Use of sulfuryl fluoride in the management of strongly phosphine-resistant insect pest populations in bulk grain storages in Australia

Sulfuryl fluoride (SF), an effective structural fumigant, is registered recently as Profume™ for controlling insect pests of stored grains and processed commodities. Information on its effectiveness in disinfestation of bulk grain, however, is limited. The ongoing problem with the strong level of resistance to phosphine has been addressed recently through deployment of SF as a ‘resistance breaker’ in bulk storages in Australia. This paper discusses important results on the efficacy of SF against key phosphine- resistant insect pests, lesser grain borer, Rhyzopertha dominca, red flour beetle, Tribolium castaneum, rice weevil, Sitophilus oryzae and the rusty grain beetle, Cryptolestes ferrugineus. We have established CT (g-hm3) profiles for SF against these insect pests at two temperature regimes 25 and 30°C, that showed that both temperature and exposure period (t) has significant influence on the effectiveness of SF than the concentration. Over a seven days fumigation period, CTs of 800 and 400 g-hm3 achieved complete control of all the target pests, including the most strongly phosphine - resistant species, C. ferrugineus at 25 and 30°C, respectively. Results from four industry scale field trials involving currently registered rate of SF (1500 g-hm3) over 2–14 d exposure period, confirmed its effectiveness in achieving complete control of the target pest species. The assessment of postfumigation grain samples across all the test storages indicated that the reinfestation occurs after three months. Monitoring resistance to phosphine in C. ferrugineus over a six year period (2009–2015), showed a significant reduction in resistant populations after the introduction of SF into the fumigation strategy at problematic storage sites. Overall our research concludes that SF is a good candidate to be used as a ‘resistance breaker’ where phosphine resistance is prevalent.

Functional studies of lignin biosynthesis genes and putative flowering gene in Miscanthus x giganteus and studies on indolyl glucosinolate biosynthesis and translocation in Brassica oleracea

The first topic area of this thesis involved studies on the accumulation and translocation of glucosinolates (GSs), bioactive secondary plant compounds, in broccoli plants. Changes in GS accumulation and gene expression levels in response to exogeneous methyl jasmonate (MeJA) treatment were analyzed in different tissue types at different developmental stages of broccoli. Greater accumulation of GSs with MeJA treatment was observed in apical leaves of broccoli seedlings and florets of plants at harvest maturity. Increases in indolyl GS in apical leaves of seedlings and florets were coupled with the up-regulation of indolyl GS biosynthesis genes. The accumulation of indolyl GSs appears to be modulated by MeJA treatment in an organ-specific manner for optimal distribution of defense substances in the plant. Metabolic profiling of hydrophilic metabolites using GC-MS demonstrated increased accumulation of various phenolics, ascorbates and amino acids in broccoli tissues after MeJA treatment. Distinct changes in carbohydrate levels observed between different tissues (vegetative leaves and floret tissues) of broccoli plants after treatment suggest that carbon metabolism is differentially modulated by MeJA treatment in different tissue types depending on sink-source relationships. Reduced levels of hexose sugars and tricarboxylic acid intermediates after MeJA treatment may reflect the increased requirement for carbon and energy needed to drive secondary product biosynthesis to accumulate metabolites for defense against insects and other herbivores. Substantial increases of indolyl and aromatic GSs after exogenous treatment with MeJA in stem and petioles of seedlings and the existence of intact indolyl-GS forms in phloem exudates suggest enhanced de novo synthesis in combination with active transport. Indoly GSs share structural similarities with the auxin, IAA, and may interact with components of the auxin transport system for intra- and extra-cellular transport or translocation. Application of the auxin efflux inhibitor, 1-naphthylphthalamic acid (NPA) reduced MeJA-mediated accumulation of indolyl GSs in broccoli florets and seedling tissues. NPA did not inhibit expression of indolyl GS biosynthesis genes shown to be upregulated by MeJA treatment or the accumulation of tryptophan, the amino acid precursor of indolyl GSs. Exogenous application of benzyl GS to Arabidopsis roots induced ectopic expression of the PIN1 protein associated with the auxin transport system similar to treatment with NPA, again suggesting GS interaction with the auxin efflux carrier system. The inhibitory effect of NPA on MeJA-mediated accumulation of GS may be due to competitive binding of NPA to auxin efflux carrier components and that GS transport is mediated by the auxin transport system. The inhibitory effect of NPA on indolyl and aromatic GS accumulation and the bioactivity of exogenous treatment of these GS compounds in PIN1 localization, Arabidopsis root growth, and gravitrophic response suggest that indolyl and aromatic GSs may be antagonistic to IAA transport and biosynthesis. Indolyl and aromatic GSs can also be potentially converted into IAA by hydrolysis. This intrinsic feature of GSs may be the part of a sophisticated regulatory process where the metabolic pathways in the plant shift from active growth to a reversible defense posture in response to biotic or abiotic stress. It seems likely that indolyl and aromatic GSs are important compounds that provide connections between jasmonate and auxin signaling. Further studies are required to reveal the regulatory mechanism for crosstalk between the two hormones. The third part of this research was to investigate effect of selenium fertilization and MeJA treatment on accumulation of GSs in broccoli florets. Increasing dietary intake of the element selenium (Se) has been shown to reduce the risk of cancer. Simultaneous enhancement of both Se and GS concentrations in broccoli floret tissue were conducted through the combined treatment of MeJA with Se fertilization. A low level of Se fertilization (concentration) with MeJA treatment displayed no significant changes in total aliphatic GS concentrations with 90% and 50% increases in indolyl and total GSs concentrations, respectively. This result suggests that Se- and GS-enriched broccoli with improved health-promoting properties can be generated by this combined treatment. The second topic of this thesis was conducted to provide basic information required to improve biomass quality and productivity and develop tools for gene transformation in Miscanthus x giganteus. The perennial rhizomatous grass, Miscanthus x giganteus is an ideal biomass crop due to its rapid vegetative growth and high biomass yield potential. As a naturally occurring sterile hybrid, M. x giganteus must be propagated vegetatively by mechanicalling divided rhizomes or from micropropagated plantlets. The effect of callus type, age and culture methods on regeneration competence was studied to improve regeneration efficiency and shorten the period of tissue culture in M. x giganteus propagation. Seven lignin biosynthesis genes and one putative flowering gene were isolated from M. x giganteus by PCR reactions using maize othologous sequences. Southern hybridization and nuclear DNA content analysis indicated that the genes isolated from M. x giganteus exist in the genome of other Miscanthus species as multiple copies. Analysis of lignin content and histological staining of lignin deposition indicated that higher lignin content is found in mature stem node tissues compared to young leaves and apical stem nodal tissues. Cell wall lignification is associated with increasing tissue maturity in Miscanthus species. RNAi and antisense constructs harboring sequences of these genes were developed to generate Miscanthus transgenic plants with suppressed of lignin biosynthesis and delayed flowering.

Effect of diet on phosphine toxicity, rate of development and reproduction of the rice weevil Sitophilus oryzae (Linnaeus)

One of the loci responsible for strong phosphine resistance encodes dihydrolipoamide dehydrogenase (DLD). The strong co-incidence of enzyme complexes that contain DLD, and enzymes that require thiamine as a cofactor, motivated us to test whether the thiamine deficiency of polished white rice could influence the efficacy of phosphine fumigation against insect pests of stored grain. Three strains of Sitophilus oryzae (susceptible, weak and strong resistance) were cultured on white rice (thiamine deficient), brown rice or whole wheat. As thiamine is an essential nutrient, we firstly evaluated the effect of white rice on developmental rate and fecundity and found that both were detrimentally affected by this diet. The mean time to reach adult stage for the three strains ranged from 40 to 43 days on brown rice and 50–52 days on white rice. The mean number of offspring for the three strains ranged from 7.7 to 10.3 per female over a three day period on brown rice and 2.1 to 2.6 on white rice. Growth and reproduction on wheat was similar to that on brown rice except that the strongly resistant strain showed a tendency toward reduced fecundity on wheat. The susceptible strain exhibited a modest increase in tolerance to phosphine on white rice as expected if thiamine deficiency could mimic the effect of the dld resistance mutation at the rph2 locus. The strongly resistant strain did not respond to thiamine deficiency, but this was expected as these insects are already strongly resistant. We failed, however, to observe the expected synergistic increase in resistance due to combining thiamine deficiency with the weakly resistant strain. The lack of interaction between thiamine content of the diet and the resistance genotype in determining the phosphine resistance phenotype suggests that the mode of inhibition of the complexes is a critical determinant of resistance.

Cadmium phytoextraction capacity of white lupine ( Lupinus albus L.) and narrow-leafed lupine ( Lupinus angustifolius L.) in three contrasting agroclimatic conditions of Chile

The phytoextraction process implies the use of plants to promote the elimination of metal contaminants in the soil. In fact, metal-accumulating plants are planted or transplanted in metal-contaminated soil and cultivated in accordance with established agricultural practices. The objective of the present study was to evaluate the productivity and Cd phytoextraction capacity of white lupine ( Lupinus albus L.) and narrow-leafed lupine ( Lupinus angustifolius L.), as well as the effect on residual Cd concentration in the soil. Both species of lupines were grown at three CdCl2 rates (0, 1, and 2 mg kg-1), under three agroclimatic conditions in Chile in 2013. In the arid zone (Pan de Azúcar, 73 mm precipitation), narrow-leafed lupine production was significantly (P < 0.05) higher than white lupine (4.55 vs. 3.26 Mg DM ha-1, respectively). In locations with higher precipitation (Santa Rosa, 670 mm; Carillanca, 880 mm), narrow-leafed lupine DM production was slightly higher than in Pan de Azúcar, but white lupine was approximately three times higher. Total plant Cd concentrations in white and narrow-leafed lupine increased as Cd rates increased in the three environments, but they were much higher in narrow-leafed lupine than white lupine; 150%, 58%, and 344% higher in Pan de Azúcar, Santa Rosa, and Carillanca, respectively. Cadmium uptake (g Cd ha-1) and apparent recovery were also higher (P < 0.05) in narrow-leafed lupine in two environments (Pan de Azúcar and Carillanca). These results suggest that narrow-leafed lupine present higher potential as phytoremediation species than white lupine.

Antiangiogenic potential of yogurts added with extracts rich in apigenin derivatives

Angiogenesis is a biological process through which there is the formation of new blood vessels from preexisting ones [I]. However, in pathological cases, the abnormal growth of new blood vessels promotes the development of various diseases including cancer [2) through the production of atypically large amounts of angiogenesis factors, e.g. the vascular endothelial growth factor (VEGF) [3]. The plant secondary metabolites have been the subject of several studies to evaluate their benefits to human health. In particular, the phenolic compounds have high potential for use in the food industry, including the development of functional foods. Among these, apigenin has been associated with chemopreventive effects related to cancer [4]. In fact, chemoprevention is a present-day concept and contemplates the use of medicines, biological compounds or nutrients as an intervention strategy of cancer prevention. In this work, an Arenaria montana L hydroethanolic extract was prepared and after characterization by HPLC-DAD-ESI/MS showed to be rich in apigenin derivatives. Furthermore, it exhibited ability to inhibit the phosphorylation of VEGFR-2 (vascular endothelium growth factor receptor) through an enzymatic assay. However, for the major protection of bioactive compounds, the extract was microencapsulated by an atomization/coagulation technique with alginate as the matrix material. Posteriorly, the hydroethanolic extract, in free and microencapsulated forms, was incorporated in yogurts in order to develop a novel chemopreventer food in relation to the angiogenesis process. The functionalized yogurts with A. montana extracts (free and microencapsulated) showed a nutritional value similar to the used control (yogurt without extract); however, the samples enriched with extracts revealed added-value regarding the VEGFR-2 phosphorylation inhibition ability. This effect was more effectively preserved over time in the samples functionalized with the protected extract. Overall, this work contributes to the valorization of plants rich in flavonoids, exploring its antiangiogenic potential with VEGFR-2 as target. Moreover, the atomization/coagulation technique allowed the production of viable microspheres enriched with the plant extract. The microspheres were effectively incorporated into yogurts, protecting the extract thus envisaging the development of novel functional foods with chemopreventive effects.

Micoencapsulation of bioactive plant extracts: a tool to functionalize cottage cheeses

Foeniculum vulgare Mill. (fennel) and Matricaria recutita L. (chamomile) are two examples of plants with reported antioxidant and antimicrobial properties, which can be related with their composition in phenolic compounds [1,2]. Furthermore, according to previous results of our research group, the direct incorporation of the aqueous extracts showed capacity to maintain the nutritional properties of the cottage cheeses, up to 7 days of storage, while improving the antioxidant potential. However, after 14 days, a decrease in the antioxidant properties was observed [1,2], which can be related with factors such as light, moisture, temperature and pH, that can cause bioactive compounds degradation. Therefore, the aim of the present study was to prepare microcapsules with the aqueous extracts of fennel and chamomile for incorporation in cottage cheese samples, in order to protect the bioactive molecules present in the extracts, such as phenolic compounds, and prevent the decrease of the antioxidant activity observed after the 14 days period. The microspheres were prepared using an atomization/coagulation technique. Sodium alginate was used as the matrix material to produce the microspheres that were characterized through optical microscopy (OM), during and after atomization, for inspecting morphology. The encapsulation efficiency (EE) was determined by HPLC-DAD by an indirect method by analysing the coagulation solution. FTIR was also used to attest the presence of the extract inside of the alginate matrix. These microencapsulated extracts were incorporated in cottage cheese samples that were further characterized in terms of nutritional properties and antioxidant potential right after incorporation, and after 7 and 14 days of storage at 4•c. The EE was estimated as -100% and the FTIR analysis confirmed the presence of the extracts inside the microspheres. The results showed that the incorporation of the microencapsulated extracts did not cause changes in the nutritional value of cottage cheeses (through a comparison with control samples without extracts). The predominant fatty acids were palmitic (C16:0) and oleic (CI8:0) acids. The order of abundance of fatty acids was as follows: saturated fatty acids (SF A)> monounsaturatcd fatty acids (MUF A)> polyunsaturated fatty acids (PUF A). Regarding free sugars, lactose was the only sugar identified and quantified in all samples. Regarding the antioxidant activity, the samples functionalized with the microencapsulated extracts showed a higher preservation of this property even after the 7th day of storage. Overall, the incorporation of the protected plant extracts in dairy foods can be a strategy to provide health benefits to consumers.

Thiamethoxam on the histological characteristics of sugarcane young roots

Thiamethoxam is a systemic insecticide from the neonicotinoid group, nitroguanidin family which affects the nicotinic receptor acetyl choline in the insect membrane, wounding the nervous system and causing the death of the insect. It was used with success in the control of initial pests of several crops. It was considered that thiamethoxam has a bioactivator effect, because in the absence of insects promoted increase in vigor, development and productivity of crops. This work was carried out to verify if thiamethoxam causes histological changes in sugarcane roots. In this work, it was used optical microscopy, images arrest, tissue biometrics and statistical analysis, in young roots of sugarcane RB 83 5486 after the treatments with different thiamethoxam concentrations. It was determined changes in histological structure of tissues 7, 14, 21 and 28 days after the treatments, establishing its effects on root plant anatomy. It was verified that thiamethoxam increased root cortex width, increasing the vascular cylinder and the metaxylem vessel elements number in the vascular tissue until 21 days after application.

Microwave-assisted solvent extraction and analysis of shikimic acid from plant tissues.

A better method for determination of shikimate in plant tissues is needed to monitor exposure of plants to the herbicide glyphosate [N-(phosphonomethyl)glycine] and to screen the plant kingdom for high levels of this valuable phytochemical precursor to the pharmaceutical oseltamivir. A simple, rapid, and efficient method using microwave-assisted extraction (MWAE) with water as the extraction solvent was developed for the determination of shikimic acid in plant tissues. High performance liquid chromatography was used for the separation of shikimic acid, and chromatographic data were acquired using photodiode array detection. This MWAE technique was successful in recovering shikimic acid from a series of fortified plant tissues at more than 90% efficiency with an interference-free chromatogram. This allowed the use of lower amounts of reagents and organic solvents, reducing the use of toxic and/or hazardous chemicals, as compared to currently used methodologies. The method was used to determine the level of endogenous shikimic acid in several species of Brachiaria and sugarcane (Saccharum officinarum) and on B. decumbens and soybean (Glycine max) after treatment with glyphosate. The method was sensitive, rapid and reliable in all cases.

Natural parasitism of lepidopteran eggs by Trichogramma species (Hymenoptera: Trichogrammatidae) in agricultural crops in Minas Gerais, Brazil.

The genus Trichogramma Westwood (Hymenoptera: Trichogrammatidae) includes insect egg parasitoids that are widely used throughout the world as control agents of pest insects. The aim of this study was to identify the species of Trichogramma naturally associated with the eggs of lepidopteran pests of the following agricultural and horticultural crops: collards, Brassica oleracea L. (Brassicales: Brassicaceae); papaya, Carica papaya L. (Capparales: Caricaceae); tomato, Lycopersicon esculentum Mill. (Solanales: Solanaceae); cassava, Manihot esculenta Crantz (Malpighiales: Euphorbiaceae); banana, Musa sp. L. (Zingiberales: Musaceae); passion fruit, Passiflora sp. Degener (Malpighiales: Passifloraceae); sugarcane, Saccharum sp. L. (Poales: Poaceae); and corn (maize), Zea mays L. (Poales: Poaceae); and an invasive species (Sodom?s apple milkweed, Calotropis procera Aiton; Gentianales: Apocynaceae) in the semiarid region of Minas Gerais, Brazil. We report natural parasitism by Trichogramma in eggs of Agraulis vanillae vanillae (L.) (Lepidoptera: Nymphalidae), Antichloris eriphia F. (Lepidoptera: Arctiidae), Danaus sp. (L.) (Lepidoptera: Nymphalidae), Diatraea saccharalis F. (Lepidoptera: Crambidae), Erinnyis ello L. (Lepidopera: Sphingidae), and Protambulyx strigilis L. (Lepidopera: Sphingidae). In total, 2,242 specimens of Trichogramma were obtained, belonging to the species T. pretiosum Riley, T. manicobai Brun, Moraes & Soares, T. marandobai Brun, Moraes & Soares, and T. galloi Zucchi. These species of Trichogramma may be candidates for biological control programs of lepidopteran pests in the semiarid region of Minas Gerais and in other semiarid regions.

Phytopharmacological and ethnomedicinal uses of the Genus Berberis (Berberidaceae): A review

Plants belonging to Berberis are reported in several folklore medicinal pharmacopeias and are used in traditional medicines in Asia and European countries. The plants have been used in the preparation of various traditional and synthetic medicines since pre-historic times for wound healing, fever, eye disease, jaundice, vomiting during pregnancy, rheumatism, kidney and gall balder stones, and several other illnesses. Their healing properties are appear to be due to the presence of secondary metabolites and important alkaloids with different pharmacological activities. Their antibacterial, antifungal, antiviral, anti-diabetic, and anti-tumor activities as well as positive effects on the cardiovascular and body immune systems have been reported. Root extracts of some species of the plant genus contain quinine which acts as a powerful anti-malarial agent. The main chemical constituents of Berberis plants are alkaloids, steroids, glycosides, flavonoids, saponins, terpenoids and reducing sugars. Of these alkaloids, berberine is the most important. The present review focuses on recent advances in phytopharmacological and ethnomedicinal uses of plants belonging to Berberis genus.

An Analysis of the Potential Risk Exposure to Lead (Pb) through Urban Community Gardens

Community gardening in cities is increasing, driven by social interaction and food security. City soils are sinks for heavy metals; including neurotoxic lead (Pb). Exposure routes are primarily through inhalation/ingestion of soil, or second by ingestion of plants that have accumulated Pb. This research evaluates soil at three Liberty City, Florida sites estimating risk of Pb exposure through primary and secondary pathways. Soil cores were collected from Liberty City, and red Malabar spinach (Basella rubra) was grown in Pb soil treatments in a greenhouse. Total soil Pb levels and plant tissues were measured after acid digestion, by ICP-OES. In Liberty City, two sites had hotspots with areas of elevated soil Pb levels. Plants grown on Pb contaminated soil all accumulated statistically significant Pb concentrations. Therefore, there is a potential risk of Pb exposure to residents in Liberty City by exposure in hotspot sites through both the primary and secondary pathways.

Heat and water stress induce unique transcriptional signatures of heat-shock proteins and transcription factors in grapevine

Grapevine is an extremely important crop worldwide.In southern Europe, post-flowering phases of the growth cycle can occur under high temperatures, excessive light, and drought conditions at soil and/or atmospheric level. In this study, we subjected greenhouse grown grapevine, variety Aragonez, to two individual abiotic stresses, water deficit stress(WDS), and heat stress (HS). The adaptation of plants to stress is a complex response triggered by cascades of molecular net works involved in stress perception, signal transduction, and the expression of specific stress-related genes and metabolites. Approaches such as array-based transcript profiling allow assessing the expression of thousands of genes in control and stress tissues. Using microarrays, we analyzed the leaf transcriptomic profile of the grapevine plants. Photosynthesis measurements verified that the plants were significantly affected by the stresses applied. Leaf gene expression was obtained using a high-throughput transcriptomic grapevine array, the 23K custom-made Affymetrix Vitis GeneChip. We identified 1,594 genes as differentially expressed between control and treatments and grouped them into ten major functional categories using MapMan software. The transcriptome of Aragonez was more significantly affected by HS when compared with WDS. The number of genes coding for heat-shock proteins and transcription factors expressed solely in response to HS suggesting their expression as unique signatures of HS. However, across-talk between the response pathways to both stresses was observed at the level of AP2/ERF transcription factors.

A machine learning approach for recognising woody plants on railway trackbeds

The purpose of this work in progress study was to test the concept of recognising plants using images acquired by image sensors in a controlled noise-free environment. The presence of vegetation on railway trackbeds and embankments presents potential problems. Woody plants (e.g. Scots pine, Norway spruce and birch) often establish themselves on railway trackbeds. This may cause problems because legal herbicides are not effective in controlling them; this is particularly the case for conifers. Thus, if maintenance administrators knew the spatial position of plants along the railway system, it may be feasible to mechanically harvest them. Primary data were collected outdoors comprising around 700 leaves and conifer seedlings from 11 species. These were then photographed in a laboratory environment. In order to classify the species in the acquired image set, a machine learning approach known as Bag-of-Features (BoF) was chosen. Irrespective of the chosen type of feature extraction and classifier, the ability to classify a previously unseen plant correctly was greater than 85%. The maintenance planning of vegetation control could be improved if plants were recognised and localised. It may be feasible to mechanically harvest them (in particular, woody plants). In addition, listed endangered species growing on the trackbeds can be avoided. Both cases are likely to reduce the amount of herbicides, which often is in the interest of public opinion. Bearing in mind that natural objects like plants are often more heterogeneous within their own class rather than outside it, the results do indeed present a stable classification performance, which is a sound prerequisite in order to later take the next step to include a natural background. Where relevant, species can also be listed under the Endangered Species Act.

Assessing the conservation status of Mediterranean temporary ponds using biodiversity: a new tool for practitioners

The assessment of the habitat condition is the first step of conservation actions and several tools are available to assess wetlands. However, only a few tools are adapted to the priority habitat Mediterranean temporary ponds. Thus, our objectives were (i) to identify biological indicators associated with the different conservation status of Mediterranean tem- porary ponds and (ii) to create an efficient evaluation tool for non-experts using indicators of conservation status. A total of 87 ponds were sampled in southwest Portugal to assess the presence of plants, large branchiopods, amphibians, threatened voles and bats. Ponds with favourable conservation status showed higher species richness of plants, large branchiopods and amphibians. We identified eighteen indicators for favourable ponds: 15 plants, one large branchiopod and two amphibian taxa. We propose a new tool to assess the conservation status of Mediterranean tem- porary ponds based on the presence of these indicators. This tool is an alternative to other common, but time- consuming, methods and can be readily used by trained practitioners. The replication and adaptation of this tool to other regions and habitats enables the collection of comparable data and the geographical scaling-up of the assessments.

Nitrogen requirements at bulb initiation for production of intermediate-day onions

Nitrogen requirements at bulb initiation for production of intermediate-day onions Article in Acta horticulturae · October 2016 DOI: 10.17660/ActaHortic.2016.1142.11 1st Rui Machado 16.44 · Universidade de Évora 2nd David R. Bryla 30.16 · United States Department of Agriculture Abstract Nitrogen requirements at bulb initiation for production of intermediate-day onions Authors: R.M.A. Machado, D.R. Bryla Keywords: Allium cepa, crop growth, nitrogen uptake, soil nitrate Abstract: The effect of nitrogen application on growth, nitrogen (N) uptake, yield, and quality of intermediate-day onion (Allium cepa 'Guimar') was evaluated in the field in southern Portugal. Plants were fertilized with 30 kg ha-1 N at transplanting, 10 kg ha-1 N at 29 days after transplanting (DAT) during early leaf growth, and with 0, 20, 40 and 60 kg ha-1 N at 51 DAT at the initiation of bulbing. The root system of plants in each treatment were concentrated in the top 0.1 m of soil and limited to 0.3 m depth but neither root length density nor rooting depth were affected by N application during later stages of bulb development. Leaf and bulb dry matter, on the other hand, increased linearly with N rate during bulb growth (85 DAT) and at harvest (114 DAT), respectively. Soil nitrate-N (NO3-N) at 0-0.3 m depth likewise increased linearly with N rate during bulb growth but declined from 15-30 mg kg-1 at bulbing to >10 mg kg-1 in each treatment by harvest. A substantial amount of N in the plants, which ranged from 302-525 mg, was taken up from the soil. Application of 60 kg ha-1 N resulted in luxury consumption. Yield (fresh bulb weight) increased from 0.19 kg plant-1 with no N at bulbing to as much as 0.28 kg plant-1 with 60 kg ha-1 N. Bulbs harvested from plants fertilized 40-60 kg ha-1 N averaged 8.2-8.5 cm in diameter, while those from plants with no N at bulbing averaged only 7.2 cm in diameter. Application of N fertilizer is thus recommended at bulbing to increase N uptake, yield, and bulb size of intermediate-day onions, particularly in dry Mediterranean climates where many onions are produced. Other components of quality, including neck diameter, bulb water content, total soluble solids, and juice pH, were not affect by N applied at bulbing.