Dipartimento di Biologia (BIO) | UniFI - The multi-level ......Elena Perrin, Michele Giovannini, Francesca Di Patti, Barbara Cardazzo, Lisa Carraro, Paola Turano, Veronica Ghini, Renato

BackgroundResults

ConclusionsAcknowledgements

The multi-level relationship betweenheterotrophic bacteria and nutrients

Diauxie and co-utilization are not exclusive during growth innutritionally complex environments

Elena Perrin, Michele Giovannini, Francesca Di Patti, Barbara Cardazzo, Lisa Carraro, PaolaTurano, Veronica Ghini, Renato Fani, Marco Fondi

[email protected]

FEMS 2019, Glasgow, July 9, 2019

Marco Fondi The multi-level relationship between heterotrophic bacteria and nutrients

BackgroundResults


The classic view of microbial growth strategy when multiple carbonsources are available states that they either metabolize them

sequentially (diauxic growth) or simultaneously (co-utilization).

”All you can eat” vs. ”a la cart” strategies result in different growthdynamics (Monod, 1942).


BackgroundResults


Diauxic shift is commonly seen as a phase in which the bacteria preparethemselves to use the second sugar. The existence of two stable cell typeswith alternative metabolic strategies emerge and coexist in a culture of the

bacterium has been demonstrated.


BackgroundResults


Our knowledge is biased by the fact that this process has been mainlyanalyzed in over-simplified laboratory settings, i.e. using a few modelmicroorganisms and growth media containing only two alternativecompounds.

The marine environment:

• Low average nutrient level (e.g., the concentration of amino acids is inthe range of about 10−9 M)

• Nutrients appear and disappear in a sporadic fashion.

• DOM as a complex medium


BackgroundResults


• Chemotaxis ofPseudoalteromonashaloplanktis TAC125(PhTAC125) toward a pulse ofphytoplankton exudates(Stocker et al. 2010, Science)

• What happens once the patchis colonized?


BackgroundResults


Growth on complex mediumAA metabolismGrowth on a 19 AA mediumA growth mathematica model

We grew PhTAC125 in a complex medium (peptone, 7 replicates) andanalysed its growth features.


BackgroundResults



• High expression of growth-related genes in the first hours• Stress related genes up between T1 and T2

• Motility-related genes up at the end of the growth


BackgroundResults



A non E. coli-like response to growth lag• 81 TF known to dcontrol central

metabolic enzymes in E. coli

• Homologs for 34 of them inPhTAC125, 8 of them differentiallyexpressed (10%)

• Only RpoS and RpoD aredifferentially expressed among 8selected E. coli global regulators


BackgroundResults



A focus on amino acids degradation. PhTAC125 is know to thrivepreferentially on amino acids (Wilmes et al. 2010).


BackgroundResults



We assembled a defined, nutritionally rich medium including 19 amino acids ([0.2 mM],Cys not included) and analysed growth and metabolic features.

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BackgroundResults




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argasnaspglngluglyhisisoleulysmetpheproserthreotrptyrval

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BackgroundResults




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BackgroundResults




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Glu Asp Leu Ala

Gro

wth

rate

(h-1)C1

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*

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E

B D F Order of consumption


BackgroundResults



What is regulating the system?

• Uptake kinetics (no regulation, dynamics explained by differentuptake kinetics).

• Active regulation The uptake of the different groups is somehowregulated.


BackgroundResults




• Uptake kinetics (no regulation, dynamics explained by differentuptake kinetics).

P + AβA ·φAkA+φA−−−−→ 2 P

P + BβB ·φBkB+φB−−−−→ 2 P

P + CβC ·φCkC+φC−−−−→ 2 P

P + DβD ·φDkD+φD−−−−→ 2 P 0.00

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AB

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BackgroundResults



P + AβA ·φAkA+φA−−−−→ 2 P

P + BβB ·φBkB+φB−−−−→ 2 P

P + CβC ·φCkC+φC−−−−→ 2 P

P + DβD ·φDkD+φD−−−−→ 2 P

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(660

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BackgroundResults




• Active regulation The uptake of the different groups issomehow regulated. A cybernetic model (Kompala et al. 1994)is implemented.

P + A E1, ρ1−−−→ 2 PP + B E2, ρ2−−−→ 2 PP + C E3, ρ3−−−→ 2 PP + C E4, ρ4−−−→ 2 P

C A, λ1−−−→ C + E1

C B, λ2−−−→ C + E2

C C, λ3−−−→ C + E3

C D, λ4−−−→ C + E4

dφEi

dt= λi∗u1−(ρ1∗v1+ρ2∗v2+ρ3∗v3+ρ4∗v4)∗φEi−βφEi

dφP

dt= (ρ1 ∗ v1 + ρ2 ∗ v2 + ρ3 ∗ v3 + ρ4 ∗ v4) ∗ φP − αφP

dφSi

dt= ρi ∗ vi ∗ φP

With i = 1,2,3,4 and S = {A, B, C, D}


BackgroundResults



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MM CyberneticBiomass 0.997 0.994Cluster A 0.9893 0.9983Cluster B 0.9967 0.9977Cluster C 0.9947 0.9904Cluster D 0.9723 0.9908

R2 calculation of the fit


BackgroundResults


• PhTAC125 growth on complex media results in a triauxic growthwith a regulation response to nutrient starvation that does notresemble the model one.

• Growth lag phases are due to nutrient switching.• Sequential and co-utilization are not exclusive. Most efficient C

sources are consumed first.• A simple mathematical model based on Michaelis-Menten

kinetics uptake does not result in a perfect fit with the data.Regulatory mechanisms must be included to explain this growthdynamic at the single cell level

• Emergent sub-populations?


BackgroundResults


DBIO, Unifi

• Elena Perrin

• Prof. Renato Fani

• Prof. Alessio Mengoni

• Camilla Fagorzi

• Michele Giovannini

CERM, Unifi

• Veronica Ghini

• Prof. Paola Turano

CSDC, Unifi

• Francesca Di Patti

Unipd

• Barbara Cardazzo

• Lisa Carraro


Documents

Dipartimento di Biologia (BIO) | UniFI - The multi-level ......Elena Perrin, Michele Giovannini, Francesca Di Patti, Barbara Cardazzo, Lisa Carraro, Paola Turano, Veronica Ghini, Renato