Une grande quantité des eaux usées, particulièrement celles provenant de l'industrie minière, des fermentations et du traitement des aliments, contiennent-elles des concentrations élevées de sulfate (SO42-). SO42- réduction est une problématique sérieuse au niveau environnemental sous conditions qui ne sont pas contrôlées, cette problématique peut résulter en la libération de sulfure toxique vers l'environnement. Les caractéristiques typiques des eaux usées riches en SO42- sont 0.4-20.8 g SO42-.L-1, faible pH, élevé potentiel oxydatif, faible o négligeable demande chimique d'oxygène (DCO) et hautes concentrations de métaux lourds (drainage minier acide) que peuvent drastiquement endommager la flore et la faune des masses d'eau. L'objectif de cette thèse est d'étudier l'effet du donneur d'électrons sur le processus biologique RS par des bactéries réductrices de sulfate dans des bioréacteurs. Le processus biologique RS a été étudié à l'aide de polymères à base d'hydrate de carbone (PBHC) et biodéchets lignocellulosiques (L) comme des donneurs de libération lente d'électrons (PBCH-DLLE et L-DLLE, respectivement) dans des bioréacteurs discontinus et des bioréacteurs fonctionnant en continu inverse à lit fluidisé (ILF). Les ILF ont été rigoureusement testé pour RS sous haute vitesse et (alimentation-pénurie de nourriture) des conditions d'alimentation transitoires. Dans une autre configuration du bioréacteur, un bioréacteur séquentiel, l'effet de la concentration initiale SO42- sur le démarrage du réacteur a été étudié. Par ailleurs, l'effet de la concentration initiale du donneur d'électrons, NH4+ et SO42- ont été évalués dans des bioréacteurs de traitement par lots aussi bien. La robustesse et la résilience de RS a été démontrée dans les ILF en utilisant le lactate comme donneur d'électrons, dans lequel l'efficacité d'élimination de SO42- (EES) était comparable dans la période d’alimentation (67 ± 15%) de ILF2 aux conditions d'alimentation stables (71 ± 4%) dans même ILF2 et ILF1, le réacteur de commande (61 ± 15%). De la modélisation des réseaux de neurones artificiels et de l'analyse de sensibilité des données de fonctionnement de ILF2, il était prévu que les concentrations de SO42- influents ont affecté le rendement d'élimination de la DCO, la production de sulfure et des changements de pH. Dans un autre ILF3 à un rapport DCO:SO42- de 2.3, RS dans des conditions de taux élevés (CTE = 0.125 d) était 4,866 mg SO42- L-1 d-1 et un EES de 79% a été remporté. Par ailleurs, le second ordre Grau et les modèles d'enlèvement de substrat Stover-Kincannon équipés de la performance du réacteur à haut débit avec r2 > 0.96. Le rapport DCO:SO42- était le principal facteur affectant la RS. Dans des bioréacteurs par lots, en utilisant du papier filtre PBCH-DLLE, la RS a été réalisée à EES > 98%. Avec l'utilisation de scourer comme L-DLLE, un EES de 95% a été remporté. Cependant, lorsque la lavette est utilisée comme matériau de support de L-DLLE dans un ILF4, le RS a montré 38% (± 14) de EES entre 10-33 d de fonctionnement. La SR était limitée par le taux d'hydrolyse-fermentation et, par conséquent, la complexité de la DLLE. L'utilisation du donneur d'électrons pour le processus RS a simultanément amélioré aux concentrations de donneurs d'électrons initial décroissants / Many industrial wastewaters, particularly from the mining, fermentation and food processing industry contain high sulphate (SO42-) concentrations. SO42- reduction (SR) is a serious environmental problem under non-controlled conditions, which can result in the release of toxic sulphide to the environment. Typical characteristics of SO42--rich wastewater are 0.4-20.8 g SO42-.L-1, low pH, high oxidative potential, low or negligible chemical oxygen demand (COD) and high heavy metals concentrations (acid mine drainage), that can dramatically damage the flora and fauna of water bodies. The aim of this PhD is to study the effect of electron donor supply on the biological SR process by sulphate reducing bacteria in bioreactors. The biological SR process was studied using carbohydrate based polymers (CBP) and lignocelulosic biowaste (L) as slow release electron donors (CBP-SRED and L-SRED, respectively) in batch bioreactors and continuously operated inverse fluidized bed bioreactors (IFBB). IFBB were vigorously tested for SR under high rate and transient (feast-famine) feeding conditions. In another bioreactor configuration, a sequencing batch bioreactor, the effect of the initial SO42- concentration on the reactor start-up was investigated. Besides, the effect of the initial concentration of electron donor, NH4+, and SO42- were evaluated in batch bioreactors as well. The robustness and resilience of SR was demonstrated in IFBB using lactate as the electron donor wherein the SO42- removal efficiency (SRE) was comparable in the feast period (67 ± 15%) of IFBB2 to steady feeding conditions (71 ± 4%) in the same IFBB2 and to IFBB1, the control reactor (61 ± 15%). From artificial neural network modeling and sensitivity analysis of data of IFBB2 operation, it was envisaged that the influent SO42- concentrations affected the COD removal efficiency, the sulphide production and pH changes. In another IFBB3 at a COD:SO42- ratio of 2.3, SR under high rate conditions (HRT=0.125 d) was 4,866 mg SO42-. L-1 d-1 and a SRE of 79% was achieved. Besides, the Grau second order and the Stover-Kincannon substrate removal models fitted the high rate reactor performance with r2 > 0.96. The COD:SO42- ratio was the major factor affecting the SR. In batch bioreactors, using filter paper as CBP-SRED, SR was carried out at > 98% SRE. Using scourer as L-SRED, a 95% SRE was achieved. However, when the scourer was used as the L-SRED carrier material in an IFBB4, the SR showed 38 (± 14) % SRE between 10-33 d of operation. The SR was limited by the hydrolysis-fermentation rate and, therefore, the complexity of the SRED. Concerning sequencing batch bioreactor operation, the SR process was affected by the initial SO42- concentration (2.5 g SO42-.L-1) during the start-up. The sequencing batch bioreactor performing at low SRE (< 70%) lead to propionate accumulation, however, acetate was the major end product when SRE was > 90%. In batch bioreactors, the NH4+ feast or famine conditions affected the SR rates with only 4% and the electron donor uptake was 16.6% greater under NH4+ feast conditions. The electron donor utilization via the SR process improved simultaneously to the decreasing initial electron donor concentrations. This PhD research demonstrated that the SR process is robust to transient and high rate feeding conditions. Moreover, SR was mainly affected by the approach how electron donor is supplied, e.g. as SRED or as easy available electron donor, independently of the COD:SO42- ratio. Besides, the electron donor and SO42- utilization were affected by the lack or presence of nutrients like NH4+

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Stormwater retention basins are an integral component of municipal stormwater management strategies in North America. The province of Ontario’s Ministry of the Environment and Climate Change obligates land developers to implement stormwater management in their land use and development plans to mitigate the effects of urbanization (Bradford and Gharabaghi, 2004). When stormwater retention ponds are improperly designed or maintained, these basins can fail at improving effluent water quality and may exasperate water quality issues. Intense H2S production events in stormwater infrastructure is a serious problem which is seldom encountered and documented in stormwater retention ponds. This study monitored two stormwater retention ponds situated in the Riverside South community, Ottawa, Ontario, Canada for a period of 15 consecutive months to thoroughly characterize intense hydrogen sulfide (H2S) production in a stormwater retention pond under ice covered conditions during winter operation and during periods of drought under non-ice covered conditions during the summer. Field experiments showed a strong relationship (p < 0.006, R > 0.58, n = 20+) between hypoxic conditions (dissolved oxygen (DO) concentration < 2 mg/L) and the intense production of H2S gas. Ice-capping of the stormwater ponds during winter severely hindered reaeration of the pond and led to significant production of total sulfides in the Riverside South Pond #2 (RSP2), which subsequently resulted in the accumulation of total sulfides in the water column (20.7 mg/L) during winter in this pond. There was a perceived lag phase between the drop in DO and the increase in total sulfides near the surface, which was potentially indicative of slow movement of total sulfides from the benthic sediment into the water column. These high-sulfide conditions persisted in RSP2 from early January 2015 until the spring thaw, in mid-April, 2015. Riverside South Pond #1 (RSP1), the reference pond studied in this work, showed significantly less production of total sulfides across a significantly shorter period of time. Analysis of the microbial communities showed that there was little change in the dominant bacterial populations present in the benthic sediment of the pond demonstrating significant total sulfide production (RSP2) and the pond that did not demonstrate significant total sulfide production (RSP1). Additionally, it was found that locations with the most accumulated sediment had the highest propensity for the production of H2S gas. Furthermore, there was no perceivable community shift in the two ponds throughout the seasons, indicating that the sulfate-reducing bacteria (SRB) in stormwater benthic sediment are ubiquitous, exist in an acclimatized microbial population and are robust. Study of the microbial abundances revealed that SRB represented approximately 5.01 ± 0.79 % of the microbes present in the benthic sediment of RSP2. Likewise, in the stormwater pond which did not experience intense H2S gas production, RSP1, 6.22 ± 2.11 % of microbes were of the SRB type, demonstrating that H2S gas production does not correspond to higher concentrations of SRB or the proliferation of dominant species, but rather is a symptom of increased bacterial activity due to favourable environmental conditions. In addition, this work also covers the kinetics of sediment oxygen demand (SOD), ammonification and sulfate-reduction, and attempts to understand the processes leading to H2S gas production events. In doing so, it was observed that kinetics obtained full-scale field studies were greater than in laboratory kinetic experiments. Laboratory experiments at 4°C identified total SOD, ammonification and sulfate-reduction kinetics to be 0.023 g/m2/day, 0.027 g N/m2/day and 0.004 g S/m2/day, respectively. Meanwhile, kinetics calculated from the field study of stormwater retention ponds for total SOD, ammonification and sulfate-reduction were of 0.491 g/m2/day, 0.120 g N/m2/day and 0.147 g S/m2/day, respectively. It is expected that this difference is due to the depth of active sediment influencing the total rates of production/consumption, making area-normalized daily rates of production/consumption (g/m2/day) unsuitable for the comparison of field and laboratory studies, without some scaling factor. This study also measured supplementary kinetic parameters such as the Arrhenius coefficients and the half-saturation coefficient, to add to existing knowledge of sulfate-reduction.

Orientador: Ana Julia Fernandes / Resumo: Atividades portuárias são consideradas fontes de contaminação por metais, que em altas concentrações são tóxicos aos microrganismos, podendo mudar o tamanho, a composição e a atividade da comunidade microbiana. Essa pressão seletiva faz com que bactérias desenvolvam mecanismos para combater o estresse causado por eles. Diversos grupos de microrganismos que desenvolvem sistemas de tolerância a metais podem fazer sua biorremediação. Bactérias redutoras de sulfato (BRS) podem, devido ao seu metabolismo, precipitar metais facilitando sua remoção. A hipótese é que em áreas portuárias exista uma alteração na comunidade microbiana onde são favorecidos microrganismos resistentes, aumentando a chance do isolamento de bactérias resistentes com potencial biotecnológico. Portanto o objetivo dessa tese foi avaliar a influência da contaminação de metais na comunidade microbiana em áreas portuárias e na seleção de cepas resistentes, e isolar cepas de BRS com potencial biotecnológico. Para isso, foram coletados sedimentos de 2 áreas portuárias (Santos e São Sebastião) e uma área pristina (Ubatuba). Foram caracterizadas variáveis ambientais do sedimento (como pH, salinidade, temperatura, oxigênio dissolvido, porcentagem de matéria orgânica, concentrações de Cr, Cu, Cd e Zn). Foi extraído DNA total do sedimento para caracterização da comunidade utilizando sequenciamento Mi-Seq. Os sedimentos serviram como inóculo para isolamento de cepas que foram submetidas a testes de resistência aos metais.... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Port activities are considered sources of metal contamination, which in high concentrations are toxic to microorganisms, and can change the size, composition and activity of the microbial community. This selective pressure forces bacteria to develop resistance mechanisms. Several microorganisms groups that develop metal tolerance systems can perform their bioremediation. One example is Sulfate-reducing bacteria (SRB). Because of their metabolism, this group can precipitate metals facilitating their removal. In this way, the hypothesis is that in port areas there is a change in the microbial community where resistant microorganisms are favored, increasing the chance of isolation of resistant bacteria with biotechnological potential. Therefore, the objective of this thesis was evaluate the influence of metal contamination in the microbial community in port areas and in the selection of resistant strains, and to isolate strains of SRB with biotechnological potential. For this, sediments were collected from two port areas (Santos and São Sebastião) and one pristine area (Ubatuba). Sediment environmental variables were characterized (such as pH, salinity, temperature, dissolved oxygen, percentage of organic matter, concentrations of Cr, Cu, Cd and Zn). Total DNA was extracted from the sediment for community characterization using Mi-Seq sequencing. The sediments was used as inoculum for strain isolation, and the metal resistance was checked. The most contaminated sediments also wa... (Complete abstract click electronic access below) / Doutor

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Iron is a common cation in biomineral sand; it is present for example in magnetite produced by magnetotactic bacteria and in iron sulfides produced by sulfate reducing microorganisms. The work presented in this thesis focused on two types of microorganisms capable of forming iron biominerals. In the first project I have studied the effect of O2 on the respiratory physiology and the formation of magnetosomes by Magnetospirillum magneticum AMB-1. In the second project I have studied the relationship between olivine and the activity of dissimilatory sulfate reducing (DSR) microorganisms. For the first project, I grew cells of AMB-1 in cultures with various concentrations of O2 and monitored growth and the formation of magnetic mineral particles (MMP). Results have shown that AMB-1 cells grew better at 100-225 uMO2(aq) than at lower [O2], yet the formation of MMP was repressed at ~45 uM O2(aq) and strongly inhibited at >100 uM O2(aq).These results have helped better understand the dissimilarity between the optimal growth conditions of magnetotactic bacteria and the conditions needed for the formation of MMPs. My results have also shown that the reaction between H2S produced by DSRs and olivine is abiotic, not catalyzed and exergonic. The pH did not vary significantly during this reaction and pH variation (in the 5-9 range) did not significantly influence this chemical reaction. Bicarbonate inhibited the reaction between H2S and olivine, but not the chemical equilibrium. Phosphate, a weak iron chelator, influenced the equilibrium of the reaction and it is assumed to help increase the rate of olivine weathering in the presence of DSRs. The activity of DSRs was positively influenced by the presence and abundance of olivine. Based on my results I propose that olivine help DSR obtain energy more efficiently, but does not represent a source of energy or nutrients for the cells. These results helped better understand the formation of iron biominerals and signatures of this activity.

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Microbial-influenced corrosion (MIC) has been widely recognized as a significant economic and environmental problem in the oil and gas industry. MIC can be classified into two types based on the mechanisms: the extracellular electron transfer MIC (EET-MIC) and the metabolite MIC (M-MIC). The first includes electroactive bacteria that facilitate EET, while the latter includes bacteria that secrete corrosive metabolites. Sulfate-reducing bacteria (SRB) is believed to cause EET-MIC in carbon steel, a widely used metal in the oil and gas industry. In previous electroactive bacteria studies, nanowires have been shown to facilitate EET by acting as electron mediators. This study investigates the use of magnetite nanowires as electron mediators to accelerate EET-MIC of C1020 by Desulfovibrio vulgaris. The addition of 40 ppm (w/w) nanowires to carbon steel incubated with D. vulgaris, corrosive SRB species, for seven days resulted in 45% weight loss and 57% deeper pitting of carbon steel. Furthermore, electrochemical measurements of open circuit potential, linear polarization resistance and potentiodynamic polarization were found to be parallel with weight loss and pitting results. Therefore, these findings highlight the possibility of using magnetic nanowires as an electron mediator with high efficiency and selectivity to EET-MIC for future MIC studies and applications.

Acid mine drainage (AMD) is an acidic, iron-rich leachate that causes the dissolution of metals. It constitutes a worldwide problem of environmental contamination detrimental to aquatic life and water quality. AMD, however, is naturally attenuated at Davis Mine in Rowe, Massachusetts. We hypothesize that sulfate-reducing bacteria (SRB) are attenuating AMD. To elucidate the mechanisms by which SRB attenuate AMD, three research projects were conducted using a suite of molecular and geochemical techniques. First we established biological influence on the attenuation of AMD by comparing the microbial community and geochemical trends of microcosms of two contrasting areas within the site: AMD attenuating (AZ) and AMD generating (GZ) zones. The differences in geochemical trends between these zones were related to differences in microbial community membership. SRB were only detected in microcosms of the AZ, while iron oxidizers were only detected in the GZ. This study indicates that biological activity contributes to the attenuation of AMD and that SRB may have a role. To further describe the role of SRB, we determined the rates of sulfate reduction, the abundance, and membership of SRB in the second project. The sulfate reduction rate was weakly correlated with the abundance of SRB. This indicates that the SRB population may be utilizing another electron acceptor. One such electron acceptor would be iron, which was investigated in the third project. When SRB are inhibited, neither accumulation of reduced iron nor the formation of reduced iron sulfide precipitates occurred. Higher concentration of sulfide produced an increase in reduced iron and pH. Therefore, iron reduction mediated by reaction with biogenic sulfide contributes to the attenuation of AMD. This is the first report of the biological enhancement of iron reduction by acidotolerant SRB. The interdisciplinary research described in this dissertation provides evidence that SRB attenuate AMD through sulfate and iron reduction and a greater understanding of SRB in acidic environments. It also demonstrates how the biogeochemical cycling of sulfur is coupled to the iron cycle. Overall, the ubiquity and metabolic versatility of SRB offers boundless potential and exciting opportunities of study in the fields of bioremediation, geomicrobiology, and microbial ecology.

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