Plant Syst. Evol. 239: 95–112 (2003) DOI 10.1007/s00606-002-0252-7 Genetic variation within and between populations of Potamogeton pusillus agg. Z. Kaplan and J. Sˇteˇpa´nek Institute of Botany, Academy of Sciences, Pru˚ honice, Czech Republic Received August 6, 2002; accepted November 22, 2002 Published online: May 15, 2003 Ó Springer-Verlag 2003 Abstract. Patterns of isozyme variation were ex- Key words: Potamogeton pusillus, Potamogeton amined in 17 populations of P. pusillus and berchtoldii, isozymes, genetic variation, population P. berchtoldii, together with one population of structure, reproductive systems, clonal growth. P. trichoides taken for comparison. Both P. pusillus and P. berchtoldii displayed low levels of variation The Potamogeton pusillus agg.1 belongs among within populations associated with high levels of the taxonomically most difficult groups in the interpopulation differentiation. This pattern of family Potamogetonaceae. The complex is partitioning of genetic variation within and be- almost cosmopolitan in its distribution; the tween populations is attributed to the founder only continent from which it is absent is effect, frequent vegetative propagation by turions, Australia. All morphotypes within this group dominant self-fertilization and limited seedling were by earlier authors referred to as a single recruitment. The mechanism of pollen transfer was investigated in cultivation. Effective pollina- species, sometimes divided into a few varieties. tion takes place in air above the water surface The first attempt to arrange all variation in this (autogamy, geitonogamy, anemogamy), on the group in Central Europe and to describe it in water surface (epihydrogamy) or below water terms of formal classification appeared in the surface (hydroautogamy). The species are self- Central-European Flora by Ascherson and compatible. The low level of infra-population Graebner (1897) who distinguished five infra- variation together with rare occurrence of hetero- specific taxa. Soon after, Hagstro¨ m (1901) zygotes suggest that selfing is the most frequent separated two species, P. pusillus (nowadays mode of pollination, although the protogynous called P. berchtoldii Fieber) and P. panormit- flowers may occasionally permit some cross-polli- anus Biv. (true P. pusillus L., see Dandy and nation. Unique enzyme markers were found for Taylor 1938, 1940). Fischer (1907) followed P. pusillus and P. berchtoldii, and also for the single Hagstrom in treating two distinct species and population of P. trichoides. All multienzyme phe- ¨ notypes were species-specific. Isozyme data support subdivided Bavarian plants into 23 and eight the separate position of P. pusillus and P. berch- infraspecific taxa, respectively, at the variety toldii. UPGMA dendogram based on enzyme data of 133 plant samples revealed three distinct main 1 In this paper, Potamogeton pusillus agg. is understood as enzymatic entities perfectly corresponding to the comprising two species in Europe, namely P. pusillus L. s. three morphologically defined species. str. (syn. P. panormitanus Biv.) and P. berchtoldii Fieber. 96 Z. Kaplan and J. Sˇ teˇ pa´ nek: Genetic variation in Potamogeton pusillus agg. and form levels. In his worldwide account, P. berchtoldii at three levels: (1) genetic vari- Graebner (1907) lumped the two species into ation within populations in order to estimate P. pusillus and recognized 14 units mostly at prevailing modes of reproduction, (2) genetic variety level within it, besides 3-4 closely variation between populations within each related South American species. Both species species to reveal possible contribution of were subdivided into several forms and varie- different genotypes to observed morphological ties also by Hagstro¨ m (1916) in his monograph. differences, and (3) inter-specific differences Besides these two basic units, he described six to test the distinction of P. pusillus and additional similar species from this complex P. berchtoldii at species level. mostly from Africa. In a revision of North American linear-leaved Potamogeton species, Material a methods Fernald (1932) adopted both species with several varieties. The concept of many infra- Plant material. Potamogeton pusillus and P. berch- specific taxa has been gradually abandoned in toldii are morphologically very similar. In fact, the literature, with the notable exception of most recent guides to identification stress only a Soo´ (1936, 1938) who proposed many new single morphological feature: structure of stipules. combinations of highly dubious value. At These are closed and tubular when young in P. pusillus but open and convolute in P. berchtoldii. present, both main taxa are generally recog- Hence in this study, sampled plant material of nized as separate species in Eurasia (e.g. Dandy P. pusillus agg. was divided into two groups solely 1980, Kasˇ ina 1988, Preston 1995) or either as according to the stipule shape. Besides similar varieties (Haynes 1974, Hellquist and Crow morphology, both species also share many basic 1980) or recently as subspecies (Haynes and biological features. The two species are largely Hellquist 1996, Haynes and Hellquist 2000) in sympatric in Eurasia and North America, but only North America. At the worldwide scale, satis- P. pusillus occurs in most of Africa. The exact factory taxonomic solution for this complex is position of South American plants that belong to still not available. In a recent global account of this complex is still insufficiently known. P. tricho- Potamogeton species (Wiegleb and Kaplan ides Cham. et Schltdl. is a species related to 1998), a broad concept of P. pusillus had to be P. pusillus agg. as delimited here, which is, howev- temporarily adopted again to cover the ex- er, taxonomically well defined and usually mor- phologically well distinguishable particularly when tremely rich mosaic of widespread phenotypes, fertile material is available (Preston 1995, Wiegleb local taxa and extreme forms of P. pusillus agg. and Kaplan 1998). A single population of this In spite of the fact that the extreme species was also included in analyses for inter- morphological variation in P. pusillus agg. specific comparison. All three species are consid- has been observed by many authors, its origin ered to be diploids with a chromosome number has little been studied systematically. Variation 2n = 26 (cf. Hollingsworth et al. 1998). Nine due to seasonal development was noted by chromosome counts have also been determined in Haynes (1974) who reported differences in plants included in this study, five in P. pusillus s. development of lacunae and shape of leaf apex str. and four in P. berchtoldii, all with chromo- between plants collected in early summer and some number 2n=26 (Jarolı´ mova´ and Kaplan, those collected later in the season. Contribu- unpubl.). tion of variation due to environment has Plants for cultivation and analyses were sam- pled from sites in Central Europe in 1997–2000. recently been investigated by Kaplan (2002), The origin of plants used for isozyme analysis is who observed considerable phenotypic plasti- summarized in Table 1. The reference population city in general appearance, branching pattern, numbers correspond to numbers of cultivated width and colour of leaves, details of venation, material and numbers of vouchers; herbarium and shape of leaf apex. specimens of individuals from the populations were The main aim of this study is to provide designated as their fractions (e.g. 307/1, 307/2 etc.). data on isozyme variation in P. pusillus and Voucher herbarium specimens from both field and Table 1. Localities of samples used for isozyme analysis, with population designations used in the text and reference population numbers Z. Kaplan and J. S corresponding to numbers of cultivated material and numbers of voucher herbarium material Species Population No. Locality P. pusillus Untersee 990 Switzerland: Distr. Thurgau: S margin of the Untersee (lake) 1 km ESE of Ermatingen village, 395 m a. s. l., 23 VI 1998 Bodensee I 987 (P) Austria: Vorarlberg: Distr. Bregenz: inlet at S margin of Fussacher Bucht (bay) of Bodensee Lake ˇ te ˇ 1.5 km N–NNE of Fußach village, near Ho¨ chst, 396 m a. s. l., 23 VI 1998 pa ´ Litomeˇ rˇ ice 959 Czech Republic: Bohemia: Distr. Litomeˇ rˇ ice: small pond in valley at N margin of Litomeˇ rˇ ice- nek: Genetic variation in Pokratice town suburb, 260 m a. s. l., 27 V 1998 Rokytnˇ any 1133 Czech Republic: Bohemia: Distr. Mlada´ Boleslav: pond 0.4 km W of Dolnı´ Rokytnˇ any village, 234 m a. s. l., 11 VI 1999 Dubovec 1212 Czech Republic: Bohemia: Distr. Jindrˇ ichu˚ v Hradec: Velky Dubovec pond 1.3 km SSE of Lomnice nad Luzˇ nicı´ village, 425 m a. s. l., 15 VI 2000 Panensky 1159 Czech Republic: Bohemia: Distr. Jindrˇ ichu˚ v Hradec: Velky Panensky fishpond 1.3 km SSE of Lomnice nad Luzˇ nicı´ village, 420 m a. s. l., 8 IX 1999 Struzˇ ky 1213 Czech Republic: Bohemia: Distr. Jindrˇ ichu˚ v Hradec: Struzˇ ky fishpond 0.8 km W of Brˇ ilice village, 445 m a. s. l., 15 VI 2000 Potamogeton pusillus Sˇ imanov 307 Czech Republic: Bohemia: Distr. Jindrˇ ichu˚ v Hradec: Sˇ imanov pond (above Thierov pond) near Trˇ ebonˇ , 1.4 km NE of Trˇ ebonˇ railway station, 428 m a. s. l., 21 VIII 1996 P. berchtoldii Thuner See 991 Switzerland: Distr. Bern: Interlaken: small pool on E bank of the Thuner See (lake) W of Unterseen town, 560 m a. s. l., 21 VI 1998 Bodensee II 987 (B) Austria: Vorarlberg: Distr. Bregenz: inlet at S margin of Fussacher Bucht (bay) of Bodensee Lake 1.5 km N–NNE of Fußach village, near Ho¨ chst, 396 m a. s. l., 23 VI 1998 Skalna´ I 1147 Czech Republic: Bohemia: Distr. Cheb: pool in bottom of disused clay-pit called Sucha´ 1.8 km E–ESE of Skalna´ , 430 m a. s. l., 29 VII 1999 agg. 97 Skalna´ II 1224 Czech Republic: Bohemia: Distr. Cheb: pool in bottom of disused clay-pit near Zelena´ settlement, 0.8 km SE of Skalna´ , 440 m a.