1. E. coli evolution in low radioactivity conditions
Pierre Micheau, Alexandre Taravaud, Thomas Hindré, Lydia Maigne,
Michel Zampaolo, Fabrice Piquemal, Vincent Breton

2. Study of long term evolution and evolution pathways
Evolution is linked to mutation events in genes and genome
Natural mutation (error prone during replication)
Induced mutation (caused by chemical alterations, radiation)
Does the level of radiation in the environment affect the evolution process ?
Use of experimental evolution with Escherichia coli
No pathogenic
Easy to cultivate
Short generation time (30-60 min)
Large population sizes
Cellular process well-known
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

3. Richard Lenski’s experimental evolution
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Richard Lenski
Michigan State Univ. -1 -2 -3 -4 -5 -6 +1 +2 +3 +4 +5 +6
Ancêtre
E. coli B
Ara-
Ara+
-80°C
> generations
1988
2012…….
Daily transfer in the same glucose limited medium
12 populations at the same time
Populations are sampled every 500 generations and conserved as a frozen fossil that can be revivified
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

4. Richard Lenski’s experimental evolution
Direct competition between ancestor and evolved bacteria to measure adaptation : Relative Fitness
Which grows up the quickest ? ancestor or evolved bacteria
Ancestor
Ara-
Evolved
Ara+
Fitness values were measured by placing evolved population in competition with the ancestral strain in the same environment used for the experimental evolution. Fitness is expressed relative to the common ancestor, and it is calculated simply as the ratio of the growth rates realized by the two competitors during their head to head competition.
24h, 37°C
Relative Fitness increases in the 12 evolved populations
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

5. Richard Lenski’s experimental evolution
Cell volume measurement for ancestral and evolved strains
Volume increases in the 12 evolved populations
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

6. Start experiments in three differents environments
Does the level of radiation in the environment impact the evolution process ?
Start experiments in three differents environments
Underground Laboratory of Modane (LSM) as « zero » radiation (around 5nSv/h) – Done
Laboratory of Corpuscular Physics (LPC) around 150nSv/h – in progress
LPC around 150nSv/h + 0.5Gy/week – not start
Protocole is similar to Lenski’s one
Same bacteria E.coli B Ara- , Ara+
12 replicates of Ara-, 12 replicates of Ara+
37°C – Daily transfer in limited glucose medium
8.23 generation per day
Biological data used for modeling bacterial evolution
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

7. Experimental evolution at LSM
Culture at LSM
No radiations come from cosmos, sun and electronic waves
Protected from others telurics radiations thanks to the lead castle
Traditional culture conditions (37°C, agitation)
Cosmics radiations
Telurics radiations
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

8. Experimental evolution at LSM
E. coli populations have been propagated for 200 Generations (8000 years at the human scale) – 45 days experiment
Comparison of 200 Generations evolved E. coli at LSM to ancestor
Fitness test
40Gy survival rate
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

9. Experimental evolution at LSM
Fitness test
Ancestor
Ara-
Evolved
Ara+
24h, 37°C
 Fitness around 1 – No increase of fitness for evolved E.coli
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

10. E. Coli behavior after high dose radiation
Purpose
Low radiation environment = less oxydative damages on E.coli genome
Are evolved bacteria lost capacity to fight against oxydative damages during strong irradiation ?
High dose radiation using medical linac
18 MV photon beam
Dose = 40 Gy
Tests and sample analysis:
Survival rates of two evolved and ancestral E. coli
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

11. E. Coli behavior after high dose radiation
Results:
Ancestor and evolved E.coli have the same resistance to irradiation
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

12. Conclusions - Perspectives
E. coli populations have been evolved during 200 generations in the low radiation environment (LSM)
Same fitness between ancestor and evolved E.coli
Same survival rate to short strong irradiation
200 generations in LSM environment not enough to characterize modifications of evolution process
Propagated E.coli during more generations at LSM
TO DO NEXT: Comparisons between E. coli in natural radiation exposure and E. coli in low radiation environement
Fitness measurement
40Gy survival rate
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012

13. Acknowledgements Fabrice Piquemal Michel Zampaolo Guillaume Warrot
Thomas Hindré
Vincent Breton
Lydia Maigne
Alexandre Taravaud
Pierre Micheau - ASPERA conference - E. coli evolution in low radioactivity conditions - december 2012