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RÉPUBLIQUE
FRANÇAISE

Liberté
Égalité
Fraternité

INRAE

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BORDEAUX
METABOLOME

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METABOHUB

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UNIVERSITÉ
Clermont
Auvergne

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# New insights into the influence of temperature and nutrients on the growth of *Microcystis* sp. and the metabolism of its phycosphere thanks to metabolomics

Thomas Forjonel,¹,² Sylvia Moreira,¹,² Charlotte Duval,³ Melissa Eon,¹,² Benjamin Marie,³ Binta Diémé,⁴ Nicolas Creusot¹,²

1 INRAE, UR1454 EABX, Gazinet Cestas, France, 2 Bordeaux Metabolome, MetaboHub, PHENOME-EMPHASIS, Villenave d'Ornon

3 Muséum National d'Histoire Naturelle, UMR 7245 CNRS-MNHN, MCAM, Paris, France, 4 Université Clermont Auvergne, INRAE, UNH, Plateforme d'Exploration du Métabolisme

## Context

* ✓ **Cyanobacterial blooms** are a phenomenon of growing international concern, due to their potential ecological, human and economic impacts [1]

* ✓ While certain **abiotic factors** are known to contribute to these phenomena, the underlying **ecological and physiological mechanisms** remain only partially understood [2].

* ✓ Recently, some studies have highlighted the potential role of **biotic interactions between heterotrophic bacteria and cyanobacteria** in the **phycosphere**, a microscale environment surrounding cyanobacterial cells [3].

* ✓ Omics approaches, particularly metabolomics and metagenomics, offer powerful tools to unravel the complex chemical and microbial interactions shaping blooms.

* ✓ In this context, the COMIC project aims to better understand the role of **cross-feeding** metabolic interactions between *Microcystis* sp. and its heterotrophic bacteria by combining **metabolomic, metagenomic and modeling strategies**. To do so, a first objective was to compare the metabolism of 12 Microcystis strains physcosphere and associated metabolic interaction. The second objectives was to investigate the influence of environmental factors in the metabolism and growth of one of the strain.

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## The phycosphere

The phycosphere can be characterized as the **microenvironment surrounding cyanobacteria**, a restricted zone of the order of a few tens of microns, but one that is biologically very active.

<figure><img src="image_8.png" /><figcaption>Cyanobacteria</figcaption></figure>

<figure><img src="image_9.png" /><figcaption>Associated bacteria
in the phycosphere</figcaption></figure>

## Aim

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This study aims to characterize the influence of temperature and nutrient content on the metabolism and the growth of *Microcystis aeruginosa* and associated heterotrophic bacteria in one selected strain and further contribute to elucidate cross-feeding interaction within the phycosphere

## Materials and methods

### Sampling and cultivation

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### Biochemistry -> Metabolomic (UPLC-HRMS-MS)

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#### Endometabolome (cells)

Characterization of **intracellular** metabolites to assess the impact of temperature and nutrients on metabolism

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#### Exometabolome (medium)

Profiling of **extracellular** metabolites likely involved in **microbial interactions** and **cross-feeding dynamics**.

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Identification of metabolites related to temperature/nutrients acclimatization

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Environmental regulation of microcystins production

### Workflow & expected outcomes

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#### Biomass & Physiology

<figure><img src="image_23.png" /><figcaption>Optical density</figcaption></figure>

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Cyanobacteria
growth kinetic

<figure><img src="image_25.png" /><figcaption>PhytoPAM®</figcaption></figure>

Photosynthetic yield
and chlorophyll a

<figure><img src="image_26.png" /><figcaption>Cell density</figcaption></figure>

<figure><img src="image_27.png" /><figcaption>Ash Free dry weigh</figcaption></figure>

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### Cross-validation

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Further implemented in collab.

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### Nutrient monitoring (ionic chromato.)

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Weekly water sampling is performed to quantify nutrient concentrations. Anions ($NO_3^-$, $NO_2^-$, $PO_4^{3-}$) and cations ($NH_4^+$, $Mg^{2+}$) are measured to characterize the experimental trophic conditions.

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#### Metagenomics

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Whole-genome sequencing to build **Genome scale Metabolic Networks** and **predict metabolic interaction** of *Microcystis aeruginosa* and its microbial consortium.

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#### Metabarcoding

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Taxonomic composition of bacterial communities associated with *Microcystis* using 16S rRNA gene sequencing.

**References:**

[1] Igwaran et al. (2024). *Cyanobacteria Harmful Algae Blooms: Causes, Impacts, and Risk Management*. https://doi.org/10.1007/s11270-023-06782-y

[2] Pal, M., Yesankar, P. J., Dwivedi, A., & Qureshi, A. (2020). Biotic control of harmful algal blooms (HABs): https://doi.org/10.1016/j.jenvman.2020.110687

[3] Louati, I. et al. (2023). The phyto-bacterioplankton couple in a shallow freshwater ecosystem: Who leads the dance? *Harmful Algae*, 126, 102436.

https://doi.org/10.1016/j.hal.2023.102436

**Acknowledgment**

The project COMIC is funded by the INRAE metaprogram HOLOFLUX