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Multi-drug resistance (MDR) is a critical problem in cancer chemotherapy. MDR causes the overexpression of ATP-binding cassette (ABC) transporters and mutations in tumor suppressor genes and oncogenes. To tackle this issue, in this study, we focused on Nuphar plants, which have been traditionally used as food. Sesquiterpene alkaloids (1–3) were isolated from N. japonicum and dimeric sesquiterpene thioalkaloids (4–10) were isolated from N. pumilum. P-glycoprotein-overexpressing CEM/ADR5000 cells were cross-resistant to 6,6′-dihydroxythiobinupharidine (10). Using in silico molecular docking, we calculated the binding energies and simulated the interactions of these compounds with the corresponding amino acid residues at the binding site of P-gp. In addition, we investigated the cytotoxicity of these compounds towards cell lines overexpressing other ABC transporters (BCRP, ABCB5), cell lines with a knocked out tumor suppressor gene TP53 or cell lines overexpressing a deletion-activated EGFR oncogene. These cell lines were sensitive or only minimally cross-resistant to these compounds compared with their corresponding wild-type cell lines.

Nuphar alkaloids, originally isolated from water lilies, induce apoptosis in mammalian cells in less than one hour, making them possibly the fastest known inducers. However, the mechanism by which this rapid apoptosis occurs remains unknown. We have investigated canonical aspects of apoptosis to determine how the nuphar alkaloid, (+)-6-hydroxythiobinupharidine (6HTBN), induces apoptosis. 6HTBN induced rapid apoptosis in various leukemia, lymphoma and carcinoma cell lines, suggesting that its mechanism is cell-type independent. It also circumvented resistance of patient-derived chronic lymphocytic leukemia cells generated by co-culture on survival-promoting stroma. Intriguingly, 6HTBN failed to induce apoptosis in platelets. The mechanism of apoptosis involves activation of caspase 9 and caspase 3, but not caspase 8 as previously reported. The release of cytochrome c from mitochondria occurred even in the absence of BAX/BAK and in cells that retained mitochondrial membrane potential. These results suggest a novel mechanism of apoptosis that has previously not been reported. The molecular target of the nuphar alkaloids remains to be determined.

The order Nymphaeales, consisting of three families with a record of eight genera, has gained significant interest from botanists, probably due to its position as a basal angiosperm. The phylogenetic relationships within the order have been well studied, however, a few controversial nodes still remain in the Nymphaeaceae. The position of the Nuphar genus and the monophyly of the Nymphaeaceae family remain uncertain. This study adds to the increasing number of the completely sequenced plastid genomes of the Nymphaeales and applies a large chloroplast gene data set in reconstructing the intergeneric relationships within the Nymphaeaceae. Five complete chloroplast genomes were newly generated, including a first for the monotypic Euryale genus. Using a set of 66 protein-coding genes from the chloroplast genomes of 17 taxa, the phylogenetic position of Nuphar was determined and a monophyletic Nymphaeaceae family was obtained with convincing statistical support from both partitioned and unpartitioned data schemes. Although genomic comparative analyses revealed a high degree of synteny among the chloroplast genomes of the ancient angiosperms, key minor variations were evident, particularly in the contraction/expansion of the inverted-repeat regions and in RNA-editing events. Genome structure, and gene content and arrangement were highly conserved among the chloroplast genomes. The intergeneric relationships defined in this study are congruent with those inferred using morphological data.

Herein, we describe the first total syntheses of five members of the dimeric nuphar alkaloids: (+)-6,6′-dihydroxythiobinupharidine (+)-1 a, (+)-6-hydroxythiobinupharidine (+)-1 b, (−)-6,6′-dihydroxythionuphlutine (−)-2 a, (−)-6,6′-dihydroxyneothiobinupharidine (−)-3 a, and (+)-6,6′-dihydroxyneothionuphlutine (+)-4 a. The latter two have not been found in nature. We have also made each of their enantiomers (−)-1 a–b, (+)-2 a, (+)-3 a, and (−)-4 a. The key step in these syntheses was the dimerization of an α-aminonitrile (a hydrolytically stable surrogate for its corresponding hemiaminal) with chiral Lewis acid complexes. We have also reassigned the literature structures of (+)-1 a–1 b—for those instances in which the NMR spectra were obtained in CD3OD—to their corresponding CD3O-adducts. Our efforts provide for the first time apoptosis data for (−)-3 a, (+)-4 a, and all five non-natural enantiomers prepared. The data indicate high apoptotic activity regardless of the enantiomer or relative stereochemical configuration at C7 and C7′.

Background Multidrug-resistant bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin resistant enterococci (VRE), cause serious infections at clinical sites, for which the development of new drugs is necessary. We screened candidates for new antibiotics and investigated its action mechanism. Methods An antimicrobial compound was isolated from an extract of Nuphar japonicum . Its chemical structure was determined by NMR, MS, and optical rotation. We measured its minimum inhibitory concentration (MIC) using the microdilution method. The effects of the compound on DNA gyrase and DNA topoisomerase IV were investigated with DNA supercoiling, decatenation, and cleavage assay. Results We isolated and identified 6,6′-dihydroxythiobinupharidine as the antimicrobial compound. The MIC of this compound was 1–4 μg/mL against various MRSA and VRE strains. We also demonstrated that this compound inhibited DNA topoisomerase IV (IC 50 was 10–15 μM), but not DNA gyrase in S. aureus , both of which are known to be the targets of quinolone antibiotics and necessary for DNA replication. However, this compound only exhibited slight cross-resistance to norfloxacin-resistant S. aureus , which indicated that DTBN might inhibit other targets besides topoisomerase IV. These results suggest that 6,6′-dihydroxythiobinupharidine may be a potent candidate or seed for novel antibacterial agents. Conclusions DTBN from N. japonicum showed anti-MRSA and anti-VRE activities. DTBN might be involved in the inhibition of DNA topoisomerase IV. General significance DTBN might be useful as a seed compound. The information on the inhibition mechanism of DTBN will be useful for the modification of DTBN towards developing novel anti-MRSA and anti-VRE drug.

Premise of research. Fossil remains of a water lily, Nuphar carlquistii DeVore, Taylor & Pigg sp. nov. (Nymphaeaceae), are described from the latest early Eocene Republic flora of north-central Washington. Specimens include several fruits, one with in situ seeds, others with remnants of stamens and stamen attachment scars, and dispersed seeds found both within a mucilaginous dispersal matrix and separately. Associated structures, including a stigmatic disk, isolated tepals, and several rhizome segments bearing nested clusters of roots and leaf scars, are included in N. cf. carlquistii.Methodology. Fossils preserved in a lacustrine shale were degaged to reveal details, photographed with LM, and compared morphologically with extant material of related plants.Pivotal results. The study documents the most complete fossil Nuphar known to date. The combination of multiple organs, including fruits, seeds, stamens, and rhizomes with leaf scars and clustered root scars, demonstrates that Eocene Nuphar had vegetati...

Ethnopharmacological relevance Various plant organs of Nuphar lutea (L.) SM. (Nymphaeaceae) are used in traditional medicine for the treatment of arthritis, fever, aches, pains and inflammation. The main purpose of this study was to determine the anti-inflammatory effect of Nuphar lutea leaf extract (NUP) in two septic shock models: (1) Survival of mice challenged with a lethal dose of LPS, determination of pro-inflammatory and anti-inflammatory cytokines in serum, as well as in peritoneal macrophages in cell culture. (2) The effect of NUP in a murine model of fecal-induced peritonitis. Materials and methods NUP pre-treatment partially protected mice in two models of acute septic shock. We concluded that NUP is anti-inflammatory by inhibiting the NF-κB pathway, modulating cytokine production and ERK phosphorylation. Results A significant average survival rate (60%) of LPS lethally-challenged mice was achieved by pre-treatment with NUP. In addition, NUP pre-treatment reduced nuclear NF-κB translocation in peritoneal macrophages. The production of pro-inflammatory cytokines, TNF-α, IL-6 and IL-12, in the sera of LPS-treated mice or in the supernatants of peritoneal macrophages stimulated with LPS for 2–6 h was also decreased by NUP. Pre-treatment with NUP caused a significant increase in the anti-inflammatory cytokine IL-10. The NUP pre-treatment reduced and delayed mortality in mice with fecal-induced peritonitis. Our studies also revealed that NUP pre-treatment induced a dose-dependent phosphorylation of ERK in peritoneal macrophages. Since most of the reports about the anti-inflammatory effect of Nuphar lutea refer to rhizome and root powder and extracts, it is important to clarify the effectiveness of leaf extract as a source for such activity. Conclusion NUP pre-treatment partially protected mice in two models of acute septic shock. We concluded that NUP is anti-inflammatory by inhibiting the NF-κB pathway, modulating cytokine production and ERK phosphorylation.

Nymphaeaceae occupies an important phylogenetic position because of their placement as one of the basal angiosperms. From this perspective, morphological studies in the family are of great value to understanding plant phylogeny and evolution. Ovule development and female gametophyte in Nymphaea amazonum, N. gardneriana (subgenus Hydrocallis) and in Victoria cruziana were analyzed in order to provide further progress in characters of potential evolutionary interest. The ovules of all species are anatropous, bitegmic, distomic, weakly crasinucellate, and present an epistase. The female gametophyte is four-celled and corresponds to the Schisandra type, distinctive of the Nymphaeales. Comparisons among ovules of the subgenera of Nymphaea and others allied genera show differences with respect to micropyle conformation, thickness of nucellus and outer integument, and its degree of development on the raphal side. The studied species of Nymphaea share an annular outer integument and linear triad of megaspores. These results fill gaps in the current incomplete knowledge of character states especially within Nymphaea. In subgenus Hydrocallis, the ovules have an outer integument not markedly cup-shaped in contrast to subgenus Nymphaea, since the micropyle is closer to the funiculus as in Nuphar. The present observations suggest that the ovule morphology has diversified in Nymphaea and the characters studied clearly show no evidences to support the hypothesis of a monophyletic genus. Fil: Zini, Lucia Melisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Nordeste. Instituto de Botánica del Nordeste (i); Argentina Fil: Galati, Beatriz Gloria. Universidad de Buenos Aires. Facultad de Agronomia; Argentina Fil: Ferrucci, Maria Silvia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Nordeste. Instituto de Botánica del Nordeste (i); Argentina

The author gives a new syntaxonomic classification of the phytocenoses included in the association Myriophyllo-Nupharetum described by W. Koch (1926) without any documentation material. After analysing 1554 phytosociological records from the territory of Poland, in which species considered as characteristic for Myriophyllo-Nupharetum (Nuphar luteum, Nymphaea alba, N. candida and Myriophyllum verticillatum) were the forming species (in this case also the dominating ones), it was considered necessary to classify these phytocenoses to three separate associations: Nuphareto-Nymphaeetum albae, Nymphaeetum candidae and Myriophylletum verticillati. A table representing the association Nuphareto-Nymphaeetum albae, 3 synthetic tables and maps of distribution of the so far recorded sites or groups of sites of the above

A new genus and species, Notonuphar antarctica, is described from the Eocene of Seymour (Marambio) Island, the Antarctic Peninsula and assigned to the Nymphaeales based on well-preserved seeds. This is the first record of a water lily from Antarctica and the first record of a Gondwanan plant with close link to the genus Nuphar (Nymphaeaceae), which is restricted today to the Northern Hemisphere. Critical features for systematic placement of Notonuphar are the presence of a germination cap with closely spaced hilar scar and micropyle, anatropous, bitegmic and exotestal seed organization, exotesta composed of one cell layer of high sclerenchymatic palisadeshape cells, mesotesta of smaller, low parenchymatic cells, a few cell layers deep, and a thin tegmen. The seeds of Notonuphar are particularly similar to seeds of extant and fossil Nuphar in the straight, unfolded anticlinal wall of the exotestal cells and the presence of a narrow zone of exotestal tissue between hilum and micropyle. Other seed features including the very tall exotestal cells and strongly thickened cell walls of exotesta also link Notonuphar to Brasenia and related fossil taxa (Cabombaceae). This character mosaic observed in Notonuphar corroborates the transitional position of Nuphar between Cabombaceae and Nymphaeaceae. Notonuphar is the only member of Nymphaeales recorded from Antarctica and so far the only fossil seeds of Nymphaeales known from the Southern Hemisphere. The discovery of this extinct Gondwanan taxon with features suggesting close relationship with extant Northern Hemisphere genus Nuphar is a further evidence for the relictual nature of the extant group., Facultad de Ciencias Naturales y Museo

A class of monomeric nuphar analogues that are either epimeric at C1 and C1′ or lack the naturally occurring methyl group at those positions were synthesized and evaluated for biological activity. The syntheses feature enantioselective vinylogous Mukaiyama–Mannich (vM–Mannich) reactions catalyzed by chiral phosphoric acids that proceed with excellent diastereoselectivity. Biological assays reveal that both the desmethyl and C1-epimeric monomeric nuphar analogous are able to induce rapid apoptosis.

An enantiospecific synthesis of a nuphar alkaloid was achieved in nine steps from N-Boc-(L)-proline. The alkaloid is a minor component of castoreum, which is secreted from the dried scent glands of a beaver. During the course of our study, the stereochemistry of three synthetic intermediates was verified by X-ray crystal structure analysis, which helped to resolve the existing discrepancies among reports regarding the structure of this particular compound. On the basis of our synthesized alkaloid, we proposed the structure of the natural product. Also, intrigued by the therapeutic uses of castoreum, which was employed for gynecological purposes in ancient Greece and Rome, we conducted a biological screening and found that the alkaloid has affinity for the oxytocin receptor.

We show a possible palaeoenvironmental change, a short warm and dry environment, in the Early Jomon period suggested by disappearance of the aquatic plant Nuphar and AMS 14C dating. The Nakayama No.2 Mire site is located near the pass at 870m a.s.l. in Hokkaido. Although the climate was cool and rainy during 9,000 - 10,000 cal BP in the Early Jomon period, a short warm and dry period of about 65 years existed at around 9700 cal BP., 北緯43度の北海道中山峠に隣接する中山第2湿原で縄文早期のころは冷涼で降雨量が多い環境がおよそ2330年間も続いていたことを池塘に生育するコウホネ属化石花粉の存在から推定した。ところがコウホネ属化石が連続して欠落する部分があり、そこではコナラ属の増加とモチノキ属の増加、そしてトウヒ属の急減がある。この温暖で乾燥した期間を求めるために上下層で測定されたAMS 14C 年代測定結果から内挿法で推定すると、約9700年前の約65年の期間にあたる。この65年間は温暖で乾燥していた。, 名古屋大学年代測定総合研究センターシンポジウム報告

In this study we present morphological and anatomical changes in the petioles of Nymphaea alba and Nuphar luteum from two lakes greatly differing from each other in respect to the oscillation of their water level (Lake Salno -a stabilized natural reservoir, used for comparison; Lake Kwiecko -a natural reservoir for the Hydroelectric Plant in Żydowo, where the changes in the water level oscillate up to 3.1 m during a day under conditions of maximum retention). Nonrhytmic and turbulent water flows in Lake Kwiecko cause the elongation of petioles, a decrease in their diameter, and to a significant degree, the reduction of supporting elements - collenchyma, vascular bundles and idioblasts. These changes take place to a lesser extent in Numphar luteum. Although this species has a very wide ecological amplitude and does not need to adapt to changing habitat conditions, through anatomical modifications the oscillations in the water level have an unfavorable effect on its population in Lake Quick.

Little attention has been paid to how aquatic habitat characteristics affect the traits of plant species. Nuphar lutea (L.) Sm. is a keystone species distributed across temperate regions of Europe, northwest Africa and western Asia. Its apparently low phenotypic variability compared to other aquatic plants led us to test whether the species exhibited significant phenotypic variability and whether trait values correlated to environmental parameters. The hypotheses were that (1) the environmental variation within our set of wetlands (both water and sediment characteristics) led to significant variation among four sets of traits related respectively to growth, reproduction, defence and storage and (2) that nutrient limitation (nitrogen and especially phosphorus) should affect plant traits towards a higher investment in storage and defence and a lower investment in growth and reproduction, thereby negatively affecting the success of N. lutea. To test these hypotheses, 11 populations of N. lutea were sampled in wetlands differing in physicochemical characteristics and spread along three rivers. A total of 15 traits, grouped into four sets (growth, reproduction, storage and defence), were measured during the growing season. Most N. lutea traits were related to the environmental characteristics of wetlands. The growth and reproduction traits were mostly positively related to habitat resource conditions, whereas the defence traits were positively correlated with both ammonium concentration and temperature, outlining possible anoxic stress (habitat adversity). Nitrogen or phosphorus limitation led to the variation of only a few traits: the rhizome starch content was higher in phosphorus-limited wetlands, while the rhizome length and volume, and the number of flowers were higher in nitrogen-limited wetlands.

The evolution of the apertural type in the ANITA-grade family Nymphaeaceae, despite the importance of pollen characters for both palaeobotanical and neobotanical investigations, has not been yet totally understood. To elucidate some unclear aspects regarding the homologies of the apertural condition in basal and core Nymphaeaceae, we examined the pollen grains from five species of this family. We offer for the first time some evidence about the presence of a proper operculum in the pollen grain of Nuphar and for the ring-like status of the aperture of Nymphaea caerulea and Victoria cruziana. Our findings clarify and reinforce the hypothesis of an opercular origin of the distal part of the pollen grain in the core Nymphaeaceae and support the paraphyly of the genus Nymphaea as emerged from recent molecular studies. The diversity and specialisation of this 'basal' family is underlined even by its pollen characters. © 2013 Copyright Collegium Palynologicum Scandinavicum.

Hidden electrophilicity in Nuphar alkaloids is responsible for their impressive bioactivity, report Shenvi and colleagues.

The aim of this study was to determine the occurrence of micromycetes associated with disease symptoms on the leaves and flowers of three plant species, Nymphaea alba (NA), Nymphaea candida (NC), and Nuphar lutea (NL), forming nympheid phytocoenoses on Lake Płociczno in Drawa National Park during the years 2009 to 2012. From all collected plant specimens, an overall number of 38 distinct taxa of fungi and chromistan fungal analogues was isolated. The largest diversity of taxa was found on NL (37 taxa), the lowest was on NC (4 taxa), and NA contained 12 taxa.Each year, anamorphic forms of Ascomycota were dominant in the taxonomic structure. For the first time in Poland, Septoria nupharis (NA, NL, NC) and Colletotrichum nymphaeae (NL, NC) were found on their spotted leaves. For both of the mentioned pathogens, Nymphaea candida is a new host plant in Poland. Botrytis cinerea, Elongisporangium undulatum (= Pythium undulatum), Epicoccum nigrum, Fusarium incarnatum (= F. semitectum), and Gibberella avenacea (= Fusarium avenaceum) were found each year in the studied phytocoenoses. The confirmation of NA and NL flower infections by Botrytis cinerea, which leads to gangrene, is an important aspect of the gray mold epidemiology. Until now, the occurrence of smut fungi on nympheids in Drawa National Park was not observed. The taxonomic structure and the predomination of asexual stages of fungi, as well as the similarity coefficients, suggest that the seasonal decomposition of nympheids run naturally and contribute to maintaining the stability of the lake ecosystem.

Concise, scalable, and enantioselective formal syntheses of eight dimeric and three monomeric nuphar alkaloids were achieved, along with the construction of a stereochemically diverse collection of the first known monomeric analogues having apoptotic activity. The syntheses involved the development of highly enantioselective Bronsted acid catalyzed vinylogous Mukaiyama-Mannich reactions, which feature the unprecedented use of a supersilyl group to control the regio-, enantio- and diastereoselectivity. Biological studies reveal that several of these novel nuphar analogues are even more potent than their dimeric natural product counterparts.

Nuphar submersa (Nymphaeaceae) is a critically endangered freshwater macrophyte indigenous to central Japan, with only four small extant populations represented across its entire range. We investigated the genotypic and genetic diversity as well as the genetic structure of all extant individuals of N. submersa based on analysis of 15 microsatellite loci. Among 278 individual ramets, 52 multilocus genotypes were detected: 30 genotypes in Nikko City (NIK), 18 in Nasukarasuyama City (NAS), 3 in Mooka City (MOK), and 1 in Sakura City (SAK). The average number of alleles per locus ranged from 1.20 to 1.93, whereas the observed and expected heterozygosities ranged from 0.11 to 0.33 and from 0.10 to 0.24, respectively. With the exception of SAK, all populations contained multiple clones, but our results indicated low levels of within-population genetic diversity. The populations NIK and NAS comprised few large or middle-sized genets and many small genets. The populations NIK and NAS were suggested to comprise large old, old fragmented, and/or young small genets resulting from seedling establishment. All four populations were differentiated, and gene flow between the populations was restricted (average level of gene flow (Nm) = 0.122, G’ ST = 0.639). Of the total genetic diversity, 67.20 and 9.13% were attributable to inter- and intra-population diversity, respectively. STRUCTURE analysis revealed two or three well-differentiated groups of populations. Cluster I comprised one population (NIK) and cluster II comprised the remaining populations at K = 2. The populations NIK, NAS, and the remaining populations were assigned to clusters I, II, and III, respectively, at K = 3. For conservation practices, we recommend that each cluster be regarded as a different management unit. We further suggest that artificial gene flow among MOK and SAK populations is an appropriate option, whereas NIK should not be reinforced with genotypes from the remaining populations.

The leaf reflectance spectra (280–887 nm) of two heterophyllous aquatic plant species Polygonum amphibium (L.) and Nuphar luteum (L.) were compared and their relation to physical properties of the leaves examined. In P. amphibium contrasting environmental conditions along water–land gradient affected the majority of anatomical and morphological properties of leaves, but less differences were observed in photosynthetic pigment and total flavonoid contents. Leaf mass per area (LMA), palisade mesophyll, leaf thickness, trichome length and anthocyanin content per dry mass were correlated to the different parts of spectra. In N. luteum natant and submerged leaves differed significantly in all measured parameters. Chlorophyll a, anthocyanin and carotenoid contents per dry mass were related to reflectance in the red region, while leaf thickness, anthocyanin and total flavonoid contents per leaf area were related to reflectance in the near infrared region. Redundancy Analysis (RDA) indicated that in P. amphibium the average length of trichomes and LMA explained 72% and 6% variability of the spectra, whereas in N. luteum anthocyanin content per dry mass, explained 57% variability of the spectra. The comparison of natant leaves of both species showed that they were more similar than different leaf types within the single species.

As entomophilous plants, water lilies (Nymphaea) and spatterdocks (Nuphar) have low pollen production, thus can be under represented in the sediment record. These macrophytes produce distinctly shaped sclerenchyma tissue referred to as stone-cells, trichosclereids, astrosclereids or simply sclereids. This study examines the utility of using sclereids from two common species from the Nymphaeaceae Family as an alternative to their pollen. Histological studies of fresh tissues of Nymphaea odorata and Nuphar lutea show that each has distinct sclereids, but there has been confusion in the terminology used to designate their morphology. Some reports have referred to Nymphaeaceae sclereids as trichosclereids, but our histological studies show that the cells are more appropriately classified as polyramous, astrosclereids, librosclereids or rhizosclereids. Tissues from both species were treated using HCL, KOH, acetolysis and HF. We found that only the sclereids from N. lutea survived chemical treatments in a detectable form, indicating that sclereids from N. lutea can be a useful indicator of presence while the chance of observing sclereids from N. odorata in pollen preparations is very low, severely limiting the utility of the latter as a paleoecological indicator. Another limitation to using sclereids as a proxy is that they originate from tissues that require extended acetolysis treatments for release from the matrix. Finally, we examined sclereid abundance in sediments from a slough in the Florida Everglades, USA to determine if abundance of Nymphaeaceae sclereids correlates with Nymphaeaceae pollen and we find no significant correlation. Additional analyses are required to determine if empirical relationships exist amongst plant populations, pollen, sclereids and environmental conditions. A clear report of chemical treatments used and processing times are critical to verify results of studies utilizing sclereids.

We developed and characterized 13 microsatellite markers for Nuphar submersa, a critically endangered freshwater macrophyte of central Japan. In all the studied populations, ten out of isolated 13 loci showed polymorphism. The 13 loci showed one to five alleles per locus, with observed and expected heterozygosities ranging from 0 to 0.40 and from 0 to 0.67, respectively. Between 5 and 11 of the 13 loci were successfully amplified across the six other Nuphar taxa. The markers described here will be useful for a conservation genetic study of the species and informing the development of effective conservation programs of N. submersa and related species.

I analyzed spores, pollen and organic-walled algal remains in Middle Miocene deposits that filled a paleosinkhole at Tarnow Opolski, in the Upper Silesian Upland, Poland. Middle Triassic limestone bedrock in the study area underwent intense karstification under relatively warm and humid climate, resulting in development of numerous sinkholes. As the corrosion reached non-karstified marls, flowing water in the vadose zone was plugged and meteoric waters thus accumulated, leading to pond development. This palaeobotanical study was undertaken to “reconstruct” the fossil freshwater algae community and vegetation in and around the water body. Chlorophyta such as Botryococcus, filamentous algae from the family Zygnemataceae, mainly Mougeotia, Spirogyra and Zygnema, and desmids were important components of the algal assemblage. Most of the identified algae prefer mesotrophic to eutrophic conditions, and are characteristic of stagnant or slow-flowing shallow water. Among the free-floating and rooted plants were Salviniaceae/Azollaceae (probably Salvinia), Nuphar, Potamogeton, and carnivorous taxa such as Utricularia and probably Aldrovanda. The water body was surrounded by swamp vegetation composed of herbs such as Typha, Sparganium, Alismataceae and others, and riparian forests. The upper portion of the sediment section contains abundant pollen of swamp forest taxa such as Taxodium/Glyptostrobus, Nyssa and Alnus. The paleobotanical remains from the sinkhole at Tarnow Opolski suggest it was a shallow water body that was probably prone to rapid warming. Presence of resting cells, e.g. zygospores of Zygnemataceae and desmids, suggests that the pond may have dried periodically. Presence of Nuphar, Utricularia and Aldrovanda pollen, however, indicates long periods with standing water.

Here we report the effect of a partially purified alkaloid fraction (NUP) of Nuphar lutea on nuclear factor kappa B (NF-κB) expression and studied its mechanism of toxicity against Leishmania major in C3H mice peritoneal macrophages. NUP was found to be a mixture of thermo-stable dimeric sesquiterpene thioalkaloids containing mainly thionupharidines. The anti-leishmanial activity was shown to be mediated through the activation of NF-κB and increased iNOS production. Additionally, the nitric oxide inhibitor, N G -monomethyl- l -arginine (0.5 mM) totally reverted the anti-leishmanial effect of NUP (0.25 and 0.5 μg/ml). NUP was also shown to act as an anti-oxidant, almost completely inhibiting the macrophage respiratory burst activity. However, no elevated lysozyme (EC3.2.1.17) or β-galactosidase (EC3.2.1.23) activities were demonstrated in macrophages treated with NUP. This study suggests, that the activity of NUP is mediated by NF-κB activation and the production of nitric oxide which is dependent on the l -arginine:NO pathway.

Premise of the study Microsatellite loci were developed for Nuphar shimadai (Nymphaeaceae) to evaluate the population genetic dynamics for conservation purposes. The species is an endemic aquatic species in Taiwan that is endangered by anthropogenic activities. Methods and results A magnetic bead enrichment protocol was used to identify 72 potential microsatellite loci and develop 39 microsatellite markers from N. shimadai. The number of alleles per locus ranged from one to 10 per locus, with levels of observed heterozygosity ranging from 0 to 1.0 within populations. As a result of inbreeding within isolated populations, 65% of loci significantly deviated from Hardy-Weinberg equilibrium within populations. Conclusions These novel markers should be valuable tools to evaluate the genetic diversity within the endangered aquatic taxon N. shimadai for conservation and reintroduction purposes in Taiwan.

Current understanding of floral developmental genetics comes primarily from the core eudicot model Arabidopsis thaliana. Here, we explore the floral transcriptome of the basal angiosperm, Nuphar advena (water lily), for insights into the ancestral developmental program of flowers. We identify several thousand Nuphar genes with significantly upregulated floral expression, including homologs of the well-known ABCE floral regulators, deployed in broadly overlapping transcriptional programs across floral organ categories. Strong similarities in the expression profiles of different organ categories in Nuphar flowers are shared with the magnoliid Persea americana (avocado), in contrast to the largely organ-specific transcriptional cascades evident in Arabidopsis, supporting the inference that this is the ancestral condition in angiosperms. In contrast to most eudicots, floral organs are weakly differentiated in Nuphar and Persea, with staminodial intermediates between stamens and perianth in Nuphar, and between stamens and carpels in Persea. Consequently, the predominantly organ-specific transcriptional programs that characterize Arabidopsis flowers (and perhaps other eudicots) are derived, and correlate with a shift towards morphologically distinct floral organs, including differentiated sepals and petals, and a perianth distinct from stamens and carpels. Our findings suggest that the genetic regulation of more spatially discrete transcriptional programs underlies the evolution of floral morphology.

Animals facilitate macrophyte seed dispersal in various ways despite spe- cializations of macrophytes for water dispersal. Previous diet analyses of freshwater aquatic turtles revealed that several North American turtle species consume a variety and abundance of seeds among other plant material and animal prey. We quantifi ed the dietary habits of Chrysemys picta picta (Eastern Painted Turtle) in a Massa- chusetts lake to examine if these animals included hydrophyte seeds in their diet and evaluate their capacity as passive seed-dispersal agents. Fifty-four turtles were trapped and housed to collect feces. Examination of feces revealed a diverse diet with comparatively high frequencies of animal, plant, and algal matter. Eight hundred fi fty-seven seeds of at least nine plant species were egested (among 87% of turtles), with all but fi ve (99%) seeds visibly intact. Seeds of Nuphar (473) and Decodon (305) were most abundant in the feces. Life-history characteristics of both C. p. picta and Nuphar suggest an effective endozoochorous seed dispersal association.

To provide insights into the floral developmental genetics of Nymphaeales (water lilies), we investigated the expression patterns of floral organ identity genes in three genera: Cabomba, Nuphar, and Nymphaea. Additionally, because of the superficial floral similarity between Nymphaea and the early‐diverging eudicot Nelumbo, we conducted the same experiments in the latter taxon. We focused on gene expression associated with (1) perianth differentiation in Nymphaeales, (2) the transition of petaloid staminodes to stamens in Nymphaea, and (3) organ identity in Nymphaea and Nelumbo. In Cabomba, the expression patterns of B‐class gene homologues fit the “sliding boundaries” model, with B‐class gene expression in sepals and petals. In contrast, Nuphar and Nymphaea exhibit broad B‐class gene expression that extends across all floral organs (i.e., “fading borders” model). The gene expression patterns observed suggest that the innermost petals of Nymphaea originated from petaloid staminodes. Also, despite the morp...

Studies on the role of nymphaeids in water bodies are uncommon, despite the fact that they often dominate in eutrophic lakes. Therefore, we decided to evaluate the effect of a nymphaeid Nuphar lutea on its environment. We hypothesised that its presence can increase the diversity and abundance of zoobenthos, as well as modify selected abiotic parameters, both during the growing season (in the presence of leaves) and outside it (with underground structures only). Three pairs of stations (within and outside Nuphar stands) were established in two shallow, eutrophic lakes and sampled monthly during an entire year. Zoobenthos taxon richness, biodiversity and total density were higher among nymphaeids throughout the year. On the other hand, total zoobenthos biomass was not affected by the presence of plants. Most macroinvertebrates reached higher densities within Nuphar stands, the most abundant being Polypedilum nubeculosum, Potamothrix bavaricus, Ephemeroptera and Ceratopogonidae. In turn, Potamothrix hammoniensis, Chaoborus sp. and Tanypus sp. were more abundant outside the stands. Other taxa, including the dominants (Chironomus sp., Procladius spp., Einfeldiadissidens, Valvatapiscinalis), were not affected by the presence of Nuphar. Nuphar did not influence water temperature, oxygen concentration, and conductivity, though sediments within the stands exhibited lower water content and higher proportion of large organic particles. Thus, Nuphar effect on macroinvertebrates was probably direct, through increased habitat heterogeneity, physical stability of substratum and food availability. Our study has shown that nymphaeids, despite their structural distinctness, can affect positively bottom fauna similarly to other morphological groups of plants.

Plant invasions represent ecological opportunities for herbivorous insects able to exploit novel host plants. The availability of new hosts and rapid adaptations may lead to host race formation and ultimately speciation. We studied potential host race formation in the water lily leaf beetle, Galerucella nymphaeae, in response to invasion by water chestnut, Trapa natans, in eastern North America. This leaf beetle is well suited for such studies because previous work showed that different herbivore populations follow different "evolutionary pathways" and specialize locally in response to differences in habitat preferences and host plant availability. We compared host preference and performance of G. nymphaeae offspring originating from T. natans and offspring of individuals originating from an ancestral host Nuphar lutea, yellow water lily, on T. natans and three native hosts (N. lutea, Nympheae odorata, and Brasenia schreberi). Regardless of origin (Trapa or Nuphar), adults strongly preferred their native host, N. lutea, over T. natans. Although laboratory survival rates (larva to pupa) were extremely high (80%) regardless of origin or host offered, survival rates in a common garden were greatly reduced, particularly for T. natans (24%) and to a lesser extent on N. lutea (54%), regardless of beetle origin. Larval drowning during more frequent leaf changes when developing on small Trapa leaves seems to be responsible for this difference. Preference of females for N. lutea is beneficial considering the much higher larval survival on the ancestral host. Abundant T. natans where the plant is invasive provides an alternative food source that beetles can use after egg/larval loads on their preferred host reach carrying capacity, but this utilization comes at a cost of high larval mortality.

Conventionally, the Klamath and Modoc Indians of south-central Oregon and northeastern California are depicted as hunter-gatherers, who took no active role in the management of plant communities. In the course of a multi-year ethnographic effort, however, tribal elders have identified a complex of interrelated plant management practices that are consistent with contemporary definitions of plant cultivation. These include the management of black huckleberry (Vaccinium membranaceum) yields in subalpine environments, the management of marsh-edge environments for yellow pond lily (Nuphar polysepalum), the tending of “epos” or yampah (Perideridia spp.) digging sites, and the selective harvest of tree cambium, sap, and wood – especially from pines (Pinus spp.) and junipers (Juniperus spp). Together, these accounts suggest that plant management was widespread, that it often served to geographically concentrate culturally preferred species, and that it was rooted as much in cosmological motivations as ec...

Several anti-leishmanial drugs of choice are of plant origin. Many of the available drugs against the disease are toxic and in certain cases parasite drug resistance is developed. The development of new compounds is urgently required. Aims of the study To determine the leishmanicidal activity of the Nuphar lutea plant extract against Leishmania major in vitro. Materials and methods The leishmanicidal activity of methanolic plant extract against L. major free living promastigotes and intracellular amastigotes was evaluated, using microscopic examinations and the enzymatic XTT assay. Results Methanolic extract of N. lutea was highly effective against both Leishmania promastigotes and L. amastigotes (IC50=2±0.12 μg/ml; ID50=0.65±0.023 μg/ml; LD50=2.1±0.096 μg/ml, STI=3.23). The extract at 1.25 μg/ml totally eliminated the intracellular parasites within 3 days of treatment. Also, a synergistic anti-leishmanial activity was demonstrated with N. lutea extract combined with the anti-leishmanial drug – paromomycin. The partially purified N. lutea active component was found to be a thermo-stable alkaloid(s) with no electrical charge and is resistant to boiling and to methanol, dichloromethane and xylene treatment. Conclusions The present study suggests that N. lutea might be a potential source of anti-leishmanial compounds.

The conventional concept of an ‘undifferentiated perianth’, implying that all perianth organs of a flower are alike, obscures the fact that individual perianth organs are sometimes differentiated into sepaloid and petaloid regions, as in the early-divergent angiosperms Nuphar, Nymphaea, and Schisandra. In the waterlilies Nuphar and Nymphaea, sepaloid regions closely coincide with regions of the perianth that were exposed when the flower was in bud, whereas petaloid regions occur in covered regions, suggesting that their development is at least partly controlled by the environment of the developing tepal. Green and colourful areas differ from each other in trichome density and presence of papillae, features that often distinguish sepals and petals. Field experiments to test whether artificial exposure can induce sepalness in the inner tepals showed that development of sepaloid patches is initiated by exposure, at least in the waterlily species examined. Although light is an important environmental cue, other important factors include an absence of surface contact. Our interpretation contradicts the unspoken rule that ‘sepal’ and ‘petal’ must refer to whole organs. We propose a novel theory (the Mosaic theory), in which the distinction between sepalness and petalness evolved early in angiosperm history, but these features were not fixed to particular organs and were primarily environmentally controlled. At a later stage in angiosperm evolution, sepaloid and petaloid characteristics became fixed to whole organs in specific whorls, thus reducing or removing the need for environmental control in favour of fixed developmental control.

Original scanning electron microscopy (SEM) observations are presented for stems of Brasenia schreberi and Cabomba caroliniana of Cabombaceae and three species of Trithuria of Hydatellaceae. End walls of stem tracheids of Brasenia have the same peculiar microstructure that we have reported in Barclaya, Euryale, Nuphar, Nymphaea (including Ondinea) and Victoria of Nymphaeaceae. This feature unites Cabombaceae with Nymphaeaceae. The minute rhomboidal crystals on the surfaces of stellate parenchyma cells of Brasenia reported by Solereder (1906. Oxford: University Press), but not noticed since, are figured. They are like the minute crystals of the often-mentioned astrosclereids of Nymphaeaceae. Neither of these two features has been observed in Hydatellaceae. If the absence of these two features can be confirmed, the reason may be more related to ecology, development, habit and anatomical organization than to degree of phylogenetic relationship as shown by molecular studies. Anatomical observations on the stem anatomy of Trithuria are offered on the basis of paraffin sections prepared for a paper by Cheadle & Kosakai (1975. American Journal of Botany62: 1017–1026); that study is notable for a discrepancy between an illustration of a specialized vessel element on the one hand and tabular data indicating long scalariform perforation plates on the other. Long scalariform perforation plates are mostly found in scalariformly pitted vessels of monocots, whereas the tracheary elements of Trithuria mostly have helical or annular thickenings. We were unable to demonstrate the presence of vessels in Hydatellaceae. © 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 159, 572–582.

We describe in detail the floral ontogeny of Nymphaea tetragona from a wild population to provide evidence regarding the phylogenetic position of Nymphaea and to reveal evolutionary trends of flowers in Nymphaeaceae by comparison with that of the other genera. Four sepals are initiated unidirectionally. The basal petals are initiated unidirectionally and alternate with the sepals. The dome-shaped floral apex continues to expand and produces more petal and stamen primordia. The remaining petals and all stamens are initiated in spirals or whorls. Later, the periphery of the floral apex grows more quickly than the centre and results in a depression in the centre of the apex after all stamens have been initiated. Carpels are simultaneously initiated in a cycle at the periphery of the depression. They are ascidiate. After all organs have been initiated, the centre of the depression on the floral apex grows and develops into a globular structure. The connected inferior ovary, stigma caps and the globular floral apex together form an extragynoecial compitum. Within Nymphaeaceae, the floral ontogeny of Nymphaea is most similar to that of Euryale and Victoria. It differs more from Ondinea and Barclaya, and differs most from Nuphar. (C) 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 159, 211-221.

Scanning electron microscopy (SEM) photographs of thick sections from liquid-preserved stems of Victoria cruziana and Euryale ferox show accretions of coarse fibrils on pit membranes of tracheids. The first-deposited fibrils are randomly orientated; on top of them (facing the tracheid lumina) are axially orientated coarse fibrils. The two systems are interconnected. Axially orientated fibrils were more extensively observed in Euryale than in Victoria and tips of fibrils in Euryale extend over the pit apertures onto secondary wall surfaces. Tracheid–parenchyma interfaces bear rudimentary coarse fibrils on the tracheid side. End walls of Victoria tracheids have highly porose pit membranes, thinner and less complex than those of the lateral intertracheid walls. The structures reported in Victoria and Euryale are consistent with those concurrently reported for stems of other Nymphaeaceae. Although also present in Cabombaceae, the coarse fibrils are otherwise not reported for stems of angiosperms and are not yet reported in roots of any species. Pit membrane remnants in perforation plates of various woody dicotyledons represent a nonhomologous phenomenon. The accretions of coarse fibrils in stem tracheids of Nymphaeaceae do not appear to enhance conduction, although they do contain porosities interconnecting tracheids. Removal of pit membrane remnants from perforation plates of primitive dicotyledon woods by hydrolysis does, on the contrary, suggest conduction enhancement. © 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 159, 52–57.

An important driver of biodiversity is habitat structural complexity: more complex habitats generally contain more species. A good measure of habitat complexity therefore could be a strong tool to predict habitat biodiversity. In this pilot study, we test the significance of the classification of aquatic macrophytes that have similar complexity and shape into vegetation types (Emergent: Phragmites, Typha; Floating-Leaved Nymphaeidae: Nuphar; Submersed: Potamogeton, Utricularia) to explain macroinvertebrate community structure. Macroinvertebrate abundance and family richness increased from floating-leaved Nuphar to emergent and submersed vegetation. Family richness corrected for abundance did not differ among the vegetation types, possibly due to the small subsamples, but larger stands of Nuphar were needed to reach similar total richness. Six families that significantly differentiated between the vegetation types were associated with submersed and emergent vegetation. Vegetation type explained ∼21 % of the variation in macroinvertebrate community structure within a pond, and performed better than any other classification of macrophyte species in explaining variation. Thus, if these results stand generalization in space and time, vegetation type can be a fast and practical indicator of a pond's invertebrate density, and probably diversity.

Perianth differentiation into distinct morphological whorls (sepals and petals) can be difficult to assess, particularly in early-divergent angiosperms. The perianth of members of Nymphaeales (Cabomba, Brasenia, Barclaya, Euryale, Nuphar, Nymphaea, Victoria, Ondinea) has been described as differentiated into sepals and petals, undifferentiated or presenting examples of both states. In this paper, we review perianth structure in Nymphaeales using morphology and ontogeny in order to determine whether their perianths are dimorphic. The results indicate that although there are differences between the outer and inner perianth series, the organs display few "typical" sepal-petal characteristics. Our observations of the perianth of Nuphar and Nymphaea, and reports on Euryale, show that sepaloid (green) and petaloid (yellow) areas can both occur on individual perianth organs, and that sepaloid areas occur mostly in the regions of the perianth organ that were exposed when the flower was in bud. SEM study revealed further morphological differences between the sepaloid and petaloid areas of Nuphar and Nymphaea and also between the exposed and covered regions of the perianth of Cabomba, Brasenia and Barclaya. We discuss the implications of these results with respect to current concepts of perianth differentiation.

Vegetative and leaf architectural characters are used to assess the phylogenetic relationships of species of Cabombaceae and Nymphaeaceae. Twenty-eight species representing all eight genera and the five subgenera of Nymphaea are analyzed. These morphological analyses give robust support to other molecular and morphological analyses of the relationships of the two families. There is strong support for the two families to be monophyletic, and that Euryale and Victoria are closely related. Strict consensus analyses show that within the Nymphaeaceae, Nuphar is the first sister clade, followed by Barclaya and finally by a polytomy of Ondinea, Euryale-Victoria and Nymphaea. The relationships of Ondinea are uncertain, in part due to missing characters. Finally, the genera of each family are monophyletic. Each genus has a suite of distinctive vegetative characteristics that are used to make an identification key of the genera. Although the value of vegetative and leaf architectural characters is shown, inclusion of atypical leaves, for example some types of submerged leaves, results in less resolved or robust trees.