Is Eucalyptus Cryptically Self-incompatible?

BACKGROUND AND AIMS: The probability that seeds will be fertilized from self- versus cross-pollen depends strongly on whether plants have self-incompatibility systems, and how these systems influence the fate of pollen tubes. METHODS: In this study of breeding systems in Eucalyptus urophylla and Eucalyptus grandis, epifluorescence microscopy was used to study pollen tube growth in styles following self- and cross-pollinations. KEY RESULTS: Pollen tubes from self-pollen took significantly longer than those from cross-pollen to grow to the base of the style in both E. urophylla (120 h vs. 96 h) and E. grandis (96 h vs. 72 h). In addition, both species exhibited reduced seed yields following self-pollination compared with cross-pollination. CONCLUSIONS: The present observations suggest that, in addition to a late-acting self-incompatibility barrier, cryptic self-incompatibility could be a mechanism responsible for the preferential out-crossing system in these two eucalypt species.     

Phase change-related variations of dome shape in Eucalyptus urophylla × Eucalyptus grandis shoot apical meristems

Shoot apical meristem (SAM) domes derived from five different outdoor and in vitro sources of juvenile and mature Eucalyptus urophylla × Eucalyptus grandis akin genotypes were compared. Overall measurements of SAM dome height H and diameter D ranged from 2 to 35 μm and 20 to 80 μm, with significant differences according to the various physiological origins of plant material investigated. SAM domes from the mature trees “Mat” were taller than those from the rejuvenated ministock plants “Rej”; from the in vitro microcuttings “IVM” of the same clone and also from the in vitro juvenile seedlings “IVJ”, whereas outdoor seedlings “Juv” exhibited intermediate SAM dome height. SAM domes from the rejuvenated material “Rej”, from the in vitro mature “IVM” and juvenile “IVJ” origins were also narrower than those from the outdoor seedlings “Juv” and to lesser extent than those from the mature trees “Mat”. Overall, the mature source “Mat” displayed bigger and somehow sharper hemispherical domes than those from “Rej” and “Juv”, physiologically more juvenile, or those from the in vitro origins “IVM” and “IVJ” which looked flatter and smaller. SAM dome height, diameter D and H/D values varied also significantly according to the plastochron. More specifically, H, D, and H/D SAM differences between the five origins were not significant during the early plastochron phase corresponding to leaf initiation, to become more salient as leaf structures started to elongate and to differentiate. This was particularly obvious for mature tree “Mat” SAM dome shapes which showed at this stage much higher H/D values than the other SAM sources. A shape index S used for characterizing more accurately dome shape confirmed these trends. These observations provide additional arguments to the view that juvenility in trees becomes more and more time- and shoot-tip restricted as ageing increases in the course of time during the ontogenetical process and could be ultimately confined to the most organogenic phase of SAM, from which shoot characteristics derive.     

Variations of DNA methylation in Eucalyptus urophylla×Eucalyptus grandis shoot tips and apical meristems of different physiological ages

Global DNA methylation was assessed by high-performance liquid chromatography (HPLC) for the first time in Eucalyptus urophylla×Eucalyptus grandis shoot tips comparing three outdoor and one in vitro sources of related genotypes differing in their physiological age. The DNA methylation levels found were consistent with those reported for other Angiosperms using the same HPLC technology. Notwithstanding noticeable time-related fluctuations within each source of plant material, methylation rate was overall higher for the mature clone (13.7%) than for the rejuvenated line of the same clone (12.6%) and for the juvenile offspring seedlings (11.8%). The in vitro microshoots of the mature clone were less methylated (11.3%) than the other outdoor origins, but the difference with the juvenile seedlings was not significant. Immunofluorescence investigations on shoot apices established that the mature source could be distinguished from the rejuvenated and juvenile origins by a higher density of cells with methylated nuclei in leaf primordia. Shoot apical meristems (SAMs) from the mature clone also showed a greater proportion and more methylated cells than SAMs from the rejuvenated and juvenile origins. The nuclei of these latter were characterized by fewer and more dispersed labeled spots than for the mature source. Our findings establish that physiological ageing induced quantitative and qualitative variations of DNA methylation at shoot tip, SAM and even cellular levels. Overall this DNA methylation increased with maturation and conversely decreased with rejuvenation to reach the lower scores and to show the immunolabeling patterns that characterized juvenile material nuclei.     

An Efficient Procedure to Stably Introduce Genes into an Economically Important Pulp Tree (Eucalyptus grandis × Eucalyptus urophylla)

Regeneration problems are one of the main limitations preventing the wider application of genetic engineering strategies to the genus Eucalyptus. Seedlings from Eucalyptus grandis × Eucalyptus urophylla were selected according to their regeneration (adventitious organogenesis) and transformation capacity. After in vitro cloning, the best genotype of 250 tested was transformed via Agrobacterium tumefaciens. A cinnamyl alcohol dehydrogenase (CAD) antisense cDNA from Eucalyptus gunnii was transferred, under the control of the 35S CaMV promoter with a double enhancer sequence, into a selected genotype. According to kanamycin resistance and PCR verification, 120 transformants were generated. 58% were significantly inhibited for CAD activity, and nine exhibited the highest down-regulation, ranging from 69 to 78% (22% residual activity). Southern blot hybridisation showed a low transgene copy number, ranging from 1 to 4, depending on the transgenic line. Northern analyses on the 5–16 and 3–23 lines (respectively one and two insertion sites) demonstrated the antisense origin of CAD gene inhibition. With respectively 26 and 22% of residual CAD activity, these two lines were considered as the most interesting and transferred to the greenhouse for further analyses.     

葡萄桉(Eucalyptus botryoides Sm.)试管育苗

葡萄桉(E.botryoides Sm.)是世界著名的速生树种之一,我国广东、四川等省均有栽培。乔木,树高12～24米,木材红棕色,硬性、强韧而耐久,纹理交错而略粗,宜供建筑、枕木、车厢等用材,其枝叶含精油0.1％。     

In vitro rooting of genetically related Eucalyptus urophylla × Eucalyptus grandis clones in relation to the time spent in culture

In vitro responsiveness of microshoots derived from three genetically related and different age Eucalyptus urophylla × Eucalyptus grandis clones kept cultivated by monthly subcultures was assessed on two rooting media in relation to the time spent in culture. Significant differences of rooting capacity were found between the two 22-year-old half sibling genotypes 147 and 149 according to the concentration of BA added to the media and also to light intensity. The contrast was even more salient with the 1-year-old clone S, which displayed the poorest rooting ability despite being full sibling with clone 147 and much younger. The various criteria observed, i.e. rooting rate, number of roots produced, root length, microshoot height and shoot tip necrosis varied greatly and differently for the three origins according to the successive time replicates of the same rooting protocols, with rooting rates of more than 80% for clone 147 at certain dates. These results were discussed, suggesting the influence of endogenous rhythms as the most rationale cause of the time-related fluctuations of responses observed in steady in vitro environment, notwithstanding possible interferences of non-optimal tissue culture conditions. Such unexpected variations of rootability between closely related genotypes, and the notable interactions pointed out between the time spent in culture and the rooting conditions deserve to be taken into consideration before stating definitive judgment on the rooting capacities of a given genotype for arborescent species.     

Hydrologic balance,net primary productivity and water use efficiency of the introduced exotic Eucalyptus grandis × Eucalyptus urophylla plantation in south-western China

Aims Land cover changes can disrupt water balance and alter the partitioning of precipitation into surface runoff, evapotranspiration and groundwater recharge. The widely plantedEucalyptustrees in south-western China have the potential to bring about hydrologic impacts. Our research aims to elucidate the hydrologic balance characteristics of the introduced exoticEucalyptus grandis×Eucalyptus urophyllaplantation and to assess whether its high productivity results from high water use efficiency (WUE) or large water consumption.     

What physiological processes permit insects to eat Eucalyptus leaves?

Many species of insects eat Eucalyptus foliage despite its relatively low nutritional value and the many plant secondary metabolites (PSMs) present, for example, terpenes, phenols and formylated phloroglucinols (FPGs). Formylated phloroglucinols are a new class of PSMs that act as antifeedants for possums and koalas. What physiological processes are present that permit insects to eat eucalypt foliage and how do PSMs influence insect feeding or digestion? Some trees seem to be repeatedly infested with eucalypt‐feeding insects, possibly as a result of previous chemosensory cues remaining from parental selection of a plant. Avoidance or storage of PSMs permit jarrah leafminers (Perthida glyphopa) and sawflies (Perga sp.) to consume eucalypt foliage without dealing with the majority of these compounds. Some PSMs can be metabolized by polysubstrate membrane oxidases as found in caterpillars or sawflies that feed on eucalypts. High midgut pH may be advantageous for nutrient extraction and PSM metabolism, and midgut pH ranges between 8.5 and 8.9 for caterpillars of Hyalarcta huebneri. Plant secondary metabolites may not be absorbed as a result of the combined presence of the peritrophic matrix and endogenous surfactants. Excretion of PSMs can be as metabolites or intact compounds. Both putative metabolites and sideroxylonal‐A, an FPG, are present in the faeces of larvae of the case moth, H. huebneri. The presence of sideroxylonal‐A in the food had an effect on the presence of 5‐hydroxytryptamine (5HT) in the central nervous system of caterpillars, as larvae fed leaves with a high concentration of sideroxylonal‐A had relatively more 5HT in the brain and central nervous system ganglia than larvae fed leaves containing a low concentration. Further work is necessary to clarify how PSMs are handled by eucalypt‐feeding insects and what effect FPGs have on feeding and digestion.     

Frequency and distribution of cyanogenic glycosides in Eucalyptus L'Hérit

In this study approximately 420 of the described species of Eucalyptus were examined for cyanogenesis. Our work has identified an additional 18 cyanogenic species, 12 from living tissues and a further six from herbarium samples. This brings the total of known cyanogenic species to 23, representing approximately 4% of the genus. The taxonomic distribution of the species within the genus is restricted to the subgenus Symphyomyrtus, with only two exceptions. Within Symphyomyrtus, the species are in three closely related sections. The cyanogenic glycoside was found to be predominantly prunasin (1) in the 11 species where this was examined. We conclude that cyanogenesis is plesiomorphic in Symphyomyrtus (i.e. a common basal trait) but has probably arisen independently in the other two subgenera, consistent with recent phylogenetic treatments of the genus. The results of this study have important implications for the selection of trees for plantations to support wildlife, and to preserve genetic diversity.     

Plant regeneration from seedling explants of Eucalyptus grandis × E. urophylla

Hypocotyls, cotyledons, cotyledonary nodes and primary leaves were used as explants to establish a regeneration protocol for Eucalyptus grandis × E. urophylla. These seedling-derived explants were incubated on a modified MS medium (SP medium), supplemented with 2.0 M TDZ. After 1 month, the calluses obtained were transferred to SP medium containing different concentrations of BA and NAA or zeatin and NAA. Shoots were induced from these calluses at a high frequency. Shoot elongation was then stimulated on SP medium supplemented with BA, NAA and GA₃ for 20 to 30 days. For rooting, 50 mm long shoots were cultivated on root induction medium containing IBA (2.5 M) for different periods and then transferred to the same medium but without auxin, for 30 days. Plantlets were then successfully transplanted to greenhouse conditions.     

Inhibitors of Trichoderma reesei β-glucosidase activity derived from autohydrolysis-exploded Eucalyptus regnans

Summary Enzymic saccharification of Eucalyptus regnans pulps pretreated by autohydrolysis-steam explosion resulted in low cellulose conversions into glucose when using trichodermal cellulase preparations. The reduced levels of glucose were attributable to the production of compounds during enzymic hydrolysis which were inhibitory to -d-glucosidase of Trichoderma reesei C-30 and in Meicelase, but not to the cellulases. Aspergillus niger -glucosidase was not inhibited, nor were -d-xylosidase(s) and 1,4--d-xylanase(s). The inhibitory compound(s) could be extracted from the enzymic hydrolyzates with ethyl acetate. The ethyl acetate extractives inhibited -glucosidase in a competitive manner, and inhibitory action was not affected by pH. Addition of the inhibitory compound(s) to trichodermal cellulase digests of cellulose resulted in reduced glucose yields compared to a control. The inhibitory effects could be overcome when cellulase digests were supplemented with A. niger -glucosidase resulting in higher cellulose-to-glucose conversions. The inhibitory compound(s) were localized mainly in the heartwood of E. regnans. An inhibitor compound of this type has not hitherto been reported. The presence of inhibitory compound(s) in the autohydrolysis liquor fraction is also reported.     

How well can common brushtail possums regulate their intake of Eucalyptus toxins?

We studied factors affecting the ability of common brushtail possums (Trichosurus vulpecula) to regulate their intake of a dietary toxin, jensenone, extracted from Eucalyptus leaves. Increasing concentrations of jensenone in the diet led to a dose-dependent decrease in food intake best described as an exponential decay. Animals that had not previously been exposed to jensenone ate significantly more when first offered food containing the compound than on subsequent days. However, when offered the same amount of food in a number of portions throughout the night, naive animals ate significantly less than animals offered the total meal at once. When offered food containing jensenone over a 13-day period, the animals' intake varied cyclically with relatively high food intakes followed by relatively low intakes. Furthermore, animals that were exposed to cold conditions (4 degrees C) ate more than those maintained at 18 degrees C but this difference was abolished when jensenone was included in the diet. We interpret these results as showing that regulation of toxin intake by common brushtail possums depends on learned responses that can override other important influences on feeding.     

Does low-intensity surface fire increase water- and nutrient-availability to overstorey Eucalyptus gomphocephala?

The objective of this study was to investigate how the management practices of prescribed fire and understorey vegetation removal affect water and nutrient relations of old, yet prematurely declining Eucalyptus gomphocephala. Long unburnt sites were established in Yalgorup National Park, Western Australia, adjacent to frequently burnt state forest sites. Trees were allocated to vegetation clearing, prescribed fire or no prescribed fire treatments. Prescribed fire was achieved in only one long unburnt national park site so that the results were pseudoreplicated but analysed accordingly. Soil chemistry, plant nutrient availability and tree foliar carbon and nitrogen isotope ratio and nutrient concentration were investigated. No effects of vegetation clearing were found. Prescribed fire sites were associated with sky exposure and bare ground whereas no prescribed fire sites were associated with shrub and litter cover and litter depth. Foliar carbon isotope ratios were significantly more negative in prescribed fire, relative to no prescribed fire, treatments on long unburnt sites. Soil exchangeable Zn and Mn and plant available (estimated by charged resin beads) Mg were higher on prescribed fire, relative to no prescribed fire, long unburnt sites. Seedling bioassays indicated elevated P and Cu availability on prescribed fire, relative to no prescribed fire, treatments. In overstorey E. gomphocephala, foliar N levels were elevated (but not to excessive levels), and there was a trend toward elevated foliar Mn, in prescribed fire relative to no prescribed fire treatments on long unburnt sites. In the context of our large-scale pseudoreplicated case study, prescribed fire provided a pulse of water and N, (with some indications towards provision of elevated Mn, Cu and Mg) availability to E. gomphocephala in decline on sites with a history of a long absence of fire that may in part underpin observations of elevated tree health on sites that have a history of relatively frequent fire.     

Phosphite stimulated histological responses of Eucalyptus marginata to infection by Phytophthora cinnamomi

Phosphite is used to protect plants from the soil borne pathogen, Phytophthora cinnamomi. Although phosphite stimulates resistance to P. cinnamomi, this is the first histological study of its effect on Eucalyptus marginata, an economically important forest tree in Western Australia. Clonal lines of E. marginata, considered resistant and susceptible to P. cinnamomi, were underbark inoculated with P. cinnamomi. 4 days later, they were treated with 0, 2.5, 5 or 10 g L⁻¹ phosphite. Transverse hand sections were stained for suberin and lignin, and histological responses to infection were examined. Defence responses were stimulated at all phosphite concentrations in both clonal lines, and the genotypic difference in lesion length was eliminated within 8 days of treatment. In the resistant line, suberin production was stimulated while in the susceptible line both lignin and suberin were stimulated. By 2 days after treatment, phosphite stimulated a faster rate of suberin production in the resistant line than the susceptible line, but by 4 days after treatment, there was no difference in the increase between the lines. Damage caused by P. cinnamomi was found to extend furthest in the cortex and outer phloem in transverse sections in both genotypes. In the presence of P. cinnamomi, phosphite stimulated mitosis as part of the defence response, with meristematic activity involved in the compartmentalisation of damaged tissue (formation of periderm) and closure of healthy tissue (callus). Phytotoxicity had a detrimental effect in healthy tissues and this was more apparent in the resistant line, where it did not provide the best protection from lesion extension and plant mortality, suggesting phytotoxicity could disrupt defence responses. Phosphite increases the capacity of susceptible and resistant E. marginata clonal lines to wall-off and contain P. cinnamomi colonisation through lignin and suberin deposition, and increased meristematic activity.     

Chemodiversity and α-Glucosidase Activity of Eucalyptus Species from Northwestern Himalaya,India

The aim of current work was to determine essential oils (EOs) composition from three Eucalyptus species, including E. citriodora, E. camaldulensis and E. globulus and assess their α-glucosidase inhibitory activity. The EOs were collected using the hydrodistillation technique and characterized by GC/MS, GC-FID and NMR. The isolated EOs from leaves parts of Eucalyptus species varied from 0.56 to 1.0 % on fresh weight basis. The content of the EOs was distinct according to the species. The most abundant metabolites were identified as citronellal (0–83.0 %), 1,8-cineole (0.2–44.8 %), spathulenol (0.4–16.1 %) α-pinene (0.4–15.9 %), p-cymene (3.7–11.9 %), citronellol (0–8.6 %), β-eudesmol (5.3–8.6 %) and β-pinene (0–7.1 %). The EOs obtained from targeted samples exhibited strong α-glucosidase inhibitory activity. These results are encouraging and underline that the EOs of Eucalyptus species may be a promising alternative source of natural antidiabetic.     

尾叶桉(Eucalyptus urophylla)第一代育种群体的遗传多样性

本研究利用中性微卫星标记分析了尾叶桉第一代育种群体的遗传多样性水平和遗传结构。Hardy-Weinberg平衡检测表明尾叶桉第一代育种群体各天然种源内存在明显的杂合子缺失现象。标记和种源的遗传多样性参数统计都表明,尾叶桉育种群体各天然种源具有很高的遗传多样性。分子方差分析显示,尾叶桉育种群体的大部分遗传变异来自家系间,种源间的遗传变异分量很低,该结论被种源间很低的遗传分化水平再次佐证。尾叶桉育种群体内的亚群体分类(Structure分析)与各天然种源的地理分布不存在联系,但群体的内亚群体分类和基于Nei遗传距离的聚类比较吻合。本研究结论可为尾叶桉高世代的群体改良和优良杂交种亲本选配提供遗传基础信息。     

Are Sebacinaceae common and widespread ectomycorrhizal associates of Eucalyptus species in Australian forests?

A molecular survey of basidiomycete ectomycorrhizal fungi colonising root tips at a site in Eucalyptus marginata (jarrah) forest revealed the presence of many fungal species which could not be identified from a database of ITS-PCR-RFLP profiles from morphologically identified species. Three of these unidentified taxa were among the six most frequently encountered profiles. Phylogenetic analyses of ITS and nuclear LSU sequences revealed a close relationship among the three fungi and that they belong to the family Sebacinaceae (sensu Weiss and Oberwinkler 2001). The possibility that DNA of non-ectomycorrhizal rhizosphere or endophytic fungi had been amplified selectively by the basidiomycete-specific primers was tested by amplification with fungal-specific primers. A single PCR fragment was amplified in all but two of the 24 samples tested and digestion with two restriction enzymes produced RFLP profiles which matched those from the Sebacinoid sequence. We conclude, therefore, that at least three species of Sebacinaceae are common ectomycorrhizal associates of E. marginata.     

A Newly Identified Passive Hyperaccumulator Eucalyptus grandis × E. urophylla under Manganese Stress

Manganese (Mn) is an essential micronutrient needed for plant growth and development, but can be toxic to plants in excess amounts. However, some plant species have detoxification mechanisms that allow them to accumulate Mn to levels that are normally toxic, a phenomenon known as hyperaccumulation. These species are excellent candidates for developing a cost-effective remediation strategy for Mn-polluted soils. In this study, we identified a new passive Mn-hyperaccumulator Eucalyptus grandis × E. urophylla during a field survey in southern China in July 2010. This hybrid can accumulate as much as 13,549 mg/kg DW Mn in its leaves. Our results from Scanning Electron Microscope (SEM) X-ray microanalysis indicate that Mn is distributed in the entire leaf and stem cross-section, especially in photosynthetic palisade, spongy mesophyll tissue, and stem xylem vessels. Results from size-exclusion chromatography coupled with ICP-MS (Inductively coupled plasma mass spectrometry) lead us to speculate that Mn associates with relatively high molecular weight proteins and low molecular weight organic acids, including tartaric acid, to avoid Mn toxicity. Our results provide experimental evidence that both proteins and organic acids play important roles in Mn detoxification in Eucalyptus grandis × E. urophylla. The key characteristics of Eucalyptus grandis × E. urophylla are an increased Mn translocation facilitated by transpiration through the xylem to the leaves and further distribution throughout the leaf tissues. Moreover, the Mn-speciation profile obtained for the first time in different cellular organelles of Eucalyptus grandis × E. urophylla suggested that different organelles have differential accumulating abilities and unique mechanisms for Mn-detoxification.     

Community analysis of undergrowth vegetation in industrial plantation of Eucalyptus grandis×E.urophylla

基于群落调查方法,对巨尾桉林下植物进行群落学分析.结果表明:巨尾桉林下植物共有维管束植物57科、124属、154种及变种,以禾本科和菊科的种类占优势;林下植物物种组成较为分散,优势属不明显;种子植物53科划分为6个分布区类型和2个变型,以热带分布科为主;种子植物120个属有14个分布区类型和7个变型,以热带分布属为主,植物区系成分较为复杂.林下植被可分为灌木层和草本层,但是分层现象不明显,草本层植物占优势,偶见有少量的层间植物分布.林下物种丰富度表现为草本层>灌木层;而Simpson指数、Shannon-Wiener指数、Pielou均匀度指数则表现为灌木层>草本层,生境空间异质性及人为干扰活动影响了巨尾桉林下植物组成及分布.