Plateforme intégrée pour l'étude de la filière microalgue - Région wallonn

Van Camp, Camille

Wattiez, Ruddy

ORBi UMONS

EXPLORATION OF EXOPOLYSACCHARIDE PRODUCTION BY CYANOTHECE SP. PCC 7822 Camille Van Camp and Ruddy Wattiez Proteomic and Microbiology laboratory, UMONS, Mons, Belgium The Algotech project was established to progress in the field of high-added value compounds production by Microalgae/Cyanobacteria completing by a circular economy consideration. In this way, we investigate the exopolysaccharide (EPS) production of Cyanothece sp. PCC 7822, well-known for its diazotrophic metabolism. Impact of C/N ratio and N source on EPS configuration and composition will be explored. Variation of N sources and concentrations are relevant because of their influence on C:N ratio and carbon metabolism. Interestingly, modification of this parameter has an impact on Cyanothece sp. PCC 7822 growth but also on EPS production and configuration. Projected experiments will go further to better understand the effect on EPS composition and structure. A mass spectrometry analysis of the protein profil is also planed to improve knowledge of cyanobacterial EPS biosynthesis pathways. This research is part of the AlgoTech project (convention 1510612 – Algotech) and supported by Walloon Region. I thank Patrick Flammang and Igor Eeckhaut from the Biology of Marine Organisms and Biomimetics Unit for the use of the fluorescence microscope.  Corresponding author : camille.vancamp@umons.ac.be A significant quantity of EPS are spotted in NaNO3 conditions (Fig 2A, 2B). The absence of growth with NH4Cl and urea 17 mM correlated with microscopy observation suggests an EPS formation as a solution to prevent the stress caused by N sources (Fig 2C, 2E). EPS configuration in these 2 conditions is also contrasting with NaNO3 conditions. In atm. N2 condition, EPS are dispersed in the medium, characterised by a visquous culture (Fig 2G).  EPS seems more abundant in urea 1 mM condition that in urea 17 mM (Fig 2E, 2F) whereas the opposite observation is done in NH4Cl condition (Fig 2C, 2D). As shown in Fig 1, the best growth is noticed in NaNO3 condition independently of the concentration. Contrarily with literature, Cyanothece grows in presence of NH4Cl at 1 and 2.5 mM. In urea condition, optimal growth is determined at 2.5 mM. Even if the strain is able to fix atmospheric N2, growth is not as best as in NaNO3 condition. It is interesting to highlight different growth profil of Cyanothece  considering a potential relation with EPS production. As an example, environmental stress could induce an EPS production by bacteria. In presence of 1 mM of N sources, the C:N ratio increases and carbon excess could be drived in EPS production even if growth is reduced. Conclusion Fig. 3 : Total carbohydrate quantification of exopolysaccharides. RPS represent the released polysaccharides and CPS represent the capsular polysaccharides (attach to cell surface), D-glucose is used for standard curve and values are standardised by chlorophyll a concentration NaNO3 17 mM Atmospheric  N2 Modification of N sources & C:N ratio Fig. 3 reveals that RPS are dominant in NaNO3 condition representing 80% of total EPS quantified (RPS + CPS). Despite the concentration fluctuation during the growth, the ratio between RPS and CPS remains stable. Globally, less EPS are formed in atm. N2 condition and the ratio is also different with a stable RPS concentration and an increase of CPS produced over the growth. Surprisingly, The maximum EPS obtained is 33 µg/mL  at the end of the growth in atm. N2 condition.  Future acid uronic and sulphated sugar quantification will help us to select various conditions for a monosaccharide composition investigation. Fig. 2 : Observation of Cyanothece sp. PCC 7822 exopolysaccharides by using a positive alcian blue staining at pH 2.5  in  presence of 4 different nitrogen sources at different concentrations.  Experiments were performed in 4 replicates and over the growth. NaNO3 atm,. N2 NH4Cl Urea C 17 mM 1 mM A C D F E G B Introduction NaNO3 & atm N2 17 mM in term of N NH4Cl Urea 2,5 mM in term of N 1 mM in term of N Fig. 1 : Growth curve of Cyanothece sp. PCC 7822 in BG 11 medium supplemented with different nitrogen sources at different concentration(NaNO3, NH4Cl, urea at 1, 2.5 and 17 mM and atmospheric N2). Concentrations are defined in term of N atom. The experiment was realised in 4 replicates.  

Exploration of exopolysaccharide production by Cyanothece sp. PCC 7822

https://orbi.umons.ac.be/bitstream/20.500.12907/31276/1/poster.pdf

Exploration of exopolysaccharide production by Cyanothece sp. PCC 7822

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