DOI: 10.5061/dryad.kd51c5bc3
title: High tolerance to zinc but no evidence for local adaptation in the aquatic plant Lemna minor

description: Duckweeds are a widely distributed and economically important aquatic
 plant family that have high potential for phytoremediation of polluted
 water bodies. We collected four ecotypes of the common duckweed (Lemna
 minor) from the four corners of Switzerland and assessed how their home
 vs. away environments influenced their growth. Additionally, we
 investigated their response to a metal pollutant (Zn) in both their home
 and away environments. Zn is found in freshwater systems and can become
 harmful to plants at elevated concentrations. We hypothesized that growing
 in their home environment would help the plants buffer the negative effect
 of the metal pollutant. To test this, we measured Lemna growth in a common
 garden experiment in a glasshouse where the four ecotypes were grown in
 each of the water environments, as well as in three different
 concentrations of Zn. To investigate whether interactions between Lemna
 and their microbial community can enhance or reduce tolerance to heavy
 metal pollution, we sampled chlorophyll-a as a proxy for algal biomass.
 Finally, we measured total nitrogen and total organic carbon to describe
 the abiotic environment in more detail. The four Lemna ecotypes exhibited
 significantly different growth rates across the water treatments. This
 difference in fitness was matched with DNA sequencing revealing genetic
 differentiation between the four ecotypes. However, the effect of the
 water and zinc treatment on Lemna growth was the same for all ecotypes. We
 did not find evidence for local adaptation; instead, we observed strong
 plastic responses. Lemna growth rates were higher under higher Zn
 concentrations. This positive effect of Zn on Lemna growth could be in
 part due to reduced competition with algae. We conclude that L. minor
 ecotypes may exhibit large differences in growth rate, but that the
 species overall have a high Zn tolerance and strong plastic adaptive
 potential in novel environments.
description: Summary The
 data are based on a glasshouse common garden experiment in which four
 <em>Lemna</em> <em>minor</em> ecotypes from
 Switzerland were matched to their home water and received the water from
 the three other ecotypes (away). Additionally, we crossed this design with
 the application of Zn (in the form of ZnSO<sub>4</sub>). We
 used three Zn treatment levels: no Zn (control), low Zn and high Zn. Each
 treatment combination was replicated four times, resulting in a total of
 192 experimental units (four ecotypes x four water environments x three Zn
 treatments x four replicates. The experiment ran for 22 days in a
 glasshouse. The data submission includes two data
 files: 1) Data_fronds_fluoroprobe.xlsx contains the
 data on the number of <em>Lemna</em> fronds counted during the
 experiment, as well as the Fluoroprobe data measured at the end of the
 experiment. A second sheet also includes the fluoroprobe data from field
 samples. Metadata are included. 2) TOC_TN.xslx contains
 the data on the final nutrient concentration in the experiment. It also
 contains the nutrient data from the field samples. Metadata are
 included. The data was then processed using R. R code
 is uploaded separately. Methods On
 7 and 8 August, 2021, we collected four <em>Lemna</em>
 <em>minor</em> populations from four geographic regions of
 Switzerland: Koblenz: 47°36’03.3” N, 8°13’32.0” E, Yverdon:
 46°47'50.8"N, 6°37'59.6"E, Motto: 46°25’43.6” N,
 8°58’03.4” E, Ramosch: 46°50'01.4"N,
 10°24'04.0"E. To create the Zn treatments for
 each site, we mixed
 ZnSO<sub>4</sub>•7H<sub>2</sub>O (Alfa Aesar) with
 the filtered source water at a concentration of 3.4 mg [Zn]/L for the low
 treatment and 11.36 mg [Zn]/L for the high treatment.
 To initiate the experiment, each cup received 100 mL of filter
 source water and 30 L. minor individuals. No nutrients were added to the
 cups. All 192 cups were spread onto four different tables, one table per
 replicate. The experiment ran for 22 days. Fronds were counted based on
 photographs taken of the cups from above using Image J
 (https://imagej.nih.gov/ij/index.html). At the end of
 the experiment (day 22), an unfiltered 50-mL water samples from each cup
 was analyzed for chlorophyll-a concentration fluorometrically through a
 Fluoroprobe (bbe Moldaenke, Germany). In addition, a
 30-mL water sample was analyzed for its inorganic carbon, total organic
 carbon (TOC) and total nitrogen (TN) concentrations (Skalar Formacs HT – I
 TOC/TN Analyzer). Since the samples contained more inorganic carbon than
 organic carbon, we were not able to analyze TOC via subtraction (i.e., TOC
 = TC – IC). Thus, we measured non-purgeable organic carbon (NPOC). NPOC is
 the carbon that remains in a solution after the sample has been acidified
 and purged with aid of a gas flow. It is often reported as TOC since most
 samples contain a negligible amount of POC (NPOC = TOC – POC). Part of
 each sample (7 mL) was acidified with 100 μL of 10% HCl and purged for two
 minutes with N2 gas prior to the analyzer measurement. TN was measured
 simultaneously in a parallel compartment of the analyzer. 
 We used two metrics to evaluate population fitness. The first
 metric was initial population growth rate calculated as
 ln(N<sub>2</sub>/N<sub>1</sub>)/(t<sub>1</sub>- t<sub>2</sub>) where N is the number of fronds, and t<sub>1</sub>=1 and t<sub>2</sub>= 8 represent the first and eighth days of the experiment. The second metric was total population growth rate where t<sub>1</sub>=1 and t<sub>2</sub>=22, which represented the first and final days of the experiment.
description: Data analyses were done in R. The R scripts can be found on
 Zenodo.