Material and Methods
Study organism and strainsParamecium caudatum is a freshwater ciliate with a world-wide
distribution, feeding on bacteria and detritus. Asexual reproduction
occurs by mitotic division and represents the main mode of population
growth. Swimming is accomplished through the coordinated movement of
ciliary bands on the cell surface (48). Previous work on P.
caudatum indicated a genetic basis of dispersal propensity (49). Here,
we used 20 Paramecium strains (i.e., clonal cultures derived from a
single individual) from various geographic origins (49, 50) and
representing different groups of COI genotypes (“mitochondrial
haplotypes”; Table S1). All cultures were reared under standard
laboratory conditions in lettuce medium with the food bacteriumSerratia marcescens at 23 °C, allowing up to 3 asexual doublings
per day (51).
Founder strains measurementsPrior to the start of the long-term experiment, we assayed the 20
founder strains for dispersal and population growth characteristics
(Table S1). For the dispersal assay, we placed aliquots of 8 mL of
culture (at equilibrium density) in a 2-patch system (additional details
below) and let the Paramecium disperse for 3h. Once connections were
blocked, we estimated the number of residents and dispersers by taking
150-600 µL samples from the two tubes and counting the number of
individuals under a dissecting microscope. Dispersal was taken as the
proportion of dispersers of the total number of individuals in the
system. Dispersal rates (gaussian posteriors) were then estimated using
a generalized linear mixed model (binomial error distribution; glmer in
“lme4” R package with bobyqa optimizer (52)) for each strain with time
and observation (to account for overdispersion) as random effects. We
tested 4 replicates per strain. For the growth assay, we placed ca. 200
individuals (from cultures at equilibrium density) in 20 mL of fresh
medium. Over the course of 6 days, we tracked population density by
counting the number of individuals in daily samples of 100-200 µL. We
tested 3 replicates per strain. Using a Bayesian approach (53), we
estimated the intrinsic population growth rate (r0) and
equilibrium density (\(\overline{N}\)) for each replicate by fitting a
Beverton-Holt population growth model to the time series data. Details
of the Bayesian fitting are given in Supplementary Information (SI
Appendix).
Evolutionary experiment The evolutionary experiment comprised a sequence of cycles, where
dispersal events alternated with periods of population growth. The
founder population was created by mixing the 20 strains at equal
proportions in a single culture, which was then divided up into 15
replicate lines, assigned to the following three treatments. First, in
the range front treatment (6 lines), we placed the Paramecium in one of
the two tubes in 2-patch dispersal systems (interconnected 15-mL tubes,
Fig. 1A). Connections were opened for 3 h, during which time individuals
were allowed to swim to the other tube. We then collected the dispersers
and cultured them for 1 week under permissive conditions in 20 mL of
fresh medium (in 50-mL plastic tubes), until we initiated a new round of
dispersal, again only retaining the dispersers and culturing them for 1
week, and so on. Second, the range core treatment (6 lines) followed the
same cycles of dispersal and growth, but only the non-dispersing
residents were retained after each dispersal episode. Third, in the
control treatment (3 lines), residents and dispersers were mixed after
each dispersal event and then cultured for 1 week, as in the other
treatments. In corollary, the range front treatment mimics the advancing
cohort of a spatially expanding population, whereas populations from the
core treatment remain in place and constantly lose emigrants. The
control treatment is similar to the core treatment, except for the loss
of emigrants. This simple 2-patch scenario can capture essential
processes occurring at range core and fronts (25), and produce results
comparable to those observed in more complex continuous landscapes as
shown in previous microcosm studies using freshwater ciliates (54, 55).
A total of 161 cycles were accomplished. Prior to each dispersal event,
ca. 1800 individuals (median; 25% / 75% quantile range: 1400 / 2700)
were placed in the dispersal systems. After dispersal, the number of
individuals starting the 1-week growth period were matched between
treatments. Because dispersal rates were low at the beginning, these
starting numbers were initially set to 200 individuals (placed in a
total volume of 20 mL of fresh medium). During the following 1-week
growth period, stable population sizes were typically reached within 3-4
days, with densities of ca. 240 individuals per ml (median; 25% / 75%
quantiles: 180 / 360). After cycle 32, when dispersal had already
reached higher levels (see Results), we adjusted the starting numbers to
ca. 1500 (median; 25% / 75% quantiles: 1100 / 2000).
Data collectionFor each line, dispersal was measured at each dispersal event and
equilibrium densities (\(\overline{N}\)) taken at the end of the 1-week
growth period at each cycle. Furthermore, growth rate
(r0) was determined in assays conducted at cycle 21
(year 1), 78 (year 2) and 160 (year 3), as described above, with 2-3
replicates per line and year. Bayesian model fitting was used to
estimate r0 (SI Appendix, Fig. S1). Measurement of
swimming behaviour were also taken in the first two years of the
experiment (SI Appendix, Fig. S2).