Interacções entre bactérias e hidrocarbonetos de petróleo em ambientes costeiros

Doutoramento em BiologiaThe ability of microorganisms to use oil hydrocarbons as a source of carbon and energy is crucial for environmental oil detoxification. However, there is still a lack of knowledge on fundamental aspects of this process on specific habitats and under different climate scenarios. In the first phase of this work, the culturable fraction of the oil hydrocarbon (OH) degrading bacteria from the sea surface microlayer (SML) of the estuarine system Ria de Aveiro was characterized. In the second phase, the impact of oil contamination on the active bacterial community was studied under climate change scenarios. Pseudomonas emerged as the prevailing genera among OH degrading bacteria in the SML. Moreover, culture-independent methods revealed that the relative abundance and diversity of Gammaproteobacteria, in which Pseudomonas is included, varies along an estuarine gradient of contamination. In order to access the impact of oil contamination on microbial communities under climate change scenarios, an experimental life support system for microcosm experiments (ELLS) was developed and validated for simulation of climate change effects on microbial communities. With the ELSS it is possible to simulate, in controlled conditions, fundamental parameters of the dynamics of coastal and estuarine systems while maintaining community structure in terms of the abundance of the most relevant members of the indigenous bacterial community. A microcosm experiment in which the independent and combined impact of ultraviolet radiation, ocean acidification and oil contamination on microbial communities was conducted. The impact on bacterial communities was accessed with a 16S RNA (cDNA) based barcode pyrosequencing approach. There was a drastic decrease of Desulfobacterales relative abundance after oil contamination under the reduced pH value estimated for 2100, when compared to present values. Since members of this order are known OH degraders, such a significant decrease may have consequences on OH detoxification of contaminated environments under the pH levels of the ocean expected for the future. Metagenome predictions based on the 16S RNA database indicated that several degradation pathways of OH could be affected under oil contamination and reduced water pH. Taken together, the results from this work bring new information on the dynamics of OH degrading bacteria in coastal and estuarine environments under present and future climate scenarios.A capacidade dos microrganismos utilizarem hidrocarbonetos de petróleo como fonte de carbono e energia é crucial na reciclagem destes compostos nos meios marinhos. Apesar da importância deste processo muitos dos aspectos da degradação microbiana de hidrocarbonetos em alguns habitats específicos e na perspetiva de diferentes cenários de alterações climáticas são ainda desconhecidos. Na primeira fase deste trabalho, a fracção cultivável das bactérias com capacidade para degradar hidrocarbonetos de petróleo na microcamada superficial marinha de um sistema estuarino (Ria de Aveiro) foi caracterizada. Na segunda fase, o impacto da contaminação por petróleo nas comunidades microbianas em diferentes cenários de alterações climáticas foi estudado. O género Pseudomonas revelou ser preponderante na fracção cultivável das bactérias degradadoras de hidrocarbonetos na microcamada superficial marinha. Além disso, os métodos independentes de cultivo revelaram que a abundancia relativa e diversidade da classe Gammaproteobacteria, na qual se inclui o género Pseudomonas, é diferente ao longo de um gradiente estuarino de contaminação. De forma a caracterizar o impacto da contaminação por petróleo nas comunidades microbianas sob diferentes cenários de alterações climáticas, um sistema de suporte de vida (ELSS) foi construído com o objectivo de realizar experiências de microcosmo. O ELSS permite simular e controlar aspectos fundamentais da dinâmica de sistemas costeiros e estuarinos preservando os elementos mais representativos da comunidade bacteriana original. Com este sistema foi possível realizar uma experiência onde o impacto independente e combinado da radiação ultravioleta, acidificação da água e contaminação por petróleo sobre comunidades bacterianas foi determinado. O impacto na estrutura das comunidades foi avaliado através de pirosequenciação do 16S RNA (cDNA). A ordem Desulfobacterales sofreu uma redução de abundância relativa muito significativa em resposta à contaminação por petróleo para uma redução de pH estimada para 2100. Uma vez que os membros desta ordem desempenham um papel crucial na degradação de hidrocarbonetos de petróleo, a reciclagem destes compostos em ambientes costeiros e estuarinos pode ser alterada em cenários ambientais futuros. A previsão do metagenoma a partir da base de dados do 16S RNA indica que várias vias de degradação de hidrocarbonetos de petróleo podem ser afectadas. Os resultados obtidos neste trabalho acescentam novas pistas sobre o papel das bactérias degradadoras de hidrocarbonetos em ambientes costeiros e estuarinos nos contextos ambientais actuais e futuro

Bacterioneuston da Ria de Aveiro: abundância, actividade e degradação de hidrocarbonetos aromáticos policíclicos

Mestrado em MicrobiologiaA microcamada superficial marinha (sea surface microlayer - SML) situa-se na interface entre a hidrosfera e a atmosfera. Esta camada é geralmente definida como o milímetro superior da coluna de água e representa um ambiente único com processos biológicos, químicos e físicos característicos. A SML está envolvida na regulação de processos ambientais e desempenha um importante papel no controlo das trocas de matéria, de origem natural e antropogénica, entre a hidrosfera e a atmosfera. As comunidades bacterianas que habitam a SML, designadas de bacterioneuston, são diferentes do bacterioplâncton que se desenvolve nas águas subjacentes e as suas actividades podem ter um impacto significativo na troca de matéria na interface ar-água. Devido à constituição química da SML, compostos de origem antropogénica de natureza lipofílica, como os hidrocarbonetos aromáticos policíclicos (polycyclic aromatic hydrocarbons - PAH), tendem a acumular neste compartimento da coluna de água. Os níveis de contaminação podem influenciar e activar as comunidades bacterianas com capacidade para metabolizar os PAH, podendo este processo ser crucial na redução da concentração destes compostos. Neste estudo foram analisadas as diferenças entre o bacterioneuston e o bacterioplâncton, quanto à sua actividade heterotrófica, abundância e composição em termos de grupos relacionados com a degradação de PAH, ao longo de um gradiente de contaminação num sistema estuarino (Ria de Aveiro, Portugal). A actividade heterotrófica bacteriana foi avaliada pelas taxas de actividade enzimática extracelular e pelo perfil de utilização de fontes únicas de carbono. A composição das comunidades bacterianas foi avaliada por FISH (Fluorescent In Situ Hybridization) e a diversidade foi analisada por DGGE (Denaturing Gradient Gel Electrophoresis). As taxas de hidrólise enzimática demonstraram uma tendência para um acréscimo na SML relativamente à UW. Não se observaram diferenças significativas entre o bacterioneuston e o bacterioplâncton quanto ao padrão de utilização de fontes únicas de carbono. Ambas as comunidades revelaram idêntica abundância total de microrganismos e abundância relativa do domínio Bacteria. A subclasse γ- Proteobacteria registou os seus valores de abundância relativos mais elevados nas estações de amostragem com maior concentração de PAH de baixo peso molecular. A análise de fragmentos de rDNA 16s por DGGE, sugere que a diversidade bacteriana global é similar entre a SML e a UW. O isolamento de bactérias degradadoras de PAH da SML, evidenciou uma predominância do género Pseudomonas. A análise dos fragmentos do gene gacA, específico de Pseudomonas sp., sugere que a diversidade deste género é superior na SML. ABSTRACT: The sea surface microlayer (SML) represents the interface between the hydrosphere and the atmosphere. This layer is usually defined as the uppermost millimeter of the water column, and configures a unique environment with characteristic biological, chemical and physical processes. It is accepted that this interface controls environmental processes and plays a major role in regulating the exchanges of natural and anthropogenic substances between the atmosphere and the hydrosphere. Bacterial communities inhabiting the SML (bacterioneuston) are distinct from bacterioplankton developing in the underlying waters (UW), and their activities can significantly impact the exchange of matter across at air–water interface. Due to its chemical nature, the amounts of autochtonous and anthropogenic recalcitrant compounds such as polycyclic aromatic hydrocarbons (PAH), shown higher concentrations at the SML when compared with the UW. Such levels of contamination may activate bacterial communities able to metabolize PAH compounds, a crucial biological process for reducing PAH contamination. In this study we attempted to compare bacterioneuston and bacterioplâncton communities in terms of heterotrophic activity, abundance, and diversity of PAH degrading bacteria, along a gradient of contamination in an estuarine system (Ria de Aveiro, Portugal). Bacterial heterotrophic activity was described by the rates of ectoenzymatic activity, as well as by the profiles of sole-carbonsource utilization. The composition of bacterial communities was assessed by FISH (Fluorescent In Situ Hybridization) and DGGE (Denaturing Gradient Gel Electrophoresis) was applied to access bacterial diversity. The rates of ectoenzymatic hydrolysis were more often higher at the SML than in the UW. The patterns of sole-carbon-source utilization were similar for both communities. Total bacterial abundance and the relative abundance of the Bacteria domain, were also similar in bacterioneuston and bacterioplankton The γ-Proteobacteria subclass, showed the highest values at the estuarine sections were the concentration of low molecular weight PAH was higher. The analysis of 16s rDNA fragments by DGGE indicates that bacterial communities from the SML and UW shown similar diversity. Pseudomonas, was the dominant PAH hydrocarbon-degrading bacteria isolated from the SML. The analysis of Pseudomonas-specific gacA gene fragments indicates a higher diversity of this group in bacterioneuston

Geographical location and habitat predict variation in prokaryotic community composition of Suberites diversicolor

Purpose: Marine lakes are unique habitats that house diverse assemblages of benthic and planktonic organisms including endemic species. In this study, we aimed to assess to what extent geographical location (Berau versus Papua) and the degree of marine lake connectivity (relatively open versus closed) to the surrounding marine environment structures the prokaryotic community composition of the sponge species Suberites diversicolor. Methods: Sponge specimens were sampled in five marine lakes in Borneo and Papua and one open sea habitat in Taiwan. Result: Prokaryotic communities of S. diversicolor were dominated by members assigned to the Proteobacteria (particularly Alphaproteobacteria and Gammaproteobacteria) and Cyanobacteria, which together made up from 78 to 87% of sequences in all samples. The dominant operational taxonomic units (OTUs) in most samples, OTUs 1 and 3, were both assigned to the alphaproteobacterial order Rhodospirillales with OTU-1 dominant in the marine lakes of Berau and Papua and OTU-3 in Taiwan. OTU-3 was also largely absent from Papuan samples but present in all Berau samples. Compositionally, S. diversicolor samples clustered according to geographical location with the main axis of variation separating marine lake samples collected in Berau from those collected in Papua and the second axis of variation separating open sea samples collected in Taiwan from all marine lake samples. In addition, our results suggest that the degree of lake connectivity to the open sea also influences prokaryotic composition. Conclusion: Although previous studies have shown that sponge-associated microbial communities tend to be stable across different geographical and environmental gradients, in the present study, both geography and local environmental conditions were significant predictors of variation in prokaryotic community composition of S. diversicolor.publishe

Bacterial composition and putative functions associated with sponges, sediment and seawater from the Tioman coral reef system, Peninsular Malaysia

Microbial communities associated with sponges play important roles in sponge health, defence and nutrient dynamics in coral reef systems. Here, we used a 16S rRNA gene high-throughput sequencing approach in order to test for differences in bacterial composition and predictive functional differences among two LMA (low microbial abundance) (Stylissa carteri and Stylissa massa), one well-known HMA (high microbial abundance) (Xestospongia testudinaria), one putative HMA (Aaptos lobata) sponge species, sediment and seawater in a coral reef ecosystem surrounding Tioman Island. Our results showed that biotope (sponge species, sediment and seawater) was a significant predictor of composition and abundance of bacterial communities. Proteobacteria was the dominant phylum in S. carteri, seawater, sediment and A. lobata, whilst bacterial communities of S. massa and X. testudinaria were dominated by Cyanobacteria and Chloroflexi, respectively. The predicted metagenome revealed differentially enriched pathways in HMA and LMA sponges. Differentially enriched subcategories in HMA sponges included ‘Signaling Molecules and Interaction’, ‘Carbohydrate Metabolism’ and ‘Excretory System’. Subcategories enriched in LMA sponges included ‘Replication and Repair’, ‘Energy Metabolism’, ‘Metabolism of Cofactors and Vitamins’ and ‘Environmental Adaptation’. Overall, the predicted functional analysis suggests that, although HMA and LMA associated bacterial communities shared core functional features, they use different strategies to defend against pathogens, obtain energy or cope with stress

Emiliania huxleyi—Bacteria Interactions under Increasing CO2 Concentrations

The interactions established between marine microbes, namely phytoplankton–bacteria, are key to the balance of organic matter export to depth and recycling in the surface ocean. Still, their role in the response of phytoplankton to rising CO2 concentrations is poorly understood. Here, we show that the response of the cosmopolitan Emiliania huxleyi (E. huxleyi) to increasing CO2 is affected by the coexistence with bacteria. Specifically, decreased growth rate of E. huxleyi at enhanced CO2 concentrations was amplified in the bloom phase (potentially also related to nutrient concentrations) and with the coexistence with Idiomarina abyssalis (I. abyssalis) and Brachybacterium sp. In addition, enhanced CO2 concentrations also affected E. huxleyi’s cellular content estimates, increasing organic and decreasing inorganic carbon, in the presence of I. abyssalis, but not Brachybacterium sp. At the same time, the bacterial isolates only survived in coexistence with E. huxleyi, but exclusively I. abyssalis at present CO2 concentrations. Bacterial species or group-specific responses to the projected CO2 rise, together with the concomitant effect on E. huxleyi, might impact the balance between the microbial loop and the export of organic matter, with consequences for atmospheric carbon dioxide

Compositional analysis of archaeal communities in high and low microbial abundance sponges in the Misool coral reef system, Indonesia

The high/low microbial abundance (HMA/LMA) dichotomy in sponges has been the subject of several studies over recent years, but few studies have analysed this dichotomy in terms of the sponge archaeal community and function. Using a 16S rRNA gene barcoded pyrosequencing approach and predictive functional analysis (PICRUSt) we compared the archaeal composition, richness and predicted function of one HMA sponge (Xestospongia testudinaria), one LMA sponge (Stylissa carteri) and one sponge species of unknown microbial abundance (Aaptos lobata). Although most of the archaeal sequences were assigned to the Crenarchaeota phylum, S. carteri had the highest percentage of sequences assigned to the Euryarchaeota phylum. Variation among sponge species explained >85% of the variation in archaeal operational taxonomic unit (OTU) composition with each sponge species forming a distinct cluster. There were significant differences in predicted PICRUSt profiles among sponge species, suggesting that archaeal communities present in the studied sponge species may perform different functions. X. testudinaria and A. lobata were similar both in terms of OTU and KEGG orthologues composition, which may indicate that A. lobata is a HMA sponge. Additionally, some of the most enriched functions seem to be related to traits associated with high and low microbial abundance sponges.publishe