Elsevier

Biomass and Bioenergy

Volume 122, March 2019, Pages 280-289
Biomass and Bioenergy

Research paper
Enhancing algal biomass and lipid production through bacterial co-culture

https://doi.org/10.1016/j.biombioe.2019.01.033Get rights and content

Highlights

  • Screening of green microalgae revealed low lipid and biomass productivities when rendered axenic.

  • Characium sp. 46-4 was selected as a moderate lipid producer with high biomass capabilities.

  • Screening of Characium sp.46-4 with 39 bacteria revealed a bacterium capable of stimulating both algal biomass and lipid content.

  • Algal-bacterial co-cultures success depends on cultivation parameters.

Abstract

Renewable energies, such as biofuels from algae, are a promising approach to deviate from fossil fuels. Mass algae cultivation requires better control over algae culture including taking advantage of co-cultivation with growth and lipid-promoting microorganisms. Integrating co-cultures into algal biotechnology can be beneficial in achieving enhanced yields and reduced expenditures. This work uses the co-cultivation of an alga with bacteria to simultaneously increase biomass and lipid productivities in algal cultures. Growth and lipid-promoting microbial cell-free filtrates were used to test for possible effects of bacterial extracellular compounds. Our results show that both the presence of bacterial cell as well as cell-free filtrate from one particular strain of bacteria, Pseudomonas composti, promoted the increase in biomass yield and lipid within freshwater Characium sp. 46-4. These results indicate that unidentified extracellular compounds released by bacteria can affect the growth rate and lipid metabolism of algae.

Introduction

Procurement and combustion of fossil fuel engenders environmental degradation as well as climate change and is projected to perpetually increase atmospheric carbon concentrations [1]. The increases in atmospheric anthropogenic carbon, pressure on the fossil fuel py, and growing demand for energy have altogether invigorated interest in biologically derived renewable fuel resources; especially microalgae biomass.

Algae are an admirable biofuel feedstock as they have high growth rates, are competent in unusable land and water (deserts, wastewaters, salt waters), and produce a wide range of fuels and by-products (diesel, jet fuel, hydrocarbons, biogas, ethanol, feed, fertilizer, nutraceuticals and pharmaceuticals, etc) [2]. The production of biofuels from algae lipids, however, faces productivity issues that impede scalability. A central issue governing algal monoculture success is that lipid accumulation usually occurs in nutrient depleted conditions. Stress conditions that induce lipid storage in algae, in turn inhibit biomass productivity. A feasible and sustainable microalgae biofuel production requires conditions that allow for both rapid lipid and cell growth rate during cultivation [3].

When cultivating algae for biofuel purposes, high lipid productivities are desired for a feasible process [4]. Optimization of conditions suitable for both high lipid content and biomass may be contradictory as both rely on opposite culture conditions. Factors that affect lipid accumulation such as salts, temperature, light intensity, and growth phase have also been considered as stress factors in optimizing lipid content but not concurrently with biomass [5,6]. With a lack of growth and lipid promoting elements, other methods of employing stimuli in algae to simultaneously increase biomass and lipid productivities remain coveted.

In using algae for biofuel production, one technological aspect is persistently neglected; that is the effect of accompanying bacteria on algae growth and lipid accumulation. Microalgae are abundant in the environment and coexist with a myriad of microorganisms. Symbiotic relationships between algae and microorganisms play key roles in natural ecosystems and are increasingly becoming of interest to industrial microbiology. Symbiotic microorganism relationships are especially useful in applications within the fermented food and water treatment industrial sectors [7]. Application of co-culture or mixtures of microorganisms have also been suggested to facilitate algal biofuels; Co-cultures may be beneficial in the reduction of culture contamination risk and operational cost and may simultaneously provide co-products[8]. In this co-culture scheme, several different microorganisms have been used to stimulate algal cell growth or oil production, including yeast, fungi, bacteria, and cyanobacteria. The details pertaining to the mechanism that facilitates these microbial relationships remain poorly understood.

Phytoplankton and heterotrophic bacteria are among the most abundant microorganisms in aquatic environments [9] and are often co-dependent. In freshwater systems, there is a positive correlation between photosynthesis and microbial activity [10]. The relationship between algae and bacteria becomes apparent when observing the algal organic carbon (DOC) released by senescent algal cells [11]. The release of algal DOC stimulates bacterial growth as the peak of microbial activity usually occurs at the end of algal blooms [12]. In the presence of bacteria, algal DOC reaches a steady state, whereas in axenic cultures, the levels persistently increase [13]. Heterotrophic bacteria can also be associated with living algae cells [12]. The phycosphere of algae is a specialized habitat for some bacteria, as bacterial populations isolated from unattached samples dramatically differ from those attached to algae [14]. In the phycosphere, EPS (Extracellular polymeric substances) from algae constitutes an organic matter base for bacterial decomposition; recognized plant phycosphere symbionts, such as Rhizobium, have already shown growth-promoting effects on algae [15]. Plant growth-promoting bacteria have also been shown to affect microalgae biomass and lipid content remotely through gas and volatile organic compounds exchange [16].

Conversely, bacteria may also stimulate the growth of algae through mutualism and commensalism [17]. With decomposition, bacteria remineralization can account for much of the assimilated organic compounds (especially phosphorus and nitrogen) in phytoplankton [18,19]. When algae demonstrate vitamin auxotrophy, including cobalamin, thiamine, and biotin requirements (vitamin B12, B1, B7, respectively), bacteria are able to supplement these through decomposition in co-culture [20].

Attempts in stimulating biomass of algae with symbiotic or mutualistic bacteria include bacteria-algae consortia and co-cultures [15,21]. Artificially constructed algae consortia with growth-promoting, attached bacteria have indicated significant biomass stimulation [22,23] with implications on cell morphology and lipid content [16,24]. The suggested mechanism for the increase in algae growth is an exchange of algal DOC for bacteria-synthesized dissolved inorganic carbon (DIC) and sugars made available to algae. Screening of growth-promoting bacteria on Dunaliella revealed that algal organic matter recycling by bacteria is a major source of mineralized compounds, especially nitrogen [25]. To further elucidate the nature of bacteria-algae relationships, the effects of a bacterial-cell free filtrate was tested on algae cultures. Park et al. [26] found that bacterial exudates accounted for much of the observed growth increase in the algae. Though it is understood that bacteria synthesize organic substances that may stimulate algal growth, it remains unclear whether these exudates can concurrently stimulate lipid production.

In using algae for production of biofuels, it is essential to choose indigenous algae that have adapted to the local climactic and aquatic conditions which avoids any invasive concerns, and capable of producing high lipid yields, and rapid growth. Cultivation schemes that allow for both high biomass and lipid accumulation are paramount for economic viability of algal biofuel production. In using co-cultures to stimulate the production of algae, we understand that bacteria may stimulate algae biomass, but the stimulus of lipid accretion and the mechanisms in which biomass and lipid content of algae are motivated in response to these microorganisms remain largely obscure.

This work proposes screening for oleaginous, indigenous, South Florida chlorophytes to implement into co-cultures with heterotrophic bacteria in order to augment the production of both biomass and lipids within algae. In investigating this relationship, the cell-free exudates of bacteria were also screened in order to elucidate mechanisms by which microorganisms trigger biomass and lipid productivity within algal cells in co-culture. Developments in ecological engineering, especially of co-cultures, facilitates the use of algae platform for production of the high valued lipid compounds.

Section snippets

Algae

Eight strains of freshwater green microalgae from the FIU culture collection previously designated [27] to be oleaginous and fourteen newly isolated strains from aquatic environments in South Florida, as described below, were used. Additionally, Chlorella sp. 155-1 obtained from the Department of Energy (DOE 1412) and Botryococcus sp. 157-1 obtained from UTEX culture collection (UTEX 572) were used as reference strains.

Isolation and purification of algae

Ten water samples from across South Florida were collected from freshwater

The freshwater microorganisms used

From the 10 South Florida freshwater samples, a total of 14 microalgae species were isolated and together with 8 cultures from our culture collection and two from UTEX and DOE (Table 1) were screened for biomass and lipid productivity. The microalgae strains represented 11 genera within Chlorophyceae (Table 1).

Biomass and lipid productivity screening of microalgae

Results from the studies involving screening of 22 strains of chlorophytes showed wide variation among strains in biomass and lipid accumulation capabilities. The average biomass content

Acknowledgments

This work was partly supported by the USDA National Institute of Food and Agriculture Hatch project # FLA-FTL-005697.

References (46)

  • M. Do Nascimento et al.

    High lipid productivity of an Ankistrodesmus–Rhizobium artificial consortium

    Bioresour. Technol.

    (2013)
  • R. Ramanan et al.

    Phycosphere bacterial diversity in green algae reveals an apparent similarity across habitats

    Algal Res

    (2015)
  • IPCC
  • M. Hannon et al.

    Biofuels from algae: challenges and potential

    Biofuels

    (2010)
  • J.M. Griffiths et al.

    Lipid Productivity as a key characteristic for choosing algal species for biodiesel production

    J. Appl. Phycol.

    (2009)
  • Q. Hu et al.

    Microalgal triacylglycerols as feedstocks for biofuel production: perspectives and advances

    Plant J.

    (2008)
  • A.M. Silva-Benavides et al.

    Nitrogen and phosphorus removal through laboratory batch cultures of microalga Chlorella vulgaris and cyanobacterium Planktothrix isothrix grown as monoalgal and as co-cultures

    J. Appl. Phycol.

    (2011)
  • G. Padmaperuma et al.

    Microbial consortia: a critical look at microalgae co-cultures for enhanced biomanufacturing

    Crit. Rev. Biotechnol.

    (2018)
  • J.J. Cole

    Interactions between bacteria and algae in aquatic ecosystems

    Annu. Rev. Ecol. Systemat.

    (1982)
  • J.E. Hobbie et al.

    Radioisotope studies of heterotrophic bacteria in aquatic ecosystems

  • W.H. Bell

    Bacterial utilization of algal extracellular products. 3. The specificity of algal-bacterial interaction

    Limnol. Oceanogr.

    (1983)
  • J.G. Jones

    Studies on freshwater bacteria: association with algae and alkaline phosphatase activity

    J. Ecol.

    (1972)
  • W.J. Wiebe et al.

    Direct measurement of dissolved organic carbon release by phytoplankton and incorporation by microheterotrophs

    Mar. Biol.

    (1977)
  • Cited by (46)

    • Microalgae biorefinery: An integrated route for the sustainable production of high-value-added products

      2022, Energy Conversion and Management: X
      Citation Excerpt :

      Additionally, research into co-culturing or microbial mixtures has been proposed as a means of facilitating algal for many biorefineries [63]. The co-cultures may be advantageous in lowering the risk of culture contamination and operational costs while simultaneously producing by-products using organisms like yeast, fungi, bacteria, cyanobacteria, etc. [64]. Table 1 provides a summary of the development in the application of some recent technologies and microorganisms for the recovery of microalgae biomass.

    View all citing articles on Scopus
    View full text