Tepary (Phaseolus actifolius Gray) beans perform better than common beans (Phaseolus vulgaris) under drought conditions. The mechanism of drought tolerance in tepary bean seedlings was explored by determining the water potential (WP), osmotic potential (OP), relative water content (RWC), free sugar concentration, and the level of K ion within the expanding leaves. Two week old tepary bean seedlings were subjected to a gradual water stress with sorbitol solutions exhibiting OP values of -0.19 MPa and -0.47 MPa. Turgor remained constant whereas WP, OP and RWC declined following the stress treatment. Osmotic adjustment (OA) occurred in each treatment but the contribution of sucrose and fructose was minor. Some sorbitol was translocated to leaves and contributed to OA. The decrease of K ions in leaves indicated that these ions did not contribute to the OA. A significant decrease in cell size was observed as determined by decrease in TW/DW ratio.

Flowering Locus T (FT ) plays a pivotal role in floral induction. It integrates the inputs from a complex network of flowering signalling pathways. Flowering is an efficiently orchestrated event that occurs in a plant at a particular time to ensure maximum reproductive success. It has been suggested that the FT protein is a long- distance mobile floral stimulus. In this report studies with a mutant version of FT (mFT) which had the start codon replaced with a stop codon to generate a non-translatable FT indicated that the mRNA was also capable of long distance movement although its physiological function as a floral stimulus was inhibited. Gene function study of FT and FT orthologues on brassica, tobacco, tomato and potato using the plant virus expression vector Potato Virus X (PVX) generated some interesting findings. In Short day Maryland Mammoth tobacco plants the overexpression of the Arabidopsis FT under non-inductive Long day condition induced early flowering while the mFT and mock control remained in the vegetative stage. In short day potato, it did not seem to have an effect on tuberization as only one from five of the inoculated plants tuberized. In brassica (broccoli) the effect on flowering time was inhibited due to Virus-induced Gene Silencing (VIGS) but the tomato FT (SP6A) had an effect on flowering time. In tomato, the overexpression of the Arabidopsis FT and FT- orthologues from tomato induced early flowering but the difference in flowering time in comparison to the controls was only a few days. Phenotypical and morphological changes such as seed production and lateral side shoot development were caused by expression of the target genes. The exact mechanism of action of these genes in the control of seed production and lateral side shoot development is unclear.

Crop protection products (CPPs) are an essential component of modern agriculture, necessary to improve crop yield to feed the ever-increasing world population. Regulation and safety testing of CPPs entering the environment is mandatory to ensure that their use is not at the detriment of environmental or human health. Regulatory laboratory studies typically over-estimate the persistence of CPPs within the environment as they are not representative of environmental conditions. This study investigated the role of non-UV light on CPP degradation and the development of soil surface communities. The inclusion of non-UV light in laboratory studies impacted the degradation of fludioxonil and cinosulfuron, increasing and decreasing the rate of transformation relative to dark conditions, respectively. Further, the inclusion of light increased non-extractable residues (NER) formation in fludioxonil, paclobutrazol and benzovindiflupyr. In a field based degradation experiment, the availability of photosynthetically active radiation (PAR) increased the transformation of benzovindiflupyr relative to when PAR was restricted. Further, the formation of paclobutrazol and benzovindiflupyr NERs was increased when PAR was not restricted, and the proportion of CPP remaining at the soil surface (0-5 mm) was higher when PAR was restricted. Targeted amplicon sequencing (Illumina MiSeq) revealed that bacterial and phototrophic communities at the soil surface changed with time, and that communities formed when PAR was available were structurally distinct relative to communities when PAR was restricted. In a further experiment, analysis of bacterial and phototrophic communities under crops with differing canopy characteristics showed that distinct communities formed at the soil surface relative to bulk soil, and that phototrophic communities of bare soil and under low-density canopies were structurally distinct to those that formed under high-density canopies. This work has potential implications for regulatory CPP degradation studies, and furthers the understanding of soil surface community development in temperate environments.

There has been an increasing emphasis placed on understanding microbial c, in order to enable the patterns and processes governing the spatial distribution of soil microbiota to be determined. Due to current food security issues, this is particularly important within agricultural systems given the fundamental role microorganisms play in the maintenance of crop health and productivity. With evidence in favour of both ubiquity and endemism, complicated by systems, scales and communities, there is a need to address the question of microbial biogeography within a single system. A range of field experimental resources were used to investigate factors controlling the assembly of soil microbial communities. Microorganisms across all three domains of life demonstrated spatial scaling, in which there was no single universal driver. Land-use management was an important driver of eukaryote distribution, but also impacted the drivers of bacterial and eukaryote taxa groups under land-use practice. When considering microbial community structure, a pan microbial relationship between abundance and distribution was shown for the first time, across all microbial groups. Furthermore, partitioning microbial communities into common and rare groups provided information on the processes operating on the community and highlighted the importance of land-use management for shaping the structure of communities. Finally, a case study on plasmodiophorids increased current estimates of plasmodiophorid diversity in the soil. Also different communities were associated with the rhizosphere compared with the bulk soil, under different hosts. Plant development stage was also an important consideration acting on this previously understudied but highly significant group of protists to crop health.

With the estimated increase in global demand for food and over-reliance on staple food crops, the exploitation of agricultural biodiversity is important to address food security challenges. The aim of this study is to develop approaches to transfer major informational and physical resources developed in model plant and major crop species to resources poor crop species, using oil palm and Bambara groundnut as two exemplar crops. XSpecies (cross-species) approach, the core approach of the study, is described as the approach which uses microarrays developed for a given species to analyse another related species. The use of the XSpecies approach (here the cross-hybridisation of DNA from oil palm onto heterologous Affymetrix microarrays for Arabidopsis and rice), is the first experiment reported in oil palm and focused on a bulked segregant analysis of different shell-thicknesses for oil palm fruit. Primers design involved screening candidate probe-pairs filtered using PIGEONS software against oil palm transcriptome sequences generated using 454 sequencing technology. The results provided an insight into the effects of sequence divergence between oil palm and the reference species (Arabidopsis and rice) onto the power of detecting single feature polymorphism (SFPs) in oil palm, implying the importance of close association between studied and model plant/crop in XSpecies approach. The XSpecies approach coupled with genetical genomics was also tested within legumes, with Bambara groundnut as the query species compared to soybean as the resource rich species (20 Mya). A mild drought experiment, conducted in a controlled environment glasshouse, used an F5 segregating population derived from a controlled cross between DipC and Tiga Nicuru in Bambara groundnut. The cross-hybridisation of Bambara groundnut leaf RNA to the soybean GeneChip individual oligonucleotide probes resulted in a total of 1,531 of good quality gene expression markers (GEMs) on the basis of the differences in the hybridisation signal strength. The first ‘expression-based’ genetic map (GEM map) was constructed using 165 GEMs spanning 920.3 cM of Bambara groundnut genome. The first high density DNA-marker genetic map of 1,341.3 cM combining dominant DArT and co-dominant SNPs, developed using the DArT Seq approach, with additional pre-existing microarray-based DArT and SSR markers, was also developed in the F3 segregating population. Both maps were combined to form the first integrated map of 1,250.7 cM with 212 markers. Morphological differences and the rapid reduction in stomatal conductance observed within the F5 segregating population in the drought experiment provided trait data for a QTL analysis. The comprehensive QTL analysis in Bambara groundnut detected significant QTLs for morphological traits using GEM map, including internode length, peduncle length, pod number per plant, pod weight per plant, seed number per plant, seed weight per plant, 100-seed weight, shoot dry weight and harvest index across four linkage groups: LG1, LG2B, LG8B and LG11A. The loci controlling internode length and peduncle length were also consistently mapped to single marker on LG1 in DArTseq map using F3 segregating population, suggesting that these two traits are probably controlled by single gene or two closely linked genes. Despite significant genotypes effects on stomatal conductance tested in ANOVA analysis, no major QTLs were detected, suggesting the contributions of a number of small genetic effects to stomatal conductance. A preliminary homology search using the LG1 linkage group markers and associated gene models showed the ability to develop a framework for identification of candidate genes in Bambara groundnut relative to soybean. The present study also developed the resources for an eQTL analysis in a cross-species context. Translation from major and model plant species to underutilised and resource poor crops is critical to be able to develop many crop species with potential for future agriculture. This study examines some of the approaches which might be adopted and replicated in various underutilised crop species.

Oil palm (Elaeis guineensis) produces over five times more oil/year/hectare than oil seed rape and accounted for 33% of world vegetable oil production in 2011. Being a cross-pollinated perennial tree crop with long breeding cycles (typically 12 years) and a large planting area requirement (usually 143 palms/hectare), utilization of molecular technology could greatly improve the efficiency of oil palm breeding. In the present study, various approaches were used to develop molecular markers for genetic linkage mapping and QTL analysis, with the ultimate goal of marker-assisted selection in oil palm. Firstly, Representational Difference Analysis (RDA) and Amplified Fragment Length Polymorphism (AFLP) were coupled with Bulked Segregant Analysis (BSA) to try to identify marker(s) closely linked to the important shell-thickness gene. A novel combination of RDA with Roche 454 pyrosequencing enabled a more comprehensive study of the enrichment profiles compared to Sanger sequencing. Identification of >35% redundant sequences, repetitive sequences and organelle DNA suggested that subtractive hybridization and target enrichment of RDA were inefficient here, with the lack of elimination of common sequences masking the real difference products. The use of the AFLP method identified 29 primer pairs that yielded 49 putative shell-thickness related-polymorphic bands. A detailed analysis will need to be carried out to fully evaluate and validate these markers. The use of the relatively new Diversity Array Technology “Genotyping-By-Sequencing” (DArTSeq) platform through genotyping of two closely-related tenera self-pollinated F2 populations, 768 (n=44) and 769 (n=57), generated a total of 11,675 DArTSeq polymorphic markers of good quality. These markers were used in the construction of the first reported DArTSeq based high-density linkage maps for oil palm. Both genetic maps consist of 16 major independent linkage groups (total map length of 1874.8 and 1720.6 cM, with an average marker density of one marker every 1.33 and 1.62 cM, respectively), corresponding well with the 16 homologous chromosome pairs of oil palm (2n = 2x = 32; 14/16 chromosomes were confirmed by known location SSR markers). Preliminary quantitative trait loci (QTL) mapping of the yield and vegetative growth traits detected four significant and 34 putative as well as two significant and 30 putative QTLs for these small 768 and 769 populations, respectively. No common significant QTL were detected between the two closely-related controlled crosses which could have allowed combination of QTL across the two populations. Saturation of the shell-thickness (Sh) region with all available DArTSeq markers, as well as map integration around the Sh regions for both populations, identified 32 Single Nucleotide Polymorphism (SNP) and DArT markers mapped within a 5 cM flanking region of the Sh gene. Homology search of the DArTSeq marker sequence tag (64 bp) against the recently published oil palm genome assembly confirmed that 23 out of the 32 (72%) DArTSeq markers were located on the p5_sc00060 scaffold in which the SHELL gene was identified. The identified shell-thickness markers could be useful as molecular screening tools. This study demonstrated the potential and feasibility of using genomic resources available for genetic improvement of oil palm breeding programmes.

The potential for viruses to be causing the plateau in the yield of UK wheat (Triticum aestivum) was investigated. Mechanical inoculation of Cynosurus mottle virus to wheat cv. Scout and cv. Gladiator caused 83% and 58% reduction in the number of grains produced, highlighting the potential of viruses to cause disease and yield loss. Viruses historically detected in cereals in the UK were not found to be prevalent following real time reverse transcriptase polymerase chain reaction testing of 1,356 UK wheat samples from 2009-2012 using eleven assasys developed in the project. This included an assay for Cynosurus mottle virus, which was based on its complete genome sequence which was obtained for the first time in this project. Viruses detected were Barley yellow dwarf virus-MAV (6 samples) (BYDV-MAV), Barley yellow dwarf virus-PAV (6 samples) (BYDV-PAV) and Soil-borne cereal mosaic virus (12 samples) (SBCMV). There was a higher prevalence of viruses in the south, thought to be due to warmer temperatures which benefitted insect vectors and the molecular processes of infection. Viruses were most commonly detected in the variety JB Diego, perhaps because this variety has no known resistance to viruses. The low prevalence of known viruses could also have been because they were outcompeted or replaced by previously unknown ones. Next generation sequencing was used to test 120 samples from an organic site, including wheat, weeds and insects, to search for novel viruses. Testing of twelve storage regimes for insect traps using BYDV-PAV infected Sitobion avenae for recovery of PCR amplifiable RNA using 18S rRNA and BYDV-PAV assays found that 0.5 M EDTA was the most successful regime which was therefore used in the collection of samples for sequencing. Known viruses such as BYDV-PAV were detected along with some additional potentially novel viruses (eight possibly novel viruses or strains of viruses with four in wheat). One such virus was apparently present in 25% of all wheat samples tested, making it potentially very significant. This could be important for unlocking the yield potential of wheat because it could be a cryptic virus which is highly prevalent. In order to control the spread of viruses their methods of transmission must be understood, therefore testing of seeds and resulting plants from Cynosurus mottle virus infected material was done. Tests did not detect the virus, therefore it was concluded that seed transmission does not occur. However, further tests are required. In conclusion this study indicates that known viruses are not currently a major problem for UK winter wheat. However, novel viruses that are a problem may be detected in the future perhaps by next generation sequencing. Additonal viruses from abroad would add to the threat. The impact of all viruses in wheat may be greater in the future due to climate change.

A study was conducted to investigate the effects of varying levels of ozone (0, 1.5, 2.5, 3.5 or 5.0 ppm) for 96 h on 1. the in vitro and in vivo growth of Colletotrichum gloeosporioides, the causal organism of anthracnose; 2. the reactive oxygen species generation and spore mitochondria of C. gloeosporioides using transmission electron microscope, fluorescence microscope and laser scanning confocal microscope; 3. the production of defence-related enzymes in papaya; 4. microbiological analysis on ozone-treated and non-treated papaya; 5. the biochemical, physiological, gas exchange and sensory characteristics of papaya fruit during storage (25 ± 3 °C, 70 ± 5 %RH) for 14 days; 6. the changes in total phenols, total carotenoids and antioxidant activity; and 7. gene expression of ozone-fumigated papaya fruit. Data were analyzed using analysis of variance and differences among treatment means were separated by Duncan Multiple Range Test (DMRT). The results of antifungal studies showed that mycelial growth of C. gloeosporioides was reduced significantly (p < 0.05) at all concentrations compared to the control. The maximum inhibition in mycelium growth (41.2 %) was obtained at 5.0 ppm ozone. Similarly, conidial germination inhibition was 100 % for 5 ppm ozone. In vivo analysis revealed that 2.5 ppm ozone was the optimal concentration for controlling anthracnose disease incidence (72.5 %) and disease severity after 10 days of storage, showing that a moderate concentration of ozone is effective in the reduction of C. gloeosporioides in artificially inoculated papaya fruit without affecting the quality aspect of the fruit. The results of scanning electron microscopy (SEM) also confirmed that ozone fumigated fungus at levels above 3.5 ppm deformed and disintegrated spore and mycelia structure. Further to that, transmission electron microscopy (TEM) illustrated that the mitochondria of ozonized fungus was disintegrated and had ruptured membrane. In spores treated with 3.5 ppm ozone, mitochondrial cristae were distorted, whereas the mitochondria were almost completely degraded in spores treated with 5.0 ppm. Meanwhile, the results from microscopy studies using laser scanning confocal microscope and fluorescence microscope showed that ozone treatment caused production of reactive oxygen species (ROS) in mitochondria of C. gloeosporioides. With increased concentration of ozone, higher levels of ROS were induced in the spores. Besides its direct antifungal activity, the study strongly suggested that ozone induces a series of defense reactions through production of compounds such as total phenols, polyphenol oxidase (PPO), peroxidase (POD) and phenylalanine ammonia-lyase (PAL) in ozone-fumigated papaya. Likewise, content of ascorbic acid, β-carotene, lycopene and antioxidant activity of papaya increased as fruit ripened and was further enhanced by exposure to ozone for 96 hours from day 4 until day 8. Twenty-four hours of ozone treatment at the level of 0.5, 2, 3.5 and 5.8 ppm reduced the total mesophilic microorganism counts of fruit with initial values of 4.48 to 2.18 log cfug-1. In addition, no coliform bacteria were initiated after 24 hours at all levels of ozone exposure. In addition, 2.5 ppm ozone treated fruit showed maximum beneficial effects in reducing weight loss, maintaining firmness, reduced rate of respiration, delaying changes in peel colour and containing the highest soluble solids concentration (SSC) as compared to the control. The titratable acidity declined throughout the storage period with slower rate in ozone-fumigated fruits. Overall sensory assessment of quality after ripening showed fruit were significantly better in quality when fumigated with 2.5 ppm ozone which were assigned highest sensory score in terms of appearance, sweetness, pulp colour, texture, aroma and overall acceptability than the control. The discovery of the gene expression of papaya in defense response induced by ozone fumigation has further clarified the understanding on how specific gene involved in controlling its expression when the plant changes during stress or in any plant lifecycle event. Among those genes, some involved in ethylene biosynthesis, generation of reactive oxygen species and stress responses of plant defense were found (mitochondrion, chloroplast, heat shock proteins, polygalacturonase-inhibiting protein, hydroxyproline-rich glycoprotein, ethylene responsive factor and acyl-CoA oxidase). Thus, the findings from all the experiments carried out during this study showed that 2.5 ppm ozone reduced anthracnose incidence and extended the storage life for up to 12 days while maintaining acceptable quality of papaya fruit. Ozone exposure at 1.5 ppm resulted in poorer quality fruit as compared to 2.5 ppm ozone treated fruit. Higher concentration of ozone exposure at 3.5 ppm and 5 ppm ozone seems non-physiological and caused phytotoxic effect on the quality of papaya fruit. As a non-toxic, biodegradable product, eco-friendly and safe sanitizer, ozone has the potential to become a natural preservative for prolonging the shelf life and retaining quality of papaya by combating fungal disease, particularly fungus C. gloeosporioides, thus promoting the marketability of the crop and minimizing postharvest losses in the papaya industry.

A study was conducted to develop an ecofriendly and sustainable fungicide based on submicron chitosan dispersions using low molecular weight chitosan and locally prepared chitosan. The physicochemical properties of both submicron dispersions were determined and their antimicrobial effect on Colletotrichum gloeosporioides was also investigated. The antifungal effect was further demonstrated by the production of fungal cell wall degrading enzymes and by the production of defence related enzymes in the plants along with the effect of submicron chitosan dispersions on the vegetative growth of plants. The effect of submicron chitosan dispersions on biochemical and physiological responses of dragon fruit during storage at 10 ± 2 °C and 80 ± 5% relative humidity for 28 days was also investigated. In vitro antifungal activity of submicron chitosan dispersions showed that low molecular weight chitosan has better antifungal properties than the locally prepared chitosan and therefore was selected for further studies. However, 600 nm droplets of 1.0% chitosan showed promising results in terms of suppressing mycelial growth (90.2%), conidial germination (93.1%) and reducing dry weight of mycelium (37.9%). Similarly, in vivo studies showed that the 600 nm droplets of 1.0% chitosan helped to reduce the disease incidence by up to 33.0 %. Beside the direct antifungal effect, this study suggests that the submicron chitosan dispersions enhanced host resistance through the production of defence related compounds, such as total phenols, peroxidase (PO), polyphenol oxidase (PPO), phenylalanine ammonia lyase (PAL) and also the activity of pathogenesis related proteins, such as β, 1-3 glucanase and chitinase. Treatments with submicron chitosan dispersions as 600 nm droplets of 1.0% chitosan showed more vigorous growth of plants as compared to the control. The submicron chitosan dispersions with 600 nm droplets of 1.0% chitosan helped to reduce the incidence of disease on fruit by up to 94% as compared to the control. The biochemical and physiological studies of dragon fruit after 28 days of storage showed that weight loss of the fruit treated with 600 nm droplets of 1.0% chitosan compared with the control reduced two fold but this was not significantly different from the conventional chitosan. The same trend was observed in all the parameters tested, including antioxidant activity. Control of disease using submicron chitosan dispersions has the potential to add market value to fresh produce. The findings from all the experiments showed that 600 nm droplets of 1.0% chitosan help to reduce anthracnose in the field and also during postharvest storage. Thus, 600 nm droplets of 1.0% chitosan could be used commercially in fields of dragon fruit as a green fungicide. Being non-toxic and biodegradable, chitosan has the potential to become an alternative to synthetic fungicides for protecting fruit crops and thus assisting sustainable agriculture.

The characteristic curd phenotype of the Brassica cauliflower consists of proliferating, arrested inflorescence and floral meristems. Recent analysis of the similar phenotype in the ap1-1/cal-1 mutant of the related crucifer Arabidopsis thaliana has led to speculation that the orthologous genes from Brassica oleracea L. may be responsible for this characteristic trait. Application of molecular genetic analysis to this hypothesis allows the presentation of a genetic model based on specific, mapped loci of BoCAL and BoAP 1. This model accounts for differences in the stage of arrest between the heading phenotypes of cauliflower (B. oleracea var. botrytis L.) and Calabrese broccoli (B. oleracea var. italica Plenck), and is also predictive in accounting for intermediate stages of arrest similar to those observed in Sicilian Purple types. Further molecular genetic analysis characterised three independent loci of the floral meristem identity gene BoAP 1. Integration of this data into the genetic model proposed for curd development, suggests a combination of point mutations and expression thresholds of several copies of the key meristem identity genes BoCAL and BoAP 1 respectively may account for the development of curd tissue in the Brassica cauliflower. The association of alleles of the BoCAL-a gene with the curding phenotypes of B. oleracea was also demonstrated through a survey of over 200 crop accessions. This reveals strong correlations between specific BoCAL-a alleles and discrete inflorescence morphologies, and allows the presentation of a possible scenario for the domestication of cauliflowers. Molecular genetic analysis of BoCAL-a utilising monosomic addition lines has also demonstrated the potential for integration of the genetic and cytogenetic maps of B. oleracea. Such analysis may have significant utility for physical characterisation of replicated loci in B. oleracea, prior to the development of a strong physical map. Further examination of inflorescence morphologies amongst the heading brassicas revealed a shared trait, termed Fused Inflorescence. Preliminary investigation suggests this trait may be under the control of multiple loci, providing a possible indication of the delineation between heading and sprouting B. oleracea crops.