Current environmental EU legislation promotes recycling and recovery from organic waste products. Compost has been identified as an alternative to inorganic fertilisers and animal slurries as a nutrient source for crop plants. This study aimed to investigate nitrogen (N) and phosphorus (P) availability from various composted waste through detailed characterisations, complemented by short term lab incubations and long term plant growth experiments. Twenty-five composts were selected and classified by their groups. The composts were characterised by multiple different analytical techniques. Two incubation studies were conducted. One investigating N and P mineralisation potential of the composts and the second on the effects or soil type on P mineralisation. Two large glasshouse based pot experiments were then conducted. A subsequent stability study of two separate methods on mechanical biological treated waste was also investigated for a potential relationship. Characterisation of the composts highlighted the difference between compost feedstocks with the manure waste being distinctly separate from the other groups. The biowaste groups identified as either food waste or brown bin waste. The incubation experiment highlighted that N mineralisation was more predictable than P. From the pot experiment only manure waste mineralised N above 10%. All of the other groups mineralised minute amounts. Prediction of N mineralisation was found to be more accurate from lignin and neutral detergent fibre content over the more traditional C/N ratio. Mineralisation of small amounts of organic N occurred in summer months. There was a distinct lack of a relationship between the P incubation and pot experiment. Plant uptake of P was higher than expected for all treatments. The manure waste composts were higher in cumulative P uptake than inorganic fertiliser. Stability in composts was found not to affect mineralisation or induce immobilisation. The effects of soil type on composts were most pronounced in composts lower in fulvic acid. The composts with higher humic acid were not found to significantly mineralise a higher amount of P. The stability study investigating oxygen uptake rate (OUR) and respiration index (AT4) on the mechanical biological treated waste showed a high correlation between the two methods where AT4 was below 40 mg O2.g DM-1. The findings point to a possible reassessment of compost quality guidelines for composts. The availability of P is far greater and therefore should be considered the primary nutrient available from compost. The stability parameter for manure waste compost may also need assessing as it is unachievable in reasonable composting time frame. The biowaste compost has proven to be high in P. There is also the potential to develop quite specifically produced composts based on their feedstock and composting length for particular agronomic or environmental needs. Compost application is a feasible source of nutrients and organic matter due to escalating fertiliser prices and more stringent regulations surrounding fertilisers and slurries.

The need to develop forest management systems other than clearfelling has resulted in a requirement for improved understanding of the potential of continuous cover forestry (CCF). One suggested method for the conversion of forest stands into CCF systems and for bringing under-performing forests into productivity is thinning in conjunction with underplanting. This study was an attempt to provide information on species suitability for underplanting of two important trees in European forestry: pedunculate oak (Quercus robur L.) and European beech (Fagus sylvatica L.). To determine the morphological, physiological and growth responses of these two species to different light conditions, beech and oak seedlings previously grown at full light for two years were covered by shading nets that provide different shade levels (62%, 51% or 28% of full light) or continued to be exposed to full light. The different shade levels were intended to mimic a range of underplanted conditions and the process of acclimation to shade was studied to provide information on the ecology and adaptation of underplanted seedlings. In addition to the controlled-shade experiment another study to determine the physiological responses of beech natural regeneration to shade was conducted under natural light conditions (from open gaps to closed canopy). Both oak and beech displayed similar acclimation in response to shade for most of the traits investigated. At the plant level, seedling acclimation to shade included higher biomass allocation to above than below-ground parts and greater energy investment on height than diameter growth. At the leaf level, seedlings grown under shade reduced their leaf thickness and photosynthetic rates per unit area and increased their specific leaf area. This increase in specific leaf area seems to be one mechanism that allows seedlings to perform well under shade conditions. Another acclimation to low light conditions was to increase the efficiency of the photosystem II under shade. Photosynthetic rates were higher and leaves were retained for longer in seedlings grown at full light than under shade. Hence, this probably led to a greater growth in the full light than under shade. Despite this greater growth at full light, the results of this study suggest that beech and oak seedlings would be able to acclimate and perform well if underplanted below overstories that reduce the available light to as low as 28% of full light without having any significant adverse effect on the quality of the final crop.

In Ireland, the pig industry is concentrated in a small number of counties. Pig farms typically have a high stocking rate. Therefore, the disposal of slurry in a cost-effective and environmentally-responsible way is a serious issue for farmers. Slurry is commonly applied to land, but this may not be possible if the land is at, or approaching, phosphorus (P) saturation. As pig farmers dispose of slurry in the vicinity of their properties, most of the nearby land is at P saturation, so alternative treatment methods need to be utilised (e.g. constructed wetlands, anaerobic digestion, filtration) or the slurry needs to be transported to another location. These alternatives are not currently financially viable in Ireland. Existing legislation (S.I. 610 of 2010) and recent changes in the implementation of legislation governing the timing and quantities of slurry that may be applied to land, means that pig farmers will no longer be able to exceed the maximum legal application rate to land (from January 2017). European policy aiming to intensify pig production will only accentuate this problem. If pig farmers are forced, in exceptional circumstances, to land apply slurry to unsuitable land, surface and subsurface losses of nutrients and suspended solids (SS) may occur. This could be potentially problematic if the land is located in a critical source area (CSA), an area that is highly likely to pollute receiving waters. In these circumstances, a possible novel solution is to chemically amend the pig slurry prior to landspreading. This would mean that pig farmers may, in exceptional circumstances, utilise the land in the vicinity of their farms for landspreading, without releasing excessive nutrients and SS into receiving waters. However, knowledge gaps exist concerning the type of amendments to be used, the characteristics of the soil on which they can be most effectively used, and their impact on incidental (short-term) and chronic (long-term) losses of nutrients, SS and greenhouse gas (GHG) to surface and subsurface water and the atmosphere. Therefore, the aims of this project were to: (1) identify the most appropriate chemical amendments, and their addition rates, to reduce P losses in runoff from pig slurry based on effectiveness, cost and feasibility; (2) investigate the impacts of these chemical amendments on nutrient losses in leachate, soil properties and GHG emissions; and (3) identify suitable soil types on which to landspread chemically-amended pig slurry. Laboratory bench-scale experiments were designed to identify the amendments which had the potential to reduce P in overland runoff and to quantify the stoichiometric rates at which to add them to the slurry. Based on effectiveness, cost and feasibility, the amendments identified were alum, which reduced dissolved reactive phosphorus (DRP) in overlying water by 86%, poly-aluminium chloride (PAC) (73%) and ferric chloride (FeCl3) (71%). Following these bench-scale experiments, rainfall simulation experiments were conducted to quantify the impact of chemical amendments to slurry on surface runoff losses at various time intervals from the time of application. Poly-aluminium chloride performed best in these experiments. For the first time, the effect of these amendments on GHG emissions, soil properties and leachate was also examined. Chemical amendment did not adversely affect GHG emissions, soil properties or leachate from pig slurry, but FeCl3 increased nitrous oxide (N2O) and carbon dioxide (CO2) losses. Finally, a 3-mo incubation experiment was conducted using a range of soil types to examine the effect of amendments on the long-term plant availability of P in soil and P solubility. Alum reduced more water extractable P than PAC, but also resulted in less plant available P. Considering cost, surface runoff and subsurface leachate losses, GHG emissions and impacts on soil chemistry, PAC was found to be the most suitable amendment with which to chemically amend pig slurry. There is the potential, in combination with existing programmes of measures, to employ chemical amendment as a measure to mitigate the environmental impact arising from the landspreading of pig slurry. This should be conducted in targeted areas of the CSA and should take into account soil type and its chemical properties. Before implementation, these tests must first be validated in long-term testing at field-scale over a wide variety of soil types, and include repeated application and incorporation. At present, there is no provision in legislation for chemical amendments to be used as a mitigation measure in the land application of pig slurry, but if they are to be utilised, a regulatory framework will need to be introduced by the relevant bodies.

Perennial ryegrass (Lolium perenne L.) is the most important forage grass of temperate regions of the world. The main objective in breeding perennial ryegrass cultivars is to increase its biomass. Chloroplasts and mitochondria are two organelles of the plant cell that are actively involved in biomass production. Chloroplasts derive from cyanobacteria and are the location of photosynthesis in plant cells. Mitochondria derive from α-proteobacteria and are involved in cell respiration. Due to their evolutionary history both organelles still contain their own genome which is in general maternally inherited. The interest in chloroplast genome sequences increased in recent years because they offer a useful option for plant genetic engineering. The risk of transgene escape via pollen flow is reduced while the expression of the transgene due to the high number of chloroplast genome copies is increased (in comparison to nuclear genome transformation). Mitochondrial genomes are of special interest because they are involved in cytoplasmic male sterility. Cytoplasmic male sterility is a very important trait in plant breeding programmes because it enables the cost efficient production of hybrid seed. Additionally, both organelle genomes can be used for molecular evolution or phylogenetic studies, as well as for population genetic approaches. Therefore the major aim of this thesis was to sequence the entire chloroplast and mitochondrial genomes of L. perenne to provide sequence information for chloroplast genetic engineering approaches, insights into the mitochondrial genome of a male fertile L. perenne cultivar and to gather knowledge about sequence variation in both genomes that can be used to design new markers for phylogenetic and population genetic studies.

Miscanthus is a highly important forage and horticultural genus of perennial grasses (Poaceae) primarily native to South East Asia. Miscanthus is under intense global investigation as a biomass source for renewable energy production and several breeding initiatives are underway to develop new genotypes optimized for improved biomass and tolerance to a range of environmental stress conditions. A collection of 128 accessions belonging to the genus Miscanthus was established in Oak Park, Teagasc, Carlow, in 2008 and was investigated for morphological and molecular variation. Morphological traits were measured at the end of the second growing season and were compared with herbarium specimens of Miscanthus. Vegetative and inflorescence traits were scored and analysed using basic summary statistics, tests of normality and Principal Components Analysis (PCA). A large degree of morphological variation was recorded in the collections. The PCA of herbarium specimens was able to separate some species from others but there was also considerable overlap among species in the ordination, especially M. sacchariflorus, M. sinensis, M. condensatus and M. floridulus. These are known to be closely related and can interbreed. The PCA of the specimens from the Oak Park collection was less informative because of missing data due to lack of inflorescences (accessions did not flower). It was clear that morphology alone is often insufficient to distinguish taxa especially when inflorescence characters and ploidy information is lacking. The ploidy level of the accessions in the collection was evaluated through flow cytometry. The ploidy included di-, tri- and tetraploids. All individuals labelled as M. ×giganteus showed a triploid status, together with the newly bred M. sacchariflorus×M. sinensis hybrids. Most M. sinensis were diploids. Miscanthus sinensis Tea-62 was triploid and comparable to the value of the M. ×giganteus. A different situation was found for other non-diploid M. sinensis, in particular four M. sinensis ‘Goliath’ and the M. sinensis ‘Zebrinus’ Tea-33. In these the ratio measured by the flowcytometer was in between the values of the triploid M. giganteus and tetraploid M. sacchariflorus standards. The ‘Goliath-like’ hybrid is likely an autotriploid with three M. sinensis haploid sets, whereas M. ×giganteus is an allotriploid that is supposed to have two genomes from M. sinensis and one from M. sacchariflorus, which has a lower amount of DNA per haploid genome. DNA sequences of the internal transcribed spacer of the nrDNA were obtained for 76 genotypes in the collection and compared for polymorphism. The SNPs were particularly VI useful for differentiating M. sinensis, M. sacchariflorus and M. ×giganteus accessions and in combination with ploidy and morphology offer high potential for taxon identification. To gather more markers for population level diversity and differentiation studies, new microsatellite markers for both plastid and nuclear genomes were developed. For the development of plastid markers the chloroplast genome information of Saccharum officinarum was used. The nuclear SSRs (nSSRs) were developed from the sequences of 192 clones obtained from microsatellite enriched library. New primer pairs for the amplification of nineteen nuclear loci and six chloroplast loci were developed. Both chloroplast (cpSSR) and nSSR primers were used to characterise DNA variation, to help establish gene pools and to better understand hybridization and introgression. Huge genotypic variation was found within the genus, mostly in the species M. sinensis. The markers showed wide utility across a large number of Miscanthus species and also some closely related genera. The analysis of the cpSSRs showed a high number of different haplotypes but with a clear bias in allele composition between M. sinensis and the two species M. sacchariflorus and M. ×giganteus, thus confirming M. sacchariflorus as the maternal lineage of the hybrid M. xgiganteus. The nSSRs were found to be highly polymorphic across the collection and transferable to closely related genera such as Saccharum. The new markers were also used in UPGMA clustering and Bayesian structuring analysis to group individuals according to their similarity. Three major clusters of individuals were defined using the Bayesian STRUCTURE analysis with nuclear markers (nSSRs) and two with plastid markers (cpSSRs). In conclusion, the morphological, ploidy, sequence and microsatellite results highlighted the high level of diversity still unexplored in the genus and have clarified taxon identity of many accessions in the collection. A large set of new markers have been developed for the plant breeding and systematics community. The newly developed markers will be useful to further explore this diversity and to select useful traits for breeding of new and improved genotypes for biomass production.

Perennial ryegrass (Lolium perenne L.) is a member of the Poaceae family, is native to Europe, the Near East and North Africa and is grown in all the temperate climate areas of the world as a forage and turf grass. Due to its persistence, palatability and nutritive value for ruminants, it is a principal component of pastures, and the most important forage species in Ireland. The primary aim of this thesis was to characterise the level of diversity in a large genetic resource collection of L. perenne germplasm held at Teagasc, Oak Park. Molecular markers, both chloroplast and nuclear SSRs, biochemical characters (water soluble carbohydrate, crude protein, and dry matter), and morphological characters (vegetative and flowering) were used to characterise this diversity, as well as population differentiation, and geographic patterns. Levels of diversity in all systems were found to be high in this collection. Primers to amplify microsatellite markers from the chloroplast genome of Lolium perenne were designed and optimized using de novo sequencing and in silico sequences. With one exception, each locus was polymorphic with a range from two to nine alleles in L. perenne. The newly developed primer pairs cross-amplified in different species of Lolium and in 50 other grass species representing nine grass subfamilies. These markers were then used to characterise chloroplast genetic diversity at allelic and haplotypic level in 104 accessions of Lolium perenne, other Lolium species, Festuca species and ´Festulolium cultivars. Furthermore, genetic relationships between the accessions and biogeographic distribution of haplotypes were investigated using a range of population genetic diversity measures and an Analysis of Molecular Variance (AMOVA). An extremely high number of 511 haplotypes was detected in 1,575 individuals possibly attributable to natural and anthropogenic migration. Much of the L. perenne European ecotype diversity (61%) could be attributed to within population variance. Plastid gene pools and maternal lineages for L. perenne could be clearly identified. Evidence was found showing a most likely migration route of L. perenne into Ireland from southern regions of Europe northwards. 5 Morphological variation of 13 vegetative and reproductive traits was characterised for 2,481 individuals from 50 L. perenne accessions, a mixture of Irish and European ecotypes and cultivars. Considerable levels of among and within population variation was found across traits. Principal component analysis and UPGMA dendrograms were able to separate ecotypes from cultivars. Cultivars generally had later dates of ear emergence, better spring and summer growth, longer rachis length and more spikelets per spike than ecotypes. Correlation and regression analysis were used to assess relationships between traits and strong positive relationships were seen between reproductive characters, i.e. rachis length with spikelets per spike, florets per spikelet and glume length. The strong relationship between rachis length and the other reproductive characters suggested that rachis length could be used as a predictor for reproductive performance. Later flowering was correlated with improved spring and summer growth. Water soluble carbohydrate (WSC; glucose and fructose determined by HPLC), crude protein (determined via LECO analysis), and dry matter contents were recorded for 1,320 individuals, pooled into 132 samples from 33 L. perenne ecotypes and cultivars at five different harvest time points across the 2004 growing season. While, in general, the cultivars had higher WSC contents than the ecotypes, individual ecotypes did show potential to be used in breeding programmes, as they showed higher values than all other accessions at particular cutting points. In correlation analyses, positive relationships were shown between dry matter and glucose both early and late in the growing season, and this was in agreement with the amount of leaves compared to stem at these times in the growing season. PCA analysis allowed the separation either between cultivars and ecotypes, or between tetraploid cultivars and the rest of the accessions at four out of five cutting points. In the ANOVA analysis, cutting point was the most significant factor influencing the variation in the traits. Eight nuclear SSR markers were used to characterize genetic diversity in 928 individuals from 40 diploid ecotypes and cultivars of L. perenne. High levels of genetic diversity (0.82, Nei’s gene diversity, over all accessions) and high numbers of alleles (22.25 average number of alleles per locus) was found. An average polymorphic information content (PIC) value of 0.81 across all loci was found. When deviations from Hardy-Weinberg equilibrium were tested, the majority of populations 6 had an excess of homozygotes. Very low levels of linkage disequilibrium were found between pairs of loci tested. AMOVA analysis and F statistics were used to test partitioning of variation, and most variation was found within populations (e.g. 31% for glume length in ecotypes). UPGMA, PCA and STRUCTURE analysis all gave similar patterns of relationships between populations, where relationships with high bootstrap support on the UPGMA dendrogram were also seen in the other analyses. The overall results of the thesis are discussed in the context of plant breeding programmes and natural population genetic variation. Strategies for incorporation of the results of the thesis (and the novel markers developed within) into plant breeding programmes are suggested.