Relationships and origins of the Dryopteris varia (L.) Kuntze species complex (Dryopteridaceae) in Korea inferred from nuclear and chloroplast DNA sequences

The Dryopteris varia species complex belongs to D. subg. Erythrovariae sect. Variae and includes eight closely related, controversial species. In Korea, six species belong to the complex; D. varia s. str., Dryopteris pacifica, Dryopteris sacrosancta, Dryopteris bissetiana, Dryopteris saxifraga, and Dryopteris saxifragivaria. The morphology of these species is highly variable due to the frequent occurrence of agamospory, hybridization, and/or polyploidization. We analyzed sequences of the nDNA pgiC and cpDNA rbcL, trnL–trnF IGS and atpF–atpH IGS regions from all six of these species to elucidate their relationships and origins, and to investigate the parentage of the putative hybrid taxon, D. saxifragivaria. Fourteen nDNA pgiC haplotypes and five cpDNA haplotypes were obtained from accessions of the D. varia complex in Korea. In particular, all accessions examined appear to have multiple pgiC haplotypes. Maximum parsimony and Bayesian inference analyses of the nDNA and cpDNA sequence data show that the agamosporous triploids D. sacrosancta and D. pacifica are of allopolyploid origin involving interspecific hybridization. The results also indicate that agamosporous triploid individuals of D. saxifragivaria in Korea were derived from hybridization between sexual diploid individuals of D. saxifraga (maternal) and agamosporous diploid individuals of D. bissetiana (paternal). In addition, our results strongly suggest that the agamosporous triploid D. varia s. str. is probably of autopolyploid origin, and cytoplasmic gene flow has occurred from D. sacrosancta to Dryopteris chinensis of D. subg. Dryopteris.     

Electrophoretic variation of the apogamousDryopteris varia group (Dryopteridaceae)

A total of 35 electrophoretic variants were detected in four apogamous species of theDryopteris varia group: 14 variants inD. bissetiana triploids, 7 inD. sacrosancta triploids, 6 inD. varia triploids, 3 inD. pacifica triploids, and 5 in apogamous diploids. The variants differed from each other in various combinations of a few alleles which were common in some of the variants, suggesting recurrent origin.Dryopteris insularis var.insularis (2x) and var.chichisimensis (3x), other members of the group, were electrophoretically monomorphic, suggesting single origin. Banding patterns infer that triploids ofD. bissetiana andD. sacrosancta may have originated from hybrids whose parent is the sexualD. saxifraga, whereas this species was not involved in the origin of the other apogamous triploids and the apogamous diploids. The other parent was also discussed and we proposed a hypothesis that diploid offsprings fromD. pacifica triploids hybridized withD. saxifraga giving rise to triploids ofD. bissetiana andD. sacrosancta.     

Reticulate evolution in the apogamous Dryopteris varia complex (Dryopteridaceae,subg. Erythrovariae,sect. Variae) and its related sexual species in Japan

Apogamous fern species are often difficult to distinguish from related species because of their continuous morphological variations. To clarify the genetic relationships among the members of the Dryopteris varia complex, we analyzed the nucleotide sequences of the plastid gene rbcL and the nuclear gene PgiC. We also analyzed the diploid sexual species D. caudipinna and D. chinensis, which have not been included in the complex, but were recently shown to be closely related to the complex in a molecular phylogenetic study. The PgiC sequences of the diploid sexual species, D. varia, D. saxifraga, D. sp. ‘protobissetiana’ (undescribed diploid sexual species), D. caudipinna, and D. chinensis, were well differentiated and hence designated A, B, C, D, and E, respectively. Thus, the PgiC constitution of apogamous species in the complex was as follows: D. bissetiana, B + C; D. kobayashii, B + C + E); D. pacifica, A + C, A + B + C, or A + C + D; D. sacrosancta, A + C + E; and D. saxifragivaria, B + C. These results suggest that these apogamous species are formed by hybridizations of species including not only the three diploid sexual species of the D. varia complex (A, B, and C) but also the two diploid sexual species D. caudipinna (D) and D. chinensis (E), which do not belong to the complex.     

Electrophoretic evidence for the origin ofDryopteris yakusilvicola (Dryopteridaceae)

To investigate the origin of the triploid agamosporous speciesD. yakusilvicola, an electrophoretic analysis was made for five enzymes of theD. sparsa complex.Dryopteris yakusilvicola showed a monomorphic banding pattern for the five enzymes and was heterozygous in all six gene loci coding them. Comparison of enzyme banding patterns suggests that the genome ofD. yakusilvicola was derived through hybridization betweenD. sabaei and either a sexual tetraploid or an agamosporous triploid ofD. sparsa. Cytological evidence (Darnaediet al., 1989) supports the idea that of the two types ofD. sparsa the sexual tetraploid is a parent. The monomorphic pattern implies thatD. yakusilvicola originated from a single hybrid between the parental species, and that it is a neo-endemic of Yakushima Island.     

Spore morphology of the genus Dryopteris Adans. (Dryopteridaceae) in Korea

Spore morphology of the Korean members of the genus Dryopteris was examined by scanning electron microscopy. In particular, spores of D. hangchowensis, D. woodsiisora, D. saxifragivaria, and D. subexaltata were examined here for the first time. Three perispore types were recognized on the basis of the shape of macro-ornamentations on perispore: these include rugate, echinate, and spinose. The rugate perispore type appeared to be the most common in the genus, and probably represents plesiomorphic condition in the genus Dryopteris. The echinate perispore type is found in D. expansa, and spinose perispore type in D. formosana. The results of this study demonstrate that the perispore characteristics, particularly the shape of macro-ornamentation and surface texture, have diagnostic value at the species level. Indeed, most of the Dryopteris taxa in Korea could be distinguished by their perispore characters. In addition, spores of sexual diploid individuals of D. saxifraga were much smaller than those of agamosporous triploid individuals, supporting the contention that the size of spores in ferns is generally related to ploidy levels.     

A review of chromosomal variation in Dugesia japonica and D. ryukyuensis in the Far East

The chromosome numbers of Dugesia japonica Ichikawa et Kawakatsu, 1964, are n = 8, 2x = 16 and 3x = 24; those of Dugesia ryukyuensis Kawakatsu, 1976, are n = 7, 2x = 14 and 3x = 21. The karyotypes of both species include diploid, triploid and mixoploid; aneuploidic and mixoaneuploidic karyotypes may occur. In 785 specimens studied of D. japonica, the occurrence rates of specimens having each karyotype are substantially the same (29–37%). Diploid sexual specimens represented nearly 10% of the total and virtually no triploid or mixoploid sexual specimens were found. The diploid karyotype can be inherited by both sexual and asexual reproduction; the triploid and mixoploid karyotypes will be inherited only by asexual reproduction. In 51 specimens studied of D. ryukyuensis, the different karyotypes are diploid (ca 39%), triploid (ca 57%) and mixoploid (ca 4%). Diploid sexual specimens represented nearly 25% of the total; sexual specimens with tripooidic karyotypes made up nearly 27%. The diploid, triploid and mixoploid karyotypes were also found in juveniles hatched from cocoons. The diploid karytyype is inherited by both sexual and asexual reproductions; the other karyotypes may be inherited by parthenogenesis or self-fertilization (including pseudogamy) and asexual reproduction.     

Diploid and triploid offspring of triploid agamosporous fernDryopteris pacifica

A cytological and reproductive study of the diploid and triploid agamosporousDryopteris pacifica was made to elucidate the origin of its infraspecific cytotypes. Some triploids produced 16 spore mother cells (SMCs) sometimes with n=41II+41I chromosomes, in addition to eight SMCs with n=123II, in each sporangium. In the former case the 16 SMCs usually underwent abnormal meiosis to give rise to some 50 spores, some of which were regular-shaped; in the latter the eight SMCs multiplied into 32 spores by normal meiosis. We found that spores from one of the triploid plants developed into either diploid or triploid gametophytes, which further apogamously produced diploid or triploid sporophytes, respectively. This novel mechanism of ploidy reduction is discussed in relation to the origin of diploid agamosporous ferns, the taxonomic complexity of the species, and the correlation of agamospory with polyploidy. The mechanism is also compared to that operating in agamospermous angiosperms.     

Cytological and Reproductive Studies of Japanese Diplazium (Woodsiaceae; Pteridophyta). II. Polyploidy and Hybridity in the Species Group with Summer-Green Bi- to Tripinnate Leaves

Diplazium with summer-green bi- to tripinnate leaves. Diplazium mesosorum and D. sibiricum var. sibiricum are sexual diploids (2n=82; n=41ll); D. chinense and D. squamigerum are sexual tetraploids (2n=164; n=82ll); and D. sibiricum var. glabrum is a sterile triploid (2n=123; meiosis irregular). Diplazium nipponicum includes both sterile triploid and sexual tetraploid populations. The triploid is larger in relation to several morphological characteristics and occurs in more southern regions than the tetraploid. Allozyme analysis suggests that the triploid is a hybrid of recurrent origin between the tetraploid and an unknown diploid. Diplazium × bittyuense is a sterile tetraploid, and the mitotic chromosome number, meiotic chromosome behavior, and allozyme analysis confirm the working hypothesis that it is a hybrid between D. nipponicum and D. chinense. Apomicts are not found within Diplazium with summer-green bi- to tripinnate leaves, and the taxonomic complexity can be attributed to polyploidy and natural hybridization. Received 3 March 2000/ Accepted in revised form 19 April 2000     

Cytotaxonomic study of genusHymenasplenium (Aspleniaceae) in Xishuangbanna, Southwestern China

The gametic chromosome numbers of sevenHymenasplenium (Aspleniaceae) species from Xishuangbanna, Yunnan Prov., China, were investigated. All the examined individuals ofH. obscurum, H. cheilosorum andH. latipinnum were sexual diploids with n=39 chromosomes. Intraspecific cytological variation was found inH. excisum, which has a sexual diploid (n=39) and a tetraploid (n=78). Only a triploid apogamous cytotype (n=ca.117) was found inH. laterepens. Hymenasplenium apogamum showed the most complicated intraspecific variation and included a sexual diploid (n=39), a sexual tetraploid (n=78) and an apogamous triploid (n=ca.117). This work reports for the first time the sexual diploids ofH. cheilosorum andH. apogamum, which are only apogamous elsewhere in east Asia, Himalayas and Indochina. These results may indicate that this area is one of the diversity centers ofHymenasplenium. Most of the above species have chromosome numbers based on x=39. In contrast,H. costarisorum contains a sexual diploid (n=36) and a sexual tetraploid (n=72), indicating that its basic number is x=36.     

STUDIES IN CRATAEGUS (ROSACEAE: MALOIDEAE) XIV. THE BREEDING SYSTEM OF CRATAEGUS CRUS-GALLI SENSU LATO IN ONTARIO

Crataegus crus-galli L. sensu lato in southern Ontario is a complex of taxa, two of which appear to consist exclusively of both triploid and tetraploid individuals. While pollen fertility is reduced in triploids, both ploidy levels are characterized by production of eight nucleate aposporous embryo-sacs and hence remain seed-fertile; in addition, single, apparently sexual, embryo-sacs are found in the ovules of tetraploids. Agamospermy is shown experimentally to be pseudogamous. Polyploidy in C. crus-galli s.l. is accompanied by self-compatibility, in contrast to the self-incompatibility of diploid Crataegus species such as C. punctata, with which comparison is made. The weediness of the component taxa of C. crus-galli s.l., in general, and their tendency toward uniparental reproduction, in particular, may explain the homogeneity of their topodemes, and the way in which these topodemes may be markedly differentiated from one another even within a single taxon.     

Mode of hybridogenesis and habitat preferences influence population composition of water frogs (Pelophylax esculentus complex,Anura: Ranidae) in a region of sympatric occurrence (western Slovakia)

Coexistence of sperm‐dependent asexual hybrids with their sexual progenitors depends on genetic and ecological interactions between sexual and asexual forms. In this study, we investigate genotypic composition, modes of hybridogenetic gametogenesis and habitat preferences of European water frogs (Pelophylax esculentus complex) in a region of sympatric occurrence. Pelophylax esculentus complex comprises parental species P. ridibundus and P. lessonae, whose primary hybridization leads to hybridogenetic lineages of P. esculentus. Hybrids clonally transmit one parental genome and mate with the other parental species, forming a new generation of hybrids. In the region of western Slovakia, we found syntopic occurrence of diploid and triploid hybrids with P. lessonae, syntopic occurrence of all three taxa as well as the existence of pure P. ridibundus populations. All triploid hybrids were exclusively male possessing one ridibundus and two different lessonae genomes (RLL). Sex ratio in diploid hybrids was substantially female‐biased. Irrespective of the population composition, diploid hybrids excluded the lessonae genome from their germ line and produced ridibundus gametes. Contrarily, RLL males unequivocally eliminated the ridibundus genome and produced diploid lessonae sperms. Perpetuation of RLL males in studied populations is most likely achieved by their mating with diploid hybrid females. The composition of water frog populations is also shaped by taxon‐specific habitat preferences. While P. ridibundus preferred larger water bodies (gravelpits, fishery ponds, dead river arms), P. lessonae was most frequently found in marshes and smaller sandpits. Pelophylax esculentus occupied predominately similar habitats as its sexual host P. lessonae.     

Causes and consequences of extreme variation in reproductive strategy and vegetative growth among invasive populations of a clonal aquatic plant, <Emphasis Type="Italic">Butomus umbellatus</Emphasis> L. (Butomaceae)

The mode of reproduction may influence the spread of invasive species by affecting evolutionary potential and dispersal ability. We sampled 51 introduced North American populations of the clonal aquatic plant Butomus umbellatus L. (flowering rush) and found extreme variation in sexual fertility caused by polyploidy. Populations consisted of diploids that produced thousands of viable seeds or of sexually sterile triploids. Although a trade-off between sexual and clonal reproduction predicts that the sexual sterility of triploids would be compensated for by greater clonal reproduction, a greenhouse experiment involving eight diploid and 10 triploid populations showed that diploid plants not only invest substantially in sexual structures but also make hundreds of tiny clonal bulbils on both rhizomes and inflorescences. In contrast, triploids do not make bulbils and have very limited scope for clonal multiplication and dispersal. Diploid populations were more frequent than triploid populations, especially in the Great Lakes region. This is probably because of the difference between cytotypes in clonal rather than sexual reproduction, as genetic analyses indicate a general lack of sexual recruitment in North America. Although triploids were less common, they have a wider geographical distribution. This could be due to a greater ecological tolerance resulting from polyploidy. However, genetic evidence suggests that triploids have become widespread via their use in and escape from horticulture. North America is being colonized by two distinct forms of B. umbellatus that differ strongly in reproductive strategy as well as the vectors and pathways of invasion.     

EVALUATION OF ELEVATED PLOIDY AND ASEXUAL REPRODUCTION AS ALTERNATIVE EXPLANATIONS FOR GEOGRAPHIC PARTHENOGENESIS IN EUCYPRIS VIRENS OSTRACODS

Transitions from sexual to asexual reproduction are often coupled with elevations in ploidy. As a consequence, the importance of ploidy per se for the maintenance and spread of asexual populations is unclear. To examine the effects of ploidy and asexual reproduction as independent determinants of the success of asexual lineages, we sampled diploid sexual, diploid asexual, and triploid asexual Eucypris virens ostracods across a European wide range. Applying nuclear and mitochondrial markers, we found that E. virens consists of genetically highly differentiated diploid sexual populations, to the extent that these sexual clades could be considered as cryptic species. All sexual populations were found in southern Europe and North Africa and we found that both diploid asexual and triploid asexual lineages have originated multiple times from several sexual lineages. Therefore, the asexual lineages show a wide variety of genetic backgrounds and very strong population genetic structure across the wide geographic range. Finally, we found that triploid, but not diploid, asexual clones dominate habitats in northern Europe. The limited distribution of diploid asexual lineages, despite their shared ancestry with triploid asexual lineages, strongly suggests that the wider geographic distribution of triploids is due to elevated ploidy rather than to asexuality.     

Cytological and Reproductive Studies on Japanese Diplazium (Woodsiaceae; Pteridophyta): Apomictic Reproduction in Diplazium with Evergreen Bi- to Tripinnate Leaves

Diplazium , including polymorphic terrestrial species with evergreen bi- to tripinnate leaves. Diplazium hachijoense, D. virescens var. virescens, var. conterminum, var. okinawaense, and two other unnamed varieties, D. kawakamii var. kawakamii, D. dilatatum var. heterolepia, D. taiwanense, D. × kawabatae (=D. dilatatum × taiwanense), D. × takii (=D. hachijoense × virescens var. virescens), and D.× nakamurae (= D. hachijoense × virescens var. conterminum) are apomictic triploids (2n=n=123). Diplazium amamianum and D. esculentum are sexual diploids (2n=82, n=41) and D. subtripinnatum is a sexual tetraploid (2n= 164, n=82). Diplazium dilatatum var. dilatatum includes both sexual diploid and apomictic triploid populations. Cultivated gametophytes of six triploid taxa produced sporophytes apogamously, confirming their apomictic reproduction. All three putative hybrids, D. × kawabatae, D. × takii, and D. × nakamurae, are triploid, apomictic, and fertile taxa, therefore they are not the result of hybridization between known pairs of Japanese Diplazium plants. Received 16 March 1999/ Accepted in revised form 30 September 1999     

Chromosome numbers ofCostus lucanusianus (Costaceae) in Nigeria

Chromosome counts onCostus lucanusianus in Nigeria were carried out. Diploid (2n=18) and triploid (2n=27) chromosome numbers were found. The diploid cytotype is reported for the first time for this taxon.     

The parthenogenetic Marmorkrebs (Malacostraca: Decapoda: Cambaridae) is a triploid organism

There is a close association between parthenogenesis and polyploidy. For this reason, we undertook a karyological analysis to test whether the parthenogenetic Marmorkrebs, Procambarus fallax forma virginalis, possesses an enlarged set of chromosomes. For this purpose, we karyotyped the Marmorkrebs, the sexual form of P. fallax (together called P. fallax complex), and the closely related species P. alleni. The latter shows 94 chromosomes in the haploid condition. In contrast to this, we found a haploid set of 92 chromosomes in individuals of the P. fallax complex. However, in mitotic metaphases the sexual form shows 184 chromosomes, whereas the Marmorkrebs possesses 276 chromosomes. Hence, the parthenogenetic Marmorkrebs reveals a triple amount of the haploid chromosome number. In addition, we detected a strikingly large subtelocentric chromosome which appears once in haploid and twice in diploid cells of sexual individuals of the P. fallax complex. In the parthenogenetic Marmorkrebs, this prominent chromosome occurs thrice. All this clearly reveals that the Marmorkrebs is a triploid organism. The applicability of the used methods, the significance of polyploidy in evolution of Decapoda, putative pathways to parthenogenetic triploidy, a possible hybrid origin and the scientific and ecological consequences of an increased chromosome set in Marmorkrebs are discussed.     

Polyploidy and aneuploidy inOphrys,Orchis, andAnacamptis (Orchidaceae)

Studies on chromosome numbers and karyotypes in Orchid taxa from Apulia (Italy) revealed triploid complements inOphrys tenthredinifera andOrchis italica. InO. tenthredinifera there is no significant difference between the diploid and the triploid karyotypes. The tetraploid cytotype ofAnacamptis pyramidalis forms 36 bivalents during metaphase I in embryo sac mother cells. Aneuploidy was noticed inOphrys bertolonii ×O. tarentina with chromosome numbers n = 19 and 2n = 38. There were diploid (2n = 2x = 36), tetraploid (2n = 4x = 72), hexaploid (2n = 6x = 108) and octoploid (2n = 8x = 144) cells in the ovary wall of the diploid hybridOphrys apulica ×O. bombyliflora. Evolutionary trends inOphrys andOrchis chromosomes are discussed.     

Cytotaxonomy of Dubyaea glaucescens (Compositae–Cichorieae)

Recent molecular phylogenetic analyses indicate that Dubyaea glaucescens (Compositae–Cichorieae) should be transferred to the genus Faberia as F. glaucescens. Here, we present cytological evidence for this transfer. Dubyaea glaucescens comprises two ploidy levels, 2n = 34 (diploid) and 2n = 51 (triploid), making the basic chromosome number x = 17. The chromosomes vary in length from 5.82 μm to 2.11 μm, and the karyotypes are 2n = 20m + 14sm (3sat) for the diploid cytotype and 2n = 30m + 21sm (3sat) for the triploid cytotype. Karyological characters of D. glaucescens, including chromosome number, size, morphology, and karyotype asymmetry, all agree remarkably with those reported previously in Faberia, but are distinct from those in other species of Dubyaea. The transfer of D. glaucescens to Faberia is thus strongly corroborated.     

Contrasting power of the factors for fission and sexuality in a polyploid planarian

One of the biotypes of the planarian Dugesia benazzii is triploid in the somatic line, hexaploid in the female line owing to a chromosome set doubling, and diploid in the male line due to a haploid set elimination. In a population of this biotype only 50% of the oocytes are hexaploid, the others being triploid as a results of the lack of set doubling; the male line is always diploid. After a long period of laboratory culture most of the individuals became asexual and fissiparous. Almost all the oocytes of the few specimens which have remained sexual showed triploid complement; B-chromosomes also appeared. These events represent the manifestation of a new genetic background which act upon the two germ lines in different ways and moments. These topics are discussed.