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Genetic and morphological comparisons of lesser celandine (Ficaria verna) invasions suggest regionally widespread sexual reproduction

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Both asexual and sexual reproduction can provide important keys to the success of invasive species. A species with potential for both is lesser celandine (Ficaria verna), a European native with multiple subspecies that have been introduced in North America as ornamentals and escaped cultivation. Asexual reproduction via bulbils is prolific in many introduced populations of lesser celandine, with sexual reproduction reportedly rare. Although genetic and morphological diversity of European celandine has been studied, few have examined invasive North American populations. We aimed to document introduced genotypic and phenotypic diversity at a regional scale. We first compared sequence-related amplified polymorphism (SRAP) genotypes of 64 individuals collected from Columbus, Cincinnati, and Cleveland, OH. In a second experiment, we phenotyped 129 individuals from the same regions and from Louisville, KY, measuring traits in an outdoor common garden experiment. The SRAP markers were highly polymorphic and revealed surprising genetic differentiation. Genetic and trait variation were both structured across regions, but we also saw high variation within regions. Cleveland populations differed the most genetically and morphologically. Nearly every individual made asexual bulbils, and many individuals that flowered produced expanded achenes. Trait data suggested subspecies verna or ficariiformis occur in these regions. Genetic admixture within regions and within individuals, along with achene expansion, suggests sexual reproduction may be widespread. Sexual and asexual propagules may spread by different vectors, and our resistance analyses indicated water dispersal and habitat availability contribute to genetic structure. These findings suggest that celandine has substantial potential for further spread and evolutionary change.

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  • Arnaud-Haond SO, Belkhir KH (2007) GENCLONE: a computer program to analyse genotypic data, test for clonality and describe spatial clonal organization. Mol Ecol Notes 7:15–17

    Article  CAS  Google Scholar 

  • Arnaud-Haond SO, Duarte CM, Alberto F, Serrão EA (2007) Standardizing methods to address clonality in population studies. Mol Ecol 16:5115–5139

    Article  CAS  Google Scholar 

  • Arrigo N, Tuszynski JW, Ehrich D, Gerdes T, Alvarez N (2009) Evaluating the impact of scoring parameters on the structure of intra-specific genetic variation using RawGeno, an R package for automating AFLP scoring. BMC Bioinform 10:33

    Article  Google Scholar 

  • Arrigo N, Holderegger R, Alvarez N (2012) Automated scoring of AFLPs using RawGeno v 2.0, a free R CRAN library. In: Pompanon F, Bonin A (eds) Data production and analysis in population genomics. Humana Press, Totowa, pp 155–175

    Chapter  Google Scholar 

  • Axtell AE, DiTommaso A, Post AR (2010) Lesser celandine (Ranunculus ficaria): a threat to woodland habitats in the Northern United States and Southern Canada. Invas Plant Sci Mana 3:190–196

    Article  Google Scholar 

  • Bailleul D, Stoeckel S, Arnaud-Haond S (2016) RClone: a package to identify MultiLocus Clonal Lineages and handle clonal data sets in R. Methods Ecol Evol 7:966–970

    Article  Google Scholar 

  • Baker HG (1955) Self-compatibility and establishment after “long-distance” dispersal. Evolution 9:347–349

    Google Scholar 

  • Barrett SCH (2015) Influences of clonality on plant sexual reproduction. P Natl Acad Sci USA 112:8859–8866

    Article  CAS  Google Scholar 

  • Barton K (2020) MuMIn: multi-model inference. R package version 1.43.17.

  • Bates D, Mächler M, Bolker B, Walker S (2015) Fitting linear mixed-effects models using lme4. J Stat Softw 67:1–48

    Article  Google Scholar 

  • Borden JB, Flory SL (2021) Urban evolution of invasive species. Front Ecol Environ 19:184–191

    Article  Google Scholar 

  • Clarke RT, Rothery P, Raybould AF (2002) Confidence limits for regression relationships between distance matrices: estimating gene flow with distance. J Agric Biol Environ Stat 7:361–372

    Article  Google Scholar 

  • Dewitz J (2019) National land cover database 2016 products (ver. 2.0, July 2020): United States Geological Survey data release. Accessed 5 Nov 2020.

  • DeWoody JA, Schupp J, Kenefic L, Busch J, Murfitt L, Keim P (2004) Universal method for producing ROX-labeled size standards suitable for automated genotyping. Biotechniques 37:348–352

    Article  CAS  Google Scholar 

  • Dice L (1945) Measures of the amount of ecologic association between species. Ecology 26:297–302

    Article  Google Scholar 

  • Dlugosch KM, Parker IM (2008) Founding events in species invasions: genetic variation, adaptive evolution, and the role of multiple introductions. Mol Ecol 17:431–449

    Article  CAS  Google Scholar 

  • Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem Bull 19:11–15

    Google Scholar 

  • Dray S, Dufour A (2007) The ade4 package: implementing the duality diagram for ecologists. J Stat Softw 22:1–20

    Article  Google Scholar 

  • Earl DA, vonHoldt BM (2012) STRUCTURE HARVESTER: a website and program for visualizing STRUCTURE output and implementing the Evanno method. Conserv Genet Resour 4:359–361

    Article  Google Scholar 

  • Emadzade K, Lehnebach C, Lockhart P, Hörandl E (2010) A molecular phylogeny, morphology and classification of genera of Ranunculeae (Ranunculaceae). Taxon 59:809–828

    Article  Google Scholar 

  • ESRI (2018) Environmental Systems Research Institute ArcGIS Pro release 2.2.0. Redlands, CA

  • Evanno G, Regnaut S, Goudet J (2005) Detecting the number of clusters of individuals using the software STRUCTURE: a simulation study. Mol Ecol 14:2611–2620

    Article  CAS  Google Scholar 

  • Falush D, Stephens M, Pritchard JK (2007) Inference of population structure using multilocus genotype data: dominant markers and null alleles. Mol Ecol Notes 7:574–578

    Article  CAS  Google Scholar 

  • Fox J, Weisberg S (2019) An R companion to applied regression, 3rd edn. Sage, Thousand Oaks

    Google Scholar 

  • Gill JJ, Jones BM, Marchant CJ, McLeish J, Ockendon DJ (1972) The distribution of chromosome races of Ranunculus ficaria L. in the British Isles. Ann Bot 36:31–47

    Article  Google Scholar 

  • Herrmann M, Holderegger R, Van Strien MJ (2013) Influence of parameter settings in automated scoring of AFLPs on population genetic analysis. Mol Ecol Resour 13:128–134

    Article  CAS  Google Scholar 

  • Heywood VH, Walker S (1961) Morphological separation of cytological races in Ranunculus ficaria L. Nature 189:604–604

    Article  Google Scholar 

  • Hijmans RJ (2019) geosphere: spherical trigonometry. R package version 1.5-10.

  • Jakobsson M, Rosenberg NA (2007) CLUMPP: a cluster matching and permutation program for dealing with label switching and multimodality in analysis of population structure. Bioinformatics 23:1801–1806

    Article  CAS  Google Scholar 

  • Jombart T, Pontier D, Dufour AB (2009) Genetic markers in the playground of multivariate analysis. Heredity 102:330–341

    Article  CAS  Google Scholar 

  • Jones BMG (1966) Variation in Ranunculus ficaria. Proc Bot Soc Br Isl 6:275

    Google Scholar 

  • Kamvar ZN, Tabima JF, Grünwald NJ (2014) poppr: an R package for genetic analysis of populations with clonal, partially clonal, and/or sexual reproduction. PeerJ 2:e281

    Article  Google Scholar 

  • Kermack JP, Rauschert ES (2019) Soil characteristics drive Ficaria verna abundance and reproductive output. Invas Plant Sci Mana 12:214–222

    Article  Google Scholar 

  • Kettenring KM, McCormick MK, Baron HM, Whigham DF (2011) Mechanisms of Phragmites australis invasion: feedbacks among genetic diversity, nutrients, and sexual reproduction. J Appl Ecol 48:1305–1313

    Article  Google Scholar 

  • Lavergne S, Molofsky J (2007) Increased genetic variation and evolutionary potential drive the success of an invasive grass. Proc Natl Acad Sci USA 104:3883–3888

    Article  CAS  Google Scholar 

  • Lawrence E (1971) Lob’s wood. Cincinnati Nature Center, Milford

    Google Scholar 

  • Lenth RV (2016) Least-squares means: the R package lsmeans. J Stat Softw 69:1–33

    Article  Google Scholar 

  • Li G, Quiros CF (2001) Sequence-related amplified polymorphism (SRAP), a new marker system based on a simple PCR reaction: its application to mapping and gene tagging in Brassica. Theor Appl Genet 103:455–461

    Article  CAS  Google Scholar 

  • Mack JJ (2008) Workplan for lesser celandine (Ranunculus ficaria) control in Rocky River and Mill Stream Run Reservations, v. 1.0. Cleveland Metroparks Technical Report 2008/NR-05. Division of Natural Resources, Cleveland Metroparks, Fairview Park, Ohio

  • Marchant CJ, Brighton CA (1973) Cytological diversity and triploid frequency in a complex population of Ranunculus ficaria L. Ann Bot 38:7–15

    Article  Google Scholar 

  • Marsden-Jones EM, Turrill WB (1952) Studies on Ranunculus ficaria. J Genet 50:522–534

    Article  Google Scholar 

  • Masters JA, Emery SM (2015) Leaf litter depth has only a small influence on Ranunculus ficaria (Ranunculaceae) biomass and reproduction. Am Midl Nat 173:30–37

    Article  Google Scholar 

  • McKinney ML (2006) Urbanization as a major cause of biotic homogenization. Biol Conserv 127:247–260

    Article  Google Scholar 

  • McRae BH (2006) Isolation by resistance. Evolution 60:1551–1561

    Google Scholar 

  • McRae BH, Dickson BG, Keitt TH, Shah VB (2008) Using circuit theory to model connectivity in ecology, evolution, and conservation. Ecology 89:2712–2724

    Article  Google Scholar 

  • Metcalfe CR (1939) The sexual reproduction of Ranunculus ficaria. Ann Bot 3:91–103

    Article  Google Scholar 

  • Miller I, Miller M (2014) John E. Freund’s mathematical statistics with applications, 8th edn. Pearson, Harlow, Essex.

  • Nei M (1978) Estimation of average heterozygosity and genetic distance from a small number of individuals. Genetics 89:583–590

    Article  CAS  Google Scholar 

  • O’Geen A, Walkinshaw M, Beaudette D (2017) SoilWeb: a multifaceted interface to soil survey information. Soil Sci Soc Am J 81:853–862.

    Article  CAS  Google Scholar 

  • OGRIP (2021) Ohio Geographically Referenced Information Program digital elevation models (DEMs). Accessed 4 March 2021.

  • Ortego J, Bonal R, Muñoz A (2010) Genetic consequences of habitat fragmentation in long-lived tree species: the case of the Mediterranean holm oak (Quercus ilex, L.). J Hered 101:717–726

    Article  CAS  Google Scholar 

  • OSU MBD (2021) Ohio State University Museum of Biological Diversity Herbarium online specimen access. Accessed 11 August 2021.

  • Peterman WE (2018) ResistanceGA: an R package for the optimization of resistance surfaces using genetic algorithms. Methods Ecol Evol 9:1638–1647

    Article  Google Scholar 

  • Popelka O, Sochor M, Duchoslav M (2019a) Reciprocal hybridization between diploid Ficaria calthifolia and tetraploid Ficaria verna subsp. verna: evidence from experimental crossing, genome size and molecular markers. Bot J Linn Soc 189:293–310

    Article  Google Scholar 

  • Popelka O, Trávníček B, Šiková P, Jandová M, Duchoslav M (2019b) Natural hybridization between diploid Ficaria calthifolia and tetraploid Ficaria verna subsp. verna in central Europe: evidence from morphology, ecology and life-history traits. Preslia 91:179–212

    Article  Google Scholar 

  • Post AR, Krings A, Wall WA, Neal JC (2009) Introduced lesser celandine (Ranunculus ficaria, Ranunculaceae) and its putative subspecies in the United States: a morphometric analysis. J Bot Res Inst Tex 3:193–209

    Google Scholar 

  • Pritchard JK, Stephens M, Donnelly P (2000) Inference of population structure using multilocus genotype data. Genetics 155:945–959

    Article  CAS  Google Scholar 

  • Pyšek P, Richardson DM (2008) Traits associated with invasiveness in alien plants: where do we stand? In: Nentwig W (ed) Ecological studies (analysis and synthesis), vol 193. Biological invasions. Springer, Berlin, pp 97–125

    Google Scholar 

  • R Core Team (2019–20) R: a language and environment for statistical computing. R Foundation for Statistical Computing. Vienna, Austria.

  • Reisch C, Scheitler S (2008) Disturbance by mowing affects clonal diversity: the genetic structure of Ranunculus ficaria (Ranunculaceae) in meadows and forests. In: van der Valk A (ed) Herbaceous plant ecology. Springer, Dordrecht, pp 335–343

    Chapter  Google Scholar 

  • Robarts DWH, Wolfe AD (2014) Sequence-related amplified polymorphism (SRAP) markers: a potential resource for studies in plant molecular biology. Appl Plant Sci 2:1–14

    Article  Google Scholar 

  • Schenk MF, Thienpont CN, Koopman WJ, Gilissen LJ, Smulders MJ (2008) Phylogenetic relationships in Betula (Betulaceae) based on AFLP markers. Tree Genet Genomes 4:911–924

    Article  Google Scholar 

  • Sell PD (1994) Ranunculus ficaria L. sensu lato. Watsonia 20:41–50

    Google Scholar 

  • Serrote CM, Reiniger LR, Silva KB, dos Santos Rabaiolli SM, Stefanel CM (2020) Determining the polymorphism information content of a molecular marker. Gene 726:144175

    Article  CAS  Google Scholar 

  • Štajerová K, Šmilauer P, Brůna J, Pyšek P (2017) Distribution of invasive plants in urban environment is strongly spatially structured. Landscape Ecol 32:681–692

    Article  Google Scholar 

  • Taylor K, Markham B (1978) Ranunculus ficaria L. (Ficaria verna Huds.; F. Ranunculoides Moench). J Ecol 66:1011–1031

    Article  Google Scholar 

  • Tyers M (2020) riverdist: river network distance computation and applications. R package version 0.15.3.

  • USCB (2019a) United States Census Bureau American community survey 5-year estimates subject tables: median income in the past 12 months (in 2019 inflation-adjusted dollars). Accessed 10 March 2021.

  • USCB (2019b) United States Census Bureau TIGER/line shapefiles: census tracts. Accessed 10 March 2021.

  • USDA FS (2019) United States Department of Agriculture Forest Service national land cover database 2016 tree canopy cover (CONUS). Salt Lake City, UT. Accessed 23 Oct 2020.

  • USDA NRCS (2021) USDA Natural Resources Conservation Service. Ranunculus ficaria L. The PLANTS database. National Plant Data Team. Greensboro, NC. Accessed 28 July 2021.

  • USGS (2014) United States Geological Survey area- and depth-weighted averages of selected SSURGO variables for the conterminous United States and District of Columbia. Accessed 8 March 2021.

  • USGS (2021) United States Geological Survey national hydrography dataset best resolution (NHD) for Hydrologic Unit (HU) 8. Accessed 14 Jan 2021.

  • Veldkamp J (2015) De nomenclatuur van Speenkruiden (Ficaria verna Huds. s.l. Ranunculaceae). Gorteria 37:84–116

    Google Scholar 

  • Whittemore AT (2020) Ranunculus ficaria. In: Flora of North America Editorial Committee 1993+ (eds) Flora of North America north of Mexico, vol. 3, New York and Oxford. Accessed 19 June 2021.

  • Wolfe AD (2005) ISSR techniques for evolutionary biology. Method Enzymol 395:134–144

    Article  CAS  Google Scholar 

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We are grateful for the help of Jennifer Hillmer and Pat Lorch of Cleveland Metroparks and Paul Pira of the Geauga Park District for assistance with sites. We thank Dr. Michael Broe, Leah Weston, Dr. Scott Martin, and Dr. Bill Peterman for assistance with analyses and Dr. Alison Bennett, Associate Editor Dr. Kristin Saltonstall, Alexis Wafer, Dr. Andi Wolfe, and two anonymous reviewers for feedback on analyses and writing. We thank the Cornell University Biotechnology Resource Center for fragment analysis services. For lab assistance, we thank Eleanor Chase. For facilities support, we thank Ohio State’s Biological Sciences and Biotechnology Support Greenhouses, Center for Applied Plant Sciences, Gary Posey, David Snodgrass, and Emily Yoders-Horn. This work was supported by the Ohio State Alumni Grant for Graduate Research and Scholarship and Presidential Fellowship to KZM and start-up funds to SMH.


This work was supported by the Ohio State Alumni Grant for Graduate Research and Scholarship and Presidential Fellowship to KZM and start-up funds to SMH.

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SMH, CTCD, and AT together designed the project. CTCD, ESJR, and AT oversaw surveys and data collection, supervised by SMH. CTCD and KZM developed genetics protocols, KZM and AT executed genetic analyses, and AT performed morphological sampling. KZM and AT analysed and interpreted data, supervised by SMH and ESJR. KZM wrote the manuscript with contributions from AT, SMH, and ESJR. All authors reviewed and approved the final manuscript.

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Correspondence to Kali Z. Mattingly.

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Fig. S1. Rasters used to create distance matrices in ResistanceGA (Peterman 2018), analysed as potential predictors of Columbus-area (n=51) genetic variation: a) slope, b) development, c) income, d) canopy, e) sand. Raster values were standardized to the same range. Raster spatial projections were NAD27 / UTM zone 17N, meters, with 30x30m cells (JPG 642 KB)


Fig. S2. Density plot of the number of genetic markers that differed among all pairs of individuals, examined to detect clones having unique genotypes (multilocus lineages, Arnaud-Haond et al. 2007). Lack of a distinct peak near zero suggested no clones were present in our genetic dataset (JPG 705 KB)


Fig. S3. Plots of top two principal coordinates from principal coordinates analyses (PCoA), which qualitatively assess how PCo clusters correspond to regions and STRUCTURE clusters. Axis labels list variation explained (%) by a PCo. Point labels are region abbreviations: Central Columbus (CCOL), East Columbus (ECOL), Cincinnati (CIN), Cleveland (CLE). Colors denote STRUCTURE clusters, with color gradations corresponding to individual assignment probabilities. a) All data (n=64) with K=2; b) Columbus samples (n=51) with K=2; c) all data (n=64) with K=3 (PNG 898 KB)


Fig. S4. Single surface optimizations (ResistanceGA::SS_optim, Peterman 2018) describing the shape of the relationship of resistance to raster values (Fig. S1) for a) slope, b) development, c) income, d) canopy, and e) sand. Raster values were standardized to the same range prior to optimization. Response curves were estimated for continuous predictors, and the histograms visualize the distributions of original raster values relative to distributions of the estimated resistance values. For the categorical predictor (b), resistance was optimized for each category. Resistance is interpreted such that relatively higher values indicate barriers to gene flow, while lower values indicate conductors of gene flow (JPG 587 KB)

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Mattingly, K.Z., Day, C.T.C., Rauschert, E.S.J. et al. Genetic and morphological comparisons of lesser celandine (Ficaria verna) invasions suggest regionally widespread sexual reproduction. Biol Invasions 25, 379–397 (2023).

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