methanogenic activity of 0.33 mmol CH4.gSVT-1 h-1 at 0.77 mmol CH4.g SVT-1 h-1. For the sulfetogenic stage, the pH considerably affected the performance of the methanogenic archaea, with a more pronounced inhibition at pH 7.5. The lowest concentration tested, 50 mg S2-.L-1, resulted in an inhibition of 31.85% inhibition pH 7.0 and 67% at pH 7.5. The highest concentration, 1000 mg S2-.L-1, the inhibition was 59.75% and 94.07% respectively. Higher concentrations of propionic and acetic acid were reached at pH 5.0 and 5.5, with 1374.66 and 1477.23 mg C3H6O2.L-1 and 993.05 and 767.80 mg CH3COOH.L-1. Higher rates of VFA production occurred at pH 6.5, yielding 8.82 mmol.d-1 gSVT-1 and 7.99 mmol.d-1 gSVT-1 for acetic acid for propionic acid. Cumulative methane production in the first 60 hours increased 2210 mL, 5300 mL, 7210 mL and 7620 mL CH4, respectively to pH 5.0, 5.5, 6.0 and 6.5.

Integrated algae pond systems (IAPS) combine the use of anaerobic and aerobic bioprocesses to effect wastewater treatment. Although, IAPS as a technology process offers many advantages including efficient and simultaneous N and P removal, no requirement for additional chemicals, O2 generation, CO2 mitigation, and a biomass with potential for valorization, a lack of technological advancement and the need for large land area, has limited the reach of this technology at industrial scale. In mitigation, peroxonation was introduced as a tertiary treatment unit and its effect on COD and TSS of IAPS treated water investigated. An effort was made to characterize the soluble but persistent COD in IAPS treated water and, productivity of the HRAOP mixed liquor was investigated to gain insight into the potential use of this biomass. Results show that peroxone treatment effectively reduced COD, TSS, and nutrient load of IAPS water without any significant impact on land area requirement. Indeed, summary data describing the effect of peroxone on quality of IAPS-treated water confirmed that it complies with the general limit values for either irrigation or discharge into a water resource that is not a listed water resource for volumes up to 2 ML of treated wastewater on any given day. Extraction followed by FT-IR spectroscopy was used to confirm albeit tentatively, the identity of the soluble but persistent COD in IAPS treated water as MaB-floc EPS. Results show that MaB-flocs from HRAOPs are assemblages of microorganisms produced as discrete aggregates as a result of microbial EPS production. A relationship between photosynthesis and EPS production was established by quantification of the EPS following exposure of MaB-flocs to either continuous light or darkness. Several novel strains of bacteria were isolated from HRAOP mixed liquor and 16S ribosomal genomic sequence analysis resulted in the molecular characterization of Planococcus maitriensis strain ECCN 45b. This is the first report of Planococcus maitriensis from a wastewater treatment process. Productivity and change in MaB-flocs concentration, measured as mixed liquor suspended solids (MLSS) between morning and evening were monitored and revealed that MLSS is composed of microalgae and bacteria but not fungi. Concentration varied from 77 mg L-1 in September (winter) to 285 mg L-1 in November (spring); pond productivity increased from 5.8 g m-2 d-1 (winter) to 21.5 g m-2 d-1 (spring); and, irrespective of MLSS concentration in late afternoon, approximately 39% was lost overnight, which presumably occurred due to passive removal by the algae settling pond. The outcomes of this research are discussed in terms of the quality of treated water, and the further development of IAPS as a platform technology for establishing a biorefinery within the wastewater treatment sector.

The main purpose of this project was an attempt to modulate and simulate two existing biogas plant, situated in Lidköping and Katrineholm, Sweden and evaluate how the process reacts to certain conditions regarding feeding, layout and substrate mixture. The main goal was to optimize the existing processes to better performance. Both the modeling and simulation were executed in SIMBA#Biogas with accordance to the real conditions at the plant in question. The simulation of each model was validated against data containing measurements of, CH4 yield, CH4 production, TS, VS, NH4-N concentration and N-total concentration. The data was obtained from each plant in accordance with scheduled follow ups. Both models were statistically validated for several predictions. Predictions of N-total and NH4-N concentration failed for some cases. Both plants were tested with new process lay outs, where promising results were obtained. The Lidköping model was provided with a post-hygienization step to handle ABPs. The Katrineholm model was provided with a dewatering unit, where 35% of the centrate was recirculated back to the system. Both setups was configured to yield the highest CH4 production. This study suggests that SIMBA#Biogas can be a tool for predictions and optimizations of the biogas process.

Conventional activated sludge-based wastewater treatment is an energy and resource-intensive process. Historically it has been successful at producing safely treated wastewater effluents in the developed world, specifically in places that have the infrastructure and space to support its operation. However, with a growing need for safe and efficient wastewater treatment across the world in both urban and rural settings, a paradigm shift in waste treatment is proving to be necessary. The sustainability of the future of wastewater treatment, in a significant way, hinges on moving towards energy neutrality and wastewater effluent reuse. This potential for reuse is threatened by the recent emergence and study of contaminants that have not been previously taken into consideration, such as antibiotics and other organic micropollutants (OMPs), antibiotic resistance genes, and persistent pathogenic bacteria. This dissertation focuses on investigating the use of anaerobic membrane bioreactor (AnMBR) technology for the sustainable treatment of municipal-type wastewaters. Specifically, a microbial approach to understanding biofouling and methane recovery potential in anaerobic MBR systems has been employed to assess different reactor systems’ efficiency. This dissertation further compares AnMBRs to their more widely used aerobic counterparts. This comparison specifically focuses on the removal and biodegradation of OMPs and antibiotics in both anaerobic and aerobic MBRs, while also investigating their effect on the proliferation of antibiotic resistance genes. Due to rising interest in wastewater effluent reuse and the lack of a comprehensive understanding of MBR systems’ effects on pathogen proliferation, this dissertation also investigates the presence of pathogens in both aerobic and anaerobic MBR effluents by using molecularbased detection methods. The findings of this dissertation demonstrate that membrane-associated anaerobic digestion processes have significant potential to improve the sustainability of wastewater treatment. This is exemplified by attributes of AnMBR systems associated with both increased system efficiency and wastewater reuse potential through methane recovery and lower abundance of effluent microbial contaminants, respectively. Overall, the studies collated in this dissertation have shown that understanding the microbial communities of AnMBRs can play a central role in further improving these attributes and in reducing the risks posed by emerging contaminants and pathogens in wastewater treatment systems.

Bacteroides fragilis strain Bf-1 produces an extracellular bacteriocin at the beginning of the stationary growth phase. Production is not inducible by either ultraviolet light or mitomycin C. The low molecular weight bacteriocin (MW estimates of 13 500 and 18 800 obtained from Sephadex G-100 chromatography and SDS-PAGE electrophoresis respecively) is stable between pH 7 - 9 and is inactivated on incubation with trypsin and pronase. An unusual feature of the Bf-1 bacteriocin is its apparent biphasic temperature stability: while the majority of the activity (97%) is destroyed by heating at 60ºC (t [subscript] 1/2 = 2.5 min at 60ºC), a small proportion (3%) is stable even after autoclaving at 121ºC for 15 min. The killing of sensitive cells occurs in 2 stages and the killing action is reversed by incubation with trypsin. The transition from stage I to stage II is dependent on the temperature of incubation and the growth state of sensitive cells. 2,4-Dinitrophenol prevents this transition. The Bf-1 bacteriocin has an unusual mode of action. It specifically inhibits RNA synthesis whilst having no effect on protein or DNA synthesis. No effect on intracellular ATP levels were observed. The heat-stable (3%) fraction had a similar biochemical effect. In vitro studies involving RNA polymerase indicated that the bacteriocin and the antibiotic rifampicin have similar effects on RNA synthesis. The bacteriocinogenic strain (Bf-1) is insensitive to its own bacteriocin both in vivo and in vitro, although this immunity is overcome in vitro by the addition of higher concentrations of the Bf-1 bacteriocin. The bacteriocinogenic strain (Bf-1) harbors a cryptic plasmid (or plasmids) which on a neutral sucrose gradient, sediments faster than the Col E1 marker plasmid DNA. Attempts to cure this strain of its bacteriocinogenic phenotype were unsuccessful.

Bacteriophytochromes (Bphs) comprise a family of protein photoreceptors that help bacteria sense changes in light. Bphs contain a chromophore that, upon absorption of red or far-red light, undergoes a cis-trans isomerization that leads to a conformational change in the holoprotein (photoconversion). In the active conformation, Bphs act as a kinase and regulate gene expression through phosphorylation of target proteins. Two putative Bph orfs (rpa0122 and rpa0990) in the Rhodopseudomonas palustris genome encode Bph-like proteins that have a conserved chromophore-binding cysteine residue. The hypothesis is that one or both of these unique Bph-like genes encode proteins that are capable of binding a chromophore and functioning to modulate the cell’s phenotype. I expressed and purified His-tagged RPA0990 in R. palustris, because proteolytic degradation was observed during overexpression in an E coli. expression system. The results show that RPA0990 contains a chromophore and is capable of photoconversion. The wavelengths of light absorbed by the Pr/Pfr forms of RPA0990, predicted to be active and inactive forms respectively, were determined to be 695 nm and 755 nm. Investigation into the phenotype of the bph mutants rpa0122 and rpa0990 revealed that both of these Bphs may have a small effect on light-harvesting complexes. Also, it was observed that the absence of O₂ does not inhibit the normal function of Bphs, although O₂ was thought to be needed to make a linear tetrapyrrole cofactor, by cleaving heme using heme oxygenase. I suggest that a linear tetrapyrrole can be made anaerobically, either through anaerobic heme cleavage by a novel enzyme, or directly from the heme precursor hydroxymethylbilane without ring cleavage. The activity of a divergent promoter region between the rpa1490 (bph3) and rpa1491 (pucBe) genes was evaluated by using the E. coli lacZ gene as a reporter. The results indicated that the pucBe promoter has much higher activity than the bph3 promoter. It was also found that double knockout of the regulatory genes ppsR1ˉ2ˉ led to an increase in bph3::lacZ expression and a decrease in pucBe::lacZ expression. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

This study evaluates the enhancement of anaerobic digestion (AD) of the organic fraction of municipal solid waste (OFMSW) by microwave pretreatment (MW) at high temperatures (115, 145 and 175°C). The highest level of solubilization was achieved at 175ºC, with a supplemental water addition of 30% (SWA30). Pretreatments combining two modalities; MW heating in presence or absence of hydrogen peroxide (H2O2) was also investigated. Biochemical methane potential (BMP) tests were conducted on the whole OFMSW, as well as on the liquid fractions. The whole OFMSW pretreated at 115 and 145 ºC showed little improvement in biogas production over control. When pretreated at 175 ºC, biogas production decreased due to formation of refractory compounds, inhibiting digestion. For the liquid fraction of OFMSW, the effect of pretreatment on the cumulative biogas production (CBP) was more pronounced for supplemental water addition of 20% (SWA20) at 145 ºC. Combining MW and H2O2 modalities did not have a positive impact on OFMSW stabilization and enhanced biogas production. Based on the BMP assay results, the effects of MW pretreatment (145 ºC) on the AD of OFMSW (SWA20) were further evaluated in single and dual stage semi-continuous digesters at hydraulic retention times (HRTs) of 20, 15, 12 and 9 days. Overall, MW pretreatment did not enhance the AD of the whole waste at the HRTs tested. However, the use of a dual stage reactor digesting non pretreated whole OFMSW had the best performance with the shortest HRT of 9 days. Conversely, for free liquid after pretreatment in two stage reactors at 20 day HRT methane production was tripled. In general, the performance of the dual stage digesters surpassed that of the single stage reactors. Cyclic BMP assays indicated that using an appropriate fraction of recycled effluent leachate can be implemented without negatively effecting methanogenic activity and biogas production. Based on the results obtained in this study, digestion of OFMSW by dual stage reactors without pretreatment appears to provide the best potential for waste stabilization in terms of biogas production and yield, process stability and volumetric loading rates.

Anaerobic digestion of waste activated sludge can be improved if hydrolysis of particulate substrates is enhanced and available substrate is made more accessible by both breakup of the sludge matrix floc and rupture of the cell wall. Microwave (MW) pretreatment was suggested and studied as a way to improve digestion efficiency. The work done focuses on the effects of MW pretreatment on the characteristics of the sludge, due to thermal and athermal effects. It also evaluates the effects some process variables in the activated sludge process have on the pretreatment efficiency as well as the effect operating conditions in the downstream anaerobic digestion process have on the biodegradability efficiency of those sludges. Effects of athermal and thermal MW radiation were measured by use of a customized MW oven capable of providing MW radiation with uncoupled thermal and athermal effects. Athermal radiation was capable of increasing substrate present in the soluble phase of sludge, and had a positive effect in the digestion of athermal samples. The increases in biogas production and substrate solubilisation were smaller in magnitude than the increases measured for MW thermal tests. Further refining of the tests with athermal and thermal sludge, involved separation by size class of the solubilized substrate by means of ultrafiltration (UF), and revealed that changes in particle size distribution were significant not only for MW thermal tests, but also for athermal tests, with a particular emphasis in proteins in athermal tests. These changes had an effect on the biodegradability of the sludges by class size, with thermally pretreated sludge producing more biogas for smaller particles size classes but also exhibiting more inhibition. Tests were made with several combinations of sludge with different ages and subject to different MW pretreatment temperatures. The work showed that sludge age or solids retention time (SRT) has a significant effect on the pretreatment efficiency with maximum biogas improvements measured at different MW pretreatment temperatures depending on the SRT of the sludge tested, and with different behaviour for mesophilic and thermophilic digestion. Mesophilic tests showed greater improvements in terms of digestion effiency on average, but thermophilic tests showed more uniform performance, with a higher baseline efficiency. The presence of an optimum of MW pretreatment temperature and sludge SRT for maximal biogas production is more defined for mesophilic conditions than for thermophilic conditions. Semi-continuous studies were conducted with several combinations of single and two stage mesophilic and thermophilic digestors treating MW pretreated sludge and non-pretreated sludge. Staging and thermophilic digestion allowed the maintenance of a stable digestion process with high biogas productions and high solids removal efficiencies with production of sludge with good bacteriological characteristics for an very low residence time (5 d).

Esta tesis doctoral se centra en la dinámica de la inhibición y la recuperación del proceso de digestión anaerobia de purines de cerdo para encontrar indicadores, predecir fallos del proceso, minimizar las pérdidas de metano (CH4) y evaluar las mejores prácticas de gestión a nivel de la planta de biogás. Para cumplir con este objetivo, cinco ensayos fueron diseñados y ejecutados. En primer lugar, se diseñó un experimento para controlar los cambios físico-químicos y de emisión de gas de dos tipos de purines envejecidos durante 15 semanas de almacenamiento en condiciones de verano. En segundo lugar, diferentes concentraciones de sulfato (SO42-) fueron evaluadas en digestión anaerobia termofílica de purines de cerdo y vacuno controlando la producción de CH4 y los cambios fisicoquímicos en un ensayo en discontinuo. Así mismo, la degradación anaeróbica de la materia orgánica (MO) y el límite de inhibición de SO42- fueron investigados. En tercer lugar, los efectos de incluir purines de cerdo acidificados con SO42- en un codigestión anaerobia con purines de cerdo convencionales se estudió a dos escalas (a escala laboratorio y a gran escala), donde se evaluó el rendimiento del proceso. Así mismo, los indicadores clave del proceso fueron identificados. En cuarto lugar, una combinación de dos métodos, reacción en cadena de la polimerasa cuantitativa en tiempo real (qPCR) y microscopía electrónica de barrido cualitativa (SEM) fueron utilizados para evaluar los cambios en la población microbiana de los digestores anaerobios durante la adición de los purines. Finalmente, el rendimiento de CH4, la composición físico-química y la estructura y dinámica de la comunidad microbiológica fueron evaluadas durante la puesta en marcha de la digestión anaerobia de purines de cerdo a escala de laboratorio. Se evaluaron cuatro estrategias de puesta en marcha: inanición y no inanición, seguida de una adición gradual o brusca de los purines. Los resultados presentados en esta tesis doctoral per / Moset Hernández, V. (2012). Monitoring anaerobic digestion of animal slurry during inhibition and recovery phases [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/18069 / Palancia