Density effects and self-thinning in even-aged pure stands ofEucalyptus camaldulensis Dehn

The competition density effect and changes of mean total tree weight (w) and stand density (ρ) during course of self-thinning were examined in even-aged pure stands ofEucalyptus camaldulensis Dehn. which were planted in the tropical monsoon region. The level of competition was controlled by changing the initial stand density from 625 trees ha⁻¹ to 40,000 trees ha⁻¹. Hozumi's model was used to describe thew-ρ trajectory with aging of each stand and thew-ρ relation between stands of different densities at each time. The higher density produced trees of smaller mean tree sizes. The higher the density, the sooner self-thinning began. The growth curve ofE. camaldulensis followed the logistic growth curve where both maximum size and intrinsic growth rate change with time. Mean intrinsic growth rate was maximized at initiation of growth after lag time and then gradually decreased as time progressed. Hozumi's model was considered to be the best model with wide applicability for describing and comparing the growth characteristics during the course of self-thinning among different species, especially in tropical forest plantations, in which many diverse species were used for reforestation.     

Gas exchange and photosynthesis of Eucalyptus camaldulensis seedlings inoculated with different ectomycorrhizal symbionts

Eucalyptus camaldulensis Dehnh. seedlings inoculated with Pisolithus tinctorius (Pers.) Coker & Couch and Thelephora terrestris Ehrl. per Fr. were grown in well watered soil (_s –0.03 MPa) or subjected to a long-term soil water stress of up to –1.0 MPa over 13-week period in a glasshouse. After 13 weeks, all seedling containers were watered to field capacity and then water was withheld from the E. camaldulensis seedlings to induce a short-term drought. Diurnal measurements of seedling photosynthesis rate (A), leaf stomatal conductance (g) and leaf water potential (_p) were completed before, during, and after the short term drought. Although they were growing in an equal soil volume, photosynthesis rate (A), leaf stomatal conductance and leaf water potential (_p) of larger seedlings with P. tinctorius ectomycorrhizae were similar to those of smaller seedlings colonized with T. terrestris during the short-term drought period. Seedlings inoculated with Pisolithus tinctorius maintained higher photosynthesis rates over the course of the short-term drought. Thus, P. tinctorius ectomycorrhizae appear to be more efficient than those of T. terrestris in assisting seedlings to maintain gas exchange and photosynthesis under limited soil moisture conditions.     

Effect of plant growth regulators and basal media onin vitro shoot proliferation and rooting ofMyrtus communis L.

The influence of macronutrients and growth regulators on in vitro shoot proliferation and rooting of an East Spanish population ofMyrtus communis L. were studied. Preincubation of field material on a medium without mineral salts prevented the browning from phenolic exudates. For multiplication, nodal segments of 5 mm fromin vitro produced shoots were cultured on Murashige and Skoog (MS), Schenk and Hildebrandt (SH) and Heller (H) media (full strength or diluted to 1/2 or 1/4), with 6-benzylaminopurine (BAP) at concentrations 4.4, 13.3 and 22.2 ΜM or kinetin (K) at concentrations 4.7, 14.0 and 23.2 ΜM. The optimum shoot proliferation was on quarter-strength MS medium with 4.4 ΜM BAP, whereas the maximum number of nodal segments was produced on half-strength MS medium with 4.4 ΜM BAP. Rooting of shoots was obtained by adding 2.5 – 24.6 ΜM indole-3-butyric acid (IBA) and broad range of macronutrients; Lloyd and McCown (WPM) and Gresshoff and Doy (GD) media both full strength or diluted to 1/2 were optimum. No rooting was obtained in the presence of α-naphthaleneacetic acid (NAA). Acknowledgements: This study was supported by a grant from Conselleria de Cultura, Educació i Ciència de Ia Generalitat Valenciana. The authors are grateful to Man Cannen Perea for her helpful comments.     

Rapid clonal propagation of Vitex trifolia

This report describes in vitro shoot induction and plant regeneration from mature nodal explants of Vitex trifolia L. on Murashige and Skoog (MS) medium fortified with benzylaminopurine (BAP), kinetin (KN), thidiazuron (TDZ), adenine (ADE), and 2-isopentenyladenine (2-iP) (0.25 – 10.0 μM). Multiple shoots differentiated directly without callus mediation within 3 weeks when explants were cultured on medium supplemented with cytokinins. The maximum number of shoots (9 shoots per explant) was developed on a medium supplemented with 5.0 μM BAP. Shoot cultures was established repeatedly subculturing the original nodal explant on the same medium. Rooting of shoots was achieved on half strength MS medium supplemented with 0.5 μM naphthaleneacetic acid (NAA). Rooted plantlets transferred to pots containing autoclaved soil and vermiculite mixture (1:1) showed 90 % survival when transferred to outdoor.     

Changes in peroxidase activity, and level of phenolic compounds during light-induced plantlet regeneration from Eucalyptus camaldulensis Dehn. nodes in vitro

Node cultures of Eucalyptus camaldulensis Dehn inPetri dishes in vitro under darkness in the presence of anauxin developed meristematic agglomerates (4 to 6 diameter),i.e. dense shoot clusters in which outgrowth of numerous successive buds islimited. Similar cultures under a 16 photoperiod produced smallgreen plantlets with reduced leaves often presenting white hypertrophiedlenticels and very short roots crowning the stem bases. The use of half-litreglass vials under light allowed direct development of well-developed rootedplantlets, either in the presence of the same auxin or in the presence of acytokinin. Light favoured an increase in phenolic compounds and a reversevariation of peroxidase activity during the culture cycles. These aspects arediscussed in terms of a possible regulation of the endogenous auxin levelthrough a light control of peroxidase activity and the level of phenoliccompounds.     

Eucalyptus stomata with occluded anterior chambers

Summary This paper describes variations of a mode of stomatal development already described in a species (E. orbifolia) ofEucalyptus L'Herit. (Carr andCarr, Protoplasma 96, 127, 1978) in which the outer part of the stomatal pore (ostiole) is formed by the creation of a break in the thin layer of cuticle lying over the stomatal chamber. In a number of species with a thick cuticle (e.g., E. cooperana) the process of breakthrough is different: additions to the guard cell upper thickenings extend from them as ridges, pressing the leaf cuticle outwards. Breakthrough of the cuticle occurs above the tips of these extensions. The anterior chamber is lined throughout by the extensions, which become heavily cutinized. This mode of stomatal development is typical of many other species of eucalypts, including those dealt with in this paper.In addition, inE. halophila thickenings develop on the end walls of the anterior chamber above the unusual upturned poles of the guard cells. Cutinized thickenings, pseudo-outer stomatal ledges are also formed on the upper guard cell walls. All these wall thickenings occlude the anterior chamber, leaving only a narrow passage in the form of a letter H.Similar occlusions are found inE. balladoniensis but here the thickenings are developed into the chamber from its lateral walls. InE. gracilis, and the related speciesE. celastroides andE. calycogona, less regular occluding thickenings develop principally from the lateral walls of the chamber. In addition, large pseudo-outer stomatal ledges may be formed.These phenomena are discussed in terms of the mechanism underlying the formation of the occluding thickenings and the possibility of their adaptive significance.     

Clonal propagation of Cinnamomum zeylanicum Breyn. by tissue culture

Multiple shoot formation was induced directly from seeds of Cinnamomum zeylanicum Breyn. and also from seedling explants on Murashige and Skoog's medium containing different concentrations and combinations of auxins and cytokinins. Individual shoots were excised and induced to root on White's liquid medium. These plantlets were then transferred to pots in the green house and were eventually grown successfully under field conditions. Explants from the nodal region of these in vitro rooted plants were also subcultured to fresh medium. They produced a new crop of multiple shoots which could again be rooted by the same procedure.Abbreviations BAP 6-Benzylamino purine - 2,4-D 2,4-Dichlorophenoxy acetic acid - IAA Indole-3-acetic acid - IBA Indole-3-butyric acid - IPA Indole-3-propionic acid - KN Kinetin - NAA -Naphthalene acetic acid     

Application of a novel disposable film culture system to photoautotrophic micropropagation of <Emphasis Type="Italic">Eucalyptus uro-grandis (Urophylia x grandis)</Emphasis>

Summary To overcome various disadvantages of conventional culture vessls for plant micropropagation, we previously developed the photoautotrophic micropropagation technique, with special mention for the first practical film culture system, the ‘Miracle Pack’ (MP), which was made of fluorocarbon polymer film (Neoflo^? PFA film) and supported by a polycarbonate frame. While the PFA film has superior thermal stability, high light transmittance and high gas permeability, making the MP system (MP-PFA) superior to conventional culture vessels for the micropropagation of various plant species, its high cost is a disadvantage. In this study, a possible alternative of lower-cost OTP^? film made of TPX (4-methyl-1-pentane polymer) and CPP (a polypropylene), which possesses similar characteristics to PFA film, is evaluated to develop a novel disposable film culture vesel, termed ‘Vitron’, for culturing Eucalyptus (urophylla x grandis), plantlets. The three film culture systems, MP-PFA, MP-OTP (MP with OTP film), and Vitron, were placed under CO₂ enrichment, low photosynthetic photon flux density (PPFD; 45 μmol m⁻² s⁻¹), and sugar-free medium, using phenol resin foam (Oasis^?) as a substrate. In vitro and ex vitro growth and development of Eucalyptus shoots from the four-leaf stage to the rooting stage were compared for all three culture systems. The effects of the duration and concentration of CO₂ enrichments on in vitro growth of Eucalyptus cultured in the Vitron film system were also examined. The best growth and quality of Eucalyptus plantlets was obtained for the Vitron vessel placed in 3000 ppm CO₂ enrichment for 24 hours per day at low PPFD with sugar-free liquid medium and Oasis as substate. Results of this study suggest that the novel Vitron culture system is suitable for the photoautotrophic micropropagation of Eucalyptus. These authors contributed equally to the research results.     

In vitro regeneration of silktree (Albizzia julibrissin) from excised roots

Root segments (1 cm long) were excised from 15–20 day old seedlings of silktree (Albizzia julibrissin) grown on B5 medium. About 50% of the control (no growth regulators added) root explants formed shoot buds within 15 days after placement on the culture medium. After 30 days, there were about 4 shoots per control explant. Addition of low levels of various auxins (0.5 M) did not influence the formation of shoot buds from the explants. Higher concentrations (5M), however, decreased shoot regeneration. Kinetin and 2iP did not influence shoot regeneration at the concentrations tested (1 & 10 M). Addition of benzyladenine, Zeatin, or thidiazuron to the culture medium increased both the percentage of explants that formed shoots and the number of shoots per explant. Thidiazuron was highly effective in stimulating shoot formation at low concentrations (<1 M). At 0.05 M thidiazuron, 95% of the explants produced shoots and about 10 shoots were formed per explant. Compared to TDZ, higher concentrations (10 M) of benzyladenine and Zeatin were required to enhance shoot formation. Upon excision and transfer to B5 medium, regenerated shoots developed into normal rooted plantlets.Abbreviations BA Benzyladenine - IAA Indoleacetic acid - IBA Indolebutyric acid - NAA Naphthaleneacetic acid - TDZ Thidiazuron - 2ip Isopentenyladenine     

Micropropagation of Calluna vulgaris cv. ‘H.E. Beale’

Axillary shoot producing cultures were obtained from microcuttings and shoot tips of Calluna vulgaris cv. H.E. Beale. For cultures derived from microcuttings the highest multiplication rate of 38 shoots (5 mm or longer) was obtained on a reduced salt medium with the addition of 0.5 mgl^-1 2-isopentenyladenine (2iP) during an 8 week subculture. For shoot tip derived cultures 0.2 mgl^-1 6-benzyladenine (BA) was the best cytokinin and led to a multiplication rate of 26 for a 6 week subculture. The addition of 1 g/l casein hydrolysate to a multiplication medium enhanced shoot proliferation in presence of 0.5 mgl^-1 BA.Despite various auxin treatments shoots formed no roots in vitro but rooted readily if transferred to a peat substrate ex vitro. A high rooting percentage (80%) was also obtained with shoots taken from the end of a multiplication phase and rooted directly. An additional subculture on low auxin containing media before transfer to peat substrate is recommended because the shoot condition can be improved in this way. A high number of rooted plantlets was produced, so the methods described will allow mass propagation.     

Cotton somatic embryo morphology affects its conversion to plant

Somatic embryos differentiated from hypocotyl explant in cotton (Gossypium hirsutum L.) exhibited very divergent morphologies. Six different types of somatic embryos based on cotyledon development were observed. The growth hormones (2,4-dichlorophenoxyacetic acid and kinetin) used in induction and maintenance media did not affect embryo rooting and germination. The 95 % conversion of normal embryos (with two cotyledons) was achieved, while an overall conversion was only 38 %. Horn shaped embryos failed to exhibit shoot growth. Poorly developed apical meristems were responsible for lower conversion percentages in some of embryo classes. However, regenerated plants phenotypically resembled to seed grown control plants regardless of somatic embryo morphology.     

Simple one-medium formulation regeneration of fingerroot [<Emphasis Type="Italic">Boesenbergia rotunda</Emphasis> (L.) mansf. Kulturpfl.] via somatic embryogenesis

Summary Most published protocols necessitate different media formulations for multistep somatic embryogenesis. This study aims to establish a simple but effective formulation for the regeneration of plantlets of the pharmaceutically active Boesenbergia rotunda (L.) Mansf. Kulturpfl, formerly Boesenbergia/Kaempferia pandurata (Schult), to ensure a superior and consistent supply of materials for commercialization purposes. In this study, a single-medium formulation of Murashige and Skoog (MS) supplemented with 13.54μM 2,4-dichlorophenoxyacetic acid (2,4-D) was found to be the only medium out of eight formulations to promote the complete somatic embryogenesis process for the culture of B. rotunda (L.). Callus cultures were initiated from a total of 280 explants of rhizome meristem. The percentage of cultures forming embryogenic callus was 23.3 ±4.3% on this MS medium augmented by 13.54μM 2,4-D. The best plantlet regeneration rate was attained from the first subcultured callus with a mean of 6.6±0.1 plantlets per 1 cm diameter aggregate of callus. Somatic embryogenesis characteristic of monocots was evident from histological studies. The regenerated plantlets have been successfully established in soil.     

Factors affecting maintenance, proliferation, and germination of secondary somatic embryos of Eucalyptus globulus Labill

The described protocol for repetitive somatic embryogenesis (SE) in Eucalyptus globulus produced more somatic embryos than the primary SE protocol. Primary somatic embryos (induced on MS_3NAA) were transferred to the same medium, leading to new cycles of somatic embryos, for at least 2 years. The influence of medium (MS and B5), plant growth regulators (auxins and cytokinins), and light on secondary SE was tested. The MS medium without growth regulators (MS_WH) was more efficient for cotyledonary embryo formation and germination than the B5 medium. Reducing auxin (NAA) levels increased the proliferation of globular somatic embryos and allowed SE competence to be maintained on medium free of plant growth regulators. The addition of two cytokinins (BAP and KIN) to the MS medium did not improve proliferation of globular secondary embryos, but was crucial during later stages of the SE process (germination and conversion). Data also show that light may influence the quality of the process, depending on its stage. Darkness should be maintained until the cotyledonary stage is reached, after which exposure to light is recommended.     

Reduction of generation time in Eucalyptus globulus

The plant growth retardant paclobutrazol was applied by foliar spray to seedlings of Eucalyptus globulus (Labill.) to determine its effects on reproductive activity. It was shown to induce flowering in vegetatively juvenile seedlings and increase reproductive activity in vegetatively adult material. The former case represented a 50% (three year) reduction in generation time for commercial seedlings. Paclobutrazol treatment did not appear to have a negative effect on any aspect of reproductive development, germination of new seed, or early seedling growth, while having a distinct positive effect on reproductive output. This suggests that paclobutrazol may be an effective tool enabling tree breeders to reduce generation time and increase reproductive output in E. globulus. Foliar spraying during autumn was shown to be an effective means and timing of paclobutrazol application, with results evident in the following flowering season. Considerable persistence of paclobutrazol effects was noted in the second season after application. Pot size and growth temperatures also play roles in determining the extent of new reproductive activity each season in E. globulus seedlings.     

Effects of root-zone solutes on Eucalyptus camaldulensis and Eucalyptus bicostata seedlings: Responses to Na+, Mg2+ and Cl-

Specific-ion effects in salt-treated eucalypts were examined with two species known to differ in salt tolerance viz. E. camaldulensis (more tolerant) and E. bicostata (less tolerant). Sand-cultured plants were irrigated with different nutrient solutions designed to impose either osmotic stress (concentrated macronutrients with balanced cations and anions) or specific ion stress from either NaCl or MgCl₂, or from nutrient solutions rich in particular ions viz. Na⁺, Mg²⁺ and Cl^- (balancing counter ions were provided in all cases). Half-strength Hoagland nutrient solution served as control. All treatments were applied at osmotic pressures of approximately 0.52 MPa by appropriate concentrations of each solution. In general, salt-induced growth reductions were greater for E. camaldulensis than for E. bicostata, although E. camaldulensis showed strongest exclusion of Na⁺, Mg²⁺ and Cl^- from shoots. Application of NaCl and concentrated macronutrients resulted in similar growth reductions. E. bicostata seedlings exposed to high Cl^- concentrations in the presence of Mg²⁺ and concentrated cations suffered significantly more shoot and root reduction than those exposed to other salts. Treatment with solution rich in Cl^- resulted in extensive leaf damage, which suggested that Cl^- may have exerted a specific effect. No specific Na⁺ effect was observed for either species, even though shoot Na⁺ concentrations were considerably higher for E. bicostata than for E. camaldulensis. Root growth was considerably less for plants treated with Mg²⁺ salts and this effect was associated with low root Ca²⁺ concentrations.     

Characterisation of two distinct HKT1-like potassium transporters from Eucalyptus camaldulensis

Potassium is an essential macronutrient in higher plants. It plays an important physiological role in stoma movements, osmoregulation, enzyme activation and cell expansion. The demand for potassium can be substantial, especially when the plant concerned is a Eucalyptus tree in excess of 50 m tall. We have isolated two cDNAs, EcHKT1 and EcHKT2, from Eucalyptus camaldulensis (river red gum) which are expressed in leaves, stems and roots. These encode potassium transporter polypeptides with homology to the wheat K⁺-Na⁺ symporter, HKT1. EcHKT1 and EcHKT2 both complemented the K⁺-limited growth of an Escherichia coli K⁺-uptake-deficient triple mutant. EcHKT1 and EcHKT2also mediated Na⁺ and K⁺ uptake when expressed in Xenopus oocytes. A comparison of the EcHKT1 and EcHKT2 sequences and their transport properties indicated that these cDNAs represent two K⁺ transporters with distinct functional characteristics. The functional and structural conservation between these two E. camaldulensis genes and the wheat HKT1 suggests that they play an important, albeit elusive, physiological role.     

Phylogenetics and the conservation of rare taxa in the Eucalyptus angustissima complex in Western Australia

Effective conservation that preservesevolutionary entities and processes requiresconservation action that is targeted atappropriate taxonomic units. Molecularphylogenetics can contribute to the resolutionof taxonomic uncertainties by defining therelationships between populations and species,and allowing the recognition of taxonomicentities that reflect evolutionary history.Some uncertainty surrounds the status of taxain the Eucalyptus angustissima complex,which includes threatened taxa with restricteddistributions. The phylogenetic relationshipsbetween populations and taxa in the E.angustissima complex were investigated usingnuclear and chloroplast RFLP analysis.Phylogenetic relationships confirmed thegenetic differentiation of two taxa, identifiedspecies level divergence between two taxacurrently recognised at subspecies level, andestablished the identity of a previouslyundetermined population. Phylogeneticrelationships between the taxa were notcongruent with presumed relationships based onmorphological characters alone.     

Iron Enhances Aluminum-induced Leaf Necrosis and Plant Growth Inhibition in Eucalyptus camaldulensis

The combined effects of excess Fe and Al on Eucalyptus camaldulensis Dehnh. were studied by investigating time course and visible symptoms of leaf necrosis, plant biomass, the status of some antioxidants and pigments and nutrient concentrations. Seedlings were grown hydroponically in nutrient solutions containing 0 or 500 μM AlCl₃, each with a FeSO₄ range of 1, 12 and 120 μM at pH 4.2. Leaf necrosis and plant growth inhibition were induced by Al and enhanced by the increase in Fe concentration. The process from the first appearance of necrotic spots to leaf death (shedding) of a leaf proceeded from a few days to about 20 days after the leaf had fully expanded. Either 120 μM Fe without Al or Al reduced plant growth to a similar extent but 120 μM Fe without Al did not cause leaf necrosis. In leaves, excess Fe (12 and 120 μM) without Al reduced concentration of ASC and GSH, while concentration of Fe, DHA and GSSG and DHA:ASC and GSSG:GSH ratios tended to increase with the increase in Fe concentration in treatment solution with or without Al. At 1 μM Fe, Al increased concentration of DHA and DHA:ASC and GSSG:GSH ratios. Catalase activity in leaves reduced with the increase in leaf Al concentration. At 1 μM Fe, Al greatly reduced concentrations of Fe and chlorophylls in leaves but increase two times Fe concentration in stems. These suggest that the enhancement effects of Fe on Al-induced leaf necrosis and plant growth inhibition can be discussed in context of the excess Fe itself weakens antioxidant capability of ASC–GSH cycle in leaves and greatly reduces plant growth; and the increase in Fe accumulation in stems is involved in Al-induced leaf chlorosis.