Improved germination of the Australian natives: Hibbertia commutata, Hibbertia amplexicaulis (Dilleniaceae), Chameascilla corymbosa (Liliaceae) and Leucopogon nutans (Epacridaceae)

Hibbertia commutata (Steudel), H. amplexicaulis (Steudel), Chameascilla corymbosa [(R.Br.) F.Muell. Ex Benth.] and Leucopogon nutans (E. Pritzel) are four Australian species that are difficult to germinate during mine-site rehabilitation. Laboratory germination trails were conducted to identify dormancy mechanisms and to improve germination response. Treatments applied to all species included scarification and scarification followed by soaking seeds in smoke water (1, 5 or 10%) or gibberellic-acid solution (50, 200 or 1000 muM). Additional treatments with kinetin solution (50, 200 or 1000 muM) and smoke water (50 or 100%) were applied to scarified or unscarified seeds of C. corymbosa. Thermal-shock treatment was applied to L. nutans fruit, some of which were subsequently scarified and subjected to both smoke water (10%) and gibberellic-acid solution (1000 muM). Significant germination increases were obtained by using dormancy-breaking treatments on H. commutata ( from 12.8 to 76.0%), H. amplexicaulis (from 6.8 to 55.1%) and C. corymbosa (from 48.5 to 86.4%). Scarification alone increased germination of both Hibbertia species, suggesting that these species display a physical seed coat-imposed dormancy mechanism. Germination of H. amplexicaulis was further increased by the application of gibberellic-acid solution, indicating a possible embryo-imposed dormancy mechanism. Scarification followed by the application of smoke water produced the highest germination response for C. corymbosa seeds. Scarification alone did not significantly increase germination, inferring the existence of a smoke-responsive embryo dormancy mechanism. Seeds of L. nutans, although viable, failed to germinate and are thought to display complex seed coat- and embryo-imposed dormancy mechanisms.

Potential to increase yield in lucerne (Medicago sativa subsp sativa) through introgression of Medicago sativa subsp falcata into Australian adapted material

The effect of interspecific heterosis in crosses between Medicago sativa subsp. sativa and M. sativa subsp. falcata was assessed. Three sativa and 3 falcata plants were crossed in a diallel design. Progeny dry matter yield and natural plant height were assessed in a replicated field experiment at Gatton, Queensland. Yield data were analysed using the method of residual maximum likelihood (REML) and Griffing's model 1. There were significant differences between the reciprocal, general combining ability (GCA), and specific combining ability (SCA) effects. As expected, S-1 populations were lower yielding than their respective intraspecific cross and falcata x falcata crosses were significantly lower yielding than sativa x sativa crosses. Some of the interspecific crosses indicated substantial SCA effects, yielding at least as well as the best sativa x sativa crosses. We have demonstrated the potential usefulness of unselected M. sativa subsp. falcata as a heterotic group in the improvement of yield in northern Australian adapted lucerne material, and discuss how it could be incorporated into future breeding to overcome the yield stagnation currently being experienced in Australian programs.

Short- and long-term consequences of thermal variation in the larval environment of anurans

To survive adverse or unpredictable conditions in the ontogenetic environment, many organisms retain a level of phenotypic plasticity that allows them to meet the challenges of rapidly changing conditions. Larval anurans are widely known for their ability to modify behaviour, morphology and physiological processes during development, making them an ideal model system for studies of environmental effects on phenotypic traits. Although temperature is one of the most important factors influencing the growth, development and metamorphic condition of larval anurans, many studies have failed to include ecologically relevant thermal fluctuations among their treatments. We compared the growth and age at metamorphosis of striped marsh frogs Limnodynastes peronii raised in a diurnally fluctuating thermal regime and a stable regime of the same mean temperature. We then assessed the long-term effects of the larval environment on the morphology and performance of post-metamorphic frogs. Larval L. peronii from the fluctuating treatment were significantly longer throughout development and metamorphosed about 5 days earlier. Frogs from the fluctuating group metamorphosed at a smaller mass and in poorer condition compared with the stable group, and had proportionally shorter legs. Frogs from the fluctuating group showed greater jumping performance at metamorphosis and less degradation in performance during a 10-week dormancy. Treatment differences in performance could not be explained by whole-animal morphological variation, suggesting improved contractile properties of the muscles in the fluctuating group.

Can we learn more from fewer seeds?

The vital role that 'design of experiment' (DOE) plays when investigating dormancy, germination and potentially influential factors is discussed. The Taguchi analysis, developed by Dr. Genichi Taguchi, is a special application and analysis of the fractional factorial aimed at achieving maximum information from minimum effort and resources.

The effects of temperature and salinity on Acacia harpophylla (brigalow) (Mimosaceae) germination

Acacia harpophylla F. Muell. (brigalow) used to naturally occur over a range of about 50 000 km(2) in Queensland and New South Wales, Australia. Large scale clearing for agriculture has reduced the area to less than 20 000 km(2) and it is estimated that 20-25% of vertebrate fauna living in brigalow communities will become locally extinct as a result of the current clearing induced loss of habitat. Some coal mining companies in central Queensland have become interested in providing habitat for the endangered bridle nail-tailed wallaby that lives in brigalow vegetation. However, there is little known about establishment techniques for brigalow on mine sites and other disturbed ground; an understanding of brigalow biology and ecology is required to assist in the conservation of this threatened vegetation community and for re-creation of bridled nail-tail wallaby habitat in the post mining landscape. Brigalow is an unusual species of Acacia because it is not hard-seeded and germinates readily without the need to break seed-coat imposed dormancy. Germination trials were undertaken to test the ability of brigalow seed to germinate with a range of temperatures and salinity levels similar to those experienced in coal mine spoil. Optimum germination was found to occur at temperatures from 15 to 38 degrees C and no germination was recorded at 45 degrees C. Brigalow was very tolerant of high salt levels and germinated at percentages greater than 50% up to the highest salinity tested, 30 dS/m. Germination of greater than 90% occurred up to an electrical conductivity of 20 dS/m. The results indicate brigalow seed can be sown in summer when rains are most likely to occur, however, shading of the seed with extra soil or mulch may ensure the ground surface does not become too hot for germination. Because of its ability to germinate at high salinity levels, brigalow may be suitable for use in saline mine wastes which are common on sites to be rehabilitated after mining.

An evolutionary framework for studies of hibernation and short-term torpor

Data from diverse studies endorse ideas that short term torpor and hibernation are expressions of ancient characters. In evolutionary terms, their basic mechanisms are probably plesiomorphic (= ancestral/primitive) and physiologically similar. This contrasts with the alternate view that they are apomorphic (= derived, specialized), arising independently in many taxa from homeothermic ancestry by numerous apparent convergences. This paper explores some of the implications of accepting the plesiomorphic interpretation. Hibernation is, of course, a complex phenomenon that has undergone variations and refinements in different mammalian lineages. The argument is not that hibernation in total is a plesiomorphic character, but that it is built upon fundamental processes that are. Taking this view provides a framework for research that emphasizes the value of comparative studies, particularly of reptiles and birds. Studies of reptiles, for example, might unravel the mystery about periodic arousals. A plesiomorphic framework also explains the most extreme examples of hibernation as derived specializations from ancestry in which heterothermy is more about energy management than escape from cold. It cautions against using low body temperature (Tb) alone to diagnose torpor, emphasizes the need to distinguish between constitutional eurythermy (plesiomorphic) and constitutional stenothermy (apomorphic), and leads to a parsimonious theory about the evolution of endothermy. The paper proposes that brown adipose tissue (BAT) is apomorphic within eutheria and highlights the conundrum posed by the occurrence of both nonshivering thermogenesis (NST) and rapid arousal from hibernation in noneutherian mammals that lack BAT and uncoupling protein 1 (UCP1). It endorses the likely existence of a different, ancient and widespread mechanism for regulatory NST in mammals.

Study of the effects of climate warming on seed germination and seed longevity of snowbed species

Understanding how climate change will affect the distribution and the phenology of plants is becoming an increasingly important topic in ecological studies. In response to climate warming, there are documented upward shift and alterations of phenology and physiology of several plant species. Despite this, the effects of climate change on plant regeneration from seeds have largely been neglected. However, regeneration from seeds, a key event in the plant life cycle, could be significantly affected by climate warming. In this regard, we investigated how climatic changes will affect the seasonal dynamics of seed germination and seedling survival in two different alpine context. The first part refers to five species inhabiting a snowbed located at the Gavia pass (Parco Naturale dello Stelvio). Here, plants were exposed, in the field, to natural conditions and to artificial warming using Open Top Chambers proposed by the ITEX (International Tundra Experiment). The germination curves of seeds produced were compared in order to highlight differences in seed germination ecology and in seed physiology induced by the climate warming. In the second part, we considered two tree species that form the treeline in Davos (Switzerland). As a surrogate of climate warming we used the natural thermal gradient driven by the altitude and we compared the germination behavior of the species studied in three sites at three different elevations in order to evaluate the likelihood of treeline shift under the predicted climate warming.

Embodying and embedding children's sleep: some sociological comments and observations

This paper, drawing on our own research findings data, explores the embodiment and embedment of sleeping in children's everyday/night lives. Key themes here include children's attitudes and feelings toward the dormant body, the processes, routines and rituals associated with going to bed and going to sleep, issues associated with bedrooms and privacy, and finally the relationship between dormancy and domicile. This in turn provides the basis, in the remainder of the paper, for a further series of reflections on the mutually informing relations between the sociology of sleep and the sociology of childhood. Remaining questions and challenges involved in researching children's sleep are also considered. Sleep, it is concluded, is not simply a rich and fascinating sociological topic in its own right it also has the potential to shed valuable new light on a significant yet hitherto under-researched part of children's lives, contributing important new insights in doing so.

Growth, sporulation and spore properties of bacillus stearothemophilus produced in chemically defined media

The nutritional requirements for the vegetative growth of B. stearothermophilus strains NCIB 8919, NCTC lO,OO3 (wild) were found to be DL-methionine, biotin, nicotinic acid, thiamin, glucose and mineral salts. Strains NCIB 8920 required in addition L-tryptophan. B. stearothermophilus NCTC lO,OO3 (mutant) grew in a medium containing only glucose and mineral salts. Separate chemically defined media for the growth of Bacillus stearothermophilus strains NCIB 8919, 8920, NCTC lO,OO3 (wild) and NCTC lO,OO3 (mutant) were developed. Optimally aerated culture of B. stearothermonhilus NCTC lO,OO3(mutant) required 1.0 x 10-4 M. Mn2+ and 2.4 x 10-3 M. glutamic acid for optimal sporulation. Specific nutrient depletion of growth affected percentage sporulation. Spore suspensions of B. stearothermophilus NCTC 10,003 (mutant) were prepared from media in which sulphate (SO4-), nitrogen (N-),phosphate (Po4-), carbon (C-), magnesium-carbon simultaneously (Ng-C-) depleted growth. The heat resistance, dormancy and chemistry of these spores varied considerably. B. stearothermophilus NCTC 10,003 10,00310,00(mutant) spores prepared from carbon depleted cultures containing high and low concentrations of calcium, iron or manganese showed variations in heat resistance,dormancy and chemical composition. Progressive increase in the concentration of medium calciumfrom 1.0 X 10-5  M to 1.4 X 10-4 M. progressively increased theheat resistance of B. stearothermophilus NCTC 10,003 (mutant) spores prepared from nitrogen depleted cultures (N-). The thermodynamic functions for germination rate, magnesium and manganese release of N- and SO4- spores were within the range expected of enzymic reactions. The thermodynamic functions for the breaking of dormancy in SO4- spores and that for the release of D.P.A. were identical. Sublethal heating of SO4- spores (96.5°C and below) induced dormancy in these spores, whereas heating above 96.5°C gave rise to heat activation. Pooled results of the chemical analyses of all spore types studied showed that the concentration of D.P.A. and calcium were positively related to heat resistance whereas magnesium concentration and Mg/Ca molar ratio were inversely proportional to heat resistance.

The costs and controls of flowering and fruiting in {\it Spondias purpurea\/}

Small potted trees of Spondias purpurea were monitored to determine the costs and controls of flowering and fruiting. The effect of photoperiod, extremes in moisture and temperature, and defoliation were examined. The carbon exchange rates of the leaves, shoots and fruits were determined. Light response curves and diurnal levels were also investigated. $\sp{13}$Carbon labeling was used to determine which plant parts are carbon sinks. Photoperiod induces dormancy and bud activity. Extremes in soil moisture and temperature induce leaf fall. Flowers, fruits, and roots are carbon sinks. The results were used to develop a phenological model with latitude, soil moisture, and air temperature as variables. ^

Aedes aegypti pharate first instar aseasonal quiescence cues anticipatory plasticity with implications for urban vector ecology and control

The eggs of the dengue fever vector Aedes aegypti possess the ability to undergo an extended quiescence period hosting a fully developed first instar larvae within its chorion. As a result of this life history stage, pharate larvae can withstand months of dormancy inside the egg where they depend on stored reserves of maternal origin. This adaptation known as pharate first instar quiescence, allows A. aegypti to cope with fluctuations in water availability. An examination of this fundamental adaptation has shown that there are trade-offs associated with it. ^ Aedes aegypti mosquitoes are frequently associated with urban habitats that may contain metal pollution. My research has demonstrated that the duration of this quiescence and the extent of nutritional depletion associated with it affects the physiology and survival of larvae that hatch in a suboptimal habitat; nutrient reserves decrease during pharate first instar quiescence and alter subsequent larval and adult fitness. The duration of quiescence compromises metal tolerance physiology and is coupled to a decrease in metallothionein mRNA levels. My findings also indicate that even low levels of environmentally relevant larval metal stress alter the parameters that determine vector capacity. ^ My research has also demonstrated that extended pharate first instar quiescence can elicit a plastic response resulting in an adult phenotype distinct from adults reared from short quiescence eggs. Extended pharate first instar quiescence affects the performance and reproductive fitness of the adult female mosquito as well as the nutritional status of its progeny via maternal effects in an adaptive manner, i.e., anticipatory phenotypic plasticity results as a consequence of the duration of pharate first instar quiescence and alternative phenotypes may exist for this mosquito with quiescence serving as a cue possibly signaling the environmental conditions that follow a dry period. M findings may explain, in part, A. aegypti's success as a vector and its geographic distribution and have implications for its vector capacity and control.^

A biotecnologia na propagação e conservação do umbuzeiro (Spondias tuberosa Arr. Cam.) e percepção sobre sua importância por agricultores da comunidade Malhada Vermelha, Campo Redondo (RN-Brasil)

The umbu tree (Spondias tuberosa Arruda) is a fruit native to the northeast of Brazil with great economic, social and ecological importance for the northeastern semi-arid region. Despite its role, the umbu tree has suffered negative pressure thanks to cluttered extractivism and to negative selection of its fruits, which as the deforestation and the dormancy of seeds contribute to the decrease of its production year after year, making necessary studies that contribute to the improvement of this specie and its conservation. Given the risks to the conservation of the specie and its usefulness to the population, the association between plant biotechnology, for being a tool that can be used to increase its production. and the perception of gathering communities, by valuing the point of view and the knowledge of the population, can facilitate its conservation. This work aimed to develop methods of propagation for umbu tree as well as contribute to its conservation by using biotechnology, with specific objectives to contribute to the conservation of this species; determine concentrations of BAP and ANA in the formation of buds; testing the efficiency of different substrates and concentrations of gibberellic acid on germination in vitro and ex vitro, as well as capture the perception of families in communities that engage in the gathering of umbu. To study the germination, the seeds were inoculated in different substrates (vermiculite, vermiculite + clay, clay, clay + manure and manure + vermiculite) and in different concentrations of gibberellic acid (0 mg, 250 g and 500 mg). For the formation of buds BAP to 0.1 mg-1 was associated with different concentrations of ANA (0.2; 0.4; 0.8mg.L-1). The study of perception was conducted by applying semi-structured questionnaire with Malhada Vermelha community. The experiments resulted in vermiculite and concentration of 500 mg gibberellic acid as the best for germination. The association of 0.1 mg.L-1 of BAP to 0.2 mg.L-1 of ANA provided better formation of buds. As to the application of questionnaires, they revealed that the population understands the decreased amount of umbu plants and umbu fruit in the region, as well as shows concern for its conservation.

Structural Basis of RNA Recognition by Mycobacterium tuberculosis MazF Homologues

The MazEF toxin-antitoxin (TA) system consists of the antitoxin MazE and the toxin MazF. MazF is a sequence-specific endoribonuclease that upon activation causes cellular growth arrest and increass the level of persisters. Moreover, MazF-induced cells are in a quasi-dormant state that cells remain metabolically active while stop dividing. The quasi-dormancy is similar to the nonreplicating state of M. tuberculosis during latent tuberculosis, thus suggesting the role of mazEF in M. tuberculosis dormancy and persistence. M. tuberculosis has nine mazEF TA modules, each with different RNA cleavage specificities and implicated in selective gene expression during stress conditions. To date only the Bacillus subtilis MazF-RNA complex structure has been determined. As M. tuberculosis MazF homologues recognize distinct RNA sequences, their molecular mechanisms of substrate specificity remain unclear. By taking advantage of X-ray crystallography, we have determined structures of two M. tuberculosis MazF-RNA complexes, MazF-mt1 (Rv2801c) and MazF-mt3 (Rv1991c) in complex with an uncleavable RNA substrate. These structures have provided the molecular basis of sequence-specific RNA recognition and cleavage by MazF toxins.

Both MazF-mt1-RNA and MazF-mt3-RNA complexes showed similar structural organization with one molecule of RNA bound to a MazF-mt1 or MazF-mt3 dimer and occupying the same pocket within the MazF dimer interface. Similar to B. subtilis MazF-RNA complex, MazF-mt1 and MazF-mt3 displayed a conserved active site architecture, where two highly conserved residues, Arg and Thr, form hydrogen bonds with the scissile phosphate group in the cleavage site of the bound RNA. The MazF-mt1-RNA complex also showed specific interactions with its three-base RNA recognition element. Compared with the B. subtilis MazF-RNA complex, our structures showed that residues involved in sequence-specific recognition of target RNA vary between the MazF homologues, therefore explaining the molecular basis for their different RNA recognition sequences. In addition, local conformational changes of the loops in the RNA binding site of MazF-mt1 appear to play a role in MazF targeting different RNA lengths and sequences. In contrast, the MazF-mt3-RNA complex is in a non-optimal RNA binding state with a symmetry-related MazF-mt3 molecule found to make interactions with the bound RNA in the crystal. The crystal-packing interactions were further examined by isothermal titration calorimetry (ITC) studies on selected MazF-mt3 mutants. Our attempts to utilize a MazF-mt3 mutant bearing mutations involved in crystal contacts all crystallized with few nucleotides, which are still found to interact with a symmetry mate. However, these different crystal forms revealed the conformational flexibility of loops in the RNA binding interface of MazF-mt3, suggesting their role in RNA binding and recognition, which will require further studies on additional MazF-mt3-RNA complex interactions.

In conclusion, the structures of the MazF-mt1-RNA and MazF-mt3-RNA complexes provide the first structural information on any M. tuberculosis MazF homologues. Supplemented with structure-guided mutational studies on MazF toxicity in vivo, this study has addressed the structural basis of different RNA cleavage specificities among MazF homologues. Our work will guide future studies on the function of other M. tuberculosis MazF and MazE-MazF homologues, and will help delineate their physiological roles in M. tuberculosis stress responses and pathogenesis.

Ecological and evolutionary effects of dispersal on freshwater zooplankton

A recent focus on contemporary evolution and the connections between communities has sought to more closely integrate the fields of ecology and evolutionary biology. Studies of coevolutionary dynamics, life history evolution, and rapid local adaptation demonstrate that ecological circumstances can dictate evolutionary trajectories. Thus, variation in species identity, trait distributions, and genetic composition may be maintained among ecologically divergent habitats. New theories and hypotheses (e.g., metacommunity theory and the Monopolization hypothesis) have been developed to understand better the processes occurring in spatially structured environments and how the movement of individuals among habitats contributes to ecology and evolution at broader scales. As few empirical studies of these theories exist, this work seeks to further test these concepts. Spatial and temporal dispersal are the mechanisms that connect habitats to one another. Both processes allow organisms to leave conditions that are suboptimal or unfavorable, and enable colonization and invasion, species range expansion, and gene flow among populations. Freshwater zooplankton are aquatic crustaceans that typically develop resting stages as part of their life cycle. Their dormant propagules allow organisms to disperse both temporally and among habitats. Additionally, because a number of species are cyclically parthenogenetic, they make excellent model organisms for studying evolutionary questions in a controlled environment. Here, I use freshwater zooplankton communities as model systems to explore the mechanisms and consequences of dispersal and to test these nascent theories on the influence of spatial structure in natural systems. In Chapter one, I use field experiments and mathematical models to determine the range of adult zooplankton dispersal over land and what vectors are moving zooplankton. Chapter two focuses on prolonged dormancy of one aquatic zooplankter, Daphnia pulex. Using statistical models with field and mesocosm experiments, I show that variation in Daphnia dormant egg hatching is substantial among populations in nature, and some of that variation can be attributed to genetic differences among the populations. Chapters three and four explore the consequences of dispersal at multiple levels of biological organization. Chapter three seeks to understand the population level consequences of dispersal over evolutionary time on current patterns of population genetic differentiation. Nearby populations of D. pulex often exhibit high population genetic differentiation characteristic of very low dispersal. I explore two alternative hypotheses that seek to explain this pattern. Finally, chapter four is a case study of how dispersal has influenced patterns of variation at the community, trait and genetic levels of biodiversity in a lake metacommunity.

Efeito de fatores ambientais e tolerância a herbicidas em três espécies de plantas daninhas da família Rubiaceae

The strong selection pressure exerted by intensive use of glyphosate in cultivated areas has selected populations of the Rubiaceae weed species Borreria latifolia (Aubl.) K.Shum. (broadleaf buttonweed), Galianthe chodatiana (Standl.) E.L. Cabral (galiante) and Richardia brasiliensis Gomes (Brazilian pusley) with differential sensitivity to this herbicide in the South region of Brazil. The control of these weeds with herbicides is troublesome and signals the need to incorporate management practices of ruderal flora and crops, more sustainable and that results in more efficient control for long-term. Therefore, it is very important to expand the information about their biology and management. This study aimed: (a) select efficient methods to overcome dormancy of B. latifolia and G. chodatiana and determine how they influence the kinetics of seeds germination; (b) analyze the effects of temperature, irradiance, pH, aluminum and salinity on seed germination and initial growth of the B. latifolia, G. chodatiana e R. brasiliensis seedlings; (c) evaluate tolerance to glyphosate levels in biotypes of B. latifolia, G. chodatiana e R. brasiliensis through dose-response curves and compare two methods to evaluate herbicidal control; (d) and evaluated the effectiveness of alternative herbicides in pre-emergence and in early and late post-emergence of the three species. The treatment with KNO3 2%/3h + gibberellic acid 400 ppm resulted in higher percentage of G. chodatiana seed germination. This treatment and also the dry heat (60°C/30 min) + KNO3 2%/3h were more effective in overcoming dormancy of B. latifolia. G. chodatiana and R. brasiliensis tolerate lower temperatures during the germination process, while B. latifolia tolerate higher temperatures. B. latifolia and R. brasiliensis are positive photoblastic while G. chodatiana is indifferent to the photoperiod. B. latifolia shows higher germination and early development in pH 3, while G. chodatiana and R. brasiliensis prefer pH range between 5 and 7. B. latifolia and G. chodatiana were more tolerant to the aluminum during the germination process than R. brasiliensis. Low salt levels were sufficient to reduce the seed germination of the three species. Some biotypes of B. latifolia and R. brasiliensis showed medium-high glyphosate tolerance, not being controlled by higher doses than recommended. The G. chodatiana specie was not controlled with the highest dose used, showing a high glyphosate tolerance. The sulfentrazone, s-metolachlor and saflufenacil herbicides sprayed in pre-emergence showed high efficacy both on B. latifolia and R. brasiliensis, while chlorimuron-ethyl and diclosulan were effective only on R. brasiliensis. In early post-emergence the fomesafen, lactofem and flumioxazin herbicides efficiently controlled plants of all species, while bentazon showed high efficacy only on B. latifolia. Noteworthy the susceptibility of the G. chodatiana specie for applications in early post-emergence, because the control effectiveness and the number of effective herbicides are reduced with increasing the plant age. Many treatments with tank mix or sequencial applications with glyphosate, were effective in controlling B. latifolia and R. brasiliensis plants in advanced stage of development.