Despite their complex evolutionary histories, aquatic plants are highly underrepresented in contemporary biosystematic studies. Of them, the genus Callitriche is particularly interesting because of such evolutionary features as wide variation in chromosome numbers and pollination systems. However, taxonomic difficulties have prevented broader investigation of this genus. In this study we applied flow cytometry to Callitriche for the first time in order to gain an insight into evolutionary processes and genome size differentiation in the genus. Flow cytometry complemented by confirmation of chromosome counts was applied to an extensive dataset of 1077 Callitriche individuals from 495 localities in 11 European countries and the USA. Genome size was determined for 12 taxa. The results suggest that many important processes have interacted in the evolution of the genus, including polyploidization and hybridization. Incongruence between genome size and ploidy level, intraspecific variation in genome size, formation of autotriploid and hybridization between species with different pollination systems were also detected. Hybridization takes place particularly in the diploid-tetraploid complex C. cophocarpa-C. platycarpa, for which the triploid hybrids were frequently recorded in the area of co-occurrence of its parents. A hitherto unknown hybrid (probably C. hamulata × C. cophocarpa) with a unique chromosome number was discovered in the Czech Republic. However, hybridization occurs very rarely among most of the studied species. The main ecological preferences were also compared among the taxa collected. Although Callitriche taxa often grow in mixed populations, the ecological preferences of individual species are distinctly different in some cases. Anyway, flow cytometry is a very efficient method for taxonomic delimitation, determination and investigation of Callitriche species, and is even able to distinguish homoploid taxa and identify introduced species.

Institute of Botany Academy of Sciences of the Czech Republic Průhonice Czech Republic

Institute of Botany Academy of Sciences of the Czech Republic Průhonice Czech Republic; Department of Botany Charles University Praha Czech Republic

Zobrazit více v PubMed

Sculthorpe CD (1967) The biology of aquatic vascular plants. London: Edward Arnold. 610 p.

Cook CDK (1988) Wind pollination in aquatic angiosperms. Ann Missouri Bot Gard 75: 768–777.

Les DH (1988) Breeding systems, population structure, and evolution in hydrophilous angiosperms. Ann Missouri Bot Gard 75: 819–835.

Philbrick CT (1988) Evolution of underwater outcrossing from aerial pollination systems: A hypothesis. Ann Missouri Bot Gard 75: 836–841.

Barrett SCH, Eckert CG, Husband BC (1993) Evolutionary processes in aquatic plant populations. Aquat Bot 44: 105–145.

Grace JB (1993) The adaptive significance of clonal reproduction in angiosperms: an aquatic perspective. Aquat Bot 44: 159–180.

Les DH, Philbrick CT (1993) Studies of hybridization and chromosome number variation in aquatic angiosperms: evolutionary implications. Aquat Bot 44: 181–228.

Philbrick CT, Les DH (1996) Evolution of aquatic angiosperm reproductive systems. Bioscience 46: 813–826.

Kaplan Z (2002) Phenotypic plasticity in

Les DH, Crawford DJ, Kimball RT, Moody ML, Landolt E (2003) Biogeography of discontinuously distributed hydrophytes: a molecular appraisal of intercontinental disjunctions. Int J Plant Sci164: 917–932.

Cook CDK (1970) Hybridization in the evolution of

Fiasson JL, Gluchoff-Fiasson K, Dahlgren G (1997) Flavonoid patterns in European

Kaplan Z (2001)

Kaplan Z, Štěpánek J (2003) Genetic variation within and between populations of

Kaplan Z, Fehrer J, Hellquist CB (2009) New hybrid combinations revealed by molecular analysis: The unknown side of North American pondweed diversity (

Kaplan Z, Fehrer J (2011) Erroneous identities of

Cook CDK (1966) A monographic study of

Turała K (1969) Cytotaxonomical studies in

Dahlgren G (1991) Karyological investigations in

Hong DY (1991) A biosystematic study of

Hollingsworth PM, Preston CD, Gornall RJ (1998) Euploid and aneuploid evolution in

Wiegleb G, Kaplan Z (1998) An account of the species of

Kaplan Z, Fehrer J (2006) Comparison of natural and artificial hybridization in

Kaplan Z, Fehrer J (2007) Molecular evidence for a natural primary triple hybrid in plants revealed from direct sequencing. Ann Bot 99: 1213–1222. PubMed PMC

Kaplan Z, Jarolímová V, Fehrer J (2013) Revision of chromosome numbers of Potamogetonaceae: a new basis for taxonomic and evolutionary implications. Preslia 85: 421–482.

Kaplan Z (2010) Hybridization of

Urbanska-Worytkiewicz K (1980) Cytological variation within the family Lemnaceae.

Landolt E (1986) The family of Lemnaceae – a monographic study. Vol. 1: Morphology, karyology, ecology, geographic distribution, systematic position, nomenclature, descriptions.

Gupta PP (1978) Cytogenetics of aquatic ornamentals. II. Cytology of Nymphaeas. Cytologia 43: 477–484.

Gupta PP (1980) Cytogenetics of aquatic ornamentals. VI. Evolutionary trends and relationships in the genus

Cook CDK, Urmi-König K (1985) A revision of the genus

Moody ML, Les DH (2010) Systematics of the aquatic angiosperm genus

Wiegleb G, Herr W (1983) Taxonomie und verbreitung von

Martinsson K (1991) Natural hybridization within the genus

Preston CD, Hollingsworth PM, Gornall RJ (1998)

King RA, Gornall RJ, Preston CD, Croft JM (2001) Molecular confirmation of

Kaplan Z, Fehrer J (2009) An orphaned clone of

Schotsman HD (1967) Les Callitriches. Espèces de France et taxa nouveaux d’Europe. In: Jovet P, editor. Flora de France, vol. 1 . Paris: Editions Paul Lechevalier, pp. 1–151.

Lansdown RV (2006) Notes on the water-starworts (

Lansdown RV (2008) Water-starworts (

Fassett NC (1951)

Schotsman HD (1954) A taxonomic spectrum of the section

Schotsman HD (1958) Beitrag zur Kenntnis der

Schotsman HD (1977) Callitriches de la région Méditerranéenne. Nouvelles observations. Bull Cent Etud Rech Sci, Biarritz 11: 241–312.

Schotsman HD (1982) Biologie florale des

Mason R (1959)

Dersch G (1986) Zur Verbreitung der

Hedberg O, Hedberg I (2001) Tropical African

Prančl J (2012) Rod

Prančl J (2013) Rod

Jones H (1955) Heterophylly in some species of

Jones H (1955) Further studies on heterophylly in

Deschamp PA, Cooke TJ (1984) Causal mechanisms of leaf dimorphism in the aquatic angiosperm

Deschamp PA, Cooke TJ (1985) Leaf dimorphism in the aquatic angiosperm

McComb AJ (1965) The control of elongation in

Wong CH, McComb AJ (1967) An anatomical investigation into the effects of gibberellic acid on the expansion of

Musgrave A, Jackson MB, Ling E (1972)

Madsen TV (1991) Inorganic carbon uptake kinetics of the stream macrophyte,

Maberly SC, Madsen TV (2002) Use of bicarbonate ions as a source of carbon in photosynthesis by

Augustynowicz J, Grosicki M, Hanus-Fajerska E, Lekka M, Waloszek A, et al. (2010) Chromium(VI) bioremediation by aquatic macrophyte PubMed

Augustynowicz J, Kołton AM, Baran AM, Kostecka-Gugała AM, Lasek WW (2013) Strategy of Cr detoxification by

Sand-Jensen K, Andersen K, Andersen T (1999) Dynamic properties of recruitment, expansion and mortality of macrophyte patches in streams. Int Rev Hydrobiol 84: 497–508.

Sand-Jensen K, Pedersen ML (2008) Streamlining of plant patches in streams. Freshwater Biol 53: 714–726.

Augustynowicz J, Tokarz K, Baran A, Płachno BJ (2014) Phytoremediation of water polluted by thallium, cadmium, zinc, and lead with the use of macrophyte PubMed

Dawson I (1998) New salad and vegetable crops. From Australia's sub-Antarctic Islands. A report fo the Rural Industries Research and Develpoment Corporation. Canberra: Rural Industries Research and Development Corporation. 35 p.

Augustynowicz J, Długosz-Grochowska OG, Kostecka-Gugała AM, Leja M, Kruczek MK, et al. (2014)

Philbrick CT, Anderson GJ (1992) Pollination biology in the

Philbrick CT, Bernardello LM (1992) Taxonomic and geographic distribution of internal geitonogamy in New World

Martinsson K (1996) Growth forms and reproductive characters in six species of

Philbrick CT, Les DH (2000) Phylogenetic studies in

Savidge JP (1960) The experimental taxonomy of European

Martinsson K (1985) Problem kring

Bączkiewicz A, Szoszkiewicz K, Cichocka J, Celińsky K, Drapikowska M, et al. (2007) Isozyme patterns of

Savidge JP (1959) An interspecific hybrid in European

Schotsman HD (1961) Contribution a l’étude des

Demars BOL, Gornall RJ (2003) Identification of British species of

Triest L, Mannaert A (2006) The relationship between

Buczkowska K, Szoszkiewicz K, Bączkiewicz A (2008) Genetic variation in Polish populations of

Gomarasca S, Cicatelli A, Maggioni LA, Castiglione S (2012) The use of

Doležel J, Greilhuber J, Suda J (2007) Estimation of nuclear DNA content in plants using flow cytometry. Nature Protocols 2: 2233–2244. PubMed

Loureiro J, Doležel J, Greilhuber J, Santos C, Suda J (2008) Plant flow cytometry – far beyond the stone age. Cytom Part A 73A: 579–580. PubMed

Kolář F, Fér T, Štech M, Trávníček P, Dušková E, et al. (2012) Bringing together evolution on serpentine and polyploidy: spatiotemporal history of the diploid-tetraploid complex of PubMed DOI PMC

Kolář F, Lučanová M, Vít P, Urfus T, Chrtek J, et al. (2013) Diversity and endemism in deglaciated areas: ploidy, relative genome size and niche differentiation in the PubMed PMC

Mandák B, Trávníček P, Paštová L, Kořínková D (2012) Is hybridization involved in the evolution of the

Pellicer J, Clermont S, Houston L, Rich TC, Fay MF (2012) Cytotype diversity in the PubMed PMC

Niketić M, Siljak-Yakovlev S, Frajman B, Lazarević M, Stevanović B, et al. (2013) Towards resolving the systematics of

Loureiro J, Trávníček P, Rauchová J, Urfus T, Vít P, et al. (2010) The use of flow cytometry in the biosystematics, ecology and population biology of homoploid plants. Preslia 82: 3–21.

Bennert HW, Horn K, Kauth M, Fuchs J, Jakobsen ISB, et al. (2011) Flow cytometry confirms reticulate evolution and reveals triploidy in Central European PubMed PMC

Kúr P, Štech M, Koutecký P, Trávníček P (2012) Morphological and cytological variation in

Winter S, Chizzola R, Kriechbaum M, Kropf M (2013) Hybridisation in

Hanušová K, Ekrt L, Vít P, Kolář F, Urfus T (2014) Continuous morphological variation correlated with genome size indicates frequent introgressive hybridization among PubMed PMC

Vít P, Wolfová K, Urfus T, Tájek P, Suda J (2014) Interspecific hybridization between rare and common plant congeners inferred from genome size data: assessing the threat to the Czech serpentine endemic

Mráz P, Chrtek J, Fehrer J (2011) Interspecific hybridization in the genus

Trávníček P, Kubátová B, Čurn V, Rauchová J, Krajníková E, et al. (2011) Remarkable coexistence of multiple cytotypes of the PubMed PMC

Trávníček P, Jersáková J, Kubátová B, Krejčíková J, Bateman RM, et al. (2012) Minority cytotypes in European populations of the PubMed PMC

Diao Y, Chen L, Yang G, Zhou M, Song Y, et al. (2006) Nuclear DNA C-values in 12 species in Nymphaeales. Caryologia 59: 25–30.

Pellicer J, Kelly LJ, Magdalena C, Leitch IJ (2013) Insights into the dynamics of genome size and chromosome evolution in the early diverging angiosperm lineage Nymphaeales (water lilies). Genome 56: 437–449. PubMed

Volkova PA, Trávníček P, Brochmann C (2010) Evolutionary dynamics across discontinuous freshwater systems: Rapid expansions and repeated allopolyploid origins in the Palearctic white water-lilies (

Kabátová K, Vít P, Suda J (2014) Species boundaries and hybridization in central-European

Vialette-Guiraud ACM, Alaux M, Legeai F, Finet C, Chambrier P, et al. (2011) PubMed PMC

McCracken A, Bainard JD, Miller MC, Husband BC (2013) Pathways of introduction of the invasive aquatic plant PubMed PMC

Stift M, Bregman R, Oostermeijer JGB, van Tienderen PH (2010) Other tetraploid species and conspecific diploids as sources of genetic variation for an autotetraploid. Am J Bot 97: 1858–1866. PubMed

Morozowska M, Czarna A, Jędrzejczyk I (2010) Estimation of nuclear DNA content in

Wang W, Kerstetter RA, Michael TP (2011) Evolution of genome size in duckweeds (Lemnaceae). Journal of Botany 2011 : ID 570319. doi:10.1155/2011/570319.

Tanaka N, Ito Y, Matsuyama R, Uehara K (2007) Chromosome numbers of

Pijnacker LP, Schotsman HD. 1988. Nuclear-DNA amounts in European

Schotsman HD, Andreas CH (1974)

Otto F (1990) DAPI staining of fixed cells for high-resolution flow cytometry of nuclear DNA. In: Crisssman HA, Darzynkiewicz Z, editors. Methods in cell biology, vol. 33 . New York: Academic Press. pp. 105–110. PubMed

Leong-Škorničková J, Šída O, Jarolímová V, Sabu M, Fér T, et al. (2007) Chromosome numbers and genome size variation in Indian species of PubMed PMC

Morton A (2008) DMAP. Distribution mapping software. Available: http://www.dmap.co.uk/ Accessed 6 February 2013.

Philbrick CT, Aakjar RA, Stuckey RL (1998) Invasion and spread of

Prančl J, Schou JC (2014) Butfrugtet Vandstjerne –

Philbrick CT (1984) Pollen-tube growth within vegetative tissues of

Francis D, Davies MS, Barlow PW (2008) A strong nucleotypic effect on the cell cycle regardless of ploidy level. Ann Bot 101: 747–757. PubMed PMC

Šímová I, Herben T (2012) Geometrical constraints in the scaling relationships between genome size, cell size and cell cycle length in herbaceous plants. Proc R Soc B 279: 867–875. PubMed PMC

Ekrt L, Holubová R. Trávníček P, Suda J (2010) Species boundaries and frequency of hybridization in the PubMed

Suda J, Trávníček P, Mandák B, Berchová-Bímová K (2010) Genome size as a marker for the recognition of invasive alien taxa in

Suda J, Krahulcová A, Trávníček P, Rosenbaumová R, Peckert T, et al. (2007) Genome size variation and species relationships in PubMed PMC

Schotsman HD (1961) Races chromosomiques chez

Soltis DE, Soltis PS (1993) Molecular data facilitate a reevaluation of traditional tenets of polyploid evolution. Crit Rev Plant Sci 12: 243–273.

Soltis DE, Soltis PS (1999) Polyploidy: Origins of species and genome evolution. Trends Ecol Evol 14: 349–351. PubMed

Soltis DE, Buggs RJA, Doyle JJ, Soltis PS (2004) What we still don’t know about polyploidy. Taxon 59: 1387–1403.

Webster SD (1988)

Kaplan Z, Fehrer J (2013) Molecular identification of hybrids from a former hot spot of

Moody ML, Les DH (2007) Geographic distribution and genotypic composition of invasive hybrid watermilfoil (

Martinsson K (1993) The pollen of Swedish

Kaplan Z, Fehrer J (2004) Evidence for the hybrid origin of

Kaplan Z, Wolff P (2004) A morphological, anatomical and isozyme study of

Kaplan Z, Uotila P (2011)

Letz DR, Dančák M, Danihelka J, Šarhanová P (2012) Taxonomy and distribution of

Ekrt L, Trávníček P, Jarolímová V, Vít P, Urfus T (2009) Genome size and morphology of the

Husák Š (2000) Callitrichaceae Link – hvězdošovité. In: Slavík B, editor. Květena České republiky [Flora of the Czech Republic], vol. 6 . Praha: Academia. pp 709–718.

Zahradníková K (1982) Callitrichaceae L. Hviezdošovité. In: Futák J, Bertová L, editors. Flóra Slovenska [Flora of Slovakia], vol. 3 . Bratislava: Veda. pp. 463–470.

New estimates and synthesis of chromosome numbers, ploidy levels and genome size variation in Allium sect. Codonoprasum: advancing our understanding of the unresolved diversification and evolution of this section

Morphological, ecological and geographic differences between diploids and tetraploids of Symphytum officinale (Boraginaceae) justify both cytotypes as separate species

Evolutionary and Taxonomic Implications of Variation in Nuclear Genome Size: Lesson from the Grass Genus Anthoxanthum (Poaceae)