Construction And Regulation Of Artificial Consortia Of Scenedesmus Obliquus And Microorganism

As a source of third-generation biodiesel, microalgae have attracted a great deal of attentions because of their excellent properties including high growth rate, high photosynthetic efficiency and high lipid content. Therefore, microalgae are considered to be ideal organisms for developing highly productive and robust strains that are essential for economically viable biodiesel production. Large-scale cultivation in open ponds is the main mode for microalgal lipid production, but it is hindered by low biomass productivity and instability during cultivation resulted from invasions by pathogens. The consortium of microalgae and microorganisms can provide a comfortable micro-enviroment for microalgal growth. To date, the effects of consortium system on the microalgal growth and lipid production are still unclear. In this study, the artificial consortia of Scenedesmus obliquus and microorganisms were constructed to study their effects on the microalgal growth and lipid production. Moreover, the syntrophic machenism between microalgae and microorganisms in the consortia was also investigated. Additionally, the stability of S. obliquus and bacterium consortia was studied under open conditions in the lab.The consortia (i.e. co-cultivation) of microalgae and eukaryotic strains, including Candida tropicalis and Saccharomyces cerevisiae, were constructed respectively. In the co-cultures of S.obliquus and C. tropicalis, microalgal biomass concentration, net photosynthetic activity and lipid content increased by 30.3% ,61% and 22.5%, respectively, compared to S. obliquus alone, but no stimulation was observed in the co-culture of S. obliquus and Saccharomyces cerevisiae. It has been demonstrated that microalgae exhibited the highest concentration of biomass and net photosynthetic activity when S. obliquus and C. tropicalis were seeded at the ratio of 3:1, compared to other seeding ratios. In the consortium of microalgae and C. tropicalis, the later could degrade and use the extracellular polymeric substances (EPS) excreted from microalgae, providing a favorable micro-environment for the microalgal growth. The co-cultivation of S. obliquus and C. tropicalis could not increase the microalgal growth under open condition, although it could happen in the closed photobioreactor.The consortia of S. obliquus and bacterial strains were constructed. Based on the analysis of bacterial diversity in culutres of xenic S. obliquus, it was found that both beneficial and harmful bacterial populations were associated with xenic microalgae. The supernatant of axenic microalgal cultures was used to subculture bacterial isolates and 5 favorably syntrophic bacterial strains were obtained, including Brevundimonas aurantiaca (2-1), Rhizobium sp. (2-2), Pseudomonas sp. (3-4), Acidovorax facilis (3-10) and Diaphorobacter sp. (3-11). When the syntrophic bacterial individuals were co-cultivated with axenic S. obliquus respectively, microalgal growth was enhanced in all consortia, among which A. facilis represented the largest biomass concentration increase of 24.8% compared to the axenic culture of S. obliquus. The consortium system also increased microalgal lipid content, lipid productivity, and the proportion of saturated fatty acids and oleic acid. According to scanning electron microscopy (SEM) analysis, the selected syntrophic bacterial strains adhered directly to the S. obliquus cell surface, which was helpful for the material exchange between microalgae and bacterium. Moreover, the participation of bacterial strains could metabolize EPS and change both concentrations and compositions of proteins and polysaccharides dissolved in the co-cultures, contributing to the increased microalgal biomass and lipid production in the consortia.Studies on the regulation of microalgae-bacterium consortia were performed. In the meantime, the effects of different regulation parameters on microalgal growth and lipid production and EPS concentrations during cultivation were investigated, including seeding ratios of microalgae and bacterial strains, concentrations of exogenousmetal ions, nitrogen deficiency and different combinations of syntrophic bacterial strains. The best S. obliquus growth was observed when microalgae and bacterial individuals were seeded in a 3:1 ratio. The EPS concentration, an important regulation parameter, increased significantly in microalgae-bacterium consortia upon addition of 100 mg/L CaCl2; while, the microalgal growth and lipid production were not affected by CaCl2 at the same concentration. After cultivation under nitrogen source deficiency for 7 days, the lipid content and lipid productivity in the consortia of S. obliquus and syntrophic bacterial strains were 33.3% and 73.9% higher than those in S. obliquus, respectively. Furthermore, the presence of mixed bacterial strains could also enhance microalgal growth and lipid production. Among the different bacterial combinations, the mixture of strain 2-2,3-4, 3-10 and 3-11 showed the most positive effects, where microalgal biomass concentration and lipid content increased by 28.5% and 14.6% compared to axenic S. obliquus.Thereafter, the gas exchange and carbon exchange between microalgae and bacteria were studied with Applikon microreactor to understand the mechanism by which microalgae-bacterium consortia enhance microalgal growth. The participation of syntrophic bacterial strains utilized the dissolved oxygen, reducing oxygen stress and thus enhancing microalgal growth. The microalgae further assimilated carbon dioxide and released oxygen that could be used by the bacteria. Meanwhile, the syntrophic bacteria degraded compounds with high molecular weight and organic carbon secreted by microalgae, which reduced the concentration of total organic carbon (TOC) and increased the concentration of total inorganic carbon (TIC). The decrease of TOC stress and the increase of TIC were helpful for microalgal photosynthesis and growth. Additionally, the favorable micro-environment due to bacterial presence could enhance the transformation of polysaccharide to lipid in microalgae.In the last, the stability of microalgae-bacterium consortia was examined when the photobioreactors were operated under open conditions in the laboratory. After 16 days of cultivation, the biomass concentration in the consortia under open condition was 17.1% and 95% higher than that in axenic S. obliquus cultures under closed conditions and under open conditions, respectively. Meanwhile, the lipid content was improved by 24% in airtificial consortia under open conditions compared to axenic S. obliquus cultivation under closed conditions. Furthermore, it was demonstrated that the participation of bacterial strains could help to keep the stability of bacterial community in consortia. That is, the species and corresponding proportion of seeded individual strains were basically stable, which contributed 74-82.9% of all bacterial populations in total, and bacteria with the proportion (no more than 1% in total) accounted for about 10% during 12 days of cultivation. On the contrary, the bacterial community in axenic microalgal cultures changed significantly, and the bacterial strains with the proportion under 1% increased to 40% after 12 days of cultivation under open conditions. In addition, under open conditions, the harmful microfauna including Colpodea and Platyophrya, which lead to reduction of miacroalgal biomass, were detected in axenic S. obliquus cultures, but not found in consortia. These preliminary results imply that the airtificial consortia of S. obliquus and bacteria improved the stability of microalgal cultivation under open conditions.