Thoughts about the taxonomical relationships within the lemnaceae

Aquatic Botany, 1 (1975) 407--416 © Elsevier Scientific Publishing Company, Amsterdam -- Printed in The Netherlands

THOUGHTS ABOUT THE TAXONOMICAL RELATIONSHIPS WITHIN THE LEMNACEAE

C. DEN HARTOG Laboratorium voor Aquatische Oecologie, Katholieke Universiteit, Nijmegen (The Netherlands) (Received August 22nd, 1975)

ABSTRACT Den Hartog, C., 1975. Thoughts about the taxonomical relationships within the Lemnaceae. Aquat. Bot., 1: 407--416. A survey is given of the taxonomy of the Lemnaceae based on classic morphological characters. This taxonomic system has been compared with the results of phytochemical research, particularly that on flavonoid chemistry by McClure and co-workers; it appears that the classical taxonomical system and the phytochemical data are largely in agreement, except in the case of Wolffia. Subdivisions within the genera based on morphological characters do not coincide with subdivisions based on phytochemical data; in the genus Lernna it appears that flavonoid data, lignin data and morphological data result in three different subdivisions. Therefore, great care is needed for the application of such characters to the taxonomy above the species level. The available cytological data are at present still insufficient to give any indication on systematic relations. It is recommended to direct future research particularly at the flavonoid chemistry and the cytology of the Wolffioideae. INTRODUCTION

The L e m n a c e a e f o r m a well-defined p l a n t f a m i l y t h e d e l i m i t a t i o n o f w h i c h is e x t r e m e l y clear. Its a b e r r a n t m o r p h o l o g i c a l and a n a t o m i c a l features, h o w ever, disguise t h e relationships with o t h e r p l a n t families. N o w a d a y s m o s t a u t h o r s agree t h a t t h e L e m n a c e a e are derived f r o m t h e Araceae via t h e link o f the r a t h e r isolated a q u a t i c genus Pistia (Hegelmaier, 1 8 6 8 ; Engler, 1 8 7 7 ; Velen o v s k y , 1 9 0 7 ; Maheswari, 1 9 5 8 ) , o f w h i c h t h e m o r p h o l o g y and a n a t o m y give ample evidence f o r such a d e v e l o p m e n t . In f a c t o n l y Lawalr~e ( 1 9 4 5 ) has ano t h e r o p i n i o n , viz. t h a t t h e y are related t o t h e Helobiae, p a r t i c u l a r l y the Najadaceae, b u t several o f his a r g u m e n t s have a p p e a r e d t o be u n t e n a b l e in the light o f m o r e r e c e n t research. A c c o r d i n g t o H e g n a u e r ( 1 9 6 3 ) the few p h y t o c h e m i c a l d a t a w h i c h are available are well in line with t h e h y p o t h e s i s t h a t t h e L e m n a ceae have been derived f r o m t h e Araceae, b u t t h e evidence is scarce, and t h e d a t a o n l y indicate t h a t negative evidence is lacking so far. It is i m p o r t a n t t o note, t h a t h a r d l y a n y p h y t o c h e m i c a i d a t a o n Pistia are available (McClure,

408

1970), which as the supposed link between the two families deserves special attention. The palynological evidence for a link between the Araceae and Lemnaceae is also very meagre. Erdtman (1952) found t h a t the pollen of the Lemnaceae shows some aracean characters. Thanikaimoni (1969), however, stated t h a t the lemnaceous type of pollen has n o t been f o u n d so far amonst Araceae, and according to him there is no affinity between the L e m n a and the Pistia pollen. In the field of cytology there appears to be no close connection between Pistia (n = 14) and the Lemnaceae (n = 10). (See Wcis~o, 1970; UrbanskaWorytkiewicz, 1975). Although generally most evidence points to a relationship between the Araceae and the Lemnaceae, this relationship is certainly n o t close and the separation of the two families must have taken place already at an early stage in the history of the Monocotyledons. There is still much research needed with respect to the links between these families. Having outlined the systematic position of the Lemnaceae within the plant kingdom I will n o w discuss the internal relationships within the families on the basis of morphological and phytochemical characters. RELATIONSHIPS WITHIN THE LEMNACEAE ON THE BASIS OF MORPHOLOGICAL CHARACTERS In spite of the clearcut delimitation of the Lemnaceae, this family is internally n o t homogeneous. Hegelmaier (1868) distinguished two very well characterized subfamilies, the Lemnoideae and the Wolffioideae, within the Lemnaceae. The Lemnoideae are characterized by the presence of roots, two budding pouches situated in basal and lateral positions one on either side of the axis, and an inflorescence which develops from one of the budding pouches and which consists of one female and two male flowers enclosed by a membranous spathe. The anthers are bilocular and transversely dehiscent. Raphides are c o m m o n within the frond. The Wolffioideae have no roots, and only one budding pouch, which is situated in a basal and median position and never gives rise to an inflorescence. The inflorescence occurs in a dorsal position and consists of one female and one male flower w i t h o u t a spathe. The anthers are unilocular and apically dehiscent. Raphides are absent. Thus the two subfamilies differ in practically all characters in which they can differ. The main similarity is in the aquatic habit, the small size, the reduced inflorescence and t h a t the fronds of two or more generations can remain connected. However, up to the present time no one has ever doubted their relationship. The Wolffioideae are generally regarded as being derived from the Lemnoideae by reduction; however, the dorsal flowering cavity cannot be considered as a reduction, but is a new formation. In m y opinion, it is n o t homo-

409 logous with a second budding pouch, b u t histological studies are needed to establish its precise ontogenetic origin. The subfamily Lemnoideae consists of t w o genera, Spirodela and Lemna, which are both well-defined. In Spirodela the fronds have a dorsal and a ventral scale, one to many roots, 3--15 nerves, and a ventrally attached stipe. Moreover, the fronds contain brown pigment cells and druses within the parenchyma. Lemna is much more reduced, as it lacks the dorsal and ventral scale, and has only one r o o t (rarely none) and one to five, often indistinct nerves. The stipe is marginally attached and there are neither brown pigment cells nor druses within the parenchyma. Both genera are n o t homogeneous. Hegelmaier (1868) distinguished within Spirodela a " T y p u s der S. polyrhiza", with 5--12 mm long fronds and primary as well as secondary roots and a " T y p u s der S. oligorrhiza" with 2--5 mm long fronds with primary roots only, and with the habit of Lemna. In their Synopsis of the Lemnaceae, Den Hartog and Van der Plas (1970) considered these morphological differences t o o insignificant to regard them as separate sections of the genus. On morphological grounds, two subgenera can be recognized within the genus Lemna. The subgenus Lemna is characterized by round, elliptic, oblong or obovate floating fronds, with one root, and the slit of the budding pouch coincides with the margin of the frond or is situated slightly ventral to the margin, In the subgenus Staurogeton the fronds are oblong to linear or lanceolate and sometimes have no root. The fronds are submerged and rise to the surface in the flowering period. The slit of the budding pouch is situated just dorsal to the margin of the frond. The subgenus Staurogeton has been recognized as a separate taxon, although mostly in the rank of section, since 1830. The subgenus Lemna, as defined here, has been subdivided in various ways; the most elaborate subdivision can be found in the works of Hegelmaier (1868, 1895), who recognized four units within this taxon, all classified in 1895 as sections: Lemna (as "Eulemna"), Telmatophace, Alatae and Uninerves. Den Hartog and Van der Plas (1970) did n o t see the need for a further subdivision of the subgenus, as the differences between these sections are small. In particular the section Telmatophace, which contains only Lemna gibba L., cannot be maintained as a separate taxon (see also Kandeler, 1975), in view of the great similarity it shows with L. minor L. of the section Lemna (see De Lange, 1975; Pieterse, 1975 and Landolt, 1975). The subfamily Wolffioideae shows a greater complexity. Den Hartog and Van der Plas (1970) recognized four genera within this subfamily, viz. Wolffia, Wolffiella, Pseudowolffia and Wolffiopsis. These taxonomic units had already been recognized by Hegelmaier (1868), b u t are recorded in his work as taxa of lower rank within the genus Wolffia. In 1895 Hegelmaier removed his s,.bgenus Wolffiella from the genus Wolffia, raising it to generic rank. Wolffia remained,

410 however, a heterogeneous genus. Monod (1949) purified Wolffia, by transferring the taxa n o w classified in Pseudowolffia and Wolffiopsis, to Wolffiella, and making this genus a heterogeneous taxon. In 1970 Den Hartog and Van der Plas made the final move, by recognizing t w o new genera, for the units not fitting in Wolffia and Wolffiella. It has to be stated here, that the definition of the four units, n o w considered to be genera, has never been disputed. The characters of the four genera can be summarized as follows. In Wolffia the fronds are globular or ellipsoid and float on the surface. The budding p o u c h is funnel-shaped with a circular opening; the stipe is extremely reduced, and can only be found by dissection under the microscope. The fronds of Pseudowolffia are flat and symmetric, and round or nearly so, and also float on the surface. The budding p o u c h is triangular, flat and symmetric. The stipe is median, long, membranous and flat, The fronds o f the two other genera occur submerged, and rise only to the surface in the period of anthesis; they are thin, membranous and have dark pigment cells in the epidermis. In Wolffiella, the fronds are asymmetric, linear-oblong to almost strap-like and often falcate. The stipe is lateral and there is only one inflorescence. In Wolffiopsis, the fronds are symmetric, broad-elliptic or ovate, and curled. The short stipe is median and there are t w o inflorescences. Thus the generic subdivision of the Wolffioideae is mainly based on shape and s y m m e t r y of the fronds, and the position of the stipe. On the basis of the morphological characters, it seems clear that there is a reduction within the Lemnaceae from the most primitive genus Spirodela via Lemna to the Wolffioideae. In the reduction from Spirodela to Lernna, only vegetative characters are involved, e.g. the loss of the ventral and the dorsal scale, the reduction of the number of roots to one, etc. The generative structures in both genera are similar. The Wolffioideae show a further reduction of the vegetative structure: the loss of roots, the loss of nervation, the loss of raphides. In comparison to the Lemnoideae, the generative structures are also reduced; the inflorescence contains only one male flower instead of two, and there is no spathe. However, apart from all reductions the Wolffioideae have evolved a new structure, the dorsal flowering cavity. The morphological reduction series Spirodela -+ Lemna ~ Wolffioideae seems thus well documented. Within the Wolffioideae, such a series cannot be indicated, because the generic subdivision of this subfamily is based on derived characters, such as shape and symmetry of the fronds. On the basis of the presently known morphological data it is almost impossible to indicate one of the four genera as the most primitive one within the subfamily. The only fact is, that Wolffiopsis and Wolffiella are more closely related to each other than to the other t w o genera. Several authors (Maheswari, 1958; McClure and Alston, 1966; Turner, 1967; Blazey and McClure, 1968), however, consider Wolffia the most derived genus because of its reduced size, and possibly the reduction of its stipe.

411 RELATIONSHIPS WITHIN THE LEMNACEAE ON THE BASIS OF

PHYTOCHEMICAL

DATA The question remains whether the morphological reduction series, whi,:h in fact is the basis of the systematic arrangement within the Lemnaceae, is '..iso an evolutionary series. In order to answer this question, phytochemical, cytological, embryological and possibly other data have to be compared and tested. Fortunately, an interesting set of phytochemical data has been provided by McClure and Alston (1966), Blazey and McClure (1968) and McClure (1970, 1975). McClure and Alston (1966) made an excellent study of the flavonoid chemistry of 21 species of Lemnaceae. Their results have an obvious significance for the systematics of this family, as it appeared that all species investigated seem to be fully characterized by their set of flavonoid compounds. Even forms which are hardly distinguishable in terms of morphology, can be recognized on the basis of their flavonoid "fingerprints". These biochemical data partly confirm the existing morphology-based systematics, and are partly at variance with it. According to McClure and Alston (1966}, the genus Spirodela possesses compounds of the following flavonoid classes: giycoflavones, anthocyanins, flavonols and flavones. It appears at the species level, however, that the species which has been considered the most primitive one by Daubs (1965} and McClure and Alston (1966), because it very closely approached the hypothetical ancestral type postulated by Hegelmaier (1868) and Brooks {1940), viz. S. intermedia Koch, does not contain flavones, while S. polyrhiza (L.) Schleid. and S. biperforata Koch do not contain flavonols. Only S. punctata (G.F.W. Meyer) Thompson (under the name S. oligorrhiza (Kurz} Hegelm.) shows the whole range of flavonoids. Remarkably, the flavonoid compounds in the latter species are for ca 60% different from those of the other Spirodela species. There is, however, a striking similarity between the "fingerprints" of S. punctata and Lemna minor with respect to the glycoflavones and also in the occurrence of petunidin-3,5 diglucoside. These data indicate that Spirodela is not so homogeneous as usually has been assumed. The flavonoid chemical data sustain the vision of Hegelmaier (1868), that two "Typen" can be distinguished within the genus. It may be justifiable for S. punctata to make a special section, which can be characterized by morphological and phytochemical data. Furthermore, the primitivity of S. intermedia needs to be questioned. Although, according to its morphology, the most primitive member of the Lemnaceae, it cannot be so from a phytochemical point of view, because it seems to have lost its capacity to store flavones in measurable quantities. Thus a new hypothetical Spirodela has to be postulated which shares the general morphological structure of S. intermedia with a set of compounds of the four flavonoid classes. The morphological and phytochemical relations within the genus Spirodela are given in Fig.1. The genus Lemna lacks the flavonols. On the basis of the data provided by

412

Hypothetical Spirodela

ancestor

I°ss lof

flavones

Spirodela intermedio

reduction in size lossof secondary

roots

I°ss lof

Spirodela punctata

'Y l

flovonol$

flavono[s Spirodela polyrhiza Spirodela biperforata

reductionof number of roots

Lemna

Fig. 1. Morphological and flavonoid chemical relations within Spirodela.

McClure and Alston (1966), three groups can be distinguished, viz. (I) a group with exclusively glycoflavones, containing L. valdiviana Phil. (2) a group with glycoflavones and anthocyanins, containing L. minor, L. gibba, L. obscura Daubs and L. trisulca L. (3) a group with glycoflavones and flavones containing L. perpusilla Torrey, L. trinervis (Austin) Small and L. rninuscula Herter (under the name L. minima auct. non Beauvois). The first group is not very well defined, because it originally also contained L. minor, in which anthocyanins in the form of petunidin-3, 5 diglucoside have now been established (McClure, 1970, 1975). This species has now been transferred to the second group. The second group contains the species of the Lemna rninor--L, gibba complex (i.e. the section Lemna Hegelm. as defined by Kandeler (1975)) and the only species of the subgenus Staurogeton, which has been investigated. The third group contains section Alatae and L. rninuscula. As is apparent from the above, the flavonoid chemical data do not support the systematic subdivision of the genus Lemna, in casu the recognition of a subgenus Staurogeton. These data show, however, that phytochemically subgenus Staurogeton and section Lemna form a rather closed group. Further it is evident, that on the ground of the flavonoid chemistry the two species L. valdiviana and L. minuscula, classified by Hegelmaier in section Uninerves, have nothing in common. Furthermore, it is remarkable that none of the Lemna species contains compounds of all three flavonoid classes known from the genus. In the Wolffioideae, McClure and Alston (1966) investigated two genera. In the genus Wolffiella only flavonols were found, two of which are also known from Spirodela. The genus is extremely homogeneous, in contrast to Wolffia, of which five species were investigated. Three species of Wolffia, viz. W. punctata Griseb., W. papulifera Thompson

413

and W. microscopica (Griff.) Kurz appeared to have only flavonols, which establishes a relationship with Wolffiella, but two other species, W. arrhiza (L.) Horkel ex W i m m e r and W. columbiana Karsten appeared to be chemically completely different by the absence of flavonols and presence of glycoflavones and flavones. Morphologically these two groups are also distinct. W.arrhiza and W. columbiana are free of brown pigment cells,while in W. punctata and W. papulifera these are very obvious; W. microscopica is morphologically rather anomalous; it seems to lack these glandular cells,but this requires further confirmation. Regrettably, representatives of Wolffiopsis and Pseudowolffia have not been investigated chemically. The firstgenus has brown glandular cells,and seems to be closely related to WolffieUa. The three species described in Pseudowolffia do not possess brown pigment cells,but Rarnayya and Rajagopal (1975) recorded from the recently discovered population of P. hyalina (Del.) Hart. & Plas in India cells with a few droplets of an oily substance. I have wondered whether the occurrence of flavonols is linked with the brown pigment cells in Lemnaceae. Flavonols are absent in Lemna, which lacks brown pigment cells. However, Spirodela polyrhiza and S. biperforata, which possess such cells seem to be also devoid of flavonols (McClure and Alston, 1966). The heterogeneity of the secondary compounds in Wolffia has led to much speculation, and even the biphiletic origin of this genus has been postulated by Turner (1967), via the lines: Spirodela ~ Lemna ~ Wolffia p.p. Spirodela ~ Wolffiella ~ Wolffia p.p. Of course this situation is not very likely. In the first place, it is extremely improbable that a new formation such as the flowering cavity in the Wolffioideae would have evolved twice, viz. once during the supposed step Lemna Wolffia and the second time during the step Spirodela -~ Wolffiella. In m y opinion, the evolution of the Wolffioideae is much less simplistic, and in order to build up the evolutionary model of this subfamily at least the data on the biochemistry of Wolffiopsis and Pseudowolffia must be incorporated. On the basis of the data now available, the postulation of a protowolffioid, which contains glycoflavones, flavonols and flavones, from which Wolffia and Wolffiella developed as independent lines, cannot be avoided. In Wolffia a further differentiation t o o k place, as one group lost its glycoflavones and flavones and the other its flavonols. Also from the flavonoid chemical data as supplied by McClure and Alston (1966), it is n o t likely t h a t part of Wolffia developed from Wolffiella, because the number of flavonol compounds in Wolffia is considerably larger than in Wolffiella. A reversed sequence would be more likely from these data. The phytochemical heterogeneity of the genus Wolffia may have consequences for the t a x o n o m y ; to what e x t e n t this is the case, depends on the attitude of the investigator and forms a subjective element. If one accepts that the biochemical characters can be superior to the classic morphological characters,

414 as e.g. Turner (1967) does, the consequence m a y be that Wolffia has to be split into two genera. When, however, the morphological homogeneity of Wolffia is considered as being more important than its biochemical heterogeneity, one can decide t h a t at the most, the genus can be subdivided into two lower-ranked taxa. In m y opinion, such a fornlal step is still premature, until more is known about other Wolffioideae. Another phytochemical character, investigated by Blazey and McClure (1968), is the degree of lignification in the various genera. Their results showed that p - h y d r o x y b e n z a l d e h y d e was found in all species tested, that vanillin was found in all species of Spirodela and in four of the eight tested Lemna species and remarkably also in the flowers of Wolffia microscopica, and t h a t the third compound, syringaldehyde, appeared to be restricted to three of the four Spirodela species. The groups with and w i t h o u t vanillin in Lemna neither coincided with the groups distinguished on the basis of the flavonoid chemistry, nor with the sections of Hegelmaier. The restriction of the c o m p o u n d vaniUin to the flowers in Wolffia microscopica points to the fact t h a t the occurrence of certain phytochemical compounds can be linked with certain morphological structures. Moreover, the physiological condition of the plants m a y quantitatively, and possibly, also qualitatively influence the occurrence of various compounds. RELATIONSHIPS WITHIN THE LEMNACEAE ON THE BASIS OF CYTOLOGICAL DATA Cytological data on Lemnaceae are very scarce. Most older data are summarized in the work of Wcisto (1970), who studied the chromosomes of the Polish Lemnaceae. These data, together with the extensive data material on Lemna of Urbanska-Worytkiewicz (1975), permit the preliminary conclusion that in the Lemnaceae the basic chromosome number is 10. Furthermore, it seems that the chromosomes o f Spirodela are smaller than those of Lemna, and that Wolffia has the largest chromosomes. As the data on Spirodela and Wolffia are very limited, further research is needed for a firm establishment of such a size sequence. In Lemna species, the chromosome size is more or less the same. At the present time, the available cytological data are insufficient to support or to defy the accepted systematic relationships within the Lemnaceae. CONCLUSIONS AND RECOMMENDATIONS The main conclusion which can be drawn from the comparison o f the morphology-based t a x o n o m y of the Lemnaceae with phytochemical data is t h a t the investigated genera Spirodela, Lemna and Wolffiella are confirmed, but that the two approaches lead to a discrepancy in the case of Wolffia. Furthermore, it appears that the subdivision of the genera according to morphological characteristics is only partly confirmed b y phytochemical data. However, in Lernna, which is the best-known genus, flavonoid chemical characters lead to a different subdivision than one would obtain by using the lignin

41.5

characters. Both subdivisions are n o t in agreement with the morphological subdivision. For this reason, it has to be stressed that great care is needed for the introduction of phytochemical data as taxonomic characters above the species level. In fact, one needs to k n o w the pathways and the physiological significance of the compounds, before one can appreciate t':~ir value as systematic characters, and one needs a full set o f data. The work of McClure and collaborators satisfies this condition, inspite of the gaps which still exist in our knowledge. As a full evaluation of the flavonoid chemical data for the t a x o n o m y of the Lemnaceae can only be obtained by the closure of these gaps, the following recommendations are proposed: (1) A study of the flavonoid chemistry of members of the genera Wolffiopsis and Pseudowolffia. (2) A study of the flavonoid chemistry of the South African Wolffiella denticulata (Hegelm.) Hegelm., which is slightly aberrant, in respect to its morphology. (3) A study of the flavonoid chemistry of other Wolf[ia species. (4) A study of the flavc:loid chemistry of Pistia stratiotes L. As Pistia is considered by several investigators to be a link in the line Araceae -* Lemnaceae on morphological grounds, it seems extremely interesting to test wether such a relationship is supported b y phytochemical data. Furthermore, a study of the chromosome numbers in particularly within the subfamily Wolffioideae must be recommended. ACKNOWLEDGEMENTS

I am much indebted to Dr L. de Lange, Amsterdam, and Professor J.W. McClure, Oxford, Ohio, U.S.A., for their critical remarks on m y paper and their stimulating comments, and to Mr F. van der Plas, Delft, for his collaboration over the years.

REFERENCES

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416 Erdtman, G., 1952. Pollen Morphology and Plant Taxonomy. I. Angiosperms. Almquist and Wiksell, Stockholm; Chronica Botanica, Waltham, Mass. Hegelmaier, G., 1868. Die Lemnaceen, eine Monographische Untersuchung. Wilhelm Engelmann, Leipzig, 169 pp. Hegelmaier, G., 1895. Systematisches Ubersicht der Lemnaceen. Englers Bot. Jahrb., 21: 268--305. Hegnauer, R., 1963. Chemotaxonomie der Pflanzen, eine Uebersicht fiber die Verbreitung und die systematische Bedeutung der Pflanzenstoffe. 2. Monocotyledonae. Birh//user, Basel und Stuttgart, 540 pp. Kandeler, R., 1975. Species delimitation in the genus Lemna. Aquat. Bot., 1: 365--376. Landolt, E., 1975. Morphological differentiation and geographical distribution of the Lemna gibba--Lemna minor group. Aquat. Bot. 1: 345--363. Lawalrde, A., 1945. La position systdmatique des Lemnacdes et leur classification. Bull. Soc. R. Bot. Belg., 77 : 27--38. McClure, J.W., 1970. Secondary constituents of aquatic angiosperms. In: J.B. Harborne (editor), Phytochemical Phylogeny. Aead. Press, New York, N.Y., pp. 233--268. McClure, J.W., 1975. The applicability of polyphennllc data to systematic problems in the Lemnaceae. Aquat. Bot., 1: 395--405. McClure, J.W., and Alston, R.E., 1966. A chemotaxonomic study of Lemnaceae. Am. J. Bot., 53: 849--860. Maheswari, S.C., 1958. Spirodela polyrhiza: The link between the aroids and the duckweeds. Nature, 181: 1745--1746. Monod, T., 1949. Sur une Lemnacde Africaine: Wolffiella welwitschii (Hegelmaier, 1865) comb. nov. -- Travaux Botaniques dddids A Rend Malre. Mem. Hors.Sdr. Soe. Hist. Nat. Afr. Nord., pp. 229--242. Pieterse, A~I-I., 1975. Physiological, morphological and anatomical aspects of gibbosity in Lemna gibba. Aquat. Bot. 1 : 333--344. Ramayya, N. and Rajagopal, T., 1975. Pseudo wolffia hyalina (Del.) Hart. & Plas. (Lemnaceae) - - A new record for Asia from Hyderabad (India). Aquat. Bot., 1 : 7 1 - - 7 3 . Thanikaimoni, G., 1969. Esquisse palynologique des Aracdes. Trav. Sect. Sci. Tech. Inst. F r Pondichery, 5 (5), 31 pp. Turner, B.L., 1967. Plant chemosystematics and phylogeny. Chem. Nat. Products, 4: 189-213. Urbanska-Worytkiewicz, K., 1975. Cytological variation within Lemna L. Aquat. Bot., 1 : 377--394. Van der Plas, F., 1971. Lemnaceae. Flora Malesiana Ser. I Spermatophyta, 7: 219--237. Velenovsky, J., 1907. Vergleichende Morphologie der Pflanzen. 2.. Verlagsbuchhandlung Von Fr. Rivnac, Prag, 731 pp. Wcigro, H., 1970. Karyological studies in Polish representatives of Spadiciflorae. Act. Biol. Cracov. Ser. Bot., 13: 79--88, PI. 14. 2. Verlagsbuchhandlung Von Fr. Rivnac,