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The Immersed Boundary (IB) method has been widely used to solve fluid-structure interaction problems, including those where the structure interacts with polymeric fluids. In this paper, we examine the convergence of one such scheme for a well known two-dimensional benchmark flow for the Oldroyd-B constitutive model, and we show that the traditional IB-based scheme fails to adequately capture the polymeric stress near to embedded boundaries. We analyze the reason for such failure, and we argue that this feature is not specific to the case study chosen, but a general feature of such methods due to lack of convergence in velocity gradients near interfaces. In order to remedy this problem, we build a different scheme for the Oldroyd-B system using the Immersed Boundary Smooth Extension (IBSE) scheme, which provides convergent viscous stresses near boundaries. We show that this modified scheme produces convergent polymeric stresses through the whole domain, including on embedded boundaries, and produces solutions in good agreement with known benchmarks.

© 2015 Elsevier Inc. The Immersed Boundary method is a simple, efficient, and robust numerical scheme for solving PDE in general domains, yet it only achieves first-order spatial accuracy near embedded boundaries. In this paper, we introduce a new high-order numerical method which we call the Immersed Boundary Smooth Extension (IBSE) method. The IBSE method achieves high-order accuracy by smoothly extending the unknown solution of the PDE from a given smooth domain to a larger computational domain, enabling the use of simple Cartesian-grid discretizations (e.g. Fourier spectral methods). The method preserves much of the flexibility and robustness of the original IB method. In particular, it requires minimal geometric information to describe the boundary and relies only on convolution with regularized delta-functions to communicate information between the computational grid and the boundary. We present a fast algorithm for solving elliptic equations, which forms the basis for simple, high-order implicit-time methods for parabolic PDE and implicit-explicit methods for related nonlinear PDE. We apply the IBSE method to solve the Poisson, heat, Burgers', and Fitzhugh-Nagumo equations, and demonstrate fourth-order pointwise convergence for Dirichlet problems and third-order pointwise convergence for Neumann problems.

Mathematical models are indispensable for disentangling the interactions through which biological components work together to generate the forces and flows that position, mix, and distribute proteins, nutrients, and organelles within the cell. To illuminate the ever more specific questions studied at the edge of biological inquiry, such models inevitably become more complex. Solving, simulating, and learning from these more realistic models requires the development of new analytic techniques, numerical methods, and scalable software. In this review, we discuss some recent developments in tools for understanding how large numbers of cytoskeletal filaments, driven by molecular motors and interacting with the cytoplasm and other structures in their environment, generate fluid flows, instabilities, and material deformations which help drive crucial cellular processes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Life in complex systems, such as cities and organisms, comes to a standstill when global coordination of mass, energy, and information flows is disrupted. Global coordination is no less important in single cells, especially in large oocytes and newly formed embryos, which commonly use fast fluid flows for dynamic reorganization of their cytoplasm. Here, we combine theory, computing, and imaging to investigate such flows in the Drosophila oocyte, where streaming has been proposed to spontaneously arise from hydrodynamic interactions among cortically anchored microtubules loaded with cargo-carrying molecular motors. We use a fast, accurate, and scalable numerical approach to investigate fluid-structure interactions of 1000s of flexible fibers and demonstrate the robust emergence and evolution of cell-spanning vortices, or twisters. Dominated by a rigid body rotation and secondary toroidal components, these flows are likely involved in rapid mixing and transport of ooplasmic components.

Ensembles of particles rotating in a two-dimensional fluid can exhibit chaotic dynamics yet develop signatures of hidden order. Such rotors are found in the natural world spanning vastly disparate length scales - from the rotor proteins in cellular membranes to models of atmospheric dynamics. Here we show that an initially random distribution of either driven rotors in a viscous membrane, or ideal vortices with minute perturbations, spontaneously self assemble into a distinct arrangement. Despite arising from drastically different physics, these systems share a Hamiltonian structure that sets geometrical conservation laws resulting in prominent structural states. We find that the rotationally invariant interactions isotropically suppress long-wavelength fluctuations - a hallmark of a disordered hyperuniform material. With increasing area fraction, the system orders into a hexagonal lattice. In mixtures of two co-rotating populations, the stronger population will gain order from the other and both will become phase enriched. Finally, we show that classical 2D point vortex systems arise as exact limits of the experimentally accessible microscopic membrane rotors, yielding a new system through which to study topological defects., (© 2022. The Author(s).)

This study describes a novel series of UDP- N -acetylglucosamine acyltransferase (LpxA) inhibitors that was identified through affinity-mediated selection from a DNA-encoded compound library. The original hit was a selective inhibitor of Pseudomonas aeruginosa LpxA with no activity against Escherichia coli LpxA. The biochemical potency of the series was optimized through an X-ray crystallography-supported medicinal chemistry program, resulting in compounds with nanomolar activity against P. aeruginosa LpxA (best half-maximal inhibitory concentration (IC 50 ) <5 nM) and cellular activity against P. aeruginosa (best minimal inhibitory concentration (MIC) of 4 μg/mL). Lack of activity against E. coli was maintained (IC 50 > 20 μM and MIC > 128 μg/mL). The mode of action of analogues was confirmed through genetic analyses. As expected, compounds were active against multidrug-resistant isolates. Further optimization of pharmacokinetics is needed before efficacy studies in mouse infection models can be attempted. To our knowledge, this is the first reported LpxA inhibitor series with selective activity against P. aeruginosa .

In the cellular phenomena of cytoplasmic streaming, molecular motors carrying cargo along a network of microtubules entrain the surrounding fluid. The piconewton forces produced by individual motors are sufficient to deform long microtubules, as are the collective fluid flows generated by many moving motors. Studies of streaming during oocyte development in the fruit fly Drosophila melanogaster have shown a transition from a spatially disordered cytoskeleton, supporting flows with only short-ranged correlations, to an ordered state with a cell-spanning vortical flow. To test the hypothesis that this transition is driven by fluid-structure interactions, we study a discrete-filament model and a coarse-grained continuum theory for motors moving on a deformable cytoskeleton, both of which are shown to exhibit a swirling instability to spontaneous large-scale rotational motion, as observed.

The spindle shows remarkable diversity, and changes in an integrated fashion, as cells vary over evolution. Here, we provide a mechanistic explanation for variations in the first mitotic spindle in nematodes. We used a combination of quantitative genetics and biophysics to rule out broad classes of models of the regulation of spindle length and dynamics, and to establish the importance of a balance of cortical pulling forces acting in different directions. These experiments led us to construct a model of cortical pulling forces in which the stoichiometric interactions of microtubules and force generators (each force generator can bind only one microtubule), is key to explaining the dynamics of spindle positioning and elongation, and spindle final length and scaling with cell size. This model accounts for variations in all the spindle traits we studied here, both within species and across nematode species spanning over 100 million years of evolution., Competing Interests: RF, CY, GF, HW, DS, MR, TM, MS, DN No competing interests declared, (© 2020, Farhadifar et al.)

As integrated care is recognized as crucial to meet the challenges of chronic conditions such as Parkinson's disease (PD), integrated care networks have emerged internationally and throughout Germany. One of these networks is the Parkinson Network Eastern Saxony (PANOS). PANOS aims to deliver timely and equal care to PD patients with a collaborative intersectoral structured care pathway. Additional components encompass personalized case management, an electronic health record, and communicative and educative measures. To reach an intersectoral consensus of the future collaboration in PANOS, a structured consensus process was conducted in three sequential workshops. Community-based physicians, PD specialists, therapists, scientists and representatives of regulatory authorities and statutory health insurances were asked to rate core pathway-elements and supporting technological, personal and communicative measures. For the majority of core elements/planned measures, a consensus was reached, defined as an agreement by >75% of participants. Additionally, six representatives from all partners involved in the network-design independently assessed PANOS based on the Development Model for Integrated Care (DMIC), a validated model addressing the comprehensiveness and maturity of integrated care concepts. The results show that PANOS is currently in an early maturation state but has the potential to comprehensively represent the DMIC if all planned activities are implemented successfully. Despite the favorable high level of consensus regarding the PANOS concept and despite its potential to become a balanced integrated care concept according to the DMIC, its full implementation remains a considerable challenge.

We show that rotating membrane inclusions can crystallize due to combined hydrodynamic and steric interactions. Alone, steric repulsion of unconfined particles, even with thermal fluctuations, does not lead to crystallization, nor do rotational hydrodynamic interactions which allow only a marginally stable lattice. Hydrodynamic interactions enable particles to explore states inaccessible to a nonrotational system, yet, unlike Brownian motion, Hamiltonian conservation confines the ensemble which, when combined with steric interactions, anneals into a stable crystal state.

The medicinal chemistry community has directed considerable efforts toward the discovery of selective inhibitors of interleukin-2 inducible T-cell kinase (ITK), given its role in T-cell signaling downstream of the T-cell receptor (TCR) and the implications of this target for inflammatory disorders such as asthma. We have previously disclosed a structure- and property-guided lead optimization effort which resulted in the discovery of a new series of tetrahydroindazole-containing selective ITK inhibitors. Herein we disclose further optimization of this series that resulted in further potency improvements, reduced off-target receptor binding liabilities, and reduced cytotoxicity. Specifically, we have identified a correlation between the basicity of solubilizing elements in the ITK inhibitors and off-target antiproliferative effects, which was exploited to reduce cytotoxicity while maintaining kinase selectivity. Optimized analogues were shown to reduce IL-2 and IL-13 production in vivo following oral or intraperitoneal dosing in mice.

Interleukin-2 inducible T-cell kinase (ITK), a member of the Tec family of tyrosine kinases, plays a major role in T-cell signaling downstream of the T-cell receptor (TCR), and considerable efforts have been directed toward discovery of ITK-selective inhibitors as potential treatments of inflammatory disorders such as asthma. Using a previously disclosed indazole series of inhibitors as a starting point, and using X-ray crystallography and solubility forecast index (SFI) as guides, we evolved a series of tetrahydroindazole inhibitors with improved potency, selectivity, and pharmaceutical properties. Highlights include identification of a selectivity pocket above the ligand plane, and identification of appropriate lipophilic substituents to occupy this space. This effort culminated in identification of a potent and selective ITK inhibitor (GNE-9822) with good ADME properties in preclinical species.

Inhibition of the non-receptor tyrosine kinase ITK, a component of the T-cell receptor signalling cascade, may represent a novel treatment for allergic asthma. Here we report the structure-based optimization of a series of benzothiazole amides that demonstrate sub-nanomolar inhibitory potency against ITK with good cellular activity and kinase selectivity. We also elucidate the binding mode of these inhibitors by solving the X-ray crystal structures of several inhibitor-ITK complexes., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Premise of the Study: Discovering missing ancestors is essential to understanding the evolutionary history of biodiversity on Earth. Evidence from extinct species can provide links for reconstructing intricate patterns of reticulate relationships among extant descendents. When fossils are unavailable and other evidence yields competing hypotheses to explain species ancestry, data from proteins and DNA can help resolve conflicts and generate novel perspectives. The identity of a parent shared by two tetraploid species in the cosmopolitan fern genus Dryopteris has remained elusive for more than 50 years. Based on available data, four hypotheses were developed previously, each providing a different resolution to this uncertainty. •, Methods: New molecular evidence from studies of isozymes and restriction site analysis of chloroplast DNA tested the competing hypotheses about the diploid ancestors of these two extant Dryopteris polyploids. •, Key Results: The results falsify two of the hypotheses, resolve the uncertainty in the third, and support the fourth. •, Conclusions: Our data validate the prior existence of Dryopteris "semicristata," which was proposed 38 years ago as a diploid progenitor of the allotetraploids D. cristata and D. carthusiana but has never been collected. After developing a phylogeny using the new molecular data, we describe a plausible morphology for D. "semicristata" by extrapolating likely character states from related extant species.

Assembly of bioactive natural compounds through the action of nonribosomal peptide synthetases (NRPSs) relies on the specific interplay of modules and domains along these multiple mega-enzymes. As the C termini of several bacterial NRPSs often harbor epimerization (E) domains that generate D-amino acids, these seem to facilitate the ordered intermolecular enzymatic interaction and the directed transfer of intermediates. To elucidate this bifunctional role, E domains in recombinant bimodular proteins derived from the tyrocidine synthetase B were investigated. By utilizing sequent tryptic proteolysis and HPLC Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), we could directly interrogate and determine the formation of intermediates attached to the TycB(3)-PCP domain of wild-type TycB(2-3) and to the E domain exchange enzyme TycB(2-3)-ATCAT/E(tycA). In addition, the two proteins and a version of TycB(2-3) fused to the communication-mediating (COM) domain of TycA were applied in product formation assays with TycB(1) to corroborate E domain impact on intermodular NRPS interaction. Significant functional differences between the C-terminal aminoacyl- and peptidyl-E domains were observed in terms of in trans interaction and misinitiation. E domains originating from elongation modules (peptidyl-E domains) seem to be optimized for regulation of the progression of peptide bond formation, epimerization, and intermediate transfer to the downstream module, whereas E domains of initiation modules (aminoacyl-E domains) impair upstream condensation and cause misinitiation. The selection of E domains is therefore decisive for successful application in biocombinatorial engineering of nonribosomal peptides.

Many pharmacologically important agents are assembled on multimodular nonribosomal peptide synthetases (NRPSs) whose modules comprise a set of core domains with all essential catalytic functions necessary for the incorporation and modification of one building block. Very often, d-amino acids are found in such products which, with few exceptions, are generated by the action of NRPS integrated epimerization (E) domains that alter the stereochemistry of the corresponding peptidyl carrier protein (PCP) bound l-intermediate. In this study we present a quantitative investigation of substrate specificity of four different E domains (two 'peptidyl-' and two 'aminoacyl-'E domains) derived from different NRPSs towards PCP bound peptides. The respective PCP-E bidomain apo-proteins (TycB(3)-, FenD(2)-, TycA- and GrsA-PCP-E) were primed with various peptidyl-CoA precursors by utilizing the promiscuous phosphopantetheinyl transferase Sfp. PCP bound peptidyl-S-Ppant epimerization products were chemically cleaved and analyzed for their l/d-ratios by LCMS. We were able to show that all four E domains tolerate a broad variety of peptidyl-S-Ppant-substrates as evaluated by k(obs) values and final l/d-product equilibria determined for each reaction. The two C-terminal amino acids of the substrate seem to be recognized by 'peptidyl-'E domains. Interestingly, the 'aminoacyl-'E domains GrsA- and TycA-E were also able to convert the elongated intermediates. All four E domains accepted an N-methylated precursor as well and epimerized this substrate with high efficiency. Finally, we could demonstrate that the condensation (C) domain of TycB(1) is also able to process peptidyl substrates transferred by TycA. In conclusion, these findings are of great impact on future engineering attempts.

We present a systematic and quantitative study of the protein-protein recognition between the three tyrocidine synthetases TycA, TycB, and TycC investigated with two artificial in trans assay systems, which had been previously developed: the "DKP assay system" for the interaction of TycA with TycB and the "L/D-Phe-L-Asn assay system" for the interaction of TycB with TycC. TycA-A(Phe)TE and TycB(3)-A(Phe)TE, which are used as donor enzymes, both provide D-Phe-S-Ppant, so that no substrate specificities interfered with the quantification of protein-protein recognition. We tested all donor/acceptor enzyme combinations between the two artificial assay systems for product formation activities as well as two hybrid enzymes, where the E-domains of TycA and TycB(3) had been exchanged against each other. Furthermore, four donor/acceptor protein fusions were constructed on gene level, resulting in dimodular proteins. We were able to show that the E-domains mediate protein-protein recognition in trans. Product formation of the different donor assayed with the two acceptor enzymes TycB(1)-CA(Pro)T and TycC(1)-CA(Asn)T/Te in trans was only obtained if the donor enzyme harbored the cognate E-domain. Interestingly, all in cis fusions (dimodular proteins) were active, giving strong evidence that unnatural protein-protein interactions can be "forced" by fusion of the distinct enzymes. Finally, we were able to detect product formation in the "DKP system" with engineered hybrid proteins where the A-domain of TycA had been exchanged against the isoleucine-activating A-domain of BacA(1) and the valine-activating A-domain of TycC(4), respectively. All of these findings are of high relevance for future NRPS engineering approaches.

This article presents eight commentaries on a case study published in the journal Pediatrics in 2001. The case study, introduced in Pediatrics to highlight the issue of adolescents' compliance with drug treatment in a "high-prevalence neurobehavioral condition," briefly describes an adolescent boy who announces that he no longer needs the methylphenidate he was prescribed for ADHD and had been taking for the last five years. We invited experienced professionals from the disciplines of psychiatry, psychology, counseling, education, and occupational therapy to comment on the case and make recommendations to the adolescent boy in the case study, his family, and professionals. Their commentaries highlight issues rarely discussed in the mainstream literature, including: the extent to which ADHD is erroneously portrayed and vigorously managed as a disease; the lack of validity of the ADHD construct in adolescence; the widespread use of stimulants as performance-enhancing drugs; the need to respect an adolescent's gut instinct and developing decisions; the importance of family dynamics in ADHD-like situations; the need to ease stimulant withdrawal effects; and the human rights of children prescribed psychotropic drugs.

The chloroplast genome of most land plants is highly conserved. In contrast, physical and gene mapping studies have revealed a highly rearranged chloroplast genome in species representing four families of ferns. In all four, there has been a rare duplication of the psbA gene and the order of the psbA, 16S, and 23S rRNA genes has been inverted. Our analysis shows that the described rearrangement results from a minimum of two inversions within the inverted repeat. This chloroplast DNA structure provides unambiguous evidence that phylogenetically links families of ferns once thought to belong to different major evolutionary lineages.

This study compared the REST (Real Economy System for Teens) program with traditional talk therapy in the treatment of oppositional defiant adolescents. The REST program uses rules for five target behaviors: room care, personal hygiene, completion of chores, abusiveness, and safety violations. Objective measures and subjective parental ratings were taken. Twenty-five adolescents were assigned to each therapy group. In the REST program the adolescent was provided with only food and shelter. The adolescent had to earn money through the REST allowance program to pay for everything else. Earning the allowance was made contingent upon compliance with all rules for the five target behaviors. The results showed significantly greater improvement on all target behaviors for the REST program than for traditional therapy. In addition, parents of adolescents in the REST group reported that their adolescents seemed happier, more relaxed, and closer to them.

Thermoplasma acidophilum, a thermophilic mycoplasma, has several unusual features suggesting a possible relationship to eukaryotic cells. One feature is a histone-like protein that is associated with the DNA, condensing it into subunits similar to those in eukaryotic chromatin. A second feature is an association of cytoplasmic proteins that resembles eukaryotic actin and myosin. These two components are widely distributed in different groups of eukaryotic cells, but are typically lacking in prokaryotic cells. Furthermore, T. acidophilum lacks cytochromes and respires by enzymes that apparently are not coupled to oxidative phosphorylation. This primitive type of respiration resembles that of microbodies, another feature which is represented in the cytoplasm of all groups of eukaryotic cells. Furthermore, since T. acidophilum lacks a cell wall and appears to have a primitive correlate of endocytosis, it would appear to be mechanically capable of acquiring a symbiotic mitochondrion. Thus, our observations are consistent with the symbiotic hypothesis for the origin of eukaryotic cells. We suggest that an organism similar to T. acidophilum was the host cell for the original symbiosis, becoming the nucleus and cytoplasm of modern eukaryotic cells.

Chloroplast DNA from the fern Osmunda einnamomea was isolated by a sucrose gradient procedure utilizing PEG to stabilize chloroplasts. Analysis with the restriction endonucleases PvuII, Sacl and BstEII indicates a chloroplast genome size of 144 kb. A physical map of the fragments produced by these three enzymes was constructed by filter hybridizations using purified PvuII fragments as hybridization probes. The Osmunda chloroplast genome is circular and contains an inverted repeat 8-13 kb in size.Gene probes from tobacco, corn and spinach were used to map the positions of six genes on the Osmunda chloroplast chromosome. The 16S and 23S ribosomal RNAs are encoded by duplicate genes which lie within the inverted repeat. Genes for the large subunit of ribulose-1,5-bisphosphate carboxylase, a photosystem II polypeptide, and the alpha and beta subunits of chloroplast coupling factor are located in three different segments of the large single copy region.The Osmunda chloroplast genome is remarkably similar in size, conformation, physical organization, and map positions of known genes, to chloroplast DNA from a number of angiosperms. The major difference between chloroplast DNA from this fern and angiosperms is that the inverted repeat is smaller in Osmunda (8-13 kb) than in angiosperms (22-25 kb).

The freeliving thermophilic mycoplasma, Thermoplasma acidophilum, has a small acid-soluble protein tightly bound to its DNA. This protein is similar to eukaryotic histones in both size and amino acid composition. Here we report that the protein condenses DNA into globular particles that are about half the size of eukaryotic nucleosomes. Our conclusions are based primarily upon the following observations: (1) Nuclease digestion produced DNA fragments of 40 base-pairs. (2) The ratio of protein to DNA was such that 4--5 molecules of protein were associated with each 40 base-pair segment of DNA. (3) Protein crosslinking reagents produced tetramers of the histone-like protein. (4) Electron microscopy revealed globular particles 5--6 nm in diameter. (5) Each globular particle reduced the apparent contour length of the DNA by 40 base-pairs. Thus, each nucleoprotein particle is apparently composed of 40 base-pairs of DNA coiled around four molecules of proteins.

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.

Histochemical detection of acetylcholinesterase (AChE) activity in Xenopus embryos was found first in primary motoneurons, Rohon-Beard neurons and somitic myotubes at early tail bud stages. At late tail bud stages all primary neurons, including primary interneurons, cranial ganglion cells and ventral brainstem cells expressed this enzyme. The onset of detectable AChE activity in some primary neurons occurred near the time of initial axon outgrowth, whereas in others it occurred at much later stages. At early independent-feeding and continuous-swimming stages nearly all seemingly postmitotic neurons began to express AChE activity, and by the beginning of limb bud stages, when many secondary neuronal populations were going through their final rounds of mitosis, nearly all CNS cells outside the ventricular zone were intensely stained. Thus, the onset of detectable AChE activity in secondary neurons occurred near the time of their final mitoses. In trunk somites the enzyme activity initially was located diffusely throughout the myotube, and with progressing development it became localized to the myocommata. From the onset of AChE activity both head somites and head muscles had discrete patches of reaction product all over their surfaces. The onset of detectable AChE activity occurred in muscles near the time that they were contacted by motor axons. These data demonstrate that the primary neurons are the first to express AChE activity, and that as the secondary neurons begin to proliferate, AChE is expressed by nearly all embryonic neuronal populations.

Indole-3-acetic acid (IAA), at concentrations of 0.01 to 1.0 millimolar, and ethephon (0.3% v/v Ethrel) promote stomatal opening when applied to epidermal peels of Vicia faba L. in light or dark. The effect of ethylene is seen by 30 minutes and maximal opening (over two times that of untreated controls) occurs after only 60 to 90 minutes in the light. Stomatal opening by IAA and Ethrel in both light and dark is prevented by 0.14 millimolar AgCl. It is suggested that the effect of added IAA, but not that of light, is linked to ethylene production. The possible role of ethylene in stomatal opening during fungal infection is discussed. The stomates of Vicia faba provide a new system to study the effects of ethylene on certain membrane-regulated processes.

The thermophilic mycoplasma Thermoplasma acidophilum has tightly bound to its DNA a protein that closely resembles the histones of eukaryotes. DNA associated with this protein is more stable than free DNA against thermal denaturation by about 40 degrees C, as shown in both native nucleoprotein and in hybrid nucleoprotein reconstituted in vitro with calf DNA. Since only about 20 percent of the DNA in this organism is associated with the histone-like protein, we suggest that its physiological function is to prevent complete separation of the DNA strands during brief exposures of the organism to denaturing conditions, and thus to facilitate rapid renaturation when normal environmental conditions return.

Phylogenetic relationships of Osmunda cinnamomea, O. claytoniana, and O. regalis were explored by means of DNA sequence comparisons. Hydroxyapatite thermal elution profiles of self-reassociated repetitive DNA fragments were very similar, indicating the absence of gross differences in the amount of recent amplification or addition of repetitive DNA in any of these three genomes. Interspecific DNA sequence comparisons showed, in contrast to our earlier interpretation, that repeated DNA sequences of O. claytoniana are nearly equally diverged from those of O. cinnamomea and O. regalis. Differences between repetitive sequences of the three species can be interpreted as reflecting amplification events which occurred subsequent to speciation. The data obtained suggest that the three Osmunda species most likely arose more or less simultaneously from a common ancestor. These findings were verified in experiments with tracer DNA preparations enriched for single copy sequences. On the basis of the hydridization data presented here and of the fossil record, the rate of single copy sequence divergence in the ferns is comparable to that in the primates, although slower than that observed in other animal taxa. From this first evaluation of rates of DNA evolution in plants it would seem that the rates for plants and animals are roughly comparable. The evidence suggests that species divergence is accompanied by further reiteration of preexisting repeat sequences. The rate of addition of repetitive sequences probably is slower in ferns than in angiosperms. This difference might be attributable to the much larger effective generation time in ferns.

Restriction site variation in chloroplast DNA was examined in the neotropical fern genus Phanerophlebia and in selected species of the related Asiatic genus Cyrtomium and the cosmopolitan progenitor of these two, Polystichum. A total of 103 restriction site mutations was identified; these were used to construct phylogenetic networks and trees based on Wagner and Dollo parsimony and Fitch-Margoliash distance algorithms. The analyses provided evidence that Phanerophlebia did not arise from Cyrtomium. Both genera are convergent descendants from different progenitor groups in Polystichum, and Asiatic Cyrtomium is more closely related to temperate New World Polystichum than it is to neotropical Phanerophlebia. Reticulate venation, previously considered an important taxonomic character for infrageneric classification in Phanerophlebia, most likely evolved independently twice within the genus. Diploid maternal progenitors are suggested for two of four tetraploid species analyzed, and species-level distinctions for two closely related taxa of Phanerophlebia are questioned.

Molecular hybridization techniques have been used to estimate the degree of DNA base sequence homology between some members of the fern genus Osmunda. Under conditions permitting extensive reassociation, measurements of the extent of inter-specific reaction and the thermal stability of the hybrid molecules indicated that O. claytoniana L. (interrupted fern) shares more DNA homology with O. cinnamomea L. (cinnamon fern) than it does with O. regalis L. (royal fern). These findings are in conflict with predictions from a recent analysis of living and fossil specimens by numerical techniques. However, they are consistent with the earlier, more traditional, taxonomic assignments.

Treatment of the homosporous fern Ceratopteris thalictroides L. sporophyte with gibberellic acid results in fertile fronds appearing 3 to 5 weeks after treatment. The hetero-blastic leaf sequence is markedly reduced, and the entire life cycle can be completed in 3 months on defined medium. Treatment with gibberellic acid also stimulates the accumulation of starch in mature leaves.