Elsevier

Bioresource Technology

Volume 281, June 2019, Pages 56-65
Bioresource Technology

Bacterial community enhances flocculation efficiency of Ettlia sp. by altering extracellular polymeric substances profile

https://doi.org/10.1016/j.biortech.2019.02.062Get rights and content

Highlights

  • Self-flocculating Ettlia sp. has high extracellular protein/carbohydrate ratio (EPSP/EPSC).

  • Growth-promoting and flocculation-inducing bacterial groups were detected.

  • Filamentous EPS matrices connecting cells appeared exclusively in xenic culture.

  • More compact EPS structure was introduced by the bacterial community.

  • Bacteria was not decisive but facilitating factor for Ettlia flocculation.

Abstract

This study examined the effects of a bacterial community and extracellular polymeric substances (EPS) on Ettlia sp. flocculation. The growth rate, flocculation efficiency (FE), bacterial community, and EPS profile of axenic and xenic Ettlia cultures were monitored during 46 days of cultivation. For the xenic culture, with a great abundance of growth-promoting and flocculation-inducing bacteria, the biomass density was 18.75% higher and its FE reached 100% in the mid-stationary phase. Moreover, microscopic observation and a quantitative analysis of the EPS revealed the exclusive presence of long filamentous EPS and more compact structure in the xenic Ettlia culture, possibly explaining its better FE. Notwithstanding, for the axenic culture, despite a lower biomass density and reduced abundance of EPS, its FE reached 92.54% in the mid-stationary phase. Thus, the role of the bacterial community was found to be supportive rather than vital for the high settleability of the self-flocculating Ettlia microalgal culture.

Introduction

The advantages of microalgae have already been recognized in a wide variety of applications (Pulz and Gross, 2004). In the case of biodiesel, when compared with terrestrial plants, microalgae grow about 10–15 times faster with approximately 10–20 times higher productivity (Wan et al., 2015, Wang et al., 2016). The use of microalgae to produce pharmaceutical and other high-value products by manipulating the cultivation conditions is also very promising. However, due to the small size of microalgae, their identical density with water, and the diluted concentrations of conventional suspension cultivation systems (0.5–10 g/L), the harvesting process—which accounts for roughly 20–30% of the total production cost—still remains the bottleneck for microalgal biotechnology (Chisti, 2007, Wan et al., 2015). Currently, centrifugation, filtration, flotation, and gravity sedimentation are the main harvesting techniques, among which the most popular ones are centrifugation and sedimentation.

Flocculation before harvesting, which can be induced by physical, chemical or bio-based flocculation have been reported to save a significant amount of time and energy (Vandamme et al., 2013). However, some of these flocculation processes might contaminate the biomass and are not environment friendly or economically feasible (Wan et al., 2015).

Currently, the idea of using self-flocculating microalgal species for harvesting the non-flocculating oleaginous cells has achieved great attention (Salim et al., 2012). Possible application in the field of water bloom treatment might also be promising, although no lab-scale research has been conducted yet. Till now, only a few self-flocculating microalgae species, such as Chlorella vulgaris JSC-7, Scenedesmus obliquus AS-6-1, Ettlia texensis, Ankistrodesmus falcatus, Tetraselmis suecica, and Chlorococcum sp. GD have been reported (Alam et al., 2014, Guo et al., 2013, Lv et al., 2016, Salim et al., 2014, Salim et al., 2013, Salim et al., 2012). Researches have implied the role of released extracellular polymeric substances (EPS) or algal organic matter (AOM) in the abovementioned auto-flocculating microalgal species; however, culture axenicity has been overlooked. Without the precise acknowledgment of bacterial role, the auto-flocculation behavior or the real producer of the bioactive EPS might be misleading (Vu et al., 2018).

Accordingly, this study examined whether the bacterial community or EPS play a vital role in the previously detected strong settling properties of Ettlia sp. YC001 (La et al., 2016). EPS can be categorized into soluble (S-) and cell surface bound (B-), and the latter further divided into outer loosely bound (LB-) and inner tightly bound (TB-) layers (Sheng et al., 2010). Since the B-EPS accumulate on the cell surface and directly change cell surface characteristics—such as surface hydrophobicity, charge density, and binding sites—only this fraction was studied in this research. The next generation sequencing (NGS) method was used to investigate the bacterial communities of Ettlia culture during different growth phases. The B-EPS of bacteria-free and xenic microalgal cultures were extracted and their compositions investigated. Thereafter, flocculation efficiency (FE) of the two Ettlia cultures were measured and explained based on the bacterial community and EPS profile.

Section snippets

Microalgal strain and cultivation conditions

The microalgal Ettlia sp. YC001 strain (KCTC 12109BP) was obtained from the Korean Collection for Type Cultures (KCTC) at the Korea Research Institute of Bioscience and Biotechnology. A xenic seed culture was maintained in the steady state as previously described (Seo et al., 2017). An axenic culture was then isolated and confirmed using several enriched agar media and scanning electron microscopic (SEM) observation (Vu et al., 2018). For easy detection of contamination during culture

Ettlia sp. cultures: growth and SEM observation

The growth of axenic and xenic Ettlia cultures were quite identical in the first 9 cultivation days with a short lag phase of approximately 5 days, followed by 11 days of an exponential growth phase (Fig. 1). The biomass content in the axenic and xenic cultures reached a maximum of 2.40 ± 0.45 g/L and 2.85 ± 0.70 g/L, respectively, at the beginning of the stationary phase. This 18.75% increase in biomass density was well-agreed with some previous researches in diatom and microalgae cultures.

Conclusions

Self-flocculating Ettlia sp. possesses a distinctly high EPSP/EPSC. A bacterial community is not mandatory, yet a facilitating factor for Ettlia sp. flocculation. In this study, the flocculation of the axenic Ettlia occurred via a patching mechanism mediated by small, dust-like EPS particles. The exclusive long filamentous EPS structure established by the bacterial community further increased the flocculation efficiency via an additional bridging mechanism. The bacterial community also

Acknowledgements

This work was supported by the Advanced Biomass R&D Center (ABC), a Global Frontier Program funded by the Korean Ministry of Science and ICT (2010-0029723).

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