The effect of viral plasticity on the persistence of host-virus systems

Phenotypic plasticity plays an important role in the survival of individuals. In microbial host-virus systems, previous studies have shown the stabilizing effect that host plasticity has on the coexistence of the system. By contrast, it remains uncertain how the dependence of the virus on the metabolism of the host (i.e. “viral plasticity”) shapes bacteria-phage population dynamics in general, or the stability of the system in particular. Moreover, bacteria-phage models that do not consider viral plasticity are now recognised as overly simplistic. For these reasons, here we focus on the effect of viral plasticity on the stability of the system under different environmental conditions. We compared the predictions from a standard bacteria-phage model, which neglects plasticity, with those of a modification that includes viral plasticity. We investigated under which conditions viral plasticity promotes coexistence, with or without oscillatory dynamics. Our analysis shows that including viral plasticity reveals coexistence in regions of the parameter space where models without plasticity predict a collapse of the system. We also show that viral plasticity tends to reduce population oscillations, although this stabilizing effect is not consistently observed across environmental conditions: plasticity may instead reinforce dynamic feedbacks between the host, the virus, and the environment, which leads to wider oscillations. Our results contribute to a deeper understanding of the dynamic control of bacteriophage on host populations observed in nature

Production of fungal lipids : kinetic modeling and process design

Finding alternatives for fossil fuels is currently urgent. One of the new processes in this field is the production of biodiesel from lipids accumulated by microorganisms. Some yeasts and fungi accumulate lipids when a component needed for growth, usually the N-source, is limiting while the C-source is in excess. These oleaginous yeasts and fungi were previously mainly used for unsaturated fatty acid production, but now also come into view for production of lipids as a source of biodiesel. This thesis takes the first steps in the development of a new process to produce lipids with an oleaginous fungus in solid-state fermentation on agro-industrial waste. Solid-state fermentation is the cultivation on solid substrate particles without (free) flowing water, and has several advantages over submerged fermentation such as less waste water production, less energy use for oxygen transfer and lower production costs. In this thesis, we focused on growth and lipid production kinetics in submerged as well as solid-state fermentation. The models developed for these systems provide insight in the lipid production mechanism, needed to develop the new process based on solid-state fermentation. The thesis starts with the selection of a model strain (Chapter 2). With this strain, the kinetics of growth and lipid accumulation were studied and modeled. We started with a steady-state model (Chapter 3 and 4) in submerged chemostat culture, and extended this to a dynamic model for submerged batch culture (Chapter 5). As the next step towards solid-state fermentation, we developed a model for growth and lipid accumulation on κ-carrageenan plates with monomers (Chapter 6). These three models were finally used to calculate potential lipid yield and energy use in a biodiesel production system (Chapter 7). For the system we want to develop, we need a fungus that can utilize different substrates and can produce lipids. For this purpose, we tested two oleaginous fungi: Mortierella alpina and Umbelopsis isabellina, which is described in Chapter 2. We cultivated both fungi on agar plates containing glucose, xylose, starch, cellulose or pectin, and on sugar beet pulp in a packed bed. M. alpina did not utilize xylose, cellulose and pectin, utilized starch much slower than glucose and only consumed approximately 40% of the sugar beet pulp in 20 days. This shows that M. alpina is not a suitable organism for our production system. U. isabellina utilized pectin and xylose with the same rate as glucose, but used starch slower and (crystalline) cellulose not at all. It consumed approximately 75% of the sugar beet pulp after 8 days and approximately 100% after 20 days. Also, it accumulated some lipids (3% of remaining dry mass) in the culture on sugar beet pulp; optimization of this process by addition of enzymes increased the lipid content to 9% of remaining dry mass. This shows that U. isabellina is a promising strain for lipid production from agro-industrial waste, and is therefore a good strain to use in our research. The lipid concentrations found in SSF culture were quite low; we therefore decided to look in more depth into the kinetics of lipid production in different model systems. The first model system was a submerged chemostat culture, because the substrate supply rates can be varied in this system by varying the dilution rate as well as the concentrations in the feed. Chapter 3 describes the development of a mathematical model that includes growth, lipid accumulation and substrate consumption of oleaginous fungi in submerged chemostat cultures. Key points of the model are: (1) If the C-source supply rate is limited, maintenance has a higher priority than growth, which has a higher priority than lipid production; (2) the maximum specific lipid production rate of the fungus is independent of the actual specific growth rate. This model was validated with chemostat cultures of U. isabellina grown on mineral media with glucose and NH4+. Because of practical problems at low dilution rates, the model could only be validated for D>0.04 h‑1. For further validation, published data sets for chemostat cultures of oleaginous yeasts and a published data set for a poly-hydroxyalkanoate accumulating bacterial species were used, which is described in Chapter 4. All data sets could be described well by the model. Analysis of all data showed that the maximum specific lipid production rate is in most cases very close to the specific production rate of membrane and other functional lipids for cells growing at their maximum specific growth rate. The limiting factor suggested by Ykema et al.(1986, Antonie van Leeuwenhoek 52: 491-506), i.e. the maximum glucose uptake rate, did not give good predictions of the maximum lipid production rate. The model shows that both the C/N-ratio of the feed as well as the dilution rate has a large influence on the lipid production rate. When these data are translated to SSF, it means that a low substrate supply rate can prevent lipid production, even when the C/N-ratio of the substrate is high. The next step towards understanding lipid accumulation was a model that also describes changes in time. Therefore, we developed a model for growth, lipid production and lipid turnover in submerged batch fermentation, which is shown in Chapter 5. This model describes three subsequent phases: exponential growth when both a C-source and an N-source are available, carbohydrate and lipid production when the N-source is exhausted, and turnover of accumulated lipids when the C-source is exhausted. The model was validated with submerged batch cultures of U. isabellina with two different initial C/N-ratios. In batch culture, the specific lipid production rate was almost four times higher than in chemostat cultures and it decreased exponentially in time. This indicates that different mechanisms for lipid production are active in batch and chemostat cultures. The model could also describe several data sets from literature very well. Furthermore, the model shows that local limitation of C-source in SSF can cause lipid turnover before the average C-source concentration in the substrate is zero. The next step towards an SSF system is the inclusion of diffusion in the batch model. We did this by developing a model that describes growth, lipid production and lipid turnover in a culture on κ-carrageenan plates containing the monomers glucose and alanine as C-source and N-source, respectively. This is described in Chapter 6. The model includes reaction kinetics and diffusion of glucose, alanine and oxygen. It was validated with U. isabellina and describes the different phases of the culture very well: exponential growth, linear growth because of oxygen limitation, accumulation of lipids and carbohydrates after local N-depletion and turnover of lipids after local C-depletion. Extending the model with an unidentified extracellular product improved the fit of the model to the data. The model shows that oxygen limitation is extremely important in solid-state cultures using monomers. Together with the low specific lipid production rate found in SSF, it explains the difference in production rate with submerged cultures. In Chapter 7, we used the models from Chapter 3, 5 and 6 together with basic engineering principles to calculate lipid yield and energy use in the modeled systems. We evaluated a process including pretreatment, cultivation and down-stream processing with sugar beet pulp and wheat straw as substrate, described different reactor types, and considered both a yeast and a fungus as microorganisms. According to the models, lipid yields on substrate were between 5% w/w and 19% w/w, depending on the culture system. With the same models, improvement of the yield to 25-30% w/w was shown to be possible, for example by genetic modification of the microorganism. The net energy ratio of the non-optimized systems varied between 0.8 and 2.5 MJ produced per MJ used; energy use for pretreatment and for oxygen transfer were most important. For the optimized systems, the net energy ratio increased to 2.9 – 5.5 MJ produced per MJ used, which can compete very well with other biofuels such as bioethanol or algal biodiesel. So although there is still quite some work to be done, microbial lipids have the potential to be tomorrow’s source of biodiesel. </p

Towards a metagenomic understanding on enhanced biomethane production from waste activated sludge after pH 10 pretreatment

BACKGROUND: Understanding the effects of pretreatment on anaerobic digestion of sludge waste from wastewater treatment plants is becoming increasingly important, as impetus moves towards the utilization of sludge for renewable energy production. Although the field of sludge pretreatment has progressed significantly over the past decade, critical questions concerning the underlying microbial interactions remain unanswered. In this study, a metagenomic approach was adopted to investigate the microbial composition and gene content contributing to enhanced biogas production from sludge subjected to a novel pretreatment method (maintaining pH at 10 for 8 days) compared to other documented methods (ultrasonic, thermal and thermal-alkaline). RESULTS: Our results showed that pretreated sludge attained a maximum methane yield approximately 4-fold higher than that of the blank un-pretreated sludge set-up at day 17. Both the microbial and metabolic consortium shifted extensively towards enhanced biodegradation subsequent to pretreatment, providing insight for the enhanced methane yield. The prevalence of Methanosaeta thermophila and Methanothermobacter thermautotrophicus, together with the functional affiliation of enzymes-encoding genes suggested an acetoclastic and hydrogenotrophic methanogenesis pathway. Additionally, an alternative enzymology in Methanosaeta was observed. CONCLUSIONS: This study is the first to provide a microbiological understanding of improved biogas production subsequent to a novel waste sludge pretreatment method. The knowledge garnered will assist the design of more efficient pretreatment methods for biogas production in the future.published_or_final_versio

A kinetic study on anaerobic sulphate reduction : effects of sulphate and temperature

Includes bibliography.The objectives of this work were to provide rigorous kinetic information on the effects of feed sulphate concentration and temperature on the anaerobic sulphate reduction process and to develop a kinetic model to explain this dependency. These objectives were addressed by performing batch and continuous sulphate reduction experiments using a mixed sulphate reducing microbial culture with acetate as the organic carbon and electron donor source. Sulphate concentration, acetate concentration and biomass concentration was used to determine the metabolic activity of the microorganisms and the rate of sulphate conversion

Understanding the Human Gut Microbiota: A Mathematical Approach

In order to explore the dynamics of the human gut microbiota, we used a system of ordinary differential equations to mathematically model the biomass of three microorganism populations: Bacteroides thetaiotaomicron, Eubacterium rectale, and Methanobrevibacter smithii. Additionally, we modeled the concentrations of relevant nutrients necessary to sustain these populations over time. This system highlights the interactions and the competition among these species in order to further understand their dynamics. These three microorganisms were specifically chosen due to the system’s end product, butyrate, which aids in developing the intestinal barrier in the human gut. The basis of the mathematical model assumes the gut acts as a chemostat, with bacteria and nutrients exiting the gut at a rate proportional to the volume of the chemostat, the rate of volumetric flow, and the biomass or concentration of the particular population or nutrient. We performed global sensitivity analysis using Sobol’ sensitivities in order to estimate the importance of model parameters and to understand our results

Biooxidation kinetics of Leptospirillum Ferriphilum attached to a defined solid substrate

Includes abstract.Includes bibliographical references.Bioleaching can be categorized as being either stirred tank type (i.e. bio-oxidation) or irrigation type (i.e. heap/dump bioleaching) yet studies investigating the kinetics of bioleaching systems mostly use empirical data determined from stirred tank type and initial rate experiments in batch cultures or using iso-potential devices. Rate equations deduced from such empirical data is then used to model both the stirred tank type and irrigation type bioleaching systems overlooking the possibility that there may be significant differences in their environments and therefore the kinetics. Tank bioleaching systems are well mixed suspension systems dominated by planktonic microorganisms (freely suspended in the liquid medium). Heap bioleaching systems on the contrary, are heterogeneous in nature with chemical and physical conditions changing over time and are dominated by sessile microorganisms (attached microorganisms to the surface of a solid). The heap bioleaching system is therefore highly complex compared to the stirred tank-type systems. Microbial growth in bioleaching systems significantly influence the overall bioleaching kinetics yet biological kinetic effects in sessile/ attached environments are not well understood. Heap and dump leaching account for about 20% of the world’s copper production and are becoming popular methods of copper production from leaching low grade ores. It is therefore important that the kinetics of irrigation type bioleaching systems are well understood. A strategy to determine the microbial kinetics of a sessile microbial population is enforced in this study. From this, empirical data determined from irrigation type environments can then be used to derive equations which can be used to accurately model heap bioleaching systems. Three sets of experiments were conducted to try and achieve this: i. planktonic experiments - investigating the microbial kinetics of a planktonic microbial population ii. attachment experiments - investigating the nature of growth of the microbial population to the surface of a solid substrate during attachment to create a sessile microbial population iii. sessile experiments - investigating the microbial kinetics of the sessile microbial population A pure culture of Leptospirillum ferriphilum (a mesophilic, ferrous iron oxidizing bioleachingmicroorganism) was used in this study. Planktonic experiments were conducted in a completely mixed, well aerated continuous stirred tank reactor (CSTR) with a 1 litre working volume, operating at a pH of about 1.3 and temperature of 37oC. Attachment and sessile experiments were conducted using a CSTR with similar conditions to the planktonic experimental, however the system was modified by introducing a packed bed vessel (PBR) attached as a closed loop to the CSTR. Solution drawn from the CSTR was then continuously pumped through the PBR and back to the CSTR

Organic Matter Capture by High-Rate Inoculum-Chemostat and MBBR Systems

RÉSUMÉ Le traitement des eaux usées biologique à charge élevée permet d'utiliser la matière organique pour la production d'énergie par un procédé méthanogène qui contribue au bilan d’énergie positif pour les stations de récupération des ressources de l’eau (StaRRE). L’objectif principal de cette recherche était de maximiser la biotransformation de la matière organique soluble et colloïdale de l’affluent en matière particulaire par l’emploi d'un bioréacteur à lit mobile (MBBR) pour que cette matière particulaire soit captée et acheminée vers un procédé de digestion anaérobie.----------ABSTRACT High-rate biological treatment processes allow the recovery of organic matter from wastewater into energy via methanogenesis contributing to the energy positive development of water resource recovery facilities (WRRFs) with lower carbon footprints. The main objective of this research was to maximize the bio-transformation of influent soluble and colloidal organic matter into particulate COD using a high-rate moving bed bioreactor (MBBR) for subsequent physico-chemical capture prior to transport to anaerobic digestion process

Produção de biodiesel a partir de microalgas heterotróficas

Orientadores: Telma Teixeira Franco, Lucas Antonius Maria van der WielenTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Química e Delft University of Technology.Resumo: Esta tese descreve os resultados da pesquisa de doutorado executada na Universidade de Campinas e na Universidade Técnica de Delft, como parte do program de Doutorado de Dupla Titulação entre as duas universidades. O projeto de pesquisa foi desenvolvido em parceria com a Petrobras S. A., que proveu a maior parte do suporte financeiro assim como suporte técnico, com o objetivo de avaliar o potencial de microalgas heterotróficas para a produção de biocombustíveis. Microalgas têm gerado muito interesse devido a seu inquestionável potencial para produção de biomassa e lipídeos através de fotossíntese. Nas últimas duas décadas, a busca por novas fontes de bio-energia causou um salto na pesquisa científica sobre cultivo de microalgas, o que impulsionou rapidamente o estado da arte. Apesar disso, a produção em larga escala ainda enfrenta obstáculos significativos, que encarecem os custos de produção and impedem que as microalgas se tornem uma fonte viável de bioenergia. A maior limitação das microalgas autotróficas é a necessidade da luz para o crescimento e o inevitável efeito de auto-sombreamento que ocorre com o aumento populacional. Quando a cultura se torna mais densamente povoada, a luz não consegue atingir camadas mais profundas, consequentemente desacelerando o crescimento. Isto limita a biomassa a baixas concentraçoes e, consequentemente, aumenta os volumes de cultivo e a demanda de grande quantidade de energy para separação da água. Apesar de extensa bibliografia sobre microalgas ter sido produzida nas últimas duas décadas, apenas uma pequena fração dos estudos se focaram no potencial heterotrófico desses versáteis microorganismos. Microalgas heterotróficas utilizam carbono orgânico como fonte energética e estrutural, ao invés de absorver carbono da atmosfera. Nesta condição, as microalgas podem crescer sem limitações pela luz e alcançar altas concentrações de biomassa e lipídeos. Porém, o cultivo heterotrófico e autotrófico não são comparáveis, já que o primeiro necessita de uma fonte de carbono orgânica e o segundo absorve carbono atmosférico. A tecnologia e os custos associados a cada um dos processos diferem fortemente. O desenvolvimento do cultivo heterotrófico inicia com a seleção de cepas adequadas para a produção de biocombustíveis e outros produtos de interesse. Este ainda é um campo de pesquisa pouco explorado, já que o cultivo heterotrófico representa apenas uma pequena fração de toda a literatura sobre algas. No capítulo 2, cepas de microalgas foram avaliadas em relação a sua capacidade de crescimento heterotrófico e produção de lipídeos. Após a análise do crescimento e composição celular, potenciais aplicações comerciais foram sugeridas para cada espécie estudada, já que diferentes composições de biomassa e lipídeos podem ser adequadas a diferentes produtos, como combustíveis, alimentos e produtos químicos. Chlorella vulgaris CPCC 90 foi identificada como uma opção adequada para a produção de biodiesel devido ao seu alto conteúdo lipídico e alta produtividade. Uma cepa produtora de ácidos graxas omega-3 poliinsaturados foi identificada e um breve estudo de otimização foi conduzido para aumentar a produção do ácido graxo de alto valor agregado. Após a seleção da cepa mais adequada para a produção de bio-combustíveis, o próximo passo foi o desenvolvimento de um cultivo altamente produtivo. A maior vantagem do cultivo heterotrófico é a possibilidade de alcançar altas concentrações de biomassa e conteúdo lipídico e, consequentemente, maiores produtividades volumétricas. Porém, o acúmulo de lipídeos ocorre quando células de microalgas são expostas a certas condições limitantes, que afetam negativamente o crescimento da biomassa. Desta forma, as condições de cultivo devem ser equilibradas de modo a promover o crescimento da biomassa e aumentar o conteúdo lipídico. Inicialmente, cultivos em batelada alimentada foram avaliados quanto ao acréscimo na concentração de biomassa e teor de lipídeos. A separação do crescimento e acúmulo de lipídeos em dois diferentes estágios permitiu a obtenção de uma cultura altamente concentrada e com elevado teor lipídico. Os lipídeos resultantes foram extraídos da biomassa e convertidos a biodiesel. Os rendimentos totais dos processos de cultivo, extração e reação foram calculados e discutidos (Capítulo 3). Apesar do cultivo em batelada alimentada ter-se mostrado altamente produtivo, o cultivo contínuo tem o potencial de reduzir o tempo ocioso da planta e aumentar a produtividade global e, consequentemente, reduzir custos de produção. Porém, manter cultivos contínuos com altas concentrações celulares não é trivial. O equilíbrio entre a vazão específica e a concentração de biomassa é crucial para a manutenção de alta produtividade. Cultivos em batelada alimentada e contínuos foram comparados quanto às produtividades totais, e o efeito da vazão específica sobre a concentração e produtividade de biomassa foi estudado (Capítulo 4). Cultivos contínuos também permitem um melhor controle da qualidade do produto final. A vazão específica e outros parâmetros, tais como a razão de alimentação de Carbono e Nitrogênio, afetam significativamente a composição de biomassa e o perfil de ácidos graxos dos lipídeos intracelulares. Através da variação destes parâmetros sob regime estacionário, tanto o conteúdo lipídico como a composição de ácidos graxos foi afetadas. Através da modelagem destes efeitos, é possível otimizar o processo, de acordo com o produto lipídico desejado (Capítulo 5). A integração de processos com outros setores da indústria pode, potencialmente, aumentar a viabilidade da produção de biocombustíveis de microalgas. Como o cultivo heterotrófico exige grande disponibilidade de fontes de carbono baratas, a integração com a indústria de cana-de-açúcar é uma opção atraente. Existem também potenciais ganhos para a industria da cana-de-açúcar, já que um terço de suas emissões de carbono resulta da queima de grandes quantidades de diesel de origem fóssil em operações agrícolas e de transporte. A produção de biodiesel de microalgas heterotróficas a partir de substratos da cana-de-açúcar representa uma oportunidade de de substituir a utilização de combustível de origem fóssil e aumentar a renovabilidade das refinarias de cana-de-açúcar. No Capítulo 6, é proposto um modelo de integração em que o melaço da cana-de-açúcar, vapor e eletricidade gerados na biorefinaria de cana-de-açúcar são utilizados para a produção de biodiesel de microalgas. Os resultados das simulações mostraram que a viabilidade do modelo proposto depende ainda da maturação da tecnologia, assim como de fatores externos, tais como o preço do petróleo e políticas e incentivos favoráveis a tecnologias sustentáveis. Esta tese representa uma contribuição ao estado da arte do desenvolvimento de biocombustíveis e outros produtos a partir de microalgas heterotróficas, especificamente focado no uso de culturas com alta densidade celular. Oferece ainda uma visão geral de alguns dos desafios que devem ser superados e das mais importantes variáveis na obtenção de um processo altamente produtivo e economicamente viávelAbstract: This thesis summarizes the results of a doctoral research executed in the State University of Campinas and in the Technical University of Delft as part of the PhD Dual Degree Program between the two universities. The research project was designed in partnership with Petrobras S. A. (Brazilian Petroleum Corporation), which provided most of the financial support as well as technical cooperation, with the goal of evaluating the potential of heterotrophic microalgae for biofuels production. Microalgae have generated a lot of interest due to their undoubted potential for the production of biomass and lipids through photosynthesis. In the last two decades, the search for new bio-energy feedstocks created a boom in scientific research on microalgae cultivation, which has improved the state of art of the technology at a rapid pace. However, large scale production still faces significant bottlenecks, which increase manufacturing costs and prevent microalgae from becoming a feasible bioenergy source. The main limitation related to autotrophic microalgae is the need of light for growth and the inevitable self-shading effect with the increase in cell population. As the culture becomes more densely populated, the light cannot reach deeper layers, thus slowing down the growth. This limits biomass to low concentrations and, consequently, increases cultivation volumes and demands high amounts of energy for water separation. Although extensive research about microalgae has been produced in the last two decades, only a small fraction of the studies aimed at the heterotrophic potential of these versatile microorganisms. Heterotrophic microalgae utilize organic carbon as energy source and building blocks rather than absorbing carbon from the atmosphere. In such circumstances, they can grow without light limitations and achieve high biomass and lipid concentrations. Nevertheless, heterotrophic and autotrophic cultivations are hardly comparable, since the former requires an organic carbon feedstock and the latter absorbs carbon from the atmosphere. The costs associated with each process are remarkably different, as well as the technology involved. The development of the heterotrophic cultivation process starts with the selection of suitable strains for the production of biofuels and other products. This is still a poorly explored field of research, as heterotrophic cultivation represents only a small fraction of all literature about algae. In Chapter 2, strains of microalgae were evaluated on their capacity for heterotrophic growth and lipid production. After the analysis of growth characteristics and cell composition, potential commercial applications for each strain were suggested, as different biomass and lipid compositions may be suitable for different final products, from biofuels to food and chemicals. Chlorella vulgaris CPCC 90 was identified as a suitable option for biodiesel production due to its high lipid content and productivity. One polyunsaturated omega-3 fatty acid producing strain was identified and a short optimization study was performed in order to enhance the production of the high value added fatty acid. After selection of the most suitable strain for biofuels production, the next step was the development of a highly productive cultivation process. The greatest advantage of heterotrophic cultivation is the possibility of reaching high biomass concentrations and lipid contents and, consequently, high volumetric productivities. However, lipid accumulation occurs when microalgae cells are exposed to certain limiting conditions, which negatively affect biomass growth. Therefore, cultivation conditions must be balanced in order to promote biomass growth and increase lipid content. After identification of the most suitable strain for biofuels production, fed-batch strategies were evaluated as means of increasing biomass concentration and lipid content. Decoupling biomass growth and lipid accumulation in two different stages allowed the production of a highly concentrated culture with increased lipid content. The resulting lipids were extracted from the produced biomass and converted into biodiesel. The overall yields of cultivation, extraction and reaction processes were calculated and discussed (Chapter 3). Although fed-batch cultivation proved itself highly productive, continuous production can potentially reduce downtime operations and increase global productivity, consequently reducing production costs. Operating continuous cultivation at high cell concentrations such as in the fed-batch process, however, is not trivial. The balance between dilution rate and biomass concentration is crucial in order to maintain high productivities. Fed-batch and continuous cultures were compared in terms of overall productivities and the effect of dilution rates was evaluated over biomass concentration and productivity (Chapter 4). Continuous cultivation also allows a better control of the final product quality. Growth rates and other parameters, such as Carbon to Nitrogen feeding ratio, significantly affect biomass composition and the fatty acid profile of intracellular lipids. By varying these parameters in steady state cultivation, lipid content and fatty acid composition were affected. By modelling these effects, it is possible to optimize the process according to the desired lipid-based product (Chapter 5). Process integration with other industry sectors may potentially increase the feasibility of microalgae biofuels production. Since heterotrophic cultivation demands a large availability of cheap carbon feedstocks, integration with the sugarcane industry is an attractive option. There are potential gains for the sugarcane industry as well, since one third of their carbon emissions result from burning large quantities of fossil-based diesel in crops and transportation operations. The production of heterotrophic microalgae biodiesel from sugarcane feedstocks offers the possibility of replacing the fossil fuel utilization and increasing the overall renewability of the sugarcane biorefinery. In Chapter 6, an integration model is proposed in which molasses, steam and electricity of sugarcane biorefinery are used for the production of microalgae biodiesel. Simulation results showed that the feasibility of the proposed model depends on the further development of the technology, as well as on external factors, such as petroleum prices and sustainability-driven policies and incentives. This thesis represents a contribution to the state of the art on the development of biofuels and other products from heterotrophic microalgae, specifically focused on the use of high cell density cultures. It offers an overview of some of the challenges that need to be overcome and provide insights on the most important variables for achieving a highly productive and economically feasible processDoutoradoDesenvolvimento de Processos QuímicosDoutor em Engenharia Químic

Nitrogen-fixing cyanobacteria for protein production: experimental and computational approach

openTo convey the food and feed industry towards a more sustainable approach, the production of vegetal-derived compounds, as proteins or pigments, represents a relevant topic of research. In that regard, nitrogen-fixing cyanobacteria are of interest thanks to their capability to fix atmospheric nitrogen into valuable N-based macromolecules. Looking for a potential industrial exploitation, laboratory experiments were conducted to assess Nostoc PCC 7120 cultivation features, both in batch and continuous configurations. Discontinuous cultivation was conducted in a 275-L pilot scale bioreactor in order to assess species performances in a larger operational scale, while 200-mL continuous photobioreactors were used to evaluate the effects of operating variables on harvested biomass and production of valuable compounds. With respect to enlightenment conditions, an optimal light intensity of 550 µmol m-2 s-1 was found, as the best compromise between biomass productivity and embedded valuable compounds. Secondly, a detailed study on diazotrophic growth performances was conducted, by varying the amount and source of nitrogen within the culture environment. The exploitation of such peculiar metabolic path seems to be a promising choice, since it contributes to lower nutrients-related capital costs as well as improve cultivation outcomes. Moreover, successful industrial exploitation relies also on the possibility of predicting such outcomes. Therefore, the development of a simulation model for two cyanobacterial species, namely Anabaena PCC 7122 and Nostoc PCC 7120, was considered. In detail, the model embeds the Droop theory based on the nutrient quota, accounting for the variation of biomass growth rate based on the amount of nutrients within the biomass itself. Compared to the various predictive models already available in literature, focusing only on overall biomass, the introduction of such peculiar kinetics allows then to simultaneously account for both the two typologies of cells characterizing the species in question, namely vegetative cells and heterocysts. According to the experimental background of this work, the model was developed for continuous operations, predicting cultivation outcomes with respect to the operating residence time. The derived computational outcomes seem in line with data collected in previous experimental campaigns, which were also used for the fitting of additional unknown model parameters.To convey the food and feed industry towards a more sustainable approach, the production of vegetal-derived compounds, as proteins or pigments, represents a relevant topic of research. In that regard, nitrogen-fixing cyanobacteria are of interest thanks to their capability to fix atmospheric nitrogen into valuable N-based macromolecules. Looking for a potential industrial exploitation, laboratory experiments were conducted to assess Nostoc PCC 7120 cultivation features, both in batch and continuous configurations. Discontinuous cultivation was conducted in a 275-L pilot scale bioreactor in order to assess species performances in a larger operational scale, while 200-mL continuous photobioreactors were used to evaluate the effects of operating variables on harvested biomass and production of valuable compounds. With respect to enlightenment conditions, an optimal light intensity of 550 µmol m-2 s-1 was found, as the best compromise between biomass productivity and embedded valuable compounds. Secondly, a detailed study on diazotrophic growth performances was conducted, by varying the amount and source of nitrogen within the culture environment. The exploitation of such peculiar metabolic path seems to be a promising choice, since it contributes to lower nutrients-related capital costs as well as improve cultivation outcomes. Moreover, successful industrial exploitation relies also on the possibility of predicting such outcomes. Therefore, the development of a simulation model for two cyanobacterial species, namely Anabaena PCC 7122 and Nostoc PCC 7120, was considered. In detail, the model embeds the Droop theory based on the nutrient quota, accounting for the variation of biomass growth rate based on the amount of nutrients within the biomass itself. Compared to the various predictive models already available in literature, focusing only on overall biomass, the introduction of such peculiar kinetics allows then to simultaneously account for both the two typologies of cells characterizing the species in question, namely vegetative cells and heterocysts. According to the experimental background of this work, the model was developed for continuous operations, predicting cultivation outcomes with respect to the operating residence time. The derived computational outcomes seem in line with data collected in previous experimental campaigns, which were also used for the fitting of additional unknown model parameters

Bio-hydrogen and biomass-supported palladium catalyst for energy production and waste-minimisation

The project objective was to advance the development of the H2 economy by improving biological H2 production in a sustainable way. Pseudo-continuous H2 production was achieved with improved efficiency, via the bacterial fermentation of sugars in a dual-bioreactor (‘upstream system’) comprising a dark fermentation coupled to a photofermentation. Excess biomass from the upstream system was used to recover palladium from solution, producing ‘palladised biomass’ (Bio-Pd(0)), which was useful in the construction of bioinorganic catalytic anodes for the electricity generation from bio-H2 using a polymer electrolyte membrane fuel cell (‘downstream system’). Furthermore, the catalytic usefulness of Bio-Pd(0) was confirmed in several reactions in comparison with other palladised biomasses and with Pd(0) made chemically. The upstream modules: Escherichia coli dark fermentation and Rhodobacter sphaeroides photofermentation, were investigated and developed separately, before coupling the two stages by the novel application of electrodialysis (accelerated membrane separation). The biorecovery and testing of palladium bionanocatalyst are described, before the production of fuel cell catalyst using waste biomass. The technical challenges and potential benefits of biohydrogen production are discussed and contrasted with those of competing biofuel technologies