Improved germination of somatic embryos and plant recovery of European chestnut

For the mass production of chestnut trees with selected, hybrid, or genetically engineered genotypes, one potentially desirable propagation strategy is based on somatic embryogenesis. Although methods exist for the initiation of embryogenic cultures of Castanea sativa from immature zygotic embryos or leaf explants, the embryos produced have had low rates of conversion into plantlets. This study explored the possible benefits for somatic embryos that have already undergone maturation and cold treatments, of (a) partial slow or fast desiccation, and (b) of the addition of plant growth regulators or glutamine to the germination medium. Germination response was evaluated in terms of both conversions to plantlets and through embryos developing only shoots (shoot germination) that could be rooted following the micropropagation protocols developed for chestnut. Two or 3 wk slow desiccation in sealed empty Petri dishes resulted in a slight reduction in water content that nevertheless increased total potential plant recovery, shoot length, and the number of leaves per plantlet. However, best results were achieved by 2 h fast drying in a laminar flow hood, which reduced embryo moisture content to 57–58% and enhanced the potential plant recovery and quality of regenerated plantlets. Plant yield was also promoted by addition of 0.44 μM benzyladenine and 200–438 mg/l of glutamine to the germination medium, and plantlet quality (as evidenced by root, shoot, and leaf growth) by the further addition of 0.49 μM indole-3-butyric acid.     

Drought can be more critical in the shade than in the sun: a field study of carbon gain and photo-inhibition in a Californian shrub during a dry El Niño year

Diurnal courses of leaf water potential (Ψ_l), gas exchange and chlorophyll fluorescence were measured in natural sun and shade populations of Heteromeles arbutifolia throughout the seasons of an unusually dry El Niño year in Central California. The onset of drought resulted in decreased stomatal conductance and net photosynthesis in both sun and shade plants. However, the decline in Ψ_l was much greater and carbon gain was much more strongly limited by the development of drought stress in the shade than in the sun. Photorespiratory energy dissipation was significantly higher in the sun than in the shade in spring and autumn, but not during the summer. Pre‐dawn photochemical efficiency (F_v/F_m) was significantly higher in the shade than in the sun during the spring but the differences disappeared during the summer and autumn. The strong irradiance in the open field site studied led to a chronic but only mild reduction in F_v/F_m, with values around 0·79. Summer sunflecks led to a sustained photo‐inhibition in shade plants, which exhibited a significant reduction in pre‐dawn F_v/F_m of 10% with the onset of drought. Photo‐inhibition became relatively more important for carbon gain in the shade than in the sun due to the low photochemical efficiency under the low light that follows sunflecks. Sun plants of H. arbutifolia exhibited a rather efficient photoprotection against strong irradiance conferred by both the architecture of the crown and the physiology of the leaves. There is evidence that El Niño events and the associated droughts have become more frequent and severe. Counter‐intuitively, the effects on plant performance of such extreme droughts could be more critical in the shade than in the sun.     

Isolation and Genotyping of Acanthamoeba Strains from Soil Sources from Jamaica,West Indies

Acanthamoeba spp. are opportunistic pathogens that are ubiquitous in nature. Many species of this genus are responsible for a fatal encephalitis and keratitis in humans and other animals. Seventy‐two soil samples were collected from the parishes across Jamaica and assessed for the presence of Acanthamoeba spp. Cultivation was carried out on non‐nutrient agar plates seeded with heat killed Escherichia coli. PCR and sequencing of the DF3 region were carried out in order to genotype the isolated strains of Acanthamoeba. Thermotolerance and osmotolerance assays were utilized to investigate the pathogenic potential of the Acanthamoeba isolates. Acanthamoeba spp. was isolated from 63.9% of soil samples. Sequencing of the DF3 region of the 18S rDNA resulted in the identification of genotypes T4, T5, and T11. T4 genotype was most frequently isolated. Most isolates were thermotolerant or both thermotolerant and osmotolerant, indicating that they may present the potential to cause disease in humans and other animals.     

Additive effects of a potentially invasive grass and water stress on the performance of seedlings of gypsum specialists

Question: What is the combined effect of two drivers of local biodiversity changes (presence of a potentially invasive species and seasonal drought) on the performance of seedlings of plants from gypsum habitats under experimental conditions? Location: A controlled microcosm reconstruction of natural assemblages of gypsum plant communities from central Spain. Methods: We evaluated the effects of a potentially invasive grass (Lolium rigidum) and water stress on the survival, height growth and biomass of five woody species (Colutea hispanica, Gypsophila struthium, Thymus lacaitae, Lepidium subulatum and Helianthemum squamatum) from semi‐arid gypsum ecosystems. Seedlings of the five species were grown with or without the potential invader and under three watering regimes: early stress — simulating an advanced summer, late stress — simulating the characteristic timing of current summer drought and well‐watered. Results: Seedling survival and performance were negatively affected by the presence of the potential invader. Early stress had larger impacts on the gypsum species than late stress. No interactions were found between factors for any of the study variables, and responses to both factors were found to be species‐specific. Conclusions: The lack of interactions between factors indicates that the presence of the potentially invasive grass and water stress had additive effects in our study system. The negative impact of early water stress draws attention to the possible consequences of the advances of summer drought predicted for Mediterranean ecosystems. Finally, the differential responses found for the study species suggest that plant communities will not respond as a unit to global change, leading to significant changes in species composition and dominance.     

The functional ecology of shoot architecture in sun and shade plants of Heteromeles arbutifolia M. Roem., a Californian chaparral shrub

The functional roles of the contrasting morphologies of sun and shade shoots of the evergreen shrub Heteromeles arbutifolia were investigated in chaparral and understory habitats by applying a three-dimensional plant architecture simulation model, YPLANT. The simulations were shown to accurately predict the measured frequency distribution of photosynthetic photon flux density (PFD) on both the leaves and a horizontal surface in the open, and gave reasonably good agreement for the more complex light environment in the shade. The sun shoot architecture was orthotropic and characterized by steeply inclined (mean = 71^o) leaves in a spiral phyllotaxy with short internodes. This architecture resulted in relatively low light absorption efficiencies (E _A) for both diffuse and direct PFD, especially during the summer when solar elevation angles were high. Shade shoots were more plagiotropic with longer internodes and a pseudo-distichous phyllotaxis caused by bending of the petioles that positioned the leaves in a nearly horizontal plane (mean = 5^o). This shade-shoot architecture resulted in higher E _A values for both direct and diffuse PFD as compared to those of the sun shoots. Differences in E _A between sun and shade shoots and between summer and winter were related to differences in projection efficiencies as determined by leaf and solar angles, and by differences in self shading resulting from leaf overlap. The leaves exhibited photosynthetic acclimation to the sun and the shade, with the sun leaves having higher photosynthetic capacities per unit area, higher leaf mass per unit area and lower respiration rates per unit area than shade leaves. Despite having 7 times greater available PFD, sun shoots absorbed only 3 times more and had daily carbon gains only double of those of shade shoots. Simulations showed that sun and shade plants performed similarly in the open light environment, but that shade shoots substantially outperformed sun shoots in the shade light environment. The shoot architecture observed in sun plants appears to achieve an efficient compromise between maximizing carbon gain while minimizing the time that the leaf surfaces are exposed to PFDs in excess of those required for light saturation of photosynthesis and therefore potentially photoinhibitory. Received: 8 June 1997 / Accepted: 2 November 1997     

Effect of luteinizing hormone on RNA synthesis in Leydig cells of immature rats

Luteotrophic hormone acts on testicular interstitial cells, promoting the activation of several cellular events that culminate in steroids synthesis. Since the interstitial tissue include several cell types, purified Leydig cells were used in this work. Isolated interstitial cells from immature rats were purified through a 0-40% metrizamide gradient. Either LH, HCG or Bt2-cAMP significantly stimulated the incorporation of [3H]uridine into RNA, when compared to control. The effect of HCG on RNA synthesis was developed within 30 min after the addition of the hormone and was dose-dependent. The maximum effect was attained with 10 mIU/ml of HCG. These results indicate that HCG/LH or Bt2-cAMP but not FSH, promote an acute stimulation of RNA synthesis by Leydig cells from immature rats.     

Cardiovascular variability after arousal from sleep: time-varying spectral analysis.

We performed time-varying spectral analyses of heart rate variability (HRV) and blood pressure variability (BPV) recorded from 16 normal humans during acoustically induced arousals from sleep. Time-varying autoregressive modeling was employed to estimate the time courses of high-frequency HRV power, low-frequency HRV power, the ratio between low-frequency and high-frequency HRV power, and low-frequency power of systolic BPV. To delineate the influence of respiration on HRV, we also computed respiratory airflow high-frequency power, the modified ratio of low-frequency to high-frequency HRV power, and the average transfer gain between respiration and heart rate. During cortical arousal, muscle sympathetic nerve activity and heart rate increased and returned rapidly to baseline, but systolic blood pressure, the ratio between low-frequency and high-frequency HRV power, low-frequency HRV power, the modified ratio of low-frequency to high-frequency HRV power, and low-frequency power of systolic BPV displayed increases that remained above baseline up to 40 s after arousal. High-frequency HRV power and airflow high-frequency power showed concommitant decreases to levels below baseline, whereas the average transfer gain between respiration and heart rate remained unchanged. These findings suggest that 1) arousal-induced changes in parasympathetic activity are strongly coupled to respiratory pattern and 2) the sympathoexcitatory cardiovascular effects of arousal are relatively long lasting and may accumulate if repetitive arousals occur in close succession.     

Catabolic pathway of arginine in Anabaena involves a novel bifunctional enzyme that produces proline from arginine

Arginine participates widely in metabolic processes. The heterocyst‐forming cyanobacterium Anabaena catabolizes arginine to produce proline and glutamate, with concomitant release of ammonium, as major products. Analysis of mutant Anabaena strains showed that this catabolic pathway is the product of two genes, agrE (alr4995) and putA (alr0540). The predicted PutA protein is a conventional, bifunctional proline oxidase that produces glutamate from proline. In contrast, AgrE is a hitherto unrecognized enzyme that contains both an N‐terminal α/β propeller domain and a unique C‐terminal domain of previously unidentified function. In vitro analysis of the proteins expressed in Escherichia coli or Anabaena showed arginine dihydrolase activity of the N‐terminal domain and ornithine cyclodeaminase activity of the C‐terminal domain, overall producing proline from arginine. In the diazotrophic filaments of Anabaena, β‐aspartyl‐arginine dipeptide is transferred from the heterocysts to the vegetative cells, where it is cleaved producing aspartate and arginine. Both agrE and putA were found to be expressed at higher levels in vegetative cells than in heterocysts, implying that arginine is catabolized by the AgrE‐PutA pathway mainly in the vegetative cells. Expression in Anabaena of a homolog of the C‐terminal domain of AgrE obtained from Methanococcus maripaludis enabled us to identify an archaeal ornithine cyclodeaminase.     

Production of antigen-specific human monoclonal antibodies: comparison of mice carrying IgH/kappa or IgH/kappa/lambda transloci

Here we compare human monoclonal antibody (MAb) production from mouse strains that carry disruptions of their endogenous mouse IgH/IgK loci and harbor human IgM + Igkappa(BABkappa) or human IgM + Igkappa + IgA transloci (BABkappa,lambda). We found that whereas both strains proved effective for the isolation of antigen-specific IgM antibodies, many of the IgM MAbs elicited from BABkappa comprise human mu chains that are associated with mouse lambda chains. In contrast, BABkappa,lambda mice gave rise to fully functional, polymeric human IgM antibodies comprising both human IgH and human IgL chains. Therefore, the inclusion of a human Iglambda translocus (in addition to the human IgH + Igkappa transloci) not only diminishes problems of endogenous mouse Iglambda expression but also provides a strain of mice that yields fully human MAbs to a wide range of antigens, as witnessed by the isolation of MAbs to human blood cells, tumor cell lines, and an immunoglobulin idiotype.     

Expression of the QrCPE gene is associated with the induction and development of oak somatic embryos

Somatic embryogenesis is a powerful tool for plant regeneration and also provides a suitable material for investigating the molecular events that control the induction and development of somatic embryos. This study focuses on expression analysis of the QrCPE gene (which encodes a glycine-rich protein) during the initiation of oak somatic embryos from leaf explants and also during the histodifferentiation of somatic embryos. Northern blot and in situ hybridization were used to determine the specific localisation of QrCPE mRNA. The results showed that the QrCPE gene is developmentally regulated during the histodifferentiation of somatic embryos and that its expression is tissue- and genotype-dependent. QrCPE was strongly expressed in embryogenic cell aggregates and in embryogenic nodular structures originated in leaf explants as well as in the protodermis of somatic embryos from which new embryos are generated by secondary embryogenesis. This suggests a role for the gene during the induction of somatic embryos and in the maintenance of embryogenic competence. The QrCPE gene was highly expressed in actively dividing cells during embryo development, suggesting that it participates in embryo histodifferentiation. The localised expression in the root cap initial cells of cotyledonary somatic embryos and in the root cap of somatic seedlings also suggests that the gene may be involved in the fate of root cap cells.