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Pteris reptans (Pteridaceae), a native fern of Sri Lanka, has its distribution in India with exclusive limited occurrence in the state of Kerala. The plants from Kerala were examined for their quantity of spores per sporangium, micromorphology using SEM, gametophyte and sporophytic developmental phases. Upon examination, it was discovered that the sporangium of all accessions contained 32 tetrahedral spores with a continuous cingulum and noticeable tubercles on both surfaces. Normal shaped spore frequencies per sporangia ranged erratically from 81% to 95%. The fern exhibited Vittaria-type of spore germination. Gametophytic development followed the Adiantum-type in early stages with further Ceratopteris-type growth. A thorough analysis of antheridial development was done, but throughout the whole investigation, no archegonium was seen. Without any gametic fusion, juvenile sporophytes were produced apogamously on the prothalli. The filamentous stage was retained until the third week after germination and the sporophytes started emerging from the 12th week. After 90–95 days no further sporophytes emerged. Developmental morphology of the sporophyte was studied by observing trends in frond characteristics. The study is the first report on detailed apogamous development of P. reptans and also describes propagation methods for ex-situ multiplication. [ABSTRACT FROM AUTHOR]

Main conclusion: A novel genomic map of the apogamous gametophyte of the fern Dryopteris affinis unlocks oldest hindrance with this complex plant group, to gain insight into evo-devo approaches. The gametophyte of the fern Dryopteris affinis ssp. affinis represents a good model to explore the molecular basis of vegetative and reproductive development, as well as stress responses. Specifically, this fern reproduces asexually by apogamy, a peculiar case of apomixis whereby a sporophyte forms directly from a gametophytic cell without fertilization. Using RNA-sequencing approach, we have previously annotated more than 6000 transcripts. Here, we selected 100 of the inferred proteins homolog to those of Arabidopsis thaliana, which were particularly interesting for a detailed study of their potential functions, protein–protein interactions, and distance trees. As expected, a plethora of proteins associated with gametogenesis and embryogenesis in angiosperms, such as FERONIA (FER) and CHROMATING REMODELING 11 (CHR11) were identified, and more than a dozen candidates potentially involved in apomixis, such as ARGONAUTE family (AGO4, AGO9, and AGO 10), BABY BOOM (BBM), FASCIATED STEM4 (FAS4), FERTILIZATION-INDEPENDENT ENDOSPERM (FIE), and MATERNAL EFFECT EMBRYO ARREST29 (MEE29). In addition, proteins involved in the response to biotic and abiotic stresses were widely represented, as shown by the enrichment of heat-shock proteins. Using the String platform, the interactome revealed that most of the protein–protein interactions were predicted based on experimental, database, and text mining datasets, with MULTICOPY SUPPRESSOR OF IRA4 (MSI4) showing the highest number of interactions: 16. Lastly, some proteins were studied through distance trees by comparing alignments with respect to more distantly or closely related plant groups. This analysis identified DCL4 as the most distant protein to the predicted common ancestor. New genomic information in relation to gametophyte development, including apomictic reproduction, could expand our current vision of evo-devo approaches. [ABSTRACT FROM AUTHOR]

Background The predominance of sex in eukaryotes, despite the high costs of meiosis and mating, remains an evolutionary enigma. Many theories have been proposed, none of them being conclusive on its own, and they are, in part, not well applicable to land plants. Sexual reproduction is obligate in embryophytes for the great majority of species. Scope This review compares the main forms of sexual and asexual reproduction in ferns and angiosperms, based on the generation cycling of sporophyte and gametophyte (leaving vegetative propagation aside). The benefits of sexual reproduction for maintenance of genomic integrity in comparison to asexuality are discussed in the light of developmental, evolutionary, genetic and phylogenetic studies. Conclusions Asexual reproduction represents modifications of the sexual pathway, with various forms of facultative sexuality. For sexual land plants, meiosis provides direct DNA repair mechanisms for oxidative damage in reproductive tissues. The ploidy alternations of meiosis–syngamy cycles and prolonged multicellular stages in the haploid phase in the gametophytes provide a high efficiency of purifying selection against recessive deleterious mutations. Asexual lineages might buffer effects of such mutations via polyploidy and can purge the mutational load via facultative sexuality. The role of organelle–nuclear genome compatibility for maintenance of genome integrity is not well understood. In plants in general, the costs of mating are low because of predominant hermaphroditism. Phylogenetic patterns in the archaeplastid clade suggest that high frequencies of sexuality in land plants are concomitant with a stepwise increase of intrinsic and extrinsic stress factors. Furthermore, expansion of genome size in land plants would increase the potential mutational load. Sexual reproduction appears to be essential for keeping long-term genomic integrity, and only rare combinations of extrinsic and intrinsic factors allow for shifts to asexuality. [ABSTRACT FROM AUTHOR]

Premise: Apomixis in ferns is relatively common and obligatory. Sterile hybrids may restore fertility via apomixis at a cost of long‐term genetic stagnation. In this study, we outlined apomixis as a possible temporary phase leading to sexuality and analyzed factors relating to transitioning to and away from apomixis, such as unreduced and reduced spore formation in apomict and apo‐sex hybrid ferns. Methods: We analyzed the genome size of 15 fern species or hybrids ("taxa") via flow cytometry. The number of reduced and unreduced gametophytes was established as a proxy for viable spore formation of either type. We also calculated the spore abortion ratio (sign of reduced spores) in several taxa, including the apo‐sex hybrid Dryopteris × critica and its 16 apomictically formed offspring. Results: Four of 15 sampled taxa yielded offspring variable in genome size. Specifically, each variable taxon formed one viable reduced plant among 12–451 sampled gametophytes per taxon. Thus, haploid spore formation in the studied apomicts was very rare but possible. Spore abortion analyses indicated gradually decreasing abortion (haploid spore formation) over time. In Dryopteris × critica, abortion decreased from 93.8% to mean 89.5% in one generation. Conclusions: Our results support apomixis as a transitionary phase toward sexuality. Newly formed apomicts hybridize with sexual relatives and continue to form haploid spores early on. Thus, they may get the genomic content necessary for regular meiosis and restore sexuality. If the missing relative goes extinct, the lineage gets locked into apomixis as may be the case with the Dryopteris affinis complex. [ABSTRACT FROM AUTHOR]

Haploid sporophytes of Anisocampium niponicum with 2n = 40, were produced artificially by induced apogamy in vitro. They were subsequently transplanted into pots and two of them have been cultivated for the investigation of sporogenesis and/or production of chimera for more than 20 years. Haploid A. niponicum is sterile, but an abnormal chimeric pinnule that developed spontaneously in a single frond produced sporangia with spores. Each sporangium bore approximately 32 spores that were almost uniform in size. Sowing of these spores resulted in 50 gametophytes. Of 20 gametophytes cultured individually, five produced sporophytes apogamously after eight months. Both the gametophytes and subsequent apogamous sporophytes showed a chromosome number of 2n = 40. Our study demonstrates that a haploid sporophyte offspring can be produced from a haploid mother sporophyte via haploid spores. Since asexual reproduction is a prominent evolutionary process in ferns, the reproduction of a haploid A. niponicum sporophyte by unreduced spore formation might help to elucidate how apogamous ferns occur and evolve. [ABSTRACT FROM AUTHOR]

This cytological study on Isoetes coromandelina L.f (Isoetaceae) from Tirunelveli District, Tamil Nadu, South India shows the presence of twenty two individual chromosomes along with a fragment (n=2n=22+1) during meiosis in megaspore mother cells without the presence of any bivalent. Cytomixis between two megaspores is very common, resulting in the formation of a high percentage of abortive spores. The present material is of an apogamous taxon with asynaptic Meiosis as reported by Abraham and Ninan from Kerala and Karnataka.

Apomixis, an asexual reproduction process that does not involve meiosis and fertilization, has shown great potential in accelerating crop breeding by producing offspring with genotypes identical to those of the parental plants. In brown algae, parthenogenesis and apogamy are types of apomixis observed in gametophytes. Saccharina japonica is a widely cultivated and highly productive marine brown algae "crop". Previously, parthenogenesis was reported in female gametophytes and apogamy in male gametophytes, respectively. However, in this study, we observed apogamy in female gametophytes and parthenogenesis in male gametophytes during S. japonica gametophyte development. Apogamy in females occurred later than parthenogenesis, with apogamous sporophytes forming occasionally and being easily overlooked due to the prevalence of partheno-sporophytes. Some male gametophytes developed into partheno-sporophytes by producing smaller egg-like cells. The reproductive ability of male gametophytes through parthenogenesis was positively correlated with the formation ability of egg-like cells. These results suggest that parthenogenesis-related reproductive traits of S. japonica are not only linked to sex traits, but are complex traits regulated by multiple genes. Gene expression analysis revealed that early feminization in male gametophytes was associated with the combined down-regulation of male-biased genes and the up-regulation of female-biased genes. Twenty-two male-biased genes and 14 female-biased genes were further identified as parthenogenesis-related candidate genes, potentially playing crucial roles in the early feminization of male gametophytes. This study deepens our understanding of the regulation of parthenogenesis and apomixis in gametophytes of S. japonica. It lays the foundations for further research on the molecular mechanism of apomixis and its applications in S. japonica breeding. • Apogamy occurs occasionally and later than parthenogenesis in female gametophytes. • Male gametophytes develop into partheno-sporophytes via smaller egg-like cells. • Male gametophyte parthenogenesis correlates with egg-like cell formation ability. • Early feminization in male gametophytes involves the regulation of sex-biased genes. [ABSTRACT FROM AUTHOR]

Ferns are a diverse evolutionary lineage, sister to the seed plants, which is of great ecological importance and has a high biotechnological potential. Fern gametophytes represent one of the simplest autotrophic, multicellular plant forms and show several experimental advantages, including a simple and space-efficient in vitro culture system. However, the molecular basis of fern growth and development has hardly been studied. Here, we report on a proteomic study that identified 417 proteins shared by gametophytes of the apogamous fern Dryopteris affinis ssp. affinis and its sexual relative Dryopteris oreades. Most proteins are predicted to localize to the cytoplasm, the chloroplast, or the nucleus, and are linked to enzymatic, binding, and structural activities. A subset of 145 proteins are involved in growth, reproduction, phytohormone signaling and biosynthesis, and gene expression, including homologs of SHEPHERD (SHD), HEAT SHOCK PROTEIN 90-5 (CR88), TRP4, BOBBER 1 (BOB1), FLAVONE 3'-O-METHYLTRANSFERASE 1 (OMT1), ZEAXANTHIN EPOXIDASE (ABA1), GLUTAMATE DESCARBOXYLASE 1 (GAD), and dsRNA-BINDING DOMAIN-LIKE SUPERFAMILY PROTEIN (HLY1). Nearly 25% of the annotated proteins are associated with responses to biotic and abiotic stimuli. As for biotic stress, the proteins PROTEIN SGT1 HOMOLOG B (SGT1B), SUPPRESSOR OF SA INSENSITIVE2 (SSI2), PHOSPHOLIPASE D ALPHA 1 (PLDALPHA1), SERINE/THREONINE-PROTEIN KINASE SRK2E (OST1), ACYL CARRIER PROTEIN 4 (ACP4), and NONHOST RESISTANCE TO P. S. PHASEOLICOLA1 (GLPK) are worth mentioning. Regarding abiotic stimuli, we found proteins associated with oxidative stress: SUPEROXIDE DISMUTASE[CU-ZN] 1 (CSD1), and GLUTATHIONE S-TRANSFERASE U19 (GSTU19), light intensity SERINE HYDROXYMETHYLTRANSFERASE 1 (SHM1) and UBIQUITIN-CONJUGATING ENZYME E2 35 (UBC35), salt and heavy metal stress included MITOCHONDRIAL PHOSPHATE CARRIER PROTEIN 3 (PHT3;1), as well as drought and thermotolerance: LEA7, DEAD-BOX ATP-DEPENDENT RNA HELICASE 38 (LOS4), and abundant heat-shock proteins and other chaperones. In addition, we identified interactomes using the STRING platform, revealing protein–protein associations obtained from co-expression, co-occurrence, text mining, homology, databases, and experimental datasets. By focusing on ferns, this proteomic study increases our knowledge on plant development and evolution, and may inspire future applications in crop species. [ABSTRACT FROM AUTHOR]

Premise: Apomixis and hybridization are two essential and complementary factors in the evolution of plants, including ferns. Hybridization combines characteristics from different species, while apomixis conserves features within a lineage. When combined, these two processes result in apo‐sex hybrids. The conditions leading to the formation of these hybrids are poorly understood in ferns. Methods: We cultivated spores from 66 fern samples (43 apomicts, 7 apo‐sex hybrids, and 16 sexuals), and measured their development in vitro over 16 weeks. We evaluated germination, lateral meristem formation rates, sexual expression, and production of sporophytes and then compared ontogenetic patterns among the three groups. Results: The three examined groups formed antheridia (male gametangia) but differed in overall gametophyte development. Sexual species created archegonia (female, 86% of viable samples), but no sporophytes. Apomicts rarely created nonfunctional archegonia (8%) but usually produced apogamous sporophytes (75%). Surprisingly, apomictic and sexual species showed similar development speed. The sexually reproducing parents of viable studied hybrids formed about twice as many meristic gametophytes as the apomictic parents (39% vs. 20%, respectively). Conclusions: We present the most thorough comparison of gametangial development of sexual and apomictic ferns, to date. Despite expectations, apomictic reproduction might not lead to earlier sporophyte formation. Apomicts produce functional sperm and thus can contribute this type of gamete to their hybrids. The development patterns found in the parents of hybrids indicate a possible increase of hybridization rates by antheridiogens. The apo‐sex hybrids always inherit the apomictic reproductive strategy and are thus capable of self‐perpetuation. [ABSTRACT FROM AUTHOR]

The gametophyte of ferns reproduces either by sexual or asexual means. In the latter, apogamy represents a peculiar case of apomixis, in which an embryo is formed from somatic cells. A proteomic and physiological approach was applied to the apogamous fern Dryopteris affinis ssp. affinis and its sexual relative D. oreades. The proteomic analysis compared apogamous vs. female gametophytes, whereas the phytohormone study included, in addition to females, three apogamous stages (filamentous, spatulate, and cordate). The proteomic profiles revealed a total of 879 proteins and, after annotation, different regulation was found in 206 proteins of D. affinis and 166 of its sexual counterpart. The proteins upregulated in D. affinis are mostly associated to protein metabolism (including folding, transport, and proteolysis), ribosome biogenesis, gene expression and translation, while in the sexual counterpart, they account largely for starch and sucrose metabolism, generation of energy and photosynthesis. Likewise, ultra-performance liquid chromatography-tandem spectrometry (UHPLC-MS/MS) was used to assess the levels of indol-3-acetic acid (IAA); the cytokinins: 6-benzylaminopurine (BA), trans-Zeatine (Z), trans-Zeatin riboside (ZR), dyhidrozeatine (DHZ), dyhidrozeatin riboside (DHZR), isopentenyl adenine (iP), isopentenyl adenosine (iPR), abscisic acid (ABA), the gibberellins GA3 and GA4, salicylic acid (SA), and the brassinosteroids: brassinolide (BL) and castasterone (CS). IAA, the cytokinins Z, ZR, iPR, the gibberellin GA4, the brassinosteoids castasterone, and ABA accumulated more in the sexual gametophyte than in the apogamous one. When comparing the three apogamous stages, BA and SA peaked in filamentous, GA3 and BL in spatulate and DHRZ in cordate gametophytes. The results point to the existence of large metabolic differences between apogamous and sexual gametophytes, and invite to consider the fern gametophyte as a good experimental system to deepen our understanding of plant reproduction. [ABSTRACT FROM AUTHOR]

The gametophyte of ferns reproduces either by sexual or asexual means. In the latter, apogamy represents a peculiar case of apomixis, in which an embryo is formed from somatic cells. A proteomic and physiological approach was applied to the apogamous fern Dryopteris affinis ssp. affinis and its sexual relative D. oreades. The proteomic analysis compared apogamous vs. female gametophytes, whereas the phytohormone study included, in addition to females, three apogamous stages (filamentous, spatulate, and cordate). The proteomic profiles revealed a total of 879 proteins and, after annotation, different regulation was found in 206 proteins of D. affinis and 166 of its sexual counterpart. The proteins upregulated in D. affinis are mostly associated to protein metabolism (including folding, transport, and proteolysis), ribosome biogenesis, gene expression and translation, while in the sexual counterpart, they account largely for starch and sucrose metabolism, generation of energy and photosynthesis. Likewise, ultra-performance liquid chromatography-tandem spectrometry (UHPLC-MS/MS) was used to assess the levels of indol-3-acetic acid (IAA); the cytokinins: 6-benzylaminopurine (BA), trans-Zeatine (Z), trans-Zeatin riboside (ZR), dyhidrozeatine (DHZ), dyhidrozeatin riboside (DHZR), isopentenyl adenine (iP), isopentenyl adenosine (iPR), abscisic acid (ABA), the gibberellins GA3 and GA4, salicylic acid (SA), and the brassinosteroids: brassinolide (BL) and castasterone (CS). IAA, the cytokinins Z, ZR, iPR, the gibberellin GA4, the brassinosteoids castasterone, and ABA accumulated more in the sexual gametophyte than in the apogamous one. When comparing the three apogamous stages, BA and SA peaked in filamentous, GA3 and BL in spatulate and DHRZ in cordate gametophytes. The results point to the existence of large metabolic differences between apogamous and sexual gametophytes, and invite to consider the fern gametophyte as a good experimental system to deepen our understanding of plant reproduction.

Leek (Allium tuberosum) is highly heterozygous owing to asynchronous maturity of the androecium and gynoecium. Haploid induction is pertinent to overcome this impediment in leek breeding. In this study, unpollinated ovaries of leek were float cultured on liquid medium to induce development of haploid plantlets. On Murashige and Skoog media supplemented with 3–12% sucrose, 0–0.4 mg/L−1 2,4-dichlorophenoxyacetic acid (2,4-D), and 0–2 mg/L−1 zeatin (Zt), many ovules produced parthenogenic and/or apogamic embryos. Microscopic observation confirmed the embryos originating from the egg and egg-like antipodal cell. The optimal concentration of the medium components is 0.1 mg/L−1 2,4-D, 0.5 mg/L−1 Zt and 9% sucrose, in which 58.3% ovules produced embryos. This method will be useful in haploid breeding of leek on account of the high frequency of autonomous haploid embryogenesis. Apogamy of antipodal cells is few to occur naturally in higher plants, therefore this report is the confirmation of antipodal apogamy. The origin and development of parthenogenesis and apogamy in leek are discussed. [ABSTRACT FROM AUTHOR]

Abstract Background Apogamy is a unique asexual reproduction in the ferns, in which somatic cells of gametophytes go through dedifferentiation and then differentiate into haploid sporophytes bypassing fertilization. Restricted to the lack of genomic information, molecular mechanisms of apogamy have remained unclear. Comparative transcriptome analysis was conducted at six stages between sexual reproduction and apogamy in the fern Adiantum reniforme var. sinense, in an effort to identify genes and pathways that might initiate the asexual reproduction. Results Approximately 928 million high-quality clean reads were assembled into 264,791 unigenes with an average length of 615 bp. A total of 147,865 (55.84%) unigenes were successfully annotated. Differential genes expression analysis indicated that transcriptional regulation was more active in the early stage of apogamy compared to sexual reproduction. Further comparative analysis of the enriched pathways between the early stages of the two reproductive modes demonstrated that starch and sucrose metabolism pathway responsible for cell wall was only significantly enriched in asexual embryonic cell initiation. Furthermore, regulation of plant hormone related genes was more vigorous in apogamy initiation. Conclusion These findings would be useful for revealing the initiation of apogamy and further understanding of the mechanisms related to asexual reproduction.

Premise: Xeric environments impose major constraints on the fern life cycle, yet many lineages overcome these limitations by evolving apomixis. Here, we synthesize studies of apomixis in ferns and present an evidence‐based model for the evolution and establishment of this reproductive strategy, focusing on genetic and environmental factors associated with its two defining traits: the production of "unreduced" spores (n = 2n) and the initiation of sporophytes from gametophyte tissue (i.e., diplospory and apogamy, respectively). Methods: We evaluated existing literature in light of the hypothesis that abiotic characteristics of desert environments (e.g., extreme diurnal temperature fluctuations, high light intensity, and water limitation) drive the evolution of obligate apomixis. Pellaeid ferns (Cheilanthoideae: Pteridaceae) were examined in detail, as an illustrative example. We reconstructed a plastid (rbcL, trnG–trnR, atpA) phylogeny for the clade and mapped reproductive mode (sexual versus apomictic) and ploidy across the resulting tree. Results: Our six‐stage model for the evolution of obligate apomixis in ferns emphasizes the role played by drought and associated abiotic conditions in the establishment of this reproductive approach. Furthermore, our updated phylogeny of pellaeid ferns reveals repeated origins of obligate apomixis and shows an increase in the frequency of apomixis, and rarity of sexual reproduction, among taxa inhabiting increasingly dry North American deserts. Conclusions: Our findings reinforce aspects of other evolutionary, physiological, developmental, and omics‐based studies, indicating a strong association between abiotic factors and the establishment of obligate apomixis in ferns. Water limitation, in particular, appears critical to establishment of this reproductive mode. [ABSTRACT FROM AUTHOR]

The Japanese apogamous fern Dryopteris yakusilvicola was previously thought to be a hybrid of D. sabaei and D. sparsa var. sparsa based on morphological characterization and enzyme electrophoresis. However, the phylogenetic relationships with D. hayatae, D. melanocarpa var. melanocarpa, D. melanocarpa var. elegans, and D. sparsa var. ryukyuensis have been unclear. We performed a molecular phylogenetic analysis of the DNA sequences of two plastid (trnL‐F, rbcL) and one low‐copy nuclear marker (AK1) of Dryopteris yakusilvicola and related species. The origin of D. yakusilvicola is not a hybrid between D. sparsa var. sparsa and D. sabaei, but the parents appear to have been undetected species closely related to D. melanocarpa var. elegans and D. sabaei. Analysis of additional species of the D. sparsa complex around Southeast Asia and additional nuclear markers are needed to clarify the relationships of this complex. [ABSTRACT FROM AUTHOR]

Cytological observations were systematically employed for detecting cellular morphogenesis and chromosomal behavior of apogamous development of new sporophytes from individually cultured male gametophytes of Pyropia haitanensis. The results showed that the apical parts of premature male gametophytes initially underwent obvious apogamy, which primarily appeared in the functionally specialized spermatangial mother cells (SSMCs) that would not develop into highly differentiated spermatangia but change their developmental fate and become brownish red in color. These brownish red spermatangial mother cells (SMCs) were released accompanying the spermatia dissociated from mature spermatangia and then rarely formed circular carpospore-like cells (CLCs) and carpospore-like germlings (CLGs) that eventually developed into normal apogamous sporophytic conchocelis. Chromosome counts indicated that spermatangia and spermatia were haploid (n = 5), whereas SSMCs, CLCs, and CLGs were diploid (2n = 10). Furthermore, filamentous conchocelis and conchosporangial branch cells of apogamous sporophytes were diploid and no haploid, aneuploid, and polyploid were observed, suggesting that spontaneous chromosome doubling occurred in the process of apogamy and initiated rarely SSMCs rather than released spermatia. Moreover, pit connections were observed in the whole development of apogamous sporophytes like in heterozygous sporophytes. Mitotic chromosomal behaviors appeared during the first two divisions of conchospores that normally grew into haploid tetrads and developed into unisexual male gametophytes with the characteristics of apogamy in the aging cultures. [ABSTRACT FROM AUTHOR]

Apomixis was originally defined as the replacement of sexual reproduction by an asexual process that does not involve fertilization but, in angiosperms, it is often used in the more restricted sense of asexual reproduction through seeds. In ferns, apomixis combines the production of unreduced spores (diplospory) and the formation of sporophytes from somatic cells of the prothallium (apogamy). The genes that control the onset of apogamy in ferns are largely unknown. In this study, we describe the gametophyte transcriptome of the apogamous fern Dryopteris affinis ssp. affinis using an RNA-Seq approach to compare the gene expression profiles of one- and two-dimensional gametophytes, the latter containing apogamic centers. After collapsing highly similar de novo transcripts, we obtained 166,191 unigenes, of which 30% could be annotated using public databases. Multiple quality metrics indicate a good quality of the de novo transcriptome with a low level of fragmentation. Our data show a total of 10,679 genes (6% of all genes) to be differentially expressed between gametophytes of filamentous (one-dimensional) and prothallial (two-dimensional) architecture. 6,110 genes were up-regulated in two-dimensional relative to one-dimensional gametophytes, some of which are implicated in the regulation of meristem growth, auxin signaling, reproduction, and sucrose metabolism. 4,570 genes were down-regulated in two-dimensional versus one-dimensional gametophytes, which are enriched in stimulus and defense genes, as well as genes involved in epigenetic gene regulation and ubiquitin degradation. Our results provide insights into free-living gametophyte development, focusing on the filamentous-to-prothallus growth transition, and provide a useful resource for further investigations of asexual reproduction. Image 1 • A total of 10679 genes expressed differentially in one- vs two-dimensional gametophytes of Dryopteris affinis spp affinis. • Genes acting on meristem growth, auxin signalling, sucrose metabolism, and reproduction upregulated at two-dimensional stage. • Orthologues of SHOOT MERISTEMLESS and NAM proteins, related to meristem growth in angiosperms, were identified. • Processes related to sugar metabolism were enriched GO terms in two-dimensional gametophytes. [ABSTRACT FROM AUTHOR]

Since the 1970s, an apomictic tetraploid beech fern (genus Phegopteris (C. Presl) Fée) has been known in northeastern North America. Previously published isozyme data suggest that this lineage is of allopolyploid origin involving long beech fern (P. connectilis (Michx.) Watt.) but not broad beech fern (P. hexagonoptera (Michx.) Fée), as originally hypothesized. Its second progenitor remains unknown. We performed a principal components analysis of the apomict and its North American congeners to elucidate morphological differences between them. We recognize the apomictic tetraploid at specific rank as P. excelsior N. R. Patel & A. V. Gilman and provide an illustration, a range map, a list of exsiccatae, and a key to Phegopteris species of North America. [ABSTRACT FROM AUTHOR]

Lessonia berteroana is one of the most exploited seaweeds in the Southeastern Pacific and its populations are recurrently facing overexploitation in northern Chile. Since germplasms are not available, we decided to start gametophyte biobanking to support conservation measures for this important resource in the future. Spores of L. berteroana from nine localities at the Atacama coast were used to establish clonal male and female gametophyte cultures. Unexpectedly, after isolation and under low light conditions, juvenile sporophytes originated from somatic cells via apogamy in most female gametophyte strains. In addition, eggs from solitary female gametophytes from Caleta Cisnes and Torres del Inca had a strong tendency to generate sporophytes by parthenogenesis, some of them even under low light regimes. Contrarily, female gametophytes of the sister species Lessonia spicata from southern Chile showed no evidence for apomixis. When one of these L. berteroana strains is cross-fertilized with L. spicata, true hybrids emerged based on the presence of eggs and subsequent sperm attraction. These observations contrast with (i) kelp recruitment assumed to be majorly by sexual reproduction and (ii) the strict reproductive separation of the two taxa reported for natural populations within their contact zone at 30°S and highlight their consideration for future repopulation and breeding programs of Lessonia. [ABSTRACT FROM AUTHOR]

A new diploid taxon within the Dryopteris affinis complex is described from the Island of Arran, Scotland: D. affinis subsp. cluthensis. Its affinities are discussed and distinguishing features outlined. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Pityrogramma calomelanos (L.) Link, popularly known as "Silver fern" has significant importance as a medicinal plant used traditionally for its astringent, analgesic, anti-haemorrhagic, anti-hypertensive, anti-pyretic and anthelminthic properties. This fern demonstrates an increased morphogenetic potential towards sporophyte formation, upon exposure to low doses of gamma radiation. Young sporophytic leaf crosier cultures were established in vitro on agar based Knop's media with and without 20 g/l sucrose. The cultures were subjected to 60Co radiations in the range of 2.5–100 Gy. Apospory (production of gametophytes on sporophytic tissue without spores) was observed on leaf tissue cultured on Knops media with and without sucrose in P. calomelanos, at the end of 60 days. 5 Gy treated explants showed high number of aposporous gametophytes and was comparable to the control. Other tested doses reduced the aposporous gametophyte production significantly. In the second phase of the experimentation, the cultures were retained on the gametophyte induction media for a period of 4 weeks. Aposporous gametophytes were observed to proliferate with occasional development of antheridia. At the end of 4 weeks, morphogenetic development on the gametophytic tissue resulted in a significantly higher number of apogamous sporophytes (production of sporophytes without fusion of gametes) were obtained on 5 Gy treated tissue as compared to control and all the other treated explants. Apogamous sporophytes thus produced were successfully grown in the greenhouse and transferred to the field. Thus the use of gamma radiation in vitro not only reduced the need for sucrose for induction of apospory in P.calomelanos , it also exhibited hormesis at 5 Gy for improved sporophyte production. [ABSTRACT FROM AUTHOR]

Asexual reproduction is widespread in land plants, including ferns where 10% of all species are obligate asexuals. In these ferns, apogamous sporophytes are generated directly from gametophytes, bypassing fertilization. In the model fern Ceratopteris richardii, a sexual species, apogamy can be induced by culture on high sugar media. BABY BOOM ( BBM) genes in angiosperms are known to promote somatic embryogenesis, which like apogamy produce sporophytes without fertilization. Here, a Brassica napus BBM ( Bn BBM) was used to investigate genetic similarity between apogamy in ferns and somatic embryogenesis in angiosperms. A C. richardii transcriptome was constructed from which one AINTEGUMENTA- LIKE unigene, Cr ANT, was identified. Whole mount in situ hybridization showed that Cr ANT is expressed in sperm and fertilized eggs. Phylogenetic analysis grouped Cr ANT with other non-seed-plant ANT genes to the eu ANT clade but in a branch separate from BBM genes. Overexpression of Cr ANT or Bn BBM promotes apogamy in C. richardii without sugar supplement. Cr ANT knockdown gametophytes responded weakly to sugar for apogamy promotion. Theses results suggest some genetic conservation between apogamy and somatic embryogenesis and that such asexual reproduction may be ancient. [ABSTRACT FROM AUTHOR]

Performing proteomic studies on non-model organisms with little or no genomic information is still difficult. However, many specific processes and biochemical pathways occur only in species that are poorly characterized at the genomic level. For example, many plants can reproduce both sexually and asexually, the first one allowing the generation of new genotypes and the latter their fixation. Thus, both modes of reproduction are of great agronomic value. However, the molecular basis of asexual reproduction is not well understood in any plant. In ferns, it combines the production of unreduced spores (diplospory) and the formation of sporophytes from somatic cells (apogamy). To set the basis to study these processes, we performed transcriptomics by next-generation sequencing (NGS) and shotgun proteomics by tandem mass spectrometry in the apogamous fern D. affinis ssp. affinis. For protein identification we used the public viridiplantae database (VPDB) to identify orthologous proteins from other plant species and new transcriptomics data to generate a "species-specific transcriptome database" (SSTDB). In total 1,397 protein clusters with 5,865 unique peptide sequences were identified (13 decoy proteins out of 1,410, protFDR 0.93% on protein cluster level). We show that using the SSTDB for protein identification increases the number of identified peptides almost four times compared to using only the publically available VPDB. We identified homologs of proteins involved in reproduction of higher plants, including proteins with a potential role in apogamy. With the increasing availability of genomic data from non-model species, similar proteogenomics approaches will improve the sensitivity in protein identification for species only distantly related to models. [ABSTRACT FROM AUTHOR]

The marine red seaweed Pyropia yezoensis has a haploid-diploid life cycle wherein two heteromorphic generations, a haploid gametophyte and a diploid sporophyte, are reciprocally generated from conchospores and carpospores, respectively. When we treated gametophytic blades of P. yezoensis with H2O2, discharge of asexual monospores was accelerated, resulting in increased numbers of gametophytic clones. Production of sporophytes without fertilization of male and female gametes was also observed. These findings indicate that oxidative stress can induce vegetative cells to develop into monospores that produce gametophytes asexually and can sometimes prompt carpospores to develop into sporophytes. The discovery of oxidative stresstriggered asexual reproduction and -apogamy will stimulate progress in studies of life-cycle regulation in P. yezoensis. [ABSTRACT FROM AUTHOR]

Los helechos y licófitas tienen diversas estrategias reproductivas que las habilitan para invadir ambientes perturbados y áreas sin vegetación. En este trabajo se estudió la presencia, abundancia y estrategias reproductivas de individuos que crecen en suelos de bosques templados, en la Sierra Nevada, Estado de México. Se eligieron cuatro zonas en las que se instaló un cuadrante permanente de 200m² para cada una, con el fin de registrar variables edáficas, ambientales y de vegetación. La relación entre variables se pudo explorar con análisis de varianza y Análisis Canónico de Correspondencia dando como resultado tres paisajes con base en el grado de alteración ambiental: baja, moderada y severa. Fueron recolectadas cinco muestras de suelo en cada zona para inducir la germinación de helechos y licofitas, obteniendo resultados que revelan una relación positiva entre el grado de alteración y la abundancia de Cheilanthes. En paisajes con baja alteración, la riqueza de especies fue mayor comparada con paisajes de alteración severa, en la que la presencia de especies se redujo a Cheilanthes bonariensis, C. marginata y Pellaea ternifolia subsp. ternifolia. El suelo fue un banco de esporas de Cheilanthes y Pellaea ternifolia subsp. ternifolia ya que las esporas permanecieron viables por diferentes intervalos de tiempo. La apogamia fue la estrategia reproductiva común para especies de Cheilanthes en ecosistemas alterados, aunque estas especies tienen reproducción sexual en ecosistemas naturales con suficiente agua. Por otro lado, Pellaea ternifolia subsp. ternifolia solo presentó reproducción sexual. La apogamia puede estar relacionada con que la alternancia de generaciones se realice en menos tiempo, comparado con un ecosistema de baja alteración.Presence, abundance and reproductive strategies of ferns in disturbed areas of Sierra Nevada, México. Diverse reproductive strategies shown by ferns and lycophytes allow them to colonize a variety of habitats, particularly after the incidence of natural or anthropogenic disturbances. This study assessed the presence, abundance and reproductive strategies of ferns growing in soils of temperate forests with different levels of disturbance at the Sierra Nevada mountain range in the State of Mexico. Vegetation, soil and environmental variables were recorded in 200m² permanent plots located in four forest stands. One-Way ANOVA and Canonical Correspondence Analysis resulted in the recognition of three landscape types defined by the degree of environmental alteration: low, moderate and severe. Also, from five soil samples collected in each stand, germination of ferns and lycophytes was induced. A positive relationship was found between the alteration degree and Cheilanthes abundance. Under a low landscape alteration regime, species richness is restricted to Cheilanthes bonariensis, C. marginata and Pellaea ternifolia subsp. ternifolia. The soil is a reservoir of spores of Cheilanthes and Pellaea ternifolia subsp. ternifolia because their spores can remain viable for different time intervals. Apogamy is the usual reproductive strategy of Cheilanthes species in disturbed ecosystems, although these species also show sexual reproduction in natural ecosystems with adequate water availability. Apogamy may be related to a shorter generation time in comparison with a low disturbed ecosystem. On the other hand, Pellaea ternifolia subsp. ternifolia only has sexual reproduction. Apogamy might be related to a faster generation in comparison with a low disturbed ecosystem. Rev. Biol. Trop. 59 (1): 417-433. Epub 2011 March 01.

Phegopteris connectilis is an apogamous species, endangered in certain regions. The ex situ conservation, through the in vitro production of the gametophyte, as well as the embryos and the plants, followed by their cryo-stocking, is necessary for ensuring the survival of the endangered species, while at pace with international methodology. The differentiation of the gametophyte is characteristic of the leptosporangiate ferns, passing through the following stages: prothallic filament, prothallic blade, cordate prothallus. The uni- or pluri-serial branchings formed on the gametophytes constitute the initial differentiation stage of a new prothallus. One-cell trichomes are differentiated on the gametophyte, which produce wax, protective against de-hydration. The in vitro culture has evinced the fast life cycle of this species, as the apogamous embryo is formed in the course of only seven weeks after culture inception. The first protophylles are formed out of the apogamous embryo and the initial stem cell is differentiated between the first two. In very few cases, however, it was noticed, at the base of the first leaf, the differentiation of the first root; the latter is formed at a stage when the young sporophyte already has two to three protophylles, as rhizogenesis is favoured by the addition of AIA and Kin in the medium. The plants thus obtained can be utilized for re-establishing the natural populations in the regions where the species is endangered.

The life cycles of eukaryotes alternate between haploid and diploid phases, which are initiated by meiosis and gamete fusion, respectively. In both ascomycete and basidiomycete fungi and chlorophyte algae, the haploid-to-diploid transition is regulated by a pair of paralogous homeodomain protein encoding genes. That a common genetic program controls the haploid-to-diploid transition in phylogenetically disparate eukaryotic lineages suggests this may be the ancestral function for homeodomain proteins. Multicellularity has evolved independently in many eukaryotic lineages in either one or both phases of the life cycle. Organisms, such as land plants, exhibiting a life cycle whereby multicellular bodies develop in both the haploid and diploid phases are often referred to as possessing an alternation of generations. We review recent progress on understanding the genetic basis for the land plant alternation of generations and highlight the roles that homeodomain-encoding genes may have played in the evolution of complex multicellularity in this lineage. [ABSTRACT FROM AUTHOR]

This review provides a synopsis of apogamous reproduction in ferns and highlights important progress made in recent studies of fern apomixis. First, a summary of the apomictic fern life cycle is provided, distinguishing between two pathways to diploid spore production that have been documented in apomictic ferns (premeiotic endomitosis and meiotic first division restitution) and briefly discussing the evolutionary implications of each. Next, recent trends in fern apomixis research are discussed, exposing a shift in focus from the observation and characterization of apomixis in ferns to more integrated studies of the evolutionary and ecological implications of this reproductive mode. Peer-reviewed contributions from the past decade are then summarized, spanning the identification of new apomictic lineages through to the developmental, phylogenetic, and population genetic insights that have been made in studies of fern apomixis during that time. Gaps in our understanding are also discussed, including the extent and implications of recombinant apomixis in ferns, the possible reversibility of reproductive mode (from apomictic to sexual) in ferns, and the genomic causes and consequences of apomixis in seed free vascular plants. To conclude, future directions for fern apomixis research are proposed in the context of modern technological advances and recent insights from studies of apomixis in other groups. [ABSTRACT FROM AUTHOR]

Background and Aims In ferns, apomixis is an important mode of asexual reproduction. Although the mechanisms of fern reproduction have been studied thoroughly, most previous work has focused on cases in which ferns reproduce either exclusively sexually or exclusively asexually. Reproduction of ferns with potentially mixed systems and inheritance of apomixis remains largely unknown. This study addresses reproduction of the pentaploid Dryopteris X critica, a hybrid of triploid apomictic D. borreri and tetraploid sexual D. filix-mas. Methods Spore size, abortion percentage and number of spores per sporangium were examined in pentaploid plants of D. X critica grown in an experimental garden. The sporangial content of leaf segments was cultivated on an agar medium, and DNA ploidy levels were estimated by DAPI flow cytometry in 259 gametophytes or sporophytes arising from the F2 generation of the pentaploid hybrid. Key Results The hybrid is partly fertile (89-94% of aborted spores) and shows unstable sporogenesis with sexual and apomictic reproduction combined. The number of spores per sporangium varied from approx. 31 to 64. Within a single sporangium it was possible to detect formation of either only aborted spores or various mixtures of aborted and well-developed reduced spores and unreduced diplospores. The spores germinated in viable gametophytes with two ploidy levels: pentaploid (5x, from unreduced spores) and half of that (approx. 2.5x, from reduced spores). Moreover, 2-15% of gametophytes (both 2.5x and 5x) formed a viable sporophyte of the same ploidy level due to apogamy. Conclusions This study documents the mixed reproductive mode of a hybrid between apomictic and sexual ferns. Both sexual reduced and apomictic unreduced spores can be produced by a single individual, and even within a single sporangium. Both types of spores give rise to viable F2 generation gametophytes and sporophytes. [ABSTRACT FROM AUTHOR]

In vitro culture of the moss Amblystegium serpens (Amblystegiaceae) was established on hormone-free Murashige and Skoog (MS) medium that contained a half amount of MS micro- and macro- mineral salts and vitamins, 100 mg/l myoinositol, 30 g/l sucrose, and 0.70% (w/v) agar. Spores were germinated and primary protonema developed on the above medium at 16 h day/8 h night 25±2ºC, 60-70% air humidity, and irradiance of 47 μmol/m2s. Three months after development of primary protonema, seven sporophytes appeared directly from primary protonema without generation alternation. The phenomenon of apogamous sporophyte formation is very rare, both in nature and under in vitro conditions. This is the first report of apogamy induced by Amblystegium serpens.

Se describe y se compara la morfogénesis de la fase sexual de seis especies mexicanas de Dryopteris. Las esporas de todas las especies estudiadas son monoletes y elipsoidales, la perina es plegada, de color pardo claro a pardo obscuro, su superficie es rugosa y de contornos tuberculados. El patrón de germinación es tipo Vittaria y el desarrollo protálico es tipo Aspidium. Los gametangios son del tipo característicos de los helechos leptosporangiados. Se desarrollaron esporofitos en cinco de las seis especies y solamente en D. pseudo-filix-mas es de origen apogámico (80 días). Se hace un análisis comparativo de las características gametofíticas presentes en las 12 especies mexicanas del género y se concluye que el tipo de germinación Vittaria y el patrón de desarrollo protálico tipo Aspidium y gametofitos con tricomas unicelulares, capitados marginales y superficiales, de aspecto irregular, son características que le dan unidad y caracterizan a los géneros que conforman la familia Dryopteridaceae (Moran, 1995) a excepción del género Didymochlaena.The development of the sexual phase of six Mexican species of Dryopteris is described and compared. Spores of all studied species are monolete, ellipsoid and have a rugose surface; the perine is folded, brown to dark brown, with a tubercled outline. Germination pattern is of the Vittaria-type and the development pattern of the prothallia is of the Aspidium-type. Gametangia are of the common type for the leptosporangiate advanced ferns. First leaves of the sporophytes appear 258-265 after sowing and apparently in Dryopteris pseudo-filix-mas the sporophyte have an apogamic origin (80 days). To make a comparative analysis of gametophytic characteristics in the twelve Mexican species and conclude of germination is of the Vittaria-type and development pattern prothallial is of the Aspidium-type, and unicelular trichomes on margin and superficial gametophytic to yield irregular aspect are characteristics to yield unit and characteristic to genera to conform Dryopteridaceae family (sensu Moran 1995) with the exception of Didymochlaena genus.

Se describe y se compara la morfogénesis de la fase sexual de seis especies de Dryopteris. Las esporas son monoletes, elipsoidales con perina de color pardo a pardo obscuro, su superficie es rugosa de contornos tuberculados y la perina en forma de pliegues. El patrón de germinación en todas las especies es tipo Vittaria y el desarrollo protálico tipo Aspidium. Los gametangios son del tipo común de los helechos leptosporangiados del tipo avanzado, los cuellos de los arquegonios tienen cuatro hileras de células con 4 a 5 células cada una, se forman entre los 53-218 días. Los anteridios de las seis especies se formaron entre los 46-234 días después de sembradas las esporas y tienen 3 células que rodean a la célula androgénica. Los esporofitos se formaron entre los 60-262 días después de sembradas las esporas, como producto de la fecundación en D. cinnamomea, D. karwinskyana, D. maxonii y, D. rossii y son de origen apogámico en D. munchii y D. wallichiana.The development of the sexual phase of six Mexican species of the fern Dryopteris is described and compared. The spores of all six species are monolete, ellipsoid, the perine is folded, brown to dark brown, with a tubercled outline. The germination pattern of the species is Vittaria-type and the development pattern of the prothallia is Aspidium-type. The gametangia are the common sort for the leptosporangiate advanced ferns, the necks of the archegonia have four rows of cells, with 4-5 cells each and appear 53-218 days after the sowing of the spores. The antheridia appear 46-234 days after the sowing, have three cells surrounding the androgenic cell. The first leaves of the sporophytes appear 60-262 after the sowing and, apparently in D. munchii and D. wallichiana the sporophytes have an apogamic origin.

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. [ABSTRACT FROM AUTHOR]

It has long been argued that related asexual and sexual taxa have different distribution patterns. In general, apomictic angiosperms are believed to occur preferentially at higher latitudes and elevations compared to their sexual relatives. It is thus expected that the frequency of apomixis increases with latitude or from warmer to colder climatic regions. However, despite the significant role played by apogamy in fern evolution and diversification, the distribution pattern of apogamous ferns and lycophytes has received little attention. To clarify the ecological diversity pattern and evolutional diversification of apogamous ferns, we analysed a variety of apogamous fern species with reference to latitude, elevation and climatic factors, and reconstructed the ancestral state and estimated the divergence time of Japanese apogamous ferns. Our results on the distribution of apogamous ferns along these two gradients suggest a decline in the proportion of apogamous ferns towards high latitudes and elevations. Temperature was correlated with the proportion of apogamous ferns along both gradients, and the seasonality of precipitation was correlated with the proportion of apogamous ferns along latitude. Reconstruction of ancestral state and estimates of divergence time showed that the crown groups of apogamous ferns diversified less than 15 Ma. The results of our ecological and phylogenetic approaches reinforce the hypothesis based on previously reported phylogenetic results in which the apogamous ferns appears to be correlated with strongly seasonal climates such as the Asia monsoon. [ABSTRACT FROM AUTHOR]

Davallia repens and its close relatives have been identified as a species complex in this study because of the existence of continuously morphological variation. To decipher its evolutionary history, integrated methodologies were applied in this study including morphology, cytology, reproductive biology and molecular phylogeny. Analysis of morphological characters reveals several important discriminating characteristics, such as the shape of stipe scales, frond and indusium. Both diploid and polyploid forms are present in the complex and reproduce sexually and by apogamy, respectively. The incongruence between cpDNA and nDNA phylogeny indicates a hybrid origin for most polyploid individuals. Based on the present results, we hypothesize that there were at least two ancestral lineages distributed in the Malesian region. Through hybridization, polyploidization and apogamy, some polyploid genotypes dispersed outwards to shape the extant distribution. [ABSTRACT FROM AUTHOR]

Gametophyte morphology and development of three species of Cyrtogonellum Ching (Dryopteridaceae) were described with the aim to increase our knowledge on the gametophytic stage of this fern genus especially the occurrence of sexual structures. Spore germination followed the Vittaria-type development, whereas prothallus growth followed the Aspidium-type development. Unicellular glandular hairs were located on both margins and surface of the prothalli. The heart-shaped prothalli of C. caducum showed irregularly lobed margins with multicellular filamentous to wing-like extensions. One to several agamously formed embryos were found on a single mature gametophyte of C. inaequale but only one of these developed into a mature sporophyte. Young sporophytes possessed scales with one to several glandular cells. Antheridia were found in all three species, but archegonia were observed only on mature prothalli of C. fraxinellum, which were always located near the midrib of the cordate-thalloid prothallus. The rarity of archegonia and lack of evidence for the functionality of archegonia in these species raised questions about sexual reproduction in Cyrtogonellum. [ABSTRACT FROM AUTHOR]

Different patterns of development can influence the strength and direction of selection of a trait. Here, it is shown how this may be the case for asexual reproduction in which eggs (animals) or spores (plants) inherit the maternal chromosome number through a precursor cell undergoing an endoduplication of its chromosomes prior to meiotic reduction. Asexuality involving premeiotic endoduplication ( APE) has a wide but uneven distribution among animals and plants. It is argued that different patterns of egg and spore production and differences in when endoduplication occurs, together with differences in breeding structure (dioecious vs. hermaphroditic) and reproductive strategy (e.g. spawning vs. vivipary), can result in APE being associated with fecundity costs that can result in an overall cost of sex or in an overall cost of asex or in APE being selectively neutral relative to sex. There is a lack of a close correlation between the taxonomic distribution of APE and its potential costs and benefits however, possible causes of which are explored. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109, 487-495. [ABSTRACT FROM AUTHOR]

Background and Aims Asexual reproduction is a prominent evolutionary process within land plant lineages and especially in ferns. Up to 10 % of the approx. 10 000 fern species are assumed to be obligate asexuals. In the Asplenium monanthes species complex, previous studies identified two triploid, apomictic species. The purpose of this study was to elucidate the phylogenetic relationships in the A. monanthes complex and to investigate the occurrence and evolution of apomixis within this group. Methods DNA sequences of three plastid markers and one nuclear single copy gene were used for phylogenetic analyses. Reproductive modes were assessed by examining gametophytic and sporophyte development, while polyploidy was inferred from spore measurements. Key Results Asplenium monanthes and A. resiliens are confirmed to be apomictic. Asplenium palmeri, A. hallbergii and specimens that are morphologically similar to A. heterochroum are also found to be apomictic. Apomixis is confined to two main clades of taxa related to A. monanthes and A. resiliens, respectively, and is associated with reticulate evolution. Two apomictic A. monanthes lineages, and two putative diploid sexual progenitor species are identified in the A. monanthes clade. Conclusions Multiple origins of apomixis are inferred, in both alloploid and autoploid forms, within the A. resiliens and A. monanthes clades. [ABSTRACT FROM AUTHOR]

In apogamous ferns, all offspring from a parent are expected to be clonal. However, apogamous 'species' frequently demonstrate a large amount of morphological and genetic variations. Cyrtomium fortunei composed of four varieties ( C. fortunei var. fortunei, var. clivicola, var. intermedium, and var. atropunctatum), is all reported to be apogamous triploids, but demonstrates large and continuous morphological variation. In previous studies, we showed that considerable genetic diversity was observed in many local populations of the apogamous fern 'species'. We hypothesized that genetic segregation has occurred, because neither sexual type nor intraspecific polyploidy have been observed in C. fortunei in Japan. Of 732 progeny examined (250 gametophytes and 482 sporophytes), obtained from a parental sporophyte whose pgiC genotype was estimated as aab, 11 (4.4%) gametophytes and 8 (1.7%) sporophytes showed a different genotype ( aaa) from that of the parent sporophyte. We showed that genetic segregation occurs in apogamous C. fortunei in relatively high frequency. Moreover, we could first show that the segregation frequency in gametophytes is significantly higher than that in sporophytes of the next generation ( χ = 4.90, P = 0.027). It may suggest the existence of deleterious genes, which are expressed during the morphogenesis and growth of sporophytes. [ABSTRACT FROM AUTHOR]