Microalgae–bacteria symbiosis in microalgal growth and biofuel production: a review

Photosynthetic microalgae can capture solar energy and convert it to bioenergy and biochemical products. In nature or industrial processes, microalgae live together with bacterial communities and may maintain symbiotic relationships. In general interactions, microalgae exude dissolved organic carbon that becomes available to bacteria. In return, the bacteria remineralize sulphur, nitrogen and phosphorous to support the further growth of microalgae. In specific interactions, heterotrophic bacteria supply B vitamins as organic cofactors or produce siderophores to bind iron, which could be utilized by microalgae, while the algae supply fixed carbon to the bacteria in return. In this review, we focus on mutualistic relationship between microalgae and bacteria, summarizing recent studies on the mechanisms involved in microalgae–bacteria symbiosis. Symbiotic bacteria on promoting microalgal growth are described and the relevance of microalgae–bacteria interactions for biofuel production processes is discussed. Symbiotic microalgae–bacteria consortia could be utilized to improve microalgal biomass production and to enrich the biomass with valuable chemical and energy compounds. The suitable control of such biological interactions between microalgae and bacteria will help to improve the microalgae-based biomass and biofuel production in the future.     

Using agro-industrial wastes for the cultivation of microalgae and duckweeds: Contamination risks and biomass safety concerns

Aquatic organisms, such as microalgae (Chlorella, Arthrospira (Spirulina), Tetrasselmis, Dunalliela etc.) and duckweed (Lemna spp., Wolffia spp. etc.) are a potential source for the production of protein-rich biomass and for numerous other high-value compounds (fatty acids, pigments, vitamins etc.). Their cultivation using agro-industrial wastes and wastewater (WaW) is of particular interest in the context of a circular economy, not only for recycling valuable nutrients but also for reducing the requirements for fresh water for the production of biomass. Recovery and recycling of nutrients is an unavoidable long-term approach for securing future food and feed production. Agro-industrial WaW are rich in nutrients and have been widely considered as a potential nutrient source for the cultivation of microalgae/duckweed. However, they commonly contain various hazardous contaminants, which could potentially taint the produced biomass, raising various concerns about the safety of their consumption. Herein, an overview of the most important contaminants, including heavy metals and metalloids, pathogens (bacteria, viruses, parasites etc.), and xenobiotics (hormones, antibiotics, parasiticides etc.) is given. It is concluded that pretreatment and processing of WaW is a requisite step for the removal of several contaminants. Among the various technologies, anaerobic digestion (AD) is widely used in practice and offers a technologically mature approach for WaW treatment. During AD, various organic and biological contaminants are significantly removed. Further removal of contaminants could be achieved by post-treatment and processing of digestates (solid/liquid separation, dilution etc.) to further decrease the concentration of contaminants. Moreover, during cultivation an additional removal may occur through various mechanisms, such as precipitation, degradation, and biotransformation. Since many jurisdictions regulate the presence of various contaminants in feed or food setting strict safety monitoring processes, it would be of particular interest to initiate a multi-disciplinary discussion whether agro-industrial WaW ought to be used to cultivate microalgae/duckweed for feed or food production and identify most feasible options for doing this safely. Based on the current body of knowledge it is estimated that AD and post-treatment of WaW can lower significantly the risks associated with heavy metals and pathogens, but it is yet unclear to what extent this is the case for certain persistent xenobiotics.     

Microalgae as sources of pharmaceuticals and other biologically active compounds

In the last decade the screening of microalgae, especially the cyanobacteria (blue-green algae), for antibiotics and pharmacologically active compounds has received ever increasing interest. A large number of antibiotic compounds, many with novel structures, have been isolated and characterised. Similarly many cyanobacteria have been shown to produce antiviral and antineoplastic compounds. A range of pharmacological activities have also been observed with extracts of microalgae, however the active principles are as yet unknown in most cases. Several of the bioactive compounds may find application in human or veterinary medicine or in agriculture. Others should find application as research tools or as structural models for the development of new drugs. The microalgae are particularly attractive as natural sources of bioactive molecules since these algae have the potential to produce these compounds in culture which enables the production of structurally complex molecules which are difficult or impossible to produce by chemical synthesis.     

Biodiesel fuel from microalgae-promising alternative fuel for the future: a review

The use of organic matter such as vegetable oil to produce biodiesel fuel has been a practical technology for a number of years. However, the search for new technologies and raw materials for biodiesel fuel production has gained increased attention recently because of financial and environmental concerns. Of particular interest are raw materials that are not food-related. Microalgae have gained a great deal of attention as a potential biodiesel raw material because of their high growth rates and ability to accumulate oil, bind carbon dioxide, and remove contaminants from wastewater. This article is a literature review of technologies for biodiesel production from microalgae. The technologies relate to microalgal cultivation, microalgal growth enhancement to simultaneously increase biomass and reduce pollution, the preparation of microalgal biomass for biodiesel production, and biodiesel production itself.     

Growth and neutral lipid synthesis in green microalgae: A mathematical model

Many green microalgae significantly increased their cellular neutral lipid content when cultured in nitrogen limited or high light conditions. Due to their lipid production potential, these algae have been suggested as promising feedstocks for biofuel production. However, no models for algal lipid synthesis with respect to nutrient and light have been developed to predict lipid production and to help improve the production process. A mathematical model is derived describing the growth dynamics and neutral lipid production of green microalgae grown in batch cultures. The model assumed that as the nitrogen was depleted, photosynthesis became uncoupled from growth, resulting in the synthesis and accumulation of neutral lipids. Simulation results were compared with experimental data for the green microalgae Pseudochlorococcum sp. For growth media with low nitrogen concentration, the model agreed closely with the data; however, with high nitrogen concentration the model overestimated the biomass. It is likely that additional limiting factors besides nitrogen could be responsible for this discrepancy.     

Microalgae as multi-functional options in modern agriculture: current trends,prospects and challenges

Algae are a group of ubiquitous photosynthetic organisms comprising eukaryotic green algae and Gram-negative prokaryotic cyanobacteria, which have immense potential as a bioresource for various industries related to biofuels, pharmaceuticals, nutraceuticals and feed. This fascinating group of organisms also has applications in modern agriculture through facilitating increased nutrient availability, maintaining the organic carbon and fertility of soil, and enhancing plant growth and crop yields, as a result of stimulation of soil microbial activity. Several cyanobacteria provide nitrogen fertilization through biological nitrogen fixation and through enzymatic activities related to interconversions and mobilization of different forms of nitrogen. Both green algae and cyanobacteria are involved in the production of metabolites such as growth hormones, polysaccharides, antimicrobial compounds, etc., which play an important role in the colonization of plants and proliferation of microbial and eukaryotic communities in soil. Currently, the development of consortia of cyanobacteria with bacteria or fungi or microalgae or their biofilms has widened their scope of utilization. Development of integrated wastewater treatment and biomass production systems is an emerging technology, which exploits the nutrient sequestering potential of microalgae and its valorisation. This review focuses on prospects and challenges of application of microalgae in various areas of agriculture, including crop production, protection and natural resource management. An overview of the recent advances, novel technologies developed, their commercialization status and future directions are also included.     

莱茵衣藻中脂质代谢过程研究进展

微藻中脂质代谢产生的化合物,可用于生物燃料、营养品和生物药品的生产,因此具有重要的经济价值。脂质代谢贯穿微藻的全部生命过程,对微藻的生长发育和应对外界胁迫都具有重要意义。微藻与研究较清楚的真菌和陆地植物在脂质代谢过程方面具有相似性。当然,随着微藻脂质代谢相关功能基因逐渐被鉴定,人们发现微藻的脂质代谢也具有区别真菌和陆地植物的独特性,因此针对微藻脂质代谢过程的分析具有重要意义。莱茵衣藻是研究脂质代谢过程的模式生物,已经通过基因组、转录组、蛋白质组和代谢组等方法,对其质体、内质网和过氧化物酶体中进行的脂质合成和分解过程进行了研究。本文总结了近年来莱茵衣藻质体、内质网和过氧化物酶体中脂质代谢过程的研究成果,并进行综合阐述。     

The dilemma for lipid productivity in green microalgae: importance of substrate provision in improving oil yield without sacrificing growth

Rising oil prices and concerns over climate change have resulted in more emphasis on research into renewable biofuels from microalgae. Unlike plants, green microalgae have higher biomass productivity, will not compete with food and agriculture, and do not require fertile land for cultivation. However, microalgae biofuels currently suffer from high capital and operating costs due to low yields and costly extraction methods. Microalgae grown under optimal conditions produce large amounts of biomass but with low neutral lipid content, while microalgae grown in nutrient starvation accumulate high levels of neutral lipids but are slow growing. Producing lipids while maintaining high growth rates is vital for biofuel production because high biomass productivity increases yield per harvest volume while high lipid content decreases the cost of extraction per unit product. Therefore, there is a need for metabolic engineering of microalgae to constitutively produce high amounts of lipids without sacrificing growth. Substrate availability is a rate-limiting step in balancing growth and fatty acid (FA) production because both biomass and FA synthesis pathways compete for the same substrates, namely acetyl-CoA and NADPH. In this review, we discuss the efforts made for improving biofuel production in plants and microorganisms, the challenges faced in achieving lipid productivity, and the important role of precursor supply for FA synthesis. The main focus is placed on the enzymes which catalyzed the reactions supplying acetyl-CoA and NADPH.     

Axenic cultures for microalgal biotechnology: Establishment,assessment, maintenance,and applications

The impact of microalgae (including blue-green algae or cyanobacteria) on human life can be both beneficiary and deleterious. While microalgae can be cultivated and used as feedstocks for the production of bioenergy and high value-added products in nutraceuticals, pharmaceuticals, and aquaculture feeds, some microalgae cause harmful algal blooms (HABs) that cause large-scale mortality in aquatic environments around the world. Thus, with the development of microalgal biotechnology and increasing concern about HABs, research on microscopic algae has increased significantly. However, this growth of academic research and application fields has been hindered by difficulties in obtaining axenic cultures. Therefore, this review provides a brief explanation of diverse establishment techniques, along with their strengths and weaknesses, with the hope of facilitating successful axenic cultures. A compilation of research fields and relevant important findings is also presented to clarify the importance of pure algal cultures. Finally, several controversial and sometimes overlooked issues related to the establishment, maintenance, and utilization of axenic cultures are discussed.     

Mechanisms of response to salinity in halotolerant microalgae

Summary A limited number of organic solutes are used by microalgae to adjust their internal osmotic pressure in response to changing external salinities. Glycerol and proline are used by the most extremely halotolerant algae. Only glycerol allows growth at salinities approaching saturation. In addition to organic osmoregulatory solutes, inorganic ions also play an important role in osmoregulation. The ability of microalgae to maintain intracellular ions at levels compatible with metabolic functions may set upper limits for their salt tolerance. Requirements for NaCl in the external medium for nutrient transport may define the lower salinity limits for growth observed for some euryhaline algae.Osmotic upshocks generally cause severe temporary inhibition of photosynthesis in euryhaline microalgae. Extensive osmotic downshocks have little effect on photosynthesis in microalgae with strong cell walls, while wall-less species appear to be more sensitive. Rapid glycerol synthesis takes place in response to increased external salinity inChlamydomonas pulsatilla both in light and dark. Starch supplies carbon for glycerol synthesis in the dark and also during the initial periods of inhibition of photosynthesis in the light. Turnover of osmoregulatory solutes such as glycerol and isofloridoside may be an important aspect of the osmoregulatory mechanism.At salinities beyond the growth limit for the green flagellateChlamydomonas pulsatilla, resting spores are formed that enable this alga to survive extreme salinities.     

Microalgal-Bacterial Consortia as Future Prospect in Wastewater Bioremediation,Environmental Management and Bioenergy Production

In the recent years, microalgae have captured researchers’ attention as the alternative feedstock for various bioenergy production such as biodiesel, biohydrogen, and bioethanol. Cultivating microalgae in wastewaters to simultaneously bioremediate the nutrient-rich wastewater and maintain a high biomass yield is a more economical and environmentally friendly approach. The incorporation of algal–bacterial interaction reveals the mutual relationship of microorganisms where algae are primary producers of organic compounds from CO₂, and heterotrophic bacteria are secondary consumers decomposing the organic compounds produced from algae. This review would provide an insight on the challenges and future development of algal–bacterial consortium and its contribution in promoting a sustainable route to greener industry. It is believed that microalgal-bacterial consortia will be implemented in the near-future for sub-sequential treatment of wastewater bioremediation, bioenergy production and CO₂ fixation, promoting sustainability and making extraordinary advancement in life sciences sectors.     

Microalgae as platforms for production of recombinant proteins and valuable compounds: progress and prospects

Over the last few years microalgae have gained increasing interest as a natural source of valuable compounds and as bioreactors for recombinant protein production. Natural high-value compounds including pigments, long-chain polyunsaturated fatty acids, and polysaccharides, which have a wide range of applications in the food, feed, cosmetics, and pharmaceutical industries, are currently produced with nontransgenic microalgae. However, transgenic microalgae can be used as bioreactors for the production of therapeutic and industrially relevant recombinant proteins. This technology shows great promise to simplify the production process and significantly decrease the production costs. To date, a variety of recombinant proteins have been produced experimentally from the nuclear or chloroplast genome of transgenic Chlamydomonas reinhardtii. These include monoclonal antibodies, vaccines, hormones, pharmaceutical proteins, and others. In this review, we outline recent progress in the production of recombinant proteins with transgenic microalgae as bioreactors, methods for genetic transformation of microalgae, and strategies for highly efficient expression of heterologous genes. In particular, we highlight the importance of maximizing the value of transgenic microalgae through producing recombinant proteins together with recovery of natural high-value compounds. Finally, we outline some important issues that need to be addressed before commercial-scale production of high-value recombinant proteins and compounds from transgenic microalgae can be realized.     

Progress in physicochemical parameters of microalgae cultivation for biofuel production

Microalgae have been exploited for biofuel generation in the current era due to its enormous energy content, fast cellular growth rate, inexpensive culture approaches, accumulation of inorganic compounds, and CO₂ sequestration. Currently, research is ongoing towards the advancement of the microalgae cultivation parameters to enhance the biomass yield. The main objective of this study was to delineate the progress of physicochemical parameters for microalgae cultivation such as gaseous transfer, mixing, light demand, temperature, pH, nutrients and the culture period. This review demonstrates the latest research trends on mass transfer coefficient of different microalgae culturing reactors, gas velocity optimization, light intensity, retention time, and radiance effects on microalgae cellular growth, temperature impact on chlorophyll production, and nutrient dosage ratios for cellulosic metabolism to avoid nutrient deprivation. Besides that, cultivation approaches for microalgae associated with mathematical modeling for different parameters, mechanisms of microalgal growth rate and doubling time have been elaborately described. Along with that, this review also documents potential lipid-carbohydrate-protein enriched microalgae candidates for biofuel, biomass productivity, and different cultivation conditions including open-pond cultivation, closed-loop cultivation, and photobioreactors. Various photobioreactor types, the microalgae strain, productivity, advantages, and limitations were tabulated. In line with microalgae cultivation, this study also outlines in detail numerous biofuels from microalgae.     

Microalgae potential as a biogas source: current status,restraints and future trends

In recent years, the world energy demands have had a recurrent increase. For this reason the alternative to the fossil fuel resources are trend topics in investigation. Microalgae have been extensively studied as a source of biofuels and as one of the most promising alternatives in this new framework. One of the possibilities of obtaining renewable energy from microalgae is biogas production using anaerobic digestion process. This process is considered a significant component for biofuels and waste management, since represent an opportunity for energy generation using different wastewater products; also, the economic viability of microalgae liquid biofuel production could be improved. However, the anaerobic digestion of microalgae biomass is still not optimized because of the numerous technical limitations such as the microalgae characteristics, low carbon:nitrogen ratio, ammonia toxicity and even salinity. The present review summarizes and compares information concerning to anaerobic digestion of microalgal biomass and future directions for research. Besides, specific operational factors and potential inhibitory parameters of the process are analyzed and compared. Additionally, the paper covers the state or art concerning in methane production enhancement from algal biomass.     

Lipid Profile Remodeling in Response to Nitrogen Deprivation in the Microalgae Chlorella sp. (Trebouxiophyceae) and Nannochloropsis sp. (Eustigmatophyceae)

Many species of microalgae produce greatly enhanced amounts of triacylglycerides (TAGs), the key product for biodiesel production, in response to specific environmental stresses. Improvement of TAG production by microalgae through optimization of growth regimes is of great interest. This relies on understanding microalgal lipid metabolism in relation to stress response in particular the deprivation of nutrients that can induce enhanced TAG synthesis. In this study, a detailed investigation of changes in lipid composition in Chlorella sp. and Nannochloropsis sp. in response to nitrogen deprivation (N-deprivation) was performed to provide novel mechanistic insights into the lipidome during stress. As expected, an increase in TAGs and an overall decrease in polar lipids were observed. However, while most membrane lipid classes (phosphoglycerolipids and glycolipids) were found to decrease, the non-nitrogen containing phosphatidylglycerol levels increased considerably in both algae from initially low levels. Of particular significance, it was observed that the acyl composition of TAGs in Nannochloropsis sp. remain relatively constant, whereas Chlorella sp. showed greater variability following N-deprivation. In both algae the overall fatty acid profiles of the polar lipid classes were largely unaffected by N-deprivation, suggesting a specific FA profile for each compartment is maintained to enable continued function despite considerable reductions in the amount of these lipids. The changes observed in the overall fatty acid profile were due primarily to the decrease in proportion of polar lipids to TAGs. This study provides the most detailed lipidomic information on two different microalgae with utility in biodiesel production and nutraceutical industries and proposes the mechanisms for this rearrangement. This research also highlights the usefulness of the latest MS-based approaches for microalgae lipid research.     

Best practices in heterotrophic high-cell-density microalgal processes: achievements,potential and possible limitations

Microalgae of numerous heterotrophic genera (obligate or facultative) exhibit considerable metabolic versatility and flexibility but are currently underexploited in the biotechnological manufacturing of known plant-derived compounds, novel high-value biomolecules or enriched biomass. Highly efficient production of microalgal biomass without the need for light is now feasible in inexpensive, well-defined mineral medium, typically supplemented with glucose. Cell densities of more than 100 g l⁻¹ cell dry weight have been achieved with Chlorella, Crypthecodinium and Galdieria species while controlling the addition of organic sources of carbon and energy in fedbatch mode. The ability of microalgae to adapt their metabolism to varying culture conditions provides opportunities to modify, control and thereby maximise the formation of targeted compounds with non-recombinant microalgae. This review outlines the critical aspects of cultivation technology and current best practices in the heterotrophic high-cell-density cultivation of microalgae. The primary topics include (1) the characteristics of microalgae that make them suitable for heterotrophic cultivation, (2) the appropriate chemical composition of mineral growth media, (3) the different strategies for fedbatch cultivations and (4) the principles behind the customisation of biomass composition. The review confirms that, although fundamental knowledge is now available, the development of efficient, economically feasible large-scale bioprocesses remains an obstacle to the commercialisation of this promising technology.     

Microalgal feeds for aquaculture

The concurrent trends of increasing consumption of seafood and decreasing natural harvests will dictate that a larger portion of seafood must be derived from aquaculture in the 21st century. The difficulty of producing economically large quantities of microalgal feeds is currently one of the major impediments to the further development of the aquaculture industry. Traditional methods, which rely on photosynthetic growth in outdoor ponds or indoors under artificial lights, suffer from the phenomenon of light-limitation of biomass density. Certain species of microalgae are capable of heterotrophic growth to high density utilizing sugars or other organic compounds for energy and cell carbon. This paper reviews work with strains of heterotrophic algae that have demonstrated potential as both nutritional feeds and for economical production by fermentation.     

Microalgae mediated photoproduction of beta-carotene in aqueous-organic two phase systems

Improving productivity is a usual requirement for most biotechnological processes, and the utilisation of two-phase aqueous organic systems has proved to be an effective way to improve the productivity of poorly water-soluble or toxic compounds. The high hydrophobicity of beta-carotene, which is highly demanded by the pharma and agrofood industry, makes it a good candidate for aqueous/organic biphasic photoproduction. In the present work we have investigated the viability of a two-phase system for the production of beta-carotene by the marine microalgae Dunaliella salina using decane as organic phase. Decane, with a logP(octanol) value of 5.6, showed no toxicity to Dunaliella cells for more than 72 h, and its ability for beta-carotene extraction is acceptable. Transferring Dunaliella cells from standard to carotenogenic conditions caused inhibition of chlorophyll production and induced a strong synthesis of beta-carotene. The two-phase aqueous/decane system was stable and beta-carotene content of the cells was increasing during 4-days. About 8% of the total carotenoids produced were excreted and extracted into the decane phase.     

Microalgae and Cyanobacteria: How Exploiting These Microbial Resources Can Address the Underlying Challenges Related to Food Sources and Sustainable Agriculture: A Review

For thousands of years, crop production has almost entirely depended on conventional agriculture. However, the reality is changing. The ever-growing population, global climate change, soil degradation and biotic/abiotic stresses are a growing threat to food production and security. Thus, sustainable alternatives to increase crop production for a population projected to reach 9.8 billion by 2050 are a major priority. In addition to vertical and soilless farming, innovative products based on bioresources, including plant growth stimulants, have been a target for sustainable food production. Such solutions have led to the exploitation of microorganisms, including microalgae and cyanobacteria as potential bioresources for food and plant biostimulant products. Microalgae (eukaryotic) and cyanobacteria (prokaryotic) are photosynthetic microorganisms with the capacity to synthesize a vast array of bioactive metabolites from atmospheric CO₂ and inorganic nutrients. The present review outlines the nutritional value of microalgae and cyanobacteria as alternative food resources. The potential aspects of microalgae and cyanobacteria as stabilizers of the net change in soil organic carbon (C) levels for reduced farmland degradation are also highlighted. The applications of microalgae and cyanobacteria as remedies for improved soil structure and fertility, and as enhancers of crop productivity and abiotic stress tolerance in agricultural settings are outlined. This review also discusses the co-cultivation of crops with microalgae or cyanobacteria in hydroponic systems to favor optimum root CO₂/O₂ levels for optimized crop production.