Fig. 1. Thirty-four sites broadly distributed across southern
Quebec, supporting natural populations of Lemna minor . Map was
produced with Datawrapper.
For each site we measured several environmental variables that we
expected to be correlated with plant phenotype. Light availability, as
percent transmittance of photosynthetically active radiation (PAR), was
estimated in situ with the use of BF5 Sunshine Sensor (Delta-T,
Burwell, Cambridge, UK) (Paquette et al. 2007). This instrument consists
of an array of seven quantum sensors under a semi-shaded hemispherical
dome to give estimates of diffused light under any meteorological
condition. We took two simultaneous paired measurements, one at the
sampling site in question, and a second reference point at a nearby open
site (field or road) under full sun. Percent transmittance PAR was then
estimated as the ratio of diffused light between the site and the
reference measurement. This method has been demonstrated as a reliable
and practical alternative to more standard measurement techniques
including hemispherical image analysis (Rich et al. 1993, Paquette et
al. 2007). Percent transmittance of PAR was estimated at each of the
three microsites as well as the center of the site. All measurements
were taken 1.5m above the water level, above any aquatic macrophytes or
riparian herbaceous plants to obtain an estimate of shading from the
canopy cover. These four measurements were then averaged to produce a
single estimate of light availability for each site. To measure water
nutrient content, we took eight water samples from the center of each
site at a depth of 30cm. Total Nitrogen (TN), total Phosphorus (TP),
dissolved Nitrogen (DN) and dissolved Phosphorus (DP) were estimated
each from two replicate samples. Acid-washed tubes were first rinsed,
and then filled with sample water, unfiltered for TN and TP samples, and
sterile filtered at 0.45um for DN and DP samples. After sampling, all
tubes were stored in a cooler on ice and brought back to the lab for
analysis. Samples were then stored at 4oC and
processed within 14 days. Water samples were analysed for TN and DN with
a continuous flow analyser (OI Analytical Flow Solution 3100 ©) using an
alkaline persulfate digestion method, coupled with a cadmium reactor
(Patton and J.R. 2003) and for DP using a standard protocol (Wetzel and
Likens 2000). TP was measured using colorimetric detection with a
spectrophotometer at 890 nm, after digestion with potassium persulfate
and the addition of an ammonium molybdate solution (Wetzel and Likens
2000). All samples were analysed at the GRIL, Université du Québec à
Montréal (UQAM) analytical laboratory. We used a YSI probe (YSI
professional plus, Xylem Inc., Yellow Sprigs, OH, USA.) to measure water
temperature and pH at the centre of each site at a depth of 30cm. Full
list of environmental correlates can be found in the supplementary
information (Table S1).