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rbcL gene sequences provide evidence for the evolutionary lineages of leptosporangiate ferns.

Pteriodophytes have a longer evolutionary history than any other vascular land plant and, therefore, have endured greater loss of phylogenetically informative information. This factor has resulted in substantial disagreements in evaluating characters and, thus, controversy in establishing a stable classification. To compare competing classifications, we obtained DNA sequences of a chloroplast gene. The sequence of 1206 nt of the large subunit of the ribulose-bisphosphate carboxylase gene (rbcL) was determined from 58 species, representing almost all families of leptosporangiate ferns. Phlogenetic trees were inferred by the neighbor-joining and the parsimony methods. The two methods produced almost identical phylogenetic trees that provided insights concerning major general evolutionary trends in the leptosporangiate ferns. Interesting findings were as follows: (i) two morphologically distinct heterosporous water ferns, Marsilea and Salvinia, are sister genera; (ii) the tree ferns (Cyatheaceae, Dicksoniaceae, and Metaxyaceae) are monophyletic; and (iii) polypodioids are distantly related to the gleichenioids in spite of the similarity of their exindusiate soral morphology and are close to the higher indusiate ferns. In addition, the affinities of several "problematic genera" were assessed.

Base Sequence↗

The fern Adiantum capillus-veneris lacks stomatal responses to blue light.

We investigated the responses of stomata to light in the fern Adiantum capillus-veneris, a typical species of Leptosporangiopsida. Stomata in the intact leaves of the sporophytes opened in response to red light, but they did not open when blue light was superimposed on the red light. The results were confirmed in the isolated Adiantum epidermis. The red light-induced stomatal response was not affected by the mutation of phy3, a chimeric protein of phytochrome and phototropin in this fern. The lack of a blue light-specific stomatal response was observed in three other fern species of Leptosporangiopsida, i.e. Pteris cretica, Asplenium scolopendrium and Nephrolepis auriculata. Fusicoccin, an activator of the plasma membrane H(+)-ATPase, induced both stomatal opening and H(+) release in the Adiantum epidermis. Adiantum phototropin genes AcPHOT1 and AcPHOT2 were expressed in the fern guard cells. The transformation of an Arabidopsis phot1 phot2 double mutant, which lost blue light-specific stomatal opening, with AcPHOT1 restored the stomatal response to blue light. Taken together, these results suggest that ferns of Leptosporangiopsida lack a blue light-specific stomatal response, although the functional phototropin and plasma membrane H(+)-ATPase are present in this species.

Adiantum↗

Changes in the phyllosphere community of the resurrection fern, Polypodium polypodioides, associated with rainfall and wetting.

A combination of analyses were used to characterize the changes that occur in a bacterial community present in the phyllosphere of the epiphytic resurrection fern, Polypodium polypodioides, as the fern rehydrates from a desiccation-resistant, physiologically inactive state. Enrichment assays showed an increase in the viable count of bacteria using labile organic substrates following rainfall. Isolates obtained from enrichments were predominantly Gram-positive bacteria affiliated with various groups of the Actinobacteria and Firmicutes. In contrast, sequencing of 16S rRNA genes clones obtained from whole community DNA revealed that much of the community was dominated by other taxa, particularly the Alphaproteobacteria. Similar isolates were obtained from both dry and hydrated P. polypodioides fronds, whereas 16S rRNA gene sequencing of community DNA revealed different ribotypes on the dry and wet fern, and an overall reduction in richness following wetting. Wetting also produced changes in phyllosphere extracellular enzyme activity, with an initial burst of activity following rainfall and a subsequent burst approximately 48 h later. These findings suggest that the resurrection fern harbors a complex phyllosphere community, and that rehydration of the fern following rainfall may act as an enrichment culture stimulating certain bacterial populations and changing the overall community structure and activity.

Acid Phosphatase↗

Clinical status, ocular surface changes and tear ferning in patients with cystic fibrosis.

Twenty-three cystic fibrosis patients and 20 controls were examined for ocular surface changes and tear fluid ferning characteristics. The patients were also evaluated systemically and given numerical scores according to Schwachman's scoring system. Frequency of blepharitis, fluorescein staining, and the Schirmer's test values did not differ between patients and controls. Patients with cystic fibrosis had a higher frequency of altered ferning pattern that corresponded to the disease severity as measured by the Schwachman score (r = -0.48, p < 0.05). Although altered ferning pattern was frequent, the rate of type I ferning was also high (30%) in patients with cystic fibrosis. We conclude that the ferning test should not be used as an aid in diagnosis of cystic fibrosis but it may be used as an indicator of clinical status during follow-up.

Adolescent↗

Quantitative tear ferning. Clinical investigations.

The tears of 225 subjects were examined for several disorders of the eye by means of a newly developed method, the quantitative tear ferning test. Of the 225 subjects, 89 were normal controls, 24 were examined before and after cataract extraction, and 112 had some disease of the external part of the eye. Using the quantitative tear ferning test, it was shown that compared to normals, the ferning area was significantly decreased in cases of keratoconjunctivitis sicca, in primary and in secondary Sjögren's syndrome. This was also the case in infectious conjunctivitis and after cataract extraction, possibly due to exudation from the conjunctival vessels. In simple conjunctivitis, however, the ferning area was normal. The ferning method has a sensitivity and a specificity of the same order as the commonly used tests for Sjögren's syndrome (Schirmer I, BUT, rose bengale, lactoferrin).

Adult↗

Relationship of impression cytology and tear ferning to reports of dry eye.

PURPOSE: This study investigated whether there is a significant correlation between mild-to-moderate patient reports of dry eye and results on the tear ferning and impression cytology tests. METHODS: One hundred and four subjects were surveyed for presence or absence of dry eye symptoms. Membrane impression cytology and tear ferning were performed on each subject. Impression cytology specimens were evaluated for the presence of goblet cells, presence of epithelial cell sheets, and epithelial cell morphology. Tear specimens were evaluated for quality of ferning. RESULTS: A significant correlation was found between subjective dry eye symptoms and impression cytology results (cc = 0.28, p = 0.003). However, there was no statistically significant correlation between dry eye symptoms and tear fern results (cc = 0.072, p = 0.458). CONCLUSIONS: Impression cytology appeared to be a better predictor of mild-to-moderate dry eye symptoms than tear ferning.

Adult↗

[Experimental trial to reproduction enzootic haematuria in cattle by feeding bracken fern [Pteris aquilina (Kuhn)] (author's transl)].

Cattle were fed with a diet containing bracken fern in order to study the development of chronic bovine haematuria. The experimental 25 animals were randomly divided into 5 groups and such groups received different amounts of the plant. All groups were fed for 36 months, while no clinical effect of the disease was noted. After that time, the supplement of bracken fern to the diet was suspended and the animals still remained under observation. One of the animals (from group I) exhibit haematuria one month after the fern was removed from the diet. In the same occasion, four animals were slaughtered and only one of them presented haemorragic lesions in the urinary bladder. After 34 days another four animals were slaughtered and 114 days later three more were killed. All animals were chosen randomly and in spite of no clinical signs of the disease, all of them presented haemorragic lesions in the urinary bladder. The results obtained in this experiment confirmed that bracken fern (Pteris aquilina) is responsable for the development of chronic bovine haematuria in cattle. The disease is challenged by continuous ingestion of small amounts of bracken fern during a long period of time.

Animals↗

Efficient induction of apospory and apogamy in vitro in silver fern (Pityrogramma calomelanos L.).

Silver fern (Pityrogramma calomelanos L.) is a terrestrial or lithophytic herbaceous fern used for ornamental and medicinal purposes. In its farina it produces the cytotoxic and anticancer compound dihydrochalcone. In vitro induction of apospory and apogamy, and direct field establishment of aposporous gametophytes and subsequent sporophyte development has been accomplished. Half-strength Murashige and Skoog (MS) medium with 3.33 microM N(6)-benzyladenine (BA) and 2.32 microM kinetin (Kn) showed earlier development and produced higher numbers of aposporous gametophytes than half-strength MS basal medium. Crozier explants developed higher numbers (mean value 29.2) of gametophytes, but were slower than frond explants (mean value 23.2). The gametophytes originated from the epidermal hairs progressed from uniseriate filamentous to cordate through bi-, tri- and multiseriate and spatulate stage with the development of antheridia. Reduction in the nutrient and sucrose concentrations in the media favoured apogamy. Sucrose-free 1/10 strength MS medium and agar plates developed a mean of 30.4 and 29.9 sporophytes, respectively in the light. The greenhouse-established gametophytes developed sporophytes. The established sporophytes ex vitro showed 95% survival rate. Apogamous sporophytes and the source plant showed the same chromosome numbers (2n=116). The established protocol accomplishes apogamy and apospory in silver fern, and the aposporous gametophytes can be used for genetic transformation and development of transgenic silver fern.

Ferns↗

Tree ferns: monophyletic groups and their relationships as revealed by four protein-coding plastid loci.

Tree ferns are a well-established clade within leptosporangiate ferns. Most of the 600 species (in seven families and 13 genera) are arborescent, but considerable morphological variability exists, spanning the giant scaly tree ferns (Cyatheaceae), the low, erect plants (Plagiogyriaceae), and the diminutive endemics of the Guayana Highlands (Hymenophyllopsidaceae). In this study, we investigate phylogenetic relationships within tree ferns based on analyses of four protein-coding, plastid loci (atpA, atpB, rbcL, and rps4). Our results reveal four well-supported clades, with genera of Dicksoniaceae (sensu ) interspersed among them: (A) (Loxomataceae, (Culcita, Plagiogyriaceae)), (B) (Calochlaena, (Dicksonia, Lophosoriaceae)), (C) Cibotium, and (D) Cyatheaceae, with Hymenophyllopsidaceae nested within. How these four groups are related to one other, to Thyrsopteris, or to Metaxyaceae is weakly supported. Our results show that Dicksoniaceae and Cyatheaceae, as currently recognised, are not monophyletic and new circumscriptions for these families are needed.

Base Sequence↗

Reconciling extreme branch length differences: decoupling time and rate through the evolutionary history of filmy ferns.

The rate of molecular evolution is not constant across the Tree of Life. Characterizing rate discrepancies and evaluating the relative roles of time and rate along branches through the past are both critical to a full understanding of evolutionary history. In this study, we explore the interactions of time and rate in filmy ferns (Hymenophyllaceae), a lineage with extreme branch length differences between the two major clades. We test for the presence of significant rate discrepancies within and between these clades, and we separate time and rate across the filmy fern phylogeny to simultaneously yield an evolutionary time scale of filmy fern diversification and reconstructions of ancestral rates of molecular evolution. Our results indicate that the branch length disparity observed between the major lineages of filmy ferns is indeed due to a significant difference in molecular evolutionary rate. The estimation of divergence times reveals that the timing of crown group diversification was not concurrent for the two lineages, and the reconstruction of ancestral rates of molecular evolution points to a substantial rate deceleration in one of the clades. Further analysis suggests that this may be due to a genome-wide deceleration in the rate of nucleotide substitution.

Evolution, Molecular↗

Uptake and distribution of selenium in different fern species.

There has been an interest in using hyperaccumulating plants for the removal of heavy metals and metalloids. High selenium (Se) concentrations in the environment are detrimental to animals, humans, and sustainable agriculture, yet selenium is also an essential nutrient for humans. This experiment was conducted to screen fern plants for their potential to accumulate selenium. Eleven fern species, Pteris vittata, P. quadriaurita, P. dentata, P. ensiformis, P. cretica, Dryopteris erythrosora, Didymochlaena truncatula, Adiantum hispidulum, Actiniopteris radiata, Davallia griffithiana, and Cyrtomium fulcatum, were grown under hydroponic conditions for one week at 20 mg L(-1) selenate or selenite. Root Se concentrations reached 245-731 and 516-1082 mg kg(-1) when treated with selenate and selenite, respectively. The corresponding numbers in the fronds were 153-745 and 74-1,028 mg kg(-1) with no visible toxicity symptoms. Only three fern species were able to accumulate more Se in the fronds than the roots, which were D. griffithiana when treated with selenate, P. vittata when treated with selenite, and A. radiata regardless of the forms of Se. A. radiata was the best species overall for Se accumulation. More research is needed to further determine the potential of the fern species identified in this study for phytoremediation of the Se contaminated soils and water.

Biodegradation, Environmental↗

Genetic map-based analysis of genome structure in the homosporous fern Ceratopteris richardii.

Homosporous ferns have extremely high chromosome numbers relative to flowering plants, but the species with the lowest chromosome numbers show gene expression patterns typical of diploid organisms, suggesting that they may be diploidized ancient polyploids. To investigate the role of polyploidy in fern genome evolution, and to provide permanent genetic resources for this neglected group, we constructed a high-resolution genetic linkage map of the homosporous fern model species, Ceratopteris richardii (n = 39). Linkage map construction employed 488 doubled haploid lines (DHLs) that were genotyped for 368 RFLP, 358 AFLP, and 3 isozyme markers. Forty-one linkage groups were recovered, with average spacing between markers of 3.18 cM. Most loci (approximately 76%) are duplicated and most duplicates occur on different linkage groups, indicating that as in other eukaryotic genomes, gene duplication plays a prominent role in shaping the architecture of fern genomes. Although past polyploidization is a potential mechanism for the observed abundance of gene duplicates, a wide range in the number of gene duplicates as well as the absence of large syntenic regions consisting of duplicated gene copies implies that small-scale duplications may be the primary mode of gene duplication in C. richardii. Alternatively, evidence of past polyploidization(s) may be masked by extensive chromosomal rearrangements as well as smaller-scale duplications and deletions following polyploidization(s).

Chromosome Mapping↗

Tear mucus ferning in patients with Sjögren's syndrome.

The purpose of this study is to evaluate the possibility of using the mucus ferning phenomenon of the tears as a diagnostic test for Keratoconjunctivitis sicca in patients with Sjögren's syndrome. Using a polarising light microscope dried samples were tested for ferning phenomenon collected from (1) 36 healthy controls, (2) 21 patients with primary Sjögren's syndrome, and (3) 15 patients with secondary Sjögren's syndrome. Patients with Sjögren's had xerostomia, abnormal salivary gland biopsy and at least two positive tests for Kerato-conjunctivitis sicca. Patterns of mucus crystallization were classified into 5 types according to uniformity, branch spreading and integrity (types I, II normal, types III, IV, V abnormal). Abnormal ferning was found in tear samples of 8 out of 72 normal control eyes, 38 out of 42 with primary Sjögren's syndrome and 25 out of 30 with secondary Sjögren's syndrome. The differences between both primary and secondary Sjögren's syndrome, in comparison to healthy controls, were significant (p < 0.001). The sensitivity of the method was found 90% for primary Sjögren's syndrome and 80% for secondary Sjögren's syndrome. According to our data, tear mucus ferning test is a simple, sensitive and specific test to estimate Keratoconjunctivitis sicca in patients with Sjögren's syndrome.

Humans↗

A fern spore storage protein is genetically similar to the 1.7 S seed storage protein of Brassica napus.

The ostrich fern, Matteuccia struthiopteris L., contains two globulin spore storage proteins of 2.2 S and 11.3 S, with physical characteristics similar to those of seed storage proteins of Brassica napus (rapeseed) and Raphanus sativus (radish). By the use of a cloned cDNA that encodes the 1.7 S B. napus storage protein (napin), gene sequences that hybridized with napin were detected in fern nuclear DNA, and a 900-nucleotide homologous mRNA was detected in developing spores. In vitro translation of this fern mRNA produced a 22-kD polypeptide comparable in size to the 21-kD precursor polypeptide identified in Brassica. No hybridizations were observed between the Brassica 12 S clone and either fern DNA or developing spore mRNA.

Blotting, Southern↗

Distribution pattern of rare earth elements in fern: implication for intake of fresh silicate particles by plants.

All the lanthanide elements (REE) in fern (Matteuccia) and in soil were determined with inductively coupled plasma mass spectrometer (ICP-MS) in order to discuss REE behavior in fern. The fern sample was divided into three parts: root, stem, and leaf; the soil of the same site was also sampled and the soil sample was divided into two parts: HCl and HNO3 soluble part (soil[HCl]) and HF soluble part (soil[HF]). REE in each part was determined by ICP-MS after solvent extraction separation. The overall variation of the REE pattern of the root does not resemble that of the whole soil, but that of the HF soluble part. A tetrad effect variation (W-type) was found in the REE patterns of root, stem and leaf; more conspicuous tetrad effect variation was observed in HREE region than in LREE region, and was so in stem and leaf than in root. Negative anomalies of Ce were observed in the REE patterns of root, stem and leaf, with bigger anomalies in stem and leaf than in root. The results of this study suggest that REE in fern has come from silicates of soil and has once been in dissolved state.

Mass Spectrometry↗

Forecasting of hydrophilic contact lens tolerance by means of tear ferning test.

BACKGROUND: The ability of the tear ferning test to predict future hydrophilic contact lens tolerance was studied. METHODS: The tear ferning test (TFT) was performed on one randomly chosen eye of each of the 116 subjects who came to our contact lens clinic for hydrophilic contact lens application. The TFT was performed at the time of enrollment (TO) and then 1 month (T1), 3 months (T2) and 6 months (T3) after contact lens fitting. The specificity and the sensitivity of the TFT in identifying future contact lens tolerance was then studied. The statistical significance of the differences in behavior through the study period among the subjects with different pre-fitting TFT results was evaluated by means of survival curves. RESULTS: When only type I ferning was considered as a marker of good tear film conditions, the ability of the TFT to forecast contact lens tolerance had 78.95% sensitivity, 78.35% specificity and 78.45% diagnostic precision. The TFT showed sensitivity of 100%, specificity of 86.6% and diagnostic precision of 97.4% when performed after 1 month of contact lens wearing (T1). Survival curve analysis showed a statistically significant difference in behavior between the group of subjects with pre-fitting ferning type I and the other three groups (P < 0.001). CONCLUSION: The TFT appears to have good sensitivity and specific for prediction of contact lens tolerance in a clinical setting.

Contact Lenses, Hydrophilic↗

Alpha-expansins in the semiaquatic ferns Marsilea quadrifolia and Regnellidium diphyllum: evolutionary aspects and physiological role in rachis elongation.

To investigate the evolutionary history of expansins and their role in cell elongation in early land plants, we isolated two alpha-expansin genes, Mq-EXP1 and Rd-EXP1, respectively, from the semiaquatic ferns Marsilea quacdrifolia L. and Regnellidium diphyllum Lindm. The deduced amino acid sequences of the fern expansins exhibit a high degree of identity to those of seed plants, showing that expansin genes were conserved during the evolution of vascular plants. Gel-blot analysis of M. quadrifolia and R. diphyllum genomic DNA indicated that, in both ferns, alpha-expansins are encoded by multigene families. Expression of alpha-expansin genes probed with Mq-EXP1 was confined to the elongating region of the Marsilea rachis. Cell-wall proteins of M. quadrifolia induced in-vitro extension of acidified cucumber cell walls. In R. diphyllum, expression of Rd-EXP1 increased when elongation of the rachis was enhanced by submergence or ethylene. These results indicate that alpha-expansins act as wall-loosening proteins in ferns, as has been proposed for angiosperms. In addition, Rd-EXP1 may play a role in mediating elongation of the rachis in submerged plants.

Amino Acid Sequence↗

H-ras immunohistochemical expression and molecular analysis of urinary bladder lesions in grazing adult cattle exposed to bracken fern.

Chronic ingestion of bracken fern (Pteridium spp.) by cattle produces upper alimentary tract and urinary bladder tumours causing a syndrome called bovine enzootic haematuria (BEH). Previous studies demonstrated ptaquiloside-DNA adducts and mutations in the h-ras gene in ileal epithelial cells of bracken fern-fed calves. Systematic inspection of the bladder mucosa of grazing cattle (n=126) from bracken-fern areas was carried out in a slaughterhouse. Of the 126 slaughterhouse cattle, 46 showed macroscopical lesions of the bladder. These bladders, together with six others known to have BEH, were examined histopathologically and by H-ras immunohistochemistry. Thirteen affected bladders were also examined by H-ras molecular analysis to detect mutations. Macroscopical and histological study of urinary bladder lesions found at the slaughterhouse revealed chronic cystitis (34.1%) and tumours (2.4%). There was significantly increased immunohistochemical expression of H-ras (P<0.05) in chronic cystitis (H-ras=53.24%) and bladder tumours (H-ras=63.60%) as compared with normal urinary bladders (H-ras=4.32%). A silent mutation (D38D) was detected in one animal with a mixed bladder tumour. The prevalence of urinary bladder lesions (chronic cystitis and tumours) obtained at the slaughterhouse was higher than expected. This study demonstrates that close inspection of urinary bladders of adult grazing cows is necessary to prevent possible human exposure to bracken-fern carcinogens. The absence of mutations in the codons of h-ras studied did not exclude the presence of polymorphisms in other regions of the gene (promoter or regulation sequences) or in other genes (belonging or not to the ras family) that significantly affect the H-ras protein.

Animals↗