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Landoltia (Lemnaceae), a New Genus of Duckweeds Donald H. Les Department of Ecology and Evolutionary Biology, The University of Connecticut, Storrs, Connecticut 06269-3043, U.S.A. Daniel J. Crawford Department of Plant Biology, The Ohio State University, Columbus, Ohio 43210, U.S.A. Abstract. Morphological, allozyme, and cpDNA ter species, but with S. punctata associating with (r6cL) sequence data provide evidence for the dis¬ Lenina (Fig. 1A). tinctness of Spirodela punctata from species in both Although Landolt’s evolutionary trees reflect a Lenina and Spirodela (Lemnaceae). We propose the keen understanding of duckweeds and a compre¬ recognition of a new genus, Landoltia, to better re¬ hensive evaluation of published taxonomic litera¬ flect current phylogenetic concepts in the Lemna¬ ture on the group, they were constructed using non¬ ceae. Landoltia is distinguished by its reduced explicit phylogenetic methods, i.e., not by cladistic frond prophyllum, frond nerves (3 to 7), roots (up analyses. We have re-analyzed results of these ear¬ to 7), root tracheids, external anther locules, and lier studies using cladistic methodologies to test hy¬ also by well-supported molecular evidence provid¬ pothetical relationships proposed by Landolt. In ad¬ ed by allozymes and cpDNA sequences. The new dition, we have generated several molecular data combination Landoltia punctata is made to accom¬ sets to supplement the clearly limited number of modate this taxonomic modification. phylogenetically informative characters available for these morphologically simple plants. Although Duckweed classification remains equivocal be¬ this work remains in progress, the results of our cause phylogenetic relationships are difficult to rec¬ preliminary analyses of morphological, biochemi¬ oncile in this diminutive, reduced family (Les et cal, allozyme, and DNA (rftcL) sequence data al., 1997b). The most comprehensive phylogenetic (Crawford & Landolt, 1993, 1995; Crawford et al., hypotheses for duckweed taxa were presented in a 1995, 1997; Crawford et al., 1996; Les et al., 1994, monograph by Landolt (1986) and form the basis 1997a, 1997b) are reasonably concordant with Lan¬ of modern classification for the family Lemnaceae. dolt’s classification, but differ in a number of details In the most recent monograph of the duckweed from his phylogenetic diagrams. In particular, ge¬ family (Lemnaceae), Landolt (1986) recognized four neric subdivisions used in his classification of Lem¬ genera: Lenina, Spirodela, Wolffia, and Wolffiella. naceae are inconsistent with results of these phy¬ Few additional genera have been proposed in past logenetic analyses and merit reconsideration. taxonomic treatments of duckweeds. One example Herein we summarize evidence that warrants the is Staurogeton Reichenbach, which was elevated taxonomic segregation of a new duckweed genus. from subgeneric to generic rank by Schur (1866) to We consider this taxonomic action essential for a accommodate the morphologically distinctive Leni¬ classification that reasonably depicts our best, cur¬ na trisulca L. (Landolt, 1986). Most contemporary rent estimate of phylogenetic relationships in the classifications continue to assign this taxon to the Lemnaceae. genus Lenina. Den Hartog and van der Plas (1970) A specific diagram of intergeneric duckweed re¬ subdivided Wolffiella to create the two genera Pseu- lationships that summarizes the phylogenetic trees dowolffia and Wolffiopsis. Few taxonomic treatments recognize either of these segregate genera as dis¬ originally appearing in Landolt (1986) was provid¬ tinct from Wolffiella. To our knowledge, division of ed to us by E. Landolt. We compared these hypo¬ either Spirodela or Wolffia into subsidiary genera thetical relationships to published allozyme studies has not yet been suggested, although some authors (Crawford & Landolt, 1993) and to previous cla¬ have transferred certain species from these genera distic analyses of morphological, anatomical, and into either Lenina or Wolffiella. Landolt s recent ge¬ biochemical data (Les et al., 1997b). Intergeneric neric concept of Spirodela recognizes a paraphylet- duckweed relationships were also examined using ie taxon, with .S’, polyrhiza and S. intermedia as sis¬ preliminary results of a phylogenetic analysis of Novon 9: 530-533. 1999. Volume 9, Number 4 Les & Crawford 531 1999 New Duckweed Genus Wolffia A. rbcL sequence data for the Lemnaceae (Les et al., Wolffiella 1997b). Non-molecular data (Fig. IB) resolve the entire Lemna genus Spirodela as paraphyletic. However, S. punc¬ Spirodela punctata tata lies distinct from the other Spirodela species Spirodela polyrhiza as a separate branch, and with good internal sup¬ Spirodela intermedia port (75% bootstrap value). Strong bootstrap values (99% and 84%, respectively) from molecular (rbcL) data (Fig. 1C) support S. polyrhiza and S. inter¬ media as sister species, and also their distinction Wolffia from S. punctata. Allozyme data (Crawford & Lan¬ B. dolt, 1993) show a moderate genetic identity be¬ Wolffiella tween S. polyrhiza and .S’, intermedia (/ = 0.404), Lemna yet they share no electrophoretically detectable al¬ Spirodela punctata leles with S. punctata (/ = 0.000). Various mor¬ phological features (Table 1) are consistent with a Spirodela polyrhiza phylogenetic position for S. punctata intermediate Spirodela intermedia between Lemna and other Spirodela species. By in¬ spection of these features, the species S. punctata is not only morphologically distinct from both Lem¬ p, Wolffia na and Spirodela, but intermediate and transitional between these genera. Wolffiella Presently, Spirodela Schleiden comprises three Lemna distinct species: Spirodela intermedia W. Koch, S. Spirodela punctata polyrhiza (L.) Schleiden, and S. punctata (G. Meyer) C. H. Thompson. Landolt (1986) placed the former Spirodela polyrhiza two species into Spirodela sect. Spirodela, and seg¬ Spirodela intermedia regated the latter in section Oligorrhizae W. Koch. Spirodela punctata has been taxonomically prob¬ Figure I. Phylogenetic position of Spirodela punctata as indicated in several recent studies. —A. Landolt (1986) lematic because it possesses features similar to hypothesized the association of 5. punctata with Lemna, both Spirodela and Lemna (Table 1; Landolt, 1986). in a paraphyletic concept of Spirodela.—B. Non-molecular Meyer (1818) originally named Spirodela punc¬ data (Les et ah, 1997b) place S. punctata in a separate tata as Lemna punctata, but it was not until 50 clade from lemna, but also apart from other Spirodela species (bootstrap % shown). —C. Molecular (rftcL) data years later that Hegelmaier (1868) transferred the provide strong support (bootstrap % shown) for the dis¬ taxon (as S. oligorrhiza) to Spirodela. In Meyer’s tinctness of S. punctata from either lemna or Spirodela time, all Lemnaceae were included in the genus (Les et al., 1997b). All evidence points to an isolated po¬ Lemna. The new genera Spirodela and Wolffia were sition of S. punctata in the Lemnaceae, and its recognition as a distinct genus is compatible systematically with any created in 1839 and 1844, respectively, and Wolf¬ ol these results. fiella was established in 1895 (Landolt, 1986). It is Table 1. Morphological features compared among species of Spirodela and Lemna. Spirodela punctata is interme¬ diate between Lemna and other Spirodela species for the character states indicated (from Landolt, 1986, 1998; Shill, 1979). Feature 5. intermedia; S. polyrhiza S. punctata lemna Prophyllum at hase of frond present present, but reduced absent No. of veins in frond 7 to 16 3 to 7 1 to 5 No. of roots 7 to 21 1 to 7 (12) 1 Boot tracheids extend to tip basal only absent Dorsal ineristem of new on one side1 on both sides on both sides fronds Lxternal anther locules do not extend above inter¬ extend slightly above inter¬ extend above internal loc¬ nal locules nal locules ales 1 Lateral on other side. 532 Novon tht- merit of Hegelmaier (1868, 1895) to have clear¬ tigations. Phylogenetically, our broad-based/wide- ly separated the genera Lenina and Spirodela. He ranging studies indicate that S. punctata is indeed also demonstrated accurately and comprehensively transitional between, but not a member of either the special position of S. punctata (called S. oli- Lemna or Spirodela. It is for this reason that we gorrhiza) within the genus Spirodela. establish a new genus to better reflect this revised Spirodela punctata is very polymorphic in rela¬ hypothesis of duckweed relationships. tion to size, pigmentation, number of roots, and veins. The high level of variability led Hegelmaier Landoltia D. H. Les & D. J. Crawford, gen. nov. (1895) to distinguish two species (S. oligorrhiza and TYPE: Lemna punctata G. Meyer: Prim. FI. 5. pusilla) and to describe three other species of Esseq. 262. 1818. = Landoltia punctata (G. questionable status. Growth experiments (Landolt, Meyer) D. H. Les & D. J. Crawford. 1986; Landolt & kandeler, 1987) and allozyme studies (Crawford & Landolt, 1993) have demon¬ Herbae ex radieibus 2 ad 7 (raro 1 vel 8 ad 12) usque strated that genetic variation in S. punctata is rath¬ ad 7 cm longis, omnibus prophyllum perforantibus; tur- er limited, and many of the differences used to dis¬ ionibus absentibus. Frondes in summa aqua natantes, ova- tae ad lanceolatae, 1.5—2.0-plo longiores quam latiores, tinguish former taxa are induced environmentally. supra nitidae viridesque serie mediana papillarum orna- Thompson transferred L. punctata to Spirodela in tae, subtus laeves rubraeque; nervis 3 ad 7. Flores infre- 1898. Landolt (1986) observed that some authors quentes; antherae loculis externis super internos positis. have merged the genera Lemna and Spirodela be¬ Fructus ala laterali in parte supera praedita; seminibus I vel 2 manifeste 10 ad 15-eostatis. cause of their similar appearance and because the features of S. punctata are transitional between the Roots 2 to 7 (rarely 1 or 8 to 12), up to 7 cm genera. Yet, he clearly differentiated Spirodela (in¬ long, all perforating the prophyllum. Turions ab¬ cluding 5’. punctata) from Lemna by the reduced sent. Fronds floating on the surface of the water, prophyllum at the base of its fronds, druse crystals, ovate to lanceolate, 1.5—2 times longer than wide, pigment cells, multiple roots, better developed tra- above shining and green with a medial series of cheids, and other anatomical/morphological fea¬ papillae, below smooth and red; veins 3 to 7. Flow¬ tures (Landolt, 1986). To our knowledge, the ge¬ ers infrequent; external locules of the anther above neric distinctness of S. punctata from both the internal locules. Upper part of fruit with a lat¬ Spirodela and Lemna has not been suggested pre¬ eral wing; seeds 1 or 2 with 10 to 15 distinct ribs. viously. We have now examined relationships of duck¬ Landoltia punctata (G. Meyer) D. H. Les & I). J. weed genera using morphological, anatomical, fla- Crawford, comb. nov. Basionym: Lemna punc¬ vonoid, allozyme, and rbcL sequence data. As sum¬ tata G. Meyer, Prim. FI. Esseq. 262. 1818. marized in Figure 1, analyses of these data sets Spirodela punctata (G. Meyer) C. H. Thomp¬ indicate that Spirodela punctata represents an iso¬ son, Rep. (Annual) Missouri Bot. Gard. 9: 28. lated clade distinct from both Spirodela and Lemna. 1898. TYPE: Chile. Tierra del Fuego Island, Cladograms constructed from either morphological Orange Harbor, leg. Wilkes expedition 1838 or flavonoid data (or their combination) show high (neotype, US not seen; isoneotypes, DS, GH, internal support (75—97% bootstrap values) for the KANU, MO not seen). distinctness of S. punctata from section Spirodela (Les et ah, 1997b) and support Landolt’s original Lemna oligorrhiza kurz, J. Finn. Soc.. Bot. 9: 267. 1866. phylogenetic concept that recognized Spirodela as Spirodela oligorrhiza (Kurz) Hegelmaier, Die l.em- paraphyletic with respect to the position of S. punc¬ naceen 147. 1868. TYPK: India. Calcutta, Kurz IH65 tata (Landolt, 1986). (holotype, CAF? not seen; isotypes, k, MEF, FI not seen). Allozyme data (Crawford & Landolt, 1993, & un¬ published) indicate a complete lack of genetic The generic name Landoltia commemorates Eli¬ identity between S. punctata and any species in as Landolt for his outstanding contributions to the either Lemna or Spirodela, yet the two species of systematies and biology of Lemnaceae in his more Spirodela sect. Spirodela do retain a moderate ge¬ than 45 years of research on duckweeds. netic; identity. Chloroplast DNA (r6cL) sequence data (Les et ah, 1997b) resolve S. punctata in a Acknowledgments. We thank D. Keil for his as¬ clade between Spirodela and Lemna but not within sistance in preparing the Latin diagnosis and R. either genus. In summary, these results echo the Rutishauser for his helpful comments. This work transitional nature and unsettled taxonomic status was supported in part by NSF grant DEB-9806537 of .S', punctata manifest in prior systematic inves¬ to the authors. Volume 9, Number 4 Les & Crawford 533 1999 New Duckweed Genus Literature Cited Pp. 1—122 in E. Landolt, I. Jager-Zurn & If. A. A. Schnell (editors), Handbuch der Pflanzenanatornie, Crawford, I). J. & E. Landolt. 1993. Allozyme studies in Band 13, Teil 4: Extreme Adaptions in Angiospermous Spirodela (Lemnaceae): Variation among nonspecific Hydrophytes. G. Borntraeger, Berlin, Germany. clones and divergence among the species. Syst. Bot. 18: -& R. Kandeler. 1987. The family of Lemnaceae— 389-394. A monographic study, vol. 2. Veroff. Geobot. Inst. ETH -& -. 1995. Allozyme divergence among Stiftung Rifbel Zurich 95: 1-638. species of Wolffia (Lemnaceae). PI. Syst. Evol. 197: 59— Les, D. H., E. Landolt & D. J. Crawford. 1994. Molecular 70. systematics of the Lemnaceae. Amer. J. Bot. 81 (6, -. -. I). H. Les & E. Tepe. 1995. Allozyme suppl.): 168-169. divergence among species of Wolffiella (Lemnaceae). -, -& -. 1997a. Systematics of Lem¬ Amer. J. Bot. 82 (6, suppl.): 122. naceae: Inferences from micromolecular and morpho¬ -. -& -. 1996. An allozyme study of two sibling species of Lemna (Lemnaceae) with com¬ logical data. PI. Syst. Evol. 204: 161—177. -, D. J. Crawford. E. Landolt, R. Aakjar & E. Tepe. ments on their morphology, ecology, and distribution. Bull. Torrey Bot. Club 123: 1-6. 1997b. Systematics of Lemnaceae revisited. Amer. J. -.-,-& E. Tepe. 1997. Allozyme var¬ Bot. 84 (6, suppl.): 211. iation and the taxonomy of Wolffiella (Lemnaceae). Meyer, G. E W. 1818. Primitiae Florae Essequeboensis Aquatic Bot. 58: 43—54. adjectis descriptionibus centum circiter stirpium nova- Den Hartog, C. & F. van der Plas. 1970. A synopsis of rum, observationibusque criticis. H. Dieterieh, Gottin¬ the Lemnaceae. Blumea 18: 355—368. gen. Hegelmaier, F. 1868. Die Lemnaceen. Eine monographis- Schur, P. J. F. 1866. Enumaratio plantarum Transylvaniae. che Untersuchung. Engelmann, Leipzig. Vindobonae. -. 1895. Systematisch Uebersicht der Lemnaceen. Shih, C. Y. 1979. SEM studies of the flowering of duck¬ Bot. Jahrb. 21: 268—305. weed, Lemna perpusilla, 6746. Scan. Electron Micros¬ Landolt, E. 1986. The family of Lemnaceae—A mono¬ cop. 1979: 479^186. graphic study, vol. I. Veroff. Geobot. Inst. ETH Stiftung Thompson, C. H. 1898. A revision of the American Lem¬ Rifbel Zurich 71: 1—566. naceae occurring north of Mexico. Rep. (Annual) Mis¬ -. 1998. Anatomy of the Lemnaceae (duckweeds). souri Bot. Gard. 9: 21—42.