# Gene flow favours local adaptation under habitat choice in ciliate microcosms

Staffan Jacob, Delphine Legrand, A. S. Chaine, Dries Bonte, Nicolas

Schtickzelle, Michèle Huet, Jean Clobert

► To cite this version:

Staffan Jacob, Delphine Legrand, A. S. Chaine, Dries Bonte, Nicolas Schtickzelle, et al. Gene flow favours local adaptation under habitat choice in ciliate microcosms. Nature Ecology & Evolution, 2017, 1 (9), pp.1407-1410. 10.1038/s41559-017-0269-5. hal-04967687

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Distributed under a Creative Commons Attribution 4.0 International License
1 Gene flow favours local adaptation under habitat choice in ciliate

2 microcosms

3

4 Staffan Jacob*¹,², Delphine Legrand¹, Alexis S. Chaine¹,³, Dries Bonte⁴, Nicolas

5 Schtickzelle², Michèle Huet¹, Jean Clobert¹

6

7 * Corresponding author

8

9 ¹Station d'Ecologie Théorique et Expérimentale du CNRS UMR5321, Evolutionary Ecology

10 Group, 2 route du CNRS, 09200 Moulis, France

11 ² Université Catholique de Louvain, Earth and Life Institute, Biodiversity Research Centre,
12 Croix du Sud 4, L7.07.04, 1348 Louvain-la-Neuve, Belgium

13 ³ Institute for Advanced Studies in Toulouse, Toulouse School of Economics, 21 allée de
14 Brienne, 31015 Toulouse, France

15 ⁴ Ghent University, Terrestrial Ecology Unit, Department Biology, K.L. Ledeganckstraat 35,
16 B-9000 Ghent, Belgium

17 Staffan Jacob: jacobstaffan@gmail.com;

18 Delphine Legrand: delphine.legrand@sete.cnrs.fr;

19 Alexis S Chaine: alexis.chaine@sete.cnrs.fr;

20 Dries Bonte: Dries.Bonte@UGent.be;

21 Nicolas Schtickzelle: nicolas.schtickzelle@uclouvain.be;

22 Michèle Huet: michele.huet@sete.cnrs.fr;

23 Jean Clobert: jean.clobert@sete.cnrs.fr;

24

25 Word count: 1632
26

**Abstract**

27 Local adaptation is assumed to occur under limited gene flow. However, habitat-matching
28 theory predicts dispersal should favour rather than hinder local adaptation when individuals
29 selectively disperse towards habitats maximizing their performance. We provide experimental
30 evidence that local adaptation to the upper margin of a species' thermal niche is favoured by
dispersal with habitat choice, while hindered under random dispersal. Our study challenges
31 the idea that high gene flow precludes local adaptation, and provides unique experimental
32 evidence of habitat choice as an overlooked mechanism responsible for adaptation under rapid
33 environmental changes.
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Classical evolutionary theory predicts that moderate to high gene flow in heterogeneous environments should hinder local adaptation and restrict speciation by homogenizing local gene pools¹,². In contrast, recent theory has shown that dispersal, the movements potentially leading to gene flow³, could facilitate local adaptation when involving habitat choice⁴⁻⁸. When individuals either stay in or disperse towards habitats that maximize their performance, the resulting gene flow would involve a non-random subset of genotypes that are locally adapted⁵⁻⁷,⁹. In contrast to the common view, even high dispersal rates might facilitate rather than hinder local adaptation⁴⁻⁸,¹⁰. By generally assuming random gene flow, we might deeply misjudge the consequences of dispersal for eco-evolutionary dynamics and underestimate species adaptive potential facing environmental changes⁴⁻⁷,¹⁰,¹¹. Despite the major impact habitat choice could have on local adaptation and the extensive empirical evidence for non-random phenotype- or context-dependent dispersal¹², experimental validation of this theoretical framework is lacking.

We experimentally test whether local adaptation can occur under high gene flow if directed by habitat choice using microcosms of *Tetrahymena thermophila*, a ciliate that performs active dispersal and habitat choice¹³,¹⁴. We focussed on adaptation to temperature at the upper margin of the species' thermal niche where growth rate is significantly reduced. Temperature is an environmental factor of great importance to the fitness of many organisms, and quantifying adaptive potential at the margins of a species' thermal niche is highly relevant in the context of current global environmental changes and species range shifts. Furthermore, this model species that naturally lives under thermal spatiotemporal variability in freshwater ponds shows genetic variability in performance along a thermal gradient (Supplementary Figure 1), a condition required for habitat choice to lead to non-random gene flow⁵,⁷.

To quantify habitat choice effects on local adaptation, we first tested whether individuals choose their habitat to match their thermal performance ability using *T. thermophila*
maintained in axenic liquid media (see Methods). Genetically variable populations composed
of 10 clonal genotypes showing variation in performance along a temperature gradient
(Supplementary Figure 1) were used to inoculate the central patch of linear 3-patch dispersal
systems and allowed to disperse toward the two adjacent empty patches (Supplementary
Figure 2a). Temperature in the central and neighbouring patches were set to either 23°C
(thermal niche core; Supplementary Figure 1) or 35°C (upper thermal niche margin) to test
for habitat choice at emigration (stay or disperse through the corridors) and immigration (join
one of the neighbouring patches; Supplementary Figure 2a). Single patch systems were
inoculated and kept either at 23°C or 35°C as controls where cells had no possibility to
disperse. After 24 hours, we transferred cells from each patch into fresh media at 35°C and
quantified growth rate as a measurement of fitness at the upper thermal niche margin
(Supplementary Figure 2a).

Our results provide support for the habitat matching hypothesis: individuals that chose to stay in or join the thermal niche margin (*red points*; Fig 1) had significantly higher growth rates at 35°C than those that chose to stay in or join 23°C patches (*blue points*; Fig 1; ANOVA: F₁,₃₄=12.66; p=0.001). Alternative hypotheses such as selection, plasticity or drift cannot explain these results (see Supplementary Material). For instance, rapid selection on a subset of genotypes that better fit the local temperature would have led to higher fitness at 35°C for cells from isolated 35°C patches than isolated 23°C patches (i.e. no dispersal allowed). However, individuals kept at either 23°C or 35°C in isolated patches did not significantly differ in growth at 35°C (*grey points*; Fig 1; F₁,₁₀=0.09; p=0.77). Our results thus demonstrate that cells showed increased growth rate at 35°C because they preferred to stay in or join the temperature that matched their thermal performance.

When individuals selectively disperse towards habitats that maximize their performance,
dispersal is predicted to favour rather than hinder local adaptation<sup>4–7,10</sup>, but this hypothesis
remains unexplored experimentally. We performed a long-term selection experiment to test
the effects of dispersal with habitat choice compared to random dispersal on local adaptation
at the upper margin of the species' thermal niche (Supplementary Figure 2b). We set up large
genetically variable populations (i.e. mixes of the 10 genotypes; ~250000 cells total)
maintained at 35°C for ~250 generations (6 weeks; 2-5h generation time; 5% weekly transfer
to fresh media; Supplementary Figure 2b) and receiving immigrants weekly from the initial
genetically variable populations, mimicking regular movement of immigrants from core to
marginal habitats. Immigrants came from a weekly reiteration of the first experiment,
generating immigrants for either random or habitat choice dispersal. A random dispersal
treatment consisted of introducing as immigrants a random subset of initial populations from
isolated control patches (“no dispersal”; Supplementary Figure 2a). Habitat choice treatments
consisted of cells who chose a 35°C patch (stayed at 35°C or joined 35°C from a 35°C or
23°C origin; Supplementary Figure 2a red patches). Control choice treatments consisted of
cells who chose a 23°C patch (Supplementary Figure 2a blue patches).

Adaptation to local conditions was estimated once a week by quantifying growth rate at
35°C of a sample of cells from each population placed in fresh media, and through “local
versus foreign” and “home versus away” approaches¹,¹⁵. Foreign populations consisted of the
same initial genetically variable populations but maintained at 23°C instead of 35°C without
immigration. Furthermore, because the intensity of gene flow should affect its consequences
for local adaptation¹,¹⁶, we conducted the whole experiment at two levels of dispersal: 10%
and 30% of the local population’s initial carrying capacity. Additional controls for temporal
changes in fitness consisted of genetically variable populations maintained at 23°C and
receiving either random dispersal or habitat choice dispersers from 23°C to 35°C.

Our results show that dispersal with habitat matching favours local adaptation: local
adaptation to the niche margin increased through time when dispersal entailed habitat choice
towards the niche margin (linear mixed model, F₁,₅₉=39.63; p<0.001; Fig 2a; Supplementary Table 1 & Figures 4&5) together with an increase of population sizes (Supplementary Figure 6). This is true regardless of whether immigrants were cells that chose to disperse toward a 35°C habitat, or did not disperse but rather stayed in a 35°C habitat (Fig 2b). Thus the positive effect of habitat choice on local adaptation resulted from the choice of habitat itself, and not from a potential higher competitive or colonization ability of dispersers that would appear only when immigrants consisted of dispersers¹⁷. In contrast, local adaptation did not occur under random dispersal (F₁,₅₉=0.49; p=0.48; Fig 2a), as predicted by classical theory¹. Finally, populations receiving immigrants consisting of cells that chose a 23°C habitat showed unchanged or even decreased local adaptation (Fig 2c). All populations started with equal population sizes and received a fixed number of immigrants each week, meaning that treatment effects are very unlikely to result from drift or differences in mutational input rates (see Supplementary Material). Cells from control populations maintained at 23°C showed no change in fitness at 35°C through time (random dispersal: F₁,₁₀ = 0.11; p = 0.75; habitat choice dispersers from 23 to 35°C: F₁,₁₀ = 2.21; p = 0.17; no dispersal: F₁,₁₀ = 0.58; p = 0.46).

While the potential for local adaptation should decrease with increasing gene flow under random dispersal in temporally stable landscapes¹,¹⁶, local adaptation can be maximal for intermediate gene flow levels under temporal variability through an increase of local genetic diversity¹⁵. In contrast, gene flow may favour local adaptation even in temporally stable landscapes when involving habitat choice⁵⁻⁷. Our results support this latter view since local adaptation increased faster with a 30% compared to a 10% immigration rate when immigrants were dispersers that chose to join a 35°C patch from a 35°C original habitat (Supplementary Table 1). Increasing immigration rate did not, however, affect the rate of local adaptation in all other treatments, suggesting that a 10% immigration rate was sufficient to completely hinder or favour local adaptation depending on the type of dispersal involved¹⁵. A next critical
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step will be to identify and compare the tipping points at which the effect of dispersal on local adaptation is reversed with and without habitat choice¹⁸.

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High dispersal has long been assumed to homogenise populations in structured landscapes, thereby hindering local adaptation and speciation¹,². In agreement with this classical theory, we found that random dispersal hinders local adaptation. However, when dispersal results from habitat matching, local adaptation is facilitated⁴⁻⁷,¹⁰. Our study demonstrates how the behaviours underlying dispersal – random vs. active habitat choice movement – can shift the effects of gene flow from constraining to facilitating local adaptation. Habitat choice should drastically increase a species’ ability to adapt to new environmental conditions, especially compared to expectations from current models on the consequences of climate change, which largely assume random dispersal. For instance, the loss of genetic variation during range expansion is typically expected to reduce adaptive potential, but empirical evidence is mixed at best¹⁹,²⁰. Habitat matching during range expansion may in contrast increase adaptive potential and thereby facilitate rather than constrain range expansion⁶,²¹, which could also explain successful range expansions in some species²². Results of our microcosm experiments stress the need to identify the environmental conditions required for the evolution of habitat choice (e.g. environmental variability, information availability and reliability, dispersal costs) and to integrate this process in current theoretical and predictive frameworks⁵⁻⁷,²².

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# Methods

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## Culture conditions and genotypes

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*T. thermophila* is a 30-50µm ciliated unicellular eukaryote naturally living in freshwater ponds in North America. The species is a model organism in cell and molecular biology, and its maintenance under laboratory conditions benefit from decades of experience. We used 10 genotypes originally sampled by F.P. Doerder in North America (genotypes D1, 2, 3, 4, 6, 10,
12, 13, 15 and 17; see Supplementary Table 2), that reproduced clonally in our culture conditions.

Cells were maintained in axenic rich liquid growth media (2% Difco proteose peptone, 0.2% yeast extract) at 23°C. All manipulations were performed in sterile conditions under a laminar flow hood. We used a standardized procedure to measure cell density and morphology in *T. thermophila* cultures based on automated analysis of digital images. From each culture, we measured 5 samples (10µL) pipetted into one chamber of a multichambered counting slide (Kima precision cell 301890), and took digital pictures under dark-field microscopy. Digital pictures were analysed using IMAGEJ software (version 1.47, National Institutes of Health, USA, http://imagej.nih.gov/ij) to obtain the overall number of cells on the picture, later transformed into density per mL.

## Dispersal and habitat choice

To quantify habitat choice, we used dispersal systems consisting of 3 linearly connected patches (5mL standard Eppendorf tubes; Supplementary Figure 6), connected by corridors (4mm internal diameter silicon tube; 5cm long) and filled with growth media. Cells were placed in the central patch (i.e. “start patch”; ~200000 cells) and corridors were opened. Cells can thus choose to either stay in the “start” patch or disperse, and if disperse choose where to go.

## Population growth and fitness

We quantified fitness of individuals from each patch in the habitat choice experiment and under different gene flow treatments in the local adaptation experiment through standard population growth analyses. Small numbers of cells (~100 cells) from each patch/population were transferred to 96-well plates (250µl wells) filled with growth media. Cultures were
maintained at 23 or 35°C and absorbance measurements at 550nm were performed every 1h
for 1 week using a microplate reader (Infinite M200, TECAN). Absorbance is significantly
and linearly correlated with cell density within the range of densities observed in this study
(F₁,₂₂ = 145.94; p < 0.001; R² = 0.87; Supplementary Figure 7). Growth rate of each
population was computed using the *gcfit* function (*grofit* R-package) with spline fit
(illustration in Figure 1).

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193 Acknowledgments

194 We thank Ophélie Ronce and three anonymous referees for helpful comments. This study was
195 supported by the ANR INDHET to SJ and JC, ANR Netselect to ASC, and ARC 10-15/031,
196 F.R.S.-FNRS and UCL-FSR to SJ, DL and NS (F.R.S FNRS Research Associate), and a
197 Move-In-Louvain Marie Curie Action fellowship to SJ. DB, SJ and NS are funded by the
198 FWO research community EVENET and by the FEEDING networking grant from UCL. DB
199 is funded by FWO research project INVADED G018017N. This work is part of the
200 Laboratoire d'Excellence (LABEX) entitled TULIP (ANR-10-LABX-41) for SJ, DL, ASC,
201 MH and JC, and is contribution BRCXXX of the Biodiversity Research Centre at UCL to
202 which NS and SJ are affiliated.

203

204 Author contribution

205 SJ, DL, ASC, DB and JC defined the research theme. SJ, DL, ASC, DB, MH and JC set up
206 the experimental protocols. SJ performed the experiments and analysed the data, with the help
207 of MH and NS. SJ wrote the manuscript, and DL, ASC, DB, NS and JC contributed
208 substantially to revisions.

209

210 Data availability

211 The data supporting the findings of this study will be deposited to Dryad upon acceptance.

212

213 Competing interests

214 The authors declare no conflicts of interest.

215

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**Figure 1:** Cells that chose to stay in or join the thermal niche margin through dispersal (in red) have higher fitness in these marginal conditions than those that chose the other habitat (i.e. thermal niche core; blue) or did not have the opportunity to disperse (grey). Mean ± SE of growth rate per hour at 35°C of cells from each population are shown (N = 6 replicates initially inoculated with 40000 cells/ml in the central patch).

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**Figure 2:** Habitat choice favours local adaptation at the thermal niche margin. **a)** Growth rate at the thermal niche margin (35°C) does not significantly change over time under random dispersal (**grey**), while it significantly increases when incoming dispersers actively choose to leave a 23°C patch to join a 35°C patch (**red**). Estimating local adaptation as "local versus foreign" and "home versus away" shows a similar pattern (Fig S3&4). **b)** Local adaptation increases when dispersal involves adaptive habitat choice, regardless of if the immigrant cells chose to disperse toward a 35°C habitat, or if they did not disperse but rather stayed in a 35°C habitat, **c)** while local adaptation to 35°C is prevented or decreases when immigrants chose to stay in or join a 23°C patch. Lines connect mean values of growth rate per hour for all populations in each treatment (N = 12 per treatment; the two immigration rates are not distinguished here since the differences are mostly not significant; see Table S1), and coloured areas represent the standard error.

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