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Wild cyclamen (Cyclamen purpurascens) is considered as a precious breeding material for the development of new cultivars. Malvidin 3,5-diglucoside is the main anthocyanin in the petals of C. purpurascens, whereas the F1 progeny of the C. persicum × C. purpurascens cultivars cross contains 3,5-diglucoside-type anthocyanins as the main pigment. The anthocyanin 5-O-glucosyltransferase (A5GT) enzyme is responsible for the glycosylation of the A ring of anthocyanin at the 5-O-position, which implies that the expression of A5GT is dominant in the petals of C. purpurascens × C. persicum cultivars. Here, we isolated the complete open reading frame of the A5GT gene from C. purpurascens (Cpur5GT). Results of qRT-PCR revealed that Cpur5GT shows tissue-specific expression, with strong expression in fully opened petals and weak expression in young petals. In vitro enzyme assay showed that when uridine diphosphate glucose was used as the sugar donor, recombinant Cpur5GT could catalyze the glycosylation of 3-glucoside-type anthocyanidins at the 5-O-position, but when uridine diphosphate galactose was served as glycosyl donor, the reaction could not be performed. These results demonstrate that Cpur5GT exhibits valid anthocyanin glucosylation activity and could be used to analyze the mechanism of A5GT-mediated flower coloration in cyclamen in future studies.

Wild cyclamen tubers (Cyclamen purpurascens Mill.) (Kukavica mountain, south­east Serbia) was used as an extraction material in this study. The aqueous extract has been obtained by reflux extraction at the boiling temperature with hydromodulus 1:20 m/v during 180 minutes. The identification of bioactive components in the extract was performed by using UHPLC-DAD-HESI-MS analysis. The concentra­tions of macro- and microelements in the extract were determined by Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES). Isocyclamin and des­glucocyclamin I were identified in the obtained extract. Potassium was in the highest concentration - 10241.65 mg/kg of the plant material, while zinc was present in the highest concentration (11.57 mg/kg of plant material) among heavy metals. Presented results have shown that the obtained extract from wild cyclamen tubers is a potential source of triterpenoide saponin components isocyclamin and desglucocy­clamin I, as well as macro- and microelements.

Cyclamen purpurascens originates from the forest understorey and has variegated leaves and red abaxial epidermis. We measured the morphological and biochemical traits, and reflectance and transmittance spectra of leaves of C. purpurascens sampled through the growing season. We determined the optical properties and differences between the light-green and dark-green sections of the variegated leaves, and investigated the role of the red abaxial epidermis. The light-green leaf sections contained lower contents of chlorophyll a and b, carotenoids (in all samples) and anthocyanins (in April only) per leaf area, compared to the dark-green leaf sections, although the ratios of chlorophyll a to b and carotenoids to total chlorophyll were the same, except in April. During the closing of the canopy, chlorophyll a and b contents increased, while contents of carotenoids and anthocyanins decreased, as did the chlorophyll a/b and carotenoid/total chlorophyll ratios. The optical properties of the leaves in February showed high reflectance for UV and violet radiation, and low reflectance for near infrared radiation, for both types of leaf sections. The transmittance spectra for leaves without the red anthocyanic epidermis showed a pronounced peak from 520 nm to 625 nm, and enhancement in the near infrared region. The relative reflectance spectra of the red and colourless abaxial epidermis alone differed only in the green and yellow range. The relative transmittance of the red abaxial epidermis was significantly lower than for the colourless epidermis over the entire spectral range. Decreased transmittance in the green and yellow was obtained only for the red coloured abaxial epidermis. The variability of transmittance spectra during the season is best explained by the levels of chlorophyll b, carotenoids and anthocyanins. Our data reveal the importance of these pigments in leaf variegation, and particularly of the red abaxial anthocyanic epidermis for absorbing light in the green, yellow and near infrared.

Cyclamen purpurascens (Alpine, European or purple cyclamen) is native to central Europe. Since decades it is discussed wether the occurrences of C. purpurascens north of the alps is native or if it was introduced. Here the nuclear ribosomal DNAs (rDNA) are sequenced in oder to obtain a phylogenetic geographic pattern. Phylogenetic analyses of ITS and NTS/ETS sequences distinguish three main clades coinciding with geographical distribution: Eastern alps (Austria), southern alps (Switzerland, Italy) and western Alps (France). The paper presents interspecific relationship of C. purpurascens based on geographic sequences of rDNA. The observed variations suggest that some plants were introduced via Benedictine gardens and the plants from Monastery gardens seem to origin from Lower Austria.

Wild cyclamen tubers (Cyclamen purpurascens Mill.) (mountain Kukavica, Southeast Serbia) were used as material for extraction in this study. Aqueous extract was obtained by reflux extraction on boiling temperature with hydromodulus 1:20 m/v during 180 minutes. The total phenolic content was determined spectrophotometrically by the method of Folin-Ciocalteu, and the total flavonoids content by method with AlCl3. The antioxidant activity of extract was investigated spectrophotometrically by DPPH and ABTS test. Disc-diffusion method was used for antimicrobial activity investigation on the following pathogenic microorganisms: Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Pseudomonas aeruginosa and Klebsiella pneumoniae. The content of total phenols was 8.27 mg GAE/g dry extract while the total flavonoid content was 11.51 mg RE/g dry extract. The extract concentrations required to neutralize 50% of the initial concentration of DPPH radicals (EC50) after 20 minutes incubation and immediately after adding DPPH radical solution were 0.413 and 2.0 mg/ml, respectively, while concentrations of extract required to neutralize 50% of the initial ABTS radicals concentration is 0.743 mg/ml. The extract showed the highest antimicrobial activity on bacteria Staphylococcus aureus. The presented results indicate that cyclamen tubers extract is a potential source of natural antioxidants and antimicrobial agents.

Common cyclamen (Cyclamen pururascens Mill.) is a very interesting species not only for various breeding programs but also as an ornamental plant. The plants possess interesting floral and foliage characteristics, nice fragrances, and a very useful flavonol profile. The last is very important from the point of view of protecting against diseases and physiological disorders. Twenty-two different genotypes originating from different regions in Slovenia were analyzed in detail, based on their floral and foliage characteristics. Anthocyanin and flavonol contents were determined using high-performance liquid chromatography (HPLC)-photodiode array-mass spectrometry (MS). Color characteristics were colorimetrically evaluated separately for petals and leaves. In terms of color measurements, significant differences among the sites of origin were shown with parameters a* and L* when petals were analyzed, and with parameter a* when leaves were analyzed. The pH of petal sap stayed within the acid range, ranging from 3.96 to 4.82. Five different anthocyanins (malvidin-3,5-diglucoside, cyanidin-3-neohesperidoside, delphinidin-3-glucoside, delphinidin-3,5-diglucoside, and delphinidine-3-rutinoside) were analyzed in flowers, and this is the first report of delphinidin metabolites naturally present in common cyclamen genotypes. In relation to plant leaves, malvidin-3,5-diglucoside, malvidin-3-rutinoside, malvidin-3-glucoside, cyanidin-3-neohesperidoside, and peonidin-3-neohesperidoside were measured. Fifteen different flavonols were determined in common cyclamen flowers and 10 different flavonols in common cyclamen leaves. Various glycosides of quercetin, myricetin, and kaempferol were analyzed, as well as isorhamnetin-3-rutinoside, laricitin-3-rutinoside, and neohesperidin. The floral and foliage characteristics of the genotypes (physical and chemical) were mainly affected by the environmental conditions of the locality of origin of the genotypes (annual disposition of temperature and precipitation, structure of the soil, etc.).

Cyclamen purpurascens is considered suitable for horticultural breeding of cyclamens because it has an attractive fragrance that is not found in other wild species. To improve the commercial value of cyclamen flowers, this fragrance has been introduced into ornamental cultivars. However, variation in flower color is somewhat limited in these cultivars, and therefore understanding the genetic networks of flower coloration in C. purpurascens is required. We previously isolated DNA fragments of anthocyanin biosynthetic genes from C. purpurascens, broadening our understanding of the biosynthetic pathway of flavonols, which are co-pigments in flower coloration. In this study, we isolated complete open reading frames of flavonol synthase genes from C. purpurascens (CpurFLS1 and CpurFLS2) and analyzed the in planta functions of the genes by molecular complementation assay using the fls mutant of Arabidopsis thaliana. Expression patterns in several organs of C. purpurascens were also determined. The results strongly suggest that the CpurFLS genes participate in flavonol synthesis. We discuss the involvement of these two FLSs in flower coloration in C. purpurascens.

The catalogue of extinct plants in Spain, comprising 27 taxa is updated. Included are four species not previously classified as extinct in Spain: Cyclamen purpurascens, Grammitis quaerenda, Stratiotes aloides and Vallisneria spiralis. We analyze the causes of extinction and explore the possible influence of botanical collections in species loss. The obtained data suggest that habitat loss has been the main cause of extinction and the impact of collecting is demonstrated to be irrelevant. We list the 21 species that have been designated as extinct in previous catalogues but which no longer have that status, 14 of them having been recovered thanks to botanical exploration and collection works.

In order to evaluate the potential of common cyclamen (Cyclamen purpurascens Mill.) for the use for foliage ornamentals sixteen genotypes were collected from different regions in Slovenia. Cyclamen plants were classified into four groups based on their foliage characteristics: green, marbled, semi-silvery and silvery group. Leaves were analyzed in autumn and winter. Colour characteristics were colorimetrically evaluated, chlorophyll and carotenoid content was measured spectrophotometrically and anthocyanin and flavonol content were determined with the use of HPLC-PDA-MSn. A detailed analysis revealed that cyclamen leaves grouped into various types differed in numerous parameters. Colour measurements exposed changes between autumn and winter leaves and also among different genotypes. Marbled and semi-silvery genotypes tended to develop more red and pink colouration in autumn, whereas during the winter period the red and pink colouration was only observed in marbled genotypes. The latter were also characterized by high levels of anthocyanins. Semi-silvery, green and marbled genotypes accumulated high levels of chlorophyll (a, b, total) particularly in autumn leaves. Leaves of green and marbled cyclamen types contained high levels of different quercetin and myricetin compounds, as well as higher laricitrin and kaempferol levels compared to semi-silvery and silvery genotypes. Oppositely, isoharmnetine-3-rutinoside and neohesperidine concentrations prevailed in marbled and silvery genotypes.

The value of plant provenance (plant origin) is well-known phenomena in woody plants, but less is known in herbaceous plants (perennials). This study with common cyclamen (Cyclamen purpurascens Mill.) was conducted to reveal the importance of specific environmental site properties of plant origin for plant growth and plant quality in the next years. The plants were observed in years 2013 and 2014, more than 10 years after removing and replanting them from the original sites. Morphological characteristics of plants were evaluated by measuring the length and the width of plant rosettes, whereby plants originated from different sites did not show any significant differences. Additionally, the pigment composition, flavonol and anthocyanin content of plant leaves were evaluated. Plants removed from sunny sites showed significantly lower chlorophyll values (total chlorophyll, chlorophyll a) in the both observed years; lower carotenoid and total pigment values were measured only in year 2013. The prevailing anthocyanin in cyclamen leaves was malvidin-3,5-diglucoside with 57.28 µg l−1 FW in the year 2013 and with 103.68 µg l−1 FW in the year 2014. Plants originated from the sunny sites accumulated in 2013 significantly more malvidin-3,5-diglucoside in comparison with plants from shady sites of origin. The major substances from the flavonol group were quercetin-3-O-rutinoside and quercetin-dirhamnosyl-glucoside in both analysed years. The cyclamen leaves originated from sunny sites contained in 2013 significant more quercetin-dirhamnosyl-glucoside than cyclamen leaves from shady sites. The results of the study show that different stress parameters (irradiation and water supply in specific year) have a significant impact on the morphological and also internal parameters of cyclamen leaves.

We previously bred fragrant cyclamen cultivars by interspecific hybridization between cultivars of Cyclamen persicum and the wild species Cyclamen purpurascens. One of these fragrant cultivars, Kaori-no-mai, blooms purple flowers containing malvidin 3,5-diglucoside as the major anthocyanin. Here, we irradiated etiolated petioles of Kaori-no-mai with a 320-MeV carbon-ion beam at 0–16 Gy to increase flower color variation by mutation. Some of the M2 plants derived from self-pollination of M1 plants irradiated at 2 Gy were flower-color mutants that retained desirable flower shape, flower size, and leaf color. One of the mutants bloomed novel red-purple flowers, the major anthocyanin of which was delphinidin 3,5-diglucoside. Loss of methylation activity at the anthocyanin 3′- and 5′-hydroxyl groups with little influence on anthocyanin concentration was attributed to the mutation. Because the major anthocyanins in flowers of Cyclamen spp. were previously restricted to malvidin, peonidin, and cyanidin types, the generation of a cyclamen containing mostly the delphinidin-type anthocyanin is an important breakthrough in cyclamen breeding. We expect this mutant to become not only a commercial cultivar itself, but also a valuable genetic resource for cyclamen breeding.

V poskusu proučujemo kalitev semen navadne ciklame. Želeli smo ugotoviti najprimernejši način setve navadne ciklame ter kakšen vpliv ima populacija na delež vzkaljenih semen ter na morfološke lastnosti sejancev. Semena so bila nabrana v naravnih populacijah na treh lokacijah, in sicer: Logarska dolina, Sele in Vodriž (Wiederdris). Semena smo sejali na 5 različnih načinov. Pri posameznem načinu setve smo vrednotili število vzkaljenih semen in morfološke lastnosti sejancev. Rezultati poskusa dokazujejo, da kraj in način setve vplivata na kalitev semena navadne ciklame ter na morfološke lastnosti sejancev. Za najboljši način setve se je izkazala direktna setev predhodno namočenega semena, kjer smo dosegli 83 % kalivost semena, nabranega v populaciji Logarska dolina. Med proučevanimi populacijami obstajajo razlike v kalivosti semena in morfoloških značilnostih sejancev. We evaluated different sowing techniques of Cyclamen purpurascens. The seeds were collected in Slovenian natural habitats.. The aim of the survey was to analyse different techniques of seed germination and to find out which one would be the most suitable for using in practice. The results of the experiment showed that environment and technique of sowing (we studied five different techniques) had the highest influence on germination as well as on morphological characteristics of plants. The most successful was direct sowing of seeds which had been soaked in the water, where we tained 85 % of germination. The investigation indicates that there were significant differences in germination and morphological characteristics of plants originating from different places.

Although interspecific hybrids between cyclamen (Cyclamen persicum) cultivars and C. purpurascens were produced by using the ovule culture, mode of inheritance of flower pigment remains obscure. Therefore, inheritance of anthocyanin in the petals in the crosses between diploid cyclamen cultivars and C. purpurascens was investigated in the present study. In every cross combination, including the crosses by using acyanic cyclamen cultivars without the eye, all the F1 progenies had a purple or reddish-purple eye in the petals, showing the eye expression was a dominant characteristic. All the F1 progenies contained anthocyanins in the petals, whereas some progenies in the crosses by using a yellow-flowered cultivar contained very small amount of anthocyanin. The main flower pigment of C. purpurascens was malvidin 3,5-diglucoside. All the F1 progenies also contained 3,5- diglucoside type anthocyanins, such as malvidin 3,5-diglucoside, peonidin 3,5-diglucoside and cyanidin 3,5-diglucoside, in the petals as the main pigments even in the crosses by using acyanic cultivars or cyanic cultivars containing malvidin 3-glucoside or peonidin 3-neohesperidoside in the petals as the main pigment. These results suggest that the expression of 5-glucosyltransferase in anthocyanin synthesis in the petals was dominant even in the crosses between cyclamen cultivars and C. purpurascens.

Headspace samples collected from entire flowers of Cyclamen persicum, C. purpurascens and their interspecific hybrids were analyzed using gas chromatograph coupled with mass selective detector. The chemical constitution of volatile compounds of C. persicum (2n=2x=48, AA or 4x=96, AAAA) was distinguished by sesquiterpene hydrocarbons, whereas that of volatile compounds of C. purpurascens (2n=2x=34, BB) was distinguished by alcohols, aldehydes and esters of monoterpene or an aromatic class. The chemical constitution of volatile compounds in the hybrid (2n=41, AB) the amphidiploid (2n=82, AABB) and the sesquidiploid (2n=58, ABB) closely resembled that of volatile compounds in C. purpurascens. By contrast, the chemical constitution of volatile compounds in the sesquidiploid (2n=65, AAB) resembled that of volatile compounds in C. persicum. Flowers of diploid or tetraploid C. persicum had a woody or powdery fragrance, whereas those of C. purpurascens had a rose- or hyacinth-like fragrance. Flowers of the hybrid AB, the amphidiploid AABB and the sesquidiploid ABB had a fragrance resembling that of C. purpurascens, whereas those of the sesquidiploid AAB had a fragrance resembling that of C. persicum. Based on the present analyses, the relationship between the expression of volatile compounds and genome constitution is discussed.

application/pdf, The effects of genotypes on the production of interspecific hybrids in cyclamen cultivars (Cyclamen persicum Mill.) × C. purpurascens Mill. crosses were investigated to establish efficient breeding protocols in the crosses. In the tetraploid cyclamen cultivars×C. purpurascens Mill. crosses, few fruits and no interspecific hybrids were obtained. Although more than half of cross combinations in the diploid cyclamen cultivars × C. purpurascens Mill. crosses produced interspecific hybrids, efficiency of the hybrid production depended on the genotypes of seed parents in the crosses., The results of the present study indicated that the ploidy level and genotypes of seed parents influenced hybrid production in the cyclamen cultivars × C. purpurascens Mill. crosses. The results also suggest that the selection of desirable genotypes as seed parents in the crosses may be effective for the efficient production of the interspecific hybrids., 本研究では、C.purpurascensを利用したシクラメンの遺伝資源開発の効率化を目的として、シクラメン園芸品種とC.purpurascensとの種間雑種作出に及ぼす種子親の遺伝子型の影響を調査した。, シクラメン四倍体品種とC.purpurascensとの交雑では、ほとんどの果実が落果した。また、二倍体品種とC.purpurascensとの交雑では、半数以上の交雑組み合わせで種間雑種が得られたものの、雑種の作出効率には品種間差異が認められた。, 以上のことから、シクラメン園芸品種×C.purpurascensの交雑においては、種子親に用いる園芸品種の倍数性および遺伝子型が、種間雑種獲得の効率に影響すると考えられた。したがって、種間雑種形成が容易な遺伝子型を種子親に選択することによって、より効率的な種間雑種作出が可能と思われた。

At present, there is no relevant information on arbuscular mycorrhiza and the effect of the symbiosis on the growth of wild populations of cyclamens. To fill this gap, two populations of Cyclamen purpurascens subsp. immaculatum, endemic in Nizke Tatry (NT) mountains and Veľka Fatra (VF) mountains, Slovakia, were studied in situ as well as in a greenhouse pot experiment. For both populations, mycorrhizal root colonization of native plants was assessed, and mycorrhizal inoculation potential (MIP) of the soils at the two sites was determined in 3 consecutive years. In the greenhouse experiment, the growth response of cyclamens to cross-inoculation with arbuscular mycorrhizal fungi (AMF) was tested: plants from both sites were grown in their native soils and inoculated with a Septoglomus constrictum isolate originating either from the same or from the other plant locality. Although the MIP of soil at the NT site was significantly higher than at the VF site, the level of AMF root colonization of C. purpurascens subsp. immaculatum plants in the field did not significantly differ between the two localities. In the greenhouse experiment, inoculation with AMF generally accelerated cyclamen growth and significantly increased all growth parameters (shoot dry weight, leaf number and area, number of flowers, tuber, and root dry weight) and P uptake. The two populations of C. purpurascens subsp. immaculatum grown in their native soils, however, differed in their response to inoculation. The mycorrhizal growth response of NT plants was one-order higher compared to VF plants, and all their measured growth parameters were stimulated regardless of the fungal isolates’ origin. In the VF plants, only the non-native (NT originating) isolate showed a significant positive effect on several growth traits. It can be concluded that mycorrhiza significantly increased fitness of C. purpurascens subsp. immaculatum, despite the differences between plant populations, implying that AMF symbionts should be taken into account in conservation programs of this endemic plant.

Anthers of a F1 hybrid (2n=41) between Cyclamen persicum (2n=2x=48) and C. purpurascens (2n=2x=34) were cultured to produce microspore-derived plants. Embryoids were produced when anthers, containing microspores at the early uninucleate stage of pollen development, were cultured in B5 medium containing sucrose (90 g l-1) and NAA (0.1, 1 mg l-1) or 2,4-D (0.1 mg l-1) in the dark at 5 °C for 4 days, then at 25 °C for 60 days. The embryoids usually developed into plantlets when cultured in B5 medium containing sucrose (30 g l-1) in the dark at 25 °C. At meiosis, the F1 hybrid, used as source for anther culture, formed some cells with restitution nuclei at telophase and dyads at the tetrad stage, which resulted in the production of viable pollen grains as unreduced gametes. Plants produced by anther culture were grouped into sterile plants with 2n=41 chromosomes and fertile plants with 2n=82 chromosomes. The present findings suggested that the sterile plants were polyhaploids originating from unreduced microspores (n=41) of the F1 hybrid and that the fertile plants were amphidiploids induced by a spontaneous doubling during culture of chromosomes of such unreduced microspores.

In order to introduce valuable traits from wild species of Cyclamen into C. persicum cultivars, crosses were made between C. persicum‘Reinweis’and C. purpurascens. Crossing barriers between C. persicum‘Reinweis’and C. purpurascens were due to late-acting incompatibility reactions. Interspecific hybrids were obtained by using ovary culture. The highest number of embryos was achieved from placentas excised 21 days and 35 days after pollination and transferred to Murashige-Skoog (M.S.)–medium containing 6% sucrose and 1% agar. The hybrids showed a habit and a chromosome number intermediate between the parents. The fragrant flowers were pale red-purple. The chromosome number in root tips was determined as 2n= 41 while in C. persicum it is 2n= 48 and in C. purpurascens 2n= 34. Due to the different parental chromosome sizes, chromosomes of distinct size were still observed in the hybrid. Pollen viability varied between 0.3 and 34.0%. Parents and interspecific hybrids also showed differences in the DNA content of leaf tissue. Flow cytometric analyses were useful in the early identification of hybrids.

cyclamin, molluscicide, saponin, schistosomiasis. 1 * ; * i Cyclamin, a monodesmosidic triterpenoid saponin isolated from the tubers of Cyclamen purpurascens Mill, was found to, pos­ sess molluscicidal activity against the schistosomiasis- transmitting snail Biomphalaria glabrata (Say). The lowest concentration showing 100 % mortality to snails was 21 mg/1. The haemolytic activity and the molluscicidal potency of cyclamin were com­ pared with those of primulic acid with the result that high haemolytic activity will not automatically imply a strong molluscici­ dal potency.

Interspecific crosses were made to introduce the scent of flowers of C. purpurascens into C. persicum cultivars and ovule culture was used to rescue the abortive hybrid embryos. Cultivars of C. persicum diploid (CPD, 2n=2×=48) and C. persicum tetraploid (CPT, 2n=4×=96) were the pistillate parents and wild species of C. purpurascens (CP, 2n=34) were staminate parents. After pollination, crossed ovaries were collected periodically and examined using paraffin sections. Histological observations suggested that both hybrid ovules of CPD x CP and CPT x CP should be transferred to culture medium 35 days after pollination. Based up on this observation, crossed ovaries were collected 28 days after pollination and ovules with placenta were transferred to MS (1962) medium containing 3% sucrose. These ovules were cultured in the dark at 25° C. The hybrids (2n=41) derived from CPD x CP had the scent of C. purpurascens, whereas the hybrids (2n=65) derived from CPT x CP had the scent of C. persicum. Although both hybrids had complete genomes from the parents and produced a few viable pollen grains, they failed to yield viable seeds by self- and cross-pollination with fertile pollen grains of C. persicum cultivars.

We cultured colchicine-treated hybrid ovules in vitro to produce fertile amphidiploids of C. persicum (2n=2x=48. referred to as AA) × C. purpurascens (2n=2x=34, referred to as BB). Seedlings and mature plants were obtained from the ovules without colchicine and those exposed to 50 mg/l colchicine for 5, 10 and 15 days, whereas they were not obtained from the ovules exposed to 50 mg/l colchicine for 20 days and 500 mg/l for 5, 10, 15 and 20 days. Although 8 mature hybrids derived from the ovules without colchicine produced a few fertile pollen grains, they failed to produce viable seeds by self-fertilization. The hybrids had 41 somatic chromosomes. Four and 3 mature plants were derived from ovules exposed to 50 mg/l colchicine for 10 and 15 days, respectively. One each among 4 and 3 mature plants showed a high frequency of pollen grain fertility, produced several seeds by self-fertilization, and had 82 somatic chromosomes which is twice the number of hybrid chromosomes (2n=41, AB). These findings indicated that these plants are amphidiploids (2n=82, AABB) between C. persicum and C. purpurascens. Three and 2 viable seeds were derived by the conventional crosses of diploid C. persicum × the amphidiploid and the amphidiploid × C. purpurascens, respectively. Flowering plants that developed from the seeds of diploid C. persicum × the amphidiploid were barely fertile and had 65 somatic chromosomes (2n=65, AAB), whereas those that developed from the seeds of the amphidiploid × C. purpurascens were barely fertile and had 58 somatic chromosomes (2n=58, ABB). The somatic chromosomes indicated that these plants are probably sesquidiploids between the amphidiploid and either C. persicum or C. purpurascens. The interspecific cross-breeding of cyclamen using the amphidiploids and the sesquidiploids is discussed.

Although theCyclamen subg.Cyclamen spp. are morphologically variable, previous studies suggest a superficial cytological uniformity. New chromosome numbers and an indication of karyotypic instability are reported inC. hederifolium andC. africanum that reveal that the cytology of the subgenus is more complicated than previous accounts suggest. The possible significance of these phenomena is discussed. The diploid status ofC. purpurascens has been confirmed and a distinction between the three diploid karyotypes has been described. The cytological variation may help to explain the well documented morphological variation exhibited in these species.

Cyclamen europaeum Linnaeus, Species Plantarum 1: 145. 1753, nom. rej. "Habitat in Austriae, Tatariae, Europae australis nemorosis." RCN: 1160. Lectotype (Cafferty & Grey-Wilson in Taxon 47: 479. 1998): Herb. Burser XVII: 89 (UPS). Generitype of Cyclamen Linnaeus (vide Hitchcock, Prop. Brit. Bot.: 129. 1929). Current name: Cyclamen purpurascens Mill. (Primulaceae)., Published as part of Jarvis, Charlie, 2007, Chapter 7: Linnaean Plant Names and their Types (part C), pp. 370-473 in Order out of Chaos. Linnaean Plant Types and their Types, London :Linnaean Society of London in association with the Natural History Museum on page 467, DOI: 10.5281/zenodo.291971

Seed size and shape, seed coat surface pattern, seed coat thickness, and endosperm structure were investigated in Androsace septentrionalis, Cortusa matthioli, Hottonia palustris, Primula elatior, Soldanella carpatica (Primulaceae), Anagallis arvensis, A. minima, Cyclamen purpurascens, Glaux maritima, Lysimachia nemorum, L. vulgaris, Trientalis europaea (Myrsinaceae), and Samolus valerandi (Theophrastaceae). Three seed size categories were distinguished on the basis of biometric measurements. Almost all seeds examined were found to be small with an angular shape classified as sectoroid or polyhedral. A new type of seed shape, suboval, was identified for H. palustris. Cyclamen purpurascens seeds differed from seeds of all other species examined because of their large size, subglobose shape, and concave hilar area. Despite the different shape types, the length/width ratio of the seeds examined was constant, while their hilum length/width ratio was a highly variable feature. Three types of seed surface patterns were observed: (1) reticulate, (2) tuberculate with secondary striations, and (3) poroid-alveolate with the presence of a spongy outer layer. For seeds of Anagallis arvensis, A. minima, Cortusa matthioli, Lysimachia nemorum, and Soldanella carpatica, secondary seed sculpture was described. The seed coats of all species examined were two-layered, and great differences in testa thickness were found (9.9–128.6 μm). In general, seeds of the Myrsinaceae species were more often characterized by thick testa. Different proportions in thickness of the inner and outer testa layers were observed. In seeds with reticulate seed patterns, the inner testa layer was twice to several times thicker than the outer layer, while the opposite proportions were observed in seeds with the tuberculate or poroid-alveolate seed sculpture pattern. In seeds of all species examined, oxalate crystals were present on the surface of the inner testa layer. Thus, the presence or absence of oxalate crystals in testa is not a feature distinguishing species with angular or subglobose seeds. Four types of endosperm structure were identified according to the thickness of the endosperm cell walls and the relief of their inner surface: (1) with evenly thickened and smooth cell walls, (2) with evenly thickened cell walls and circular or helical thickenings on their inside surfaces, (3) with very thick, but not evenly thickened cell walls with constrictions (“pitted”), and (4) with very thin papery and undulate cell walls. There is no rule concerning the seed shape, type of the seed sculpture, testa thickness, or endosperm structure that corresponds to the family affiliation of the species examined.