School of BioSciences - Theses
Now showing items 1-12 of 202
An Investigation of flavanone O- and C-methylation in Eucalyptus
Flavonoid compounds are well recognized for their diverse health-promoting properties in humans. Methylation of flavonoids catalysed by plant methyltransferases is an important modification technique that alters the biochemical properties of the compounds, thereby enabling extension of their promising medicinal effects from in vitro to in vivo. Some Eucalyptus trees accumulate high levels of O-and C-methylated flavanones in their leaves. These compounds have potential medicinal and antimicrobial use. Despite the potential use of Eucalyptus species as a commercial source of these flavonoids, little is known about the underlying mechanism of in planta methylation and biosynthesis of these compounds. Through the isolation and characterization of flavanone C-and O- methyltransferases from Eucalyptus, this research aims to provide a detailed insight into the underlying mechanism of different types of in planta methylation and the process by which these compounds are modified. An integrated analysis of transcriptome and metabolome coupled with in silico methods resulted in a cDNA clone (EnOMT1), catalysing the position-specific O-methylation of a single hydroxyl group in the flavanone pinocembrin. The biochemical characterization showed that EnOMT1 is a regiospecific methyltransferase with strict positional specificity to the 7-hydroxyl of flavonoids. The reduction/absence of activity of EnOMT1 with B-ring substituted flavonoids such as luteolin, apigenin, and quercetin demonstrates the pronounced effect B-ring hydroxyl configuration has on the activity of the enzyme. Several Eucalyptus species belonging to subgenus Eucalyptus (monocalypts) show accumulation of different complements of methylated flavonoids. To further understand the genetic basis of this differential accumulation, three homologous genes of EnOMT1 were isolated and characterised from various species. A homologue (ExOMT1) from a species that does not accumulate any methylated flavanones showed 94% identity to EnOMT1; however, the homologue showed no methylation activity with flavanones in vitro. Gene expression analysis using qPCR showed a comparably lower expression of ExOMT1 to that of EnOMT1.Therefore, the observed difference in activity is likely due to the amino acid difference of the EnOMT1 sequence to ExOMT1. Homology modelling along with site-directed mutagenesis of EnOMT1 further identified residues important for activity. The residues -- namely Trp252, Val253, and Asn256 --play critical roles for EnOMT1 activity. SAM binding residues --namely, Trp148, Phe161, Met165, Asn169, Gln194, Phe216, Asp217, Arg218, Asp237, Met238, Phe239, Lyts251, Trp252, Val253, and Trp257 –– were also predicted with a higher degree of confidence. Genetic basis of flavonoid C-methylation is an understudied area and the limited published information on plant CMT sequences makes the research progress difficult. The work presented in chapter 4 provides interesting insights into flavanone C-methylation in Eucalyptus. Extensive flavanone profiling of ten species from subgenus Eucalyptus was conducted to identify a C-methyl flavanone rich species. Two known C-methyl flavanones, cryptostrobin (monomethylated at position C6) and desmethoxymatteucinol (dimethylated at C6 and C8), were identified in E diversifolia in relative abundance. The sequenced transcriptome of E diversifolia was analysed for the identification of secondary metabolite pathway genes putatively involved in C-methylation. Six putative CMT genes were cloned successfully and analysed for activity. Despite our attempts, no in vitro C-methylation was achieved on the flavanone substrates tested. The data presented in chapter 4 provide a solid platform of knowledge for future research on flavanone C-methylation. The results presented in this thesis represent an important step towards obtaining a greater understanding of flavonoid biosynthesis in Eucalyptus and provide clear insights into the reaction mechanism as well as the active site residues of the enzyme.
Investigating DEFECTIVE KERNEL1 regulation of primary cell wall biosynthesis and mechanical properties during plant growth in Arabidopsis thaliana
Plants possess cell wall, a polysaccharide exoskeleton which encompasses all plant cells. Cell wall gives plant cells mechanical support, defines their shape, enables growth and water transport through a plant. It also has important role in communication with the external environment. Regulation of plant cell wall biosynthesis and cell and organ morphogenesis depends on cell’s ability to detect mechanical signals originating both from the external environment and from internal plant tissues. Thanks to the presence of the cell wall, all living plant cells develop constant internal pressure generated by the active water uptake, known as turgor pressure, which enables them to grow. Thus, actively growing cells in the tissue are exerting mechanical stress to each other. In order to properly coordinate cell growth, tissue morphogenesis and maintain cell-to-cell adhesion, plant cell have to detect these mechanical signals. That is performed by a group of still not well enough characterized plant mechanosensitive proteins. Mechanosensors are proteins capable of detecting changes in mechanical stress patterns and translating them into physiological and developmental outputs. One of plant mechanosensitive proteins, DEFECTIVE KERNEL1 (DEK1) has shown to be a very important in proper plant development. DEK1 bears similarity with animal cysteine proteases of Calpain superfamily. DEK1 is very important for plant development since all null alleles are embryo lethal. During the last 20 years of DEK1 studies, this protein has proven to be a very difficult for different molecular and biochemical manipulations. As a consequence, very little is known about its direct target proteins. Wang and co-workers (2003) and Johnson and co-workers (2008) have given a valuable contribution to biochemical understanding of DEK1 by determining that it functions as Cys-protease in similar way as animal calpains. However, a lot of indirect knowledge was gathered about the effects of disruption and modulation of DEK1 activity. DEK1 is important for proper organ development, epidermal specification, and maintenance. However, some studies have inferred that DEK1 affects expression of different cell wall related genes, and it regulates cell-to-cell adhesion in epidermal cells. This led to two extensive studies (Amanda et al., 2016, 2017) which demonstrated importance of DEK1 in regulation leaf epidermal cell walls in A. thaliana mature leaves and inflorescence stems. These studies demonstrated that DEK1 also influences cell wall thickness and cell-to-cell adhesion and that it could potentially regulate cell growth and expansion. Building up on this research, we decided to try to further characterize molecular and biomechanical aspects of DEK1 mediated cell wall regulation, with special emphasis on regulation of cellulose synthesis. We used two mutant lines, with modulated DEK1 activity, a constitutive overexpressor for DEK1 CALPAIN domain and a point mutant in CALPAIN domain, dek1-4. In Chapter 3 we demonstrated that DEK1 regulates dynamics of Cellulose Synthase Complexes (CSCs). Both lines showed decreased crystalline cellulose contents. This led us to investigate if velocity of CSCs in cotyledons, was affected, since it is known that changes in cellulose contents are often caused by defects in CSC. We found that bothDEK1 modulated lines we used have significantly decreased velocity of CSCs. We have also examined plasma membrane turnover rates of CSCs and found out that after photo-bleaching OE CALPAIN has much faster recovery rates compared to Col-0 wild type, while dek1-4 has lower exocytotic rates of CSCs, and much longer life-time of CSCs inserted into the plasma membrane. These results suggested that DEK1 regulates different aspects of CSC dynamics, possibly through interaction with different regulatory proteins. Decrease in cellulose contents we observed in DEK1 modulated lines, prompted us to investigate how this reflects biomechanics and structural properties of epidermal cotyledon cell walls of DEK1 modulated lines, which is described in Chapter 4. To achieve this, we developed a novel microdissection method for isolation and mechanical and structural characterization of native epidermal cell wall monolayers using atomic force microscopy (AFM). AFM force spectroscopy assays showed that both DEK1 modulated lines had stiffer cell walls compared to Col-0. This was awkward since we initially detected decrease in crystalline cellulose which implied decrease in cell wall stiffness. However, subsequent high-resolution AFM imaging has revealed that DEK1 modulate lines cells walls have their cellulose microfibrils organized in thicker bundles than Col-0. Also, polysaccharide composition analysis has revealed that DEK1 modulated lines have increased abundance of pectins, which could also be responsible for the observed increase in cell wall stiffness. Previous work has shown that different dek1 mutants and modulated lines have defects in cell-to-cell adhesion. This implied that DEK1 may be involved in sensing and/or maintaining cell wall integrity (CWI). We performed several growth assays to determine role of DEK1 in CWI, which is described in Chapter 5. We performed cellulose synthesis perturbation assays with cellulose synthesis inhibitor Isoxaben and obtained very interesting results. While OE CALPAIN plants were hypersensitive to Isoxaben, dek1-4 has shown complete insensitivity. Furthermore, a regular CWI maintenance response, reported in A. thaliana as result of compromised CWI, ectopic lignification in seedlings’ roots was absent in both DEK1 modulated lines we examined. We detected interesting growth response of DEK1 lines to NaCl and mannitol treatments as well. Although these findings are pointing out that DEK1 could be part of CWI signalling pathways, more experiments are necessary to fully elucidate possible role of DEK1 in CWI sensing and/or maintenance pathways, especially to check if DEK1 is interacting with Catharanthus roseus Receptor Like Kinase group of CWI sensors. Studies on 4-month old short day grown DEK1 modulated lines, have shown defects in branching, with development of fasciated stem branches in a DEK1 modulated line overexpressing CALPAIN domain (Amanda et al., 2017). This result pointed out to a possibility that DEK1 may regulate organ morphogenesis and patterning at the level of shoot apical meristem (SAM). Work towards elucidating role of DEK1 in SAM maintenance and organ patterning is detailed in Chapter 6. We determined that OE CALPAIN had significantly larger central zone of SAM as well as larger individual SAM cells in central zone, as well as higher distribution of cell sizes, implying possible cell expansion defects. dek1-4 did not exhibited changes in SAM central zone size or individual stem cell size, but it seemed that it had increased number of stem cells in SAM central zone. Both DEK1 lines had perturbation of phyllotaxis on SAM level, with disturbed divergence angles between floral primordia. Disturbed phyllotaxis was also observed between siliques, in mature plants. In addition to this, OE CALPAIN has exhibited occurrence of multiple (up to four) siliques growing from a single stem node. All this is pointing out that DEK1 might participate in hormone-signalling in the SAM. DEK1 is a highly intriguing protein. However, since it is a unigene, and in addition to that, a regulatory protease, it probably participates in multiple signalling pathways, which makes understanding its function much more complicated.
Adaptations for temperature control in bird eggs: the role of reflectance
Early life stages are highly vulnerable in many animals. In birds, egg temperature must be maintained within an optimal range for successful development and survival. Chilling and especially overheating can lead to abnormal embryos and even be lethal. Bird embryos are effectively ectotherms because heat from incubation by parents is essential for embryo development. Therefore, parental behaviours are crucial for egg temperature regulation during the incubation period. Parents, however, face tradeoffs between incubation and self-maintenance. There are often periods of time when bird parents will leave the nest for foraging or territory defense etc., and eggs will be then left unattended. During this time, egg temperature will not be regulated by parents and will be affected by both extrinsic factors such as environmental conditions (air temperature, solar radiation and humidity) and the architecture of the nest; and intrinsic factors such as eggshell reflectivity and egg size. In this thesis I focused on studying the relationship between these factors and egg thermoregulation. Eggshell reflectivity is one of the most important intrinsic properties that may affect egg temperature. Wisocki et al. (2020) and Gomez et al. (2018) provided evidence that eggs tend to be darker in colder climates and higher latitudes. They suggested that darker colours are adaptive for eggs to retain heat in colder environments for thermoregulation. These studies, however, looked only at ultraviolet-visible wavelengths (300 – 700 nm). These wavelengths correspond to egg colour patterns and may play a role in camouflage, egg recognition, protection from DNA damaging light, in addition to temperature regulation. Due to the multiple potential functions of egg coloration, it can be difficult to isolate the role of specific selective pressures. By contrast, reflectance of near infrared (NIR) wavelengths (700 – 2500 nm) primarily affects temperature. NIR wavelengths account for more than half of the solar energy. In 1978 Bakken et al. (1978) found a uniformly high near infrared reflectance among 25 species (19 of them are ground nesting birds with cryptic eggs) and pointed out the potential importance of NIR reflectance for egg thermoregulation. However, since then there have been no subsequent studies examining the importance of NIR reflectance on bird eggs on a broader scale. In this thesis, I first aimed to study the macro-evolutionary relationships between eggshell reflectivity, climate and nest type in birds. We predicted that eggs would have higher eggshell reflectivity in hot climates, and species with eggs that are exposed would have higher eggshell reflectivity. Using a phylogenetic comparative approach among 186 species of Australian birds we found that eggs tend to have higher total reflectivity in hotter climates with higher solar radiation. Moreover, species with open nests (i.e. with potentially higher exposure to direct sunlight) had higher NIR reflectivity than species with closed nests. This suggests that eggshell reflectivity, especially NIR reflectivity, may play an important role in egg thermoregulation. Second, we investigated how egg size, reflectivity and exposure interact to affect egg temperature. We predicted that high eggshell reflectivity and shade can protect eggs from reaching lethal temperature compared to eggs with low eggshell reflectivity and without shade. We used a biophysical model to predict egg temperatures in a hot environment (Kimberley, Western Australia). The results showed that eggs with higher reflectivity exceeded the critical thermal limit for less time and did not reach temperatures as high as eggs with lower reflectivity. The amount of sunlight eggs are exposed to (shade levels in the model) also strongly affected egg temperature. These results indicate that eggshell reflectivity and exposure to sunlight are likely to be important for egg temperature regulation during incubation, particularly in hot environments. Reflectivity had a greater effect on the temperature reached by large eggs than small eggs, even though small eggs had a higher heating rate. Together, this thesis highlights the importance of eggshell reflectivity on egg thermal control and contributes to our understanding of the macro-evolution of reflectivity in bird eggs.
Characterising gene expression diversity in Indonesia
Genomic studies are disproportionately composed of populations of European ancestry. Given the diverse demographic history of modern humans, this fails to capture the full landscape of human genetic variation. In turn, this has implications for understanding complex disease and accurately assessing disease risk across a diverse range of populations. Indonesia, the world’s fourth largest country by population, is a country that is incredibly ethnically diversity, yet is severely underrepresented in genomic studies. Because of this lack of diversity, this thesis utilises whole blood transcriptomes---a molecular phenotype which can provide information about the activity of a cell and its response to the environment---from three remote island populations spanning the genomic and geographical axes of the country. In the first part of this thesis, I characterise whole blood transcriptomes of these remote populations and investigate contributions of ancestry on molecular phenotypic patterns. The second part of this thesis characterises the ways that these populations respond to their environment, in particular to pathogens in the region. I do this first by investing the effects of malaria, a disease estimated to infect 2.5 million people every year in Indonesia, on Plasmodium-infected individuals. Due to the high pathogenic diversity within the country, I also identify other pathogens within whole blood transcriptomes and explore how they influence human blood expression profiles. By doing so, I find that there are island-level differences correlating with geography, with individuals of Papuan ancestry being most distinct from populations of Asian ancestry. I also find that environmental variables are important contributors to molecular phenotypes, with malaria and flavivirus having clear contributions to shaping the immune response. By characterising molecular phenotypes across Indonesia, this study provides a valuable step in understanding human genetic diversity and genome function outside of populations of European ancestry. It also adds to ongoing literature about how populations differ in their response to disease, ultimately aiding in better diagnostics and therapeutics.
The role of cell wall related genes of plant pathogenic fungus Leptosphaeria maculans in causing blackleg disease of canola
Canola (Brassica napus) is the second largest oilseed crop globally, providing 10-15% of the world’s supply of vegetable oils. In Australia, canola is the largest oil crop and represents over half of total oilseed production in the last 10 years. Blackleg is the major disease of canola, caused by the plant pathogenic fungus Leptosphaeria maculans, restricting canola production worldwide. Yield losses of 50 per cent or even higher have been recorded in some seasons due to blackleg disease. There is an urgent need to reduce the economic damage caused by blackleg disease. The fungal cell wall is the outermost cellular component, thus regulates the host-pathogen interactions like adhesion or phagocytosis, which influences the infection process. The role of cell wall composition in pathogenicity of L. maculans towards canola is unidentified. This study takes a genetics-based analysis of key components in the synthesis of the L. maculans cell wall, by creating mutants of L. maculans with altered cell wall composition to identify pathogenicity genes. The cell wall composition and its role in virulence of L. maculans is unknown. Therefore, based on current information in the literature I selected cell wall genes in other fungi which have shown a role in pathogenicity to identify L. maculans homologs and then utilised a novel CRISPR-Cas9 tool to mutate these genes. Ten cell wall related genes were mutated, including the solo hydrophobin gene (Lema_T007750.1) present in L. maculans and all nine predicted nucleotide sugar transporter genes. The hydrophobin gene mutant exhibited a severe impact on pathogenicity of L. maculans as the mutant could hardly cause any lesion both in canola cotyledon (Brassica napus cv. Westar and Brassica juncea cv. Aurea) and stem (Brassica napus cv. Westar). The hydrophobin mutants had poor conidiation, poor spore germination rate and highly reduced spore attachment capability which likely led to perturbed pathogenicity. It is also hypothesised that hydrophobin plays a role in masking L. maculans from host recognition and to survive under stressed conditions. The substrate specificity of Lema_T023100.1 as GDP-mannose transporter, Lema_T022990.1 as UDP-galactopyranose/UDP-glucose transporter, Lema_T018380.1 as UDP-galactopyranose transporter and Lema_T030160.1 as UDP-galactofuranose transporter were confirmed by performing heterologous expression in a Saccharomyces cerevisiae assay. The substrate of the remaining five proteins could not be determined. Among the nucleotide sugar transporter genes mutants of GDP mannose transporter (Lema_T023100.1), UDP-galactopyranose/UDP-glucose transporter (Lema_T022990.1), Lema_T099510.1 (unknown substrate), Lema_T016830.1 (unknown substrate), Lema_T029030.1 (unknown substrate) and Lema_T030160.1 (UDP-galactofuranose transporter) perturbed the pathogenicity of L. maculans in causing blackleg disease. Mutation of the other genes did not exhibit any impact on L. maculans pathogenicity. In vitro characterisation was performed for all the gene mutants compared to wild type and their complemented strains, that includes growth assay, sporulation, spore germination competency, spore attachment proficiency, ability to endure under stressed conditions as these criterions were considered fundamental pathogenicity factors by mycologists. Though the nucleotide sugar transporter mutants varied in in vitro phenotypes, among all the nucleotide sugar transporter mutants that showed reduced pathogenicity a common feature was highly reduced spore attachment capability. That indicates that in plant pathogenic fungus L. maculans, which does not produce specialised infections structures but directly invades into the host by natural openings, adhesion is the commonly shared mechanism for pathogenesis that is lost in all the non-pathogenic strains. Thus, targeting different stages of the cell wall biosynthesis process in plant pathogenic fungi can be motive for developing novel anti-fungal agents. The outcomes of this research contribute fundamental new knowledge towards the protection against blackleg disease of canola, and potentially other plant diseases caused by fungi.
Anthropogenic Triple Jeopardy: Exploring the effects of multiple stressors on waterbirds
Urbanisation is occurring at unprecedented rates, exposing animals to many anthropogenic stressors. Noise, light and chemical pollution are three major sensory pollutants that often co-occur, and this simultaneous exposure to the mixture of all three could be considered an ‘anthropogenic triple jeopardy’. Yet, the combined effects of these stressors on animals are not well understood. In this thesis, I addressed three important, but previously understudied questions related to the effects of noise, light and chemical pollution on urban waterbirds. First, I reviewed the current state of knowledge and focus of work on the three stressors through a systematic review, to examine the frequency with which these stressors are studied independently or in combination. I compared the results of this literature review to a case study I conducted examining the likelihood of habitats in Greater Melbourne in south-eastern Australia being exposed to one or more stressors at levels likely to cause biological impacts. Melbourne would provide a conservative estimation given that it is relatively less urbanised than many urban centres elsewhere (i.e., 90% of the metropolitan cities have a higher population density than Melbourne). I found that while less than 4% of the research focused on multiple stressors, animals were likely to be exposed to multiple stressors at 33-46% of the habitats around Melbourne, highlighting a mismatch between the research focus and the realistic likelihood that animals are exposed to multiple stressors. This lack of multiple stressor focus was true across a range of biological endpoints, even though individual stressor studies showed that most endpoints were impacted by more than one stressor. Secondly, I tested for individual and interactive effects of noise and artificial light at night (ALAN) on the abundance of a common and widely distributed waterbird, the Eurasian coot (Fulica atra) around Melbourne. I found that coot abundance was individually and interactively negatively impacted by noise and ALAN. There was a detrimental synergistic effect of noise and ALAN on coot abundance, with coots predicted to be absent at the sites with highest noise and ALAN. Thirdly, I explored the interactive effects of noise and ALAN at the community level, testing how the abundance of 30 species varied in relation to the two stressors, and the degree to which foraging and nesting habitat use and behaviour might explain variability in effects. Abundances of 50% of the species (n=15) were impacted by noise or ALAN in isolation, and 60% of those (n=9) were impacted by interactive effects, indicating possible shifts in community composition driven by noise and ALAN. Yet, community parameters (species richness, Shannon-Weiner index, or Simpson’s index) did not capture this change, suggesting the importance of monitoring community effects at a species-level to detect changes by anthropogenic stressors. Predictions based on foraging and nesting habitats did not consistently align with the observed trends, hinting at a myriad of other factors that may influence how species are impacted by noise and ALAN. My thesis provides insight into the interactive effects of noise and ALAN on abundance of birds and demonstrates potential detrimental outcomes on the distribution of individual species as well as community composition through shifts in individual species abundances. My findings also highlight that most current research is not being optimally directed towards the anthropogenic threats and complex forms of exposure that urban animals currently experience.
From nucleotide sugars to polysaccharides: How do plants control the delivery of substrates for cell wall biosynthesis and protein glycosylation?
Plant cell walls constitute one of the most abundant raw biomaterials on Earth. The synthesis of long-chain olysaccharides, the main components of plant cell walls starts ab ovo in the cytoplasm where most of the building blocks for polysaccharide synthesis, so-called nucleotide sugars, are produced. The monosaccharide moieties of nucleotide sugars are incorporated into growing polysaccharide chains either directly at the plasma membrane by lycosyltransferases (GTs) that form cellulose synthase complexes or by those residing in the Golgi apparatus. In the latter case, nucleotide sugars have to pass the Golgi membranes with the help of nucleotide sugar transporters (NSTs). Once inside, they are used by Golgi GTs which assemble polysaccharide chains and decorate proteins and lipids with sugar residues. Recent evidence suggests that in plants nucleotide sugars can be guided to specific polysaccharides and/or glycan decorations, yet the molecular mechanisms of these processes are not fully understood. This PhD research attempts to explore the phenomenon of the targeted substrate delivery by investigating two possible hypotheses: the spatiotemporal distribution of proteins within the Golgi apparatus and the occurrence of direct interactions between NSTs and GTs. The author of this dissertation has tested both of those hypotheses by investigating protein-protein interactions, localising the individual components to their specific sub-compartments within the cell and tracking changes in mutant plants where these processes are modulated. The bifunctional UDP-RHAMNOSE/UDP-GALACTOSE TRANSPORTER (URGT) family from the model plant Arabidopsis thaliana was selected for this study. While this family has six members which in vitro are capable of transporting the same substrates, plant mutant studies indicate substrate preference and targeted delivery to specific cell wall components in vivo. This thesis presents the first evidence that members of the URGT family localise to specific sub-Golgi compartments. The colocalisation studies undertaken as part of this thesis place URGT1 and URGT5 in the cis-Golgi, URGT2, URGT4 and URGT6 – in the medial Golgi, while URGT3 seems to localise to trans-Golgi stacks. Protein-protein interactions studies have identified multiple interaction partners for the six URGTs. Notably, many of those are known galactosyltransferases, which aligns with the transport function of the URGTs. It is therefore highly likely that the identified candidates are true interactors, which use the proximity of the transporter to increase the process efficiency by substrate channelling. This finding is further supported by the fact that observed interactions between URGT family members and GTs often localise to the same sub-Golgi compartment. The study identified new potential galactosyl- and rhamnosyltransferases, including two putative arabinogalactan protein galactosyltransferases. The data obtained during this project suggest, that URGTs may determine the flow of substrate through both spatial separation within the sub-Golgi stacks and direct interactions with GTs. The study presents new insight into the sugar substrate delivery processes in plants, suggests that similar processes may take place in other NST families and that their specificity may be similarly tuned by localisation and interactions.
Persistence of marine species of the Australian South-east Marine Region under alternative fisheries management strategies
The marine environment is heavily affected by fishing pressure and knowing how different management strategies affect the persistence of marine species is crucial for ensuring sustainable fishing. Models used within the fisheries management strategies evaluation context have been implemented in the past decades from traditional single species methods to an ecosystem-based approach, but there are still uncertainties on the effectiveness of these models. This thesis aims to understand the drivers of marine species persistence coupling network-based methods and spatially explicit population viability analyses (PVA). This thesis focuses on marine species of south-eastern Australian waters, a region characterised by rich biodiversity and oceanographic complexity, supporting important Australian commercial fisheries sectors. PVA is an approach not yet widely investigated in the context of fisheries management strategies evaluation, however understanding species viability and ensuring metapopulation persistence is fundamental to avoid overexploitation of the stocks. In this thesis is examined how spatially explicit PVA compares to other commonly used models, demonstrating that PVA is a powerful alternative, suitable to inform fisheries management. PVA is applied to estimate metapopulation persistence in a spatial context, analysing how persistence changes when modelling alternative fisheries management scenarios for important commercial fisheries species. The knowledge of species distribution is very important for any spatially explicit population modelling and is needed when studying population viability and dynamics. A recent challenge in understanding and predicting species’ distributions has been focused on the influence of population connectivity. This thesis explores how to integrate graphbased metrics, representative of seascape connectivity, into marine-based species distribution models (SDMs) to understand the contribution of connectivity to predict marine species spatial distribution. Connectivity is a key determinant of metapopulation persistence of species inhabiting fragmented habitats. Larval connectivity is quantified using a biophysical model. Connectivity is visualized as networks, where habitat patches are represented by nodes and dispersal connections are represented by linkages. In this thesis graph-based metrics are calculated because of their significant role as indicator of metapopulation persistence, identifying key habitat patches for the overall persistence of the species. The results of this thesis demonstrate how PVA is a valuable tool helping to inform fisheries managers on the effects of fishing on marine species. Habitat heterogeneity and movement ecology demonstrate to be critical model parameters, strongly influencing metapopulation persistence, suggesting that fisheries managers might benefit from better understanding of movement behaviour and habitat characteristics. This thesis results also provide insight on the role of connectivity to determine marine populations persistence and distribution. Hotspots of connectivity reveal to be key habitats, often enhancing habitat suitability, and strongly influencing metapopulation persistence. Managers and ecologists would benefit by employing similar approaches in making more efficient and more ecologically informed decisions and focusing more on local connectivity patterns to better understand and protect marine species.
Quantifying biodiversity and maximising species persistence during offsetting
Biodiversity is declining globally, with habitat clearing playing a major role in perpetuating these declines. Offsets aim to achieve no net loss of biodiversity by implementing conservation actions that deliver equal biodiversity gains to those lost through development. Broad habitat-based biodiversity metrics are commonly used to quantify development impacts and assign offset requirements. However, it is unclear what reliance on habitat-based offset metrics means for biodiversity long-term. In this thesis I explore the current state of biodiversity metrics within the scientific literature and how these metrics are applied in policies around the world. I also quantitatively test nine biodiversity metrics and explore the long-term ramifications of their use in offsetting on species persistence. In a sample of 255 publications from ecology (n = 158), conservation planning (n = 54), and offsetting (n = 43), I identify 24 categories of metrics used to quantify biodiversity. The offsetting literature focused largely on habitat attributes and area-based approaches for measuring biodiversity. Similarly, consistent use of habitat attributes, habitat condition, and area in 21 offset policies demonstrates that offsets do not adequately capture individual species or communities. Many offset policies measure species presence, however, less than half of those I reviewed measured any species-specific attributes which could be used to help inform conservation actions for those species. These two reviews demonstrate that the metrics currently used to measure biodiversity don’t reflect some of the key biodiversity attributes offsets aim to protect. To understand the long-term consequences of relying on such habitat-based metrics for offsetting I developed a spatial simulation tool and present a methodological framework that quantitatively tests the effects of metric choice on long-term persistence. The R-based simulation tool uses raster data to simulate development impacts and restoration efforts. I tested nine biodiversity metrics spanning an axis of pattern versus process and capturing different levels of biodiversity: area of habitat, condition of habitat, vegetation area, vegetation condition, area * habitat suitability, condition * habitat suitability, abundance, metapopulation connectivity and rarity-weighted richness. The resulting raster outputs are used in population viability analyses for five species. Most metrics achieved no net loss or net gains in the metric values, species’ habitat suitability or abundance, but only when impacts avoided high suitability areas. When impacts cannot avoid high suitability areas, species-specific metrics deliver higher gains in abundance per hectare restored than habitat-based metrics alone. Failing to improve current offset practices ensures species will continue to decline under development-offset trading programs and should provide an incentive for policy makers to properly enforce impact avoidance measures where possible.
Birds in the sky, fish in the sea, money in the bank: quantitative methods for more effective conservation
My approach in this thesis was to explore how to wring more information out of existing data to reduce uncertainty, improve decision-making and hope to generate better conservation outcomes. I explore and develop a range of quantitative tools to this end. I look at three key areas: dealing with uncertainty, structuring decision making, and improving the use of existing information. I consider these concepts over three thematic case studies: monitoring the abundance of three vulture species in Cambodia, trading-off the costs and benefits of releasing information publicly when a new species or population is discovered, and comparing use of optimisation and project prioritisation protocols to allocate funding to species conservation efforts. In the first case-study, I develop new Bayesian hierarchical model to estimate vulture abundance, and compare the inferences available from this approach with less specialised approaches previously used. In the second case-study I develop a decision-making framework to allow decision-makers to explicitly trade-off costs and benefits, and apply the method to data collected from informants who have made these types of decisions themselves. In the final section, I explore whether additional information can improve optimisation to allocate funding, and compare performance in terms of expected avoided extinctions of the optimisation approach with a project prioritisation protocol. I find that there is indeed much more we can learn from the information we have. But this is not a free lunch – work needs to be done to uncover opportunities, and technical skills are often needed to make best use of them, and assumptions must often be made to draw conclusions.
The impact of beneficial microbes on root architecture and metabolism of the model grass Brachypodium distachyon grown under non optimal temperatures and low phosphorus availability
Cereal crops are exposed to different abiotic stresses thus impacting agricultural productivity, but soil beneficial microorganisms have the potential to improve the adaptation of crops to suboptimal environments. This study resolves the time-specific dynamics between the plant growth promoting bacteria Azospirillum brasilense and the model cereal species Brachypodium distachyon grown under suboptimal temperatures and low P availability. Results contribute to our current understanding of the phenotypic and biochemical adaptative rearrangements occurring in plants during interaction with beneficial bacteria.
Understanding and incorporating aphid parasitoids within IPM strategies in Australian grain crops
Aphids (Hemiptera: Aphididae) can be particularly devastating to grain crops, with their economic importance weighted on their ability to cause significant yield losses through a variety of methods. From feeding damage alone in 2012, cereal aphids caused an average annual loss of $14 million in Australian wheat crops. For over a century, growers have relied upon host plant resistance and chemical treatments to control invertebrate pests, however suppression of beneficial organisms and increased resistance within targeted species has created an ongoing battle with pest control. For example, the polyphagous green peach aphid (Myzus persicae (Sulzer)), often a pest of canola crops, has developed resistance to over 74 insecticides including carbamates, pyrethroids, and organophosphates around the world. Due to these issues, control of agricultural pests is now focussed on Integrated Pest Management (IPM) strategies, within which natural enemies can play a role as biological controls. Parasitoid wasps have had the most success as biological control organisms in the past, likely due to their host specificity. I spent three years collecting data on grain aphid pests and their associated natural enemies, paying particular attention to the hymenopteran parasitoids. I determined the distribution of grain aphids and their associated Aphidiinae within grain production landscapes around Australia, utilising historic data and citizen science. Additionally, I determined how aphid abundance and diversity, along with their associated parasitoids changed throughout the growing seasons. I created a key of aphid parasitoids (Hymenoptera: Aphidiinae) parasitizing aphids in Australian grain production landscapes. Finally, I determined the effects of seed treatments on specific natural enemies associated with M. persicae, identifying the difference between parasitoid and predator effects. My findings are informative for developing strategies to conserve those Aphidiinae species of particular importance in controlling aphid pests. Additionally, these results can assist with pest management decisions, enabling growers to implement IPM based on a greater breadth of knowledge.