Host-delivered RNAi: An effective strategy to silence genes in plant parasitic nematodes

Root-knot nematodes (Meloidogyne spp.) are obligate, sedentary endoparasites that infect many plant species causing large economic losses worldwide. Available nematicides are being banned due to their toxicity or ozone-depleting properties and alternative control strategies are urgently required. We have produced transgenic tobacco (Nicotiana tabacum) plants expressing different dsRNA hairpin structures targeting a root-knot nematode (Meloidogyne javanica) putative transcription factor, MjTis11. We provide evidence that MjTis11 was consistently silenced in nematodes feeding on the roots of transgenic plants. The observed silencing was specific for MjTis11, with other sequence-unrelated genes being unaffected in the nematodes. Those transgenic plants able to induce silencing of MjTis11, also showed the presence of small interfering RNAs. Even though down-regulation of MjTis11 did not result in a lethal phenotype, this study demonstrates the feasibility of silencing root-knot nematode genes by expressing dsRNA in the host plant. Host-delivered RNA interference-triggered (HD-RNAi) silencing of parasite genes provides a novel disease resistance strategy with wide biotechnological applications. The potential of HD-RNAi is not restricted to parasitic nematodes but could be adapted to control other plant-feeding pests.

Short-term responses of leaf growth rate to water deficit scale up to whole-plant and crop levels: An integrated modelling approach in maize.

Physiological and genetic studies of leaf growth often focus on short-term responses, leaving a gap to whole-plant models that predict biomass accumulation, transpiration and yield at crop scale. To bridge this gap, we developed a model that combines an existing model of leaf 6 expansion in response to short-term environmental variations with a model coordinating the development of all leaves of a plant. The latter was based on: (1) rates of leaf initiation, appearance and end of elongation measured in field experiments; and (2) the hypothesis of an independence of the growth between leaves. The resulting whole-plant leaf model was integrated into the generic crop model APSIM which provided dynamic feedback of environmental conditions to the leaf model and allowed simulation of crop growth at canopy level. The model was tested in 12 field situations with contrasting temperature, evaporative demand and soil water status. In observed and simulated data, high evaporative demand reduced leaf area at the whole-plant level, and short water deficits affected only leaves developing during the stress, either visible or still hidden in the whorl. The model adequately simulated whole-plant profiles of leaf area with a single set of parameters that applied to the same hybrid in all experiments. It was also suitable to predict biomass accumulation and yield of a similar hybrid grown in different conditions. This model extends to field conditions existing knowledge of the environmental controls of leaf elongation, and can be used to simulate how their genetic controls flow through to yield.

Bitou bush and boneseed eradication and containment in Australia

Bitou bush and boneseed (Chrysanthemoides monilifera subsp. rotundata (DC.) T.Norl. and C. monilifera subsp. monilifera (L.) T.Norl., respectively) are highly invasive environmental weeds that pose a serious threat to Australia’s natural ecosystems and biota. Bitou bush threatens coastal plant communities in New South Wales (NSW), eastern Victoria and southeast Queensland (Qld), while boneseed threatens inland and coastal native plant communities across NSW, South Australia (SA), Tasmania, Victoria, and Western Australia (WA). Over 200 plant species and ecological communities in Australia are negatively impacted by these weeds (ARMCANZ et al. 2000, DEC 2006) and over 15% (approx. 120 million ha) of Australia is susceptible to invasion (see maps in Weiss et al 2008). In 2000, the National Bitou Bush and Boneseed Strategic Plan (ARMCANZ et al. 2000) was approved as part of the Commonwealth’s Weeds of National Significance initiative. A key goal of this plan is to prevent the spread of bitou bush and boneseed in Australia. A national program sponsored by the Australian Government and the affected states has resulted in the development of national containment and eradication zones that prevent the spread of bitou bush and boneseed. This paper presents an overview of these bitou bush and boneseed containment and eradication programs.

Heterologous signals allow efficient targeting of a nuclear-encoded fusion protein to plastids and endoplasmic reticulum in diverse plant species

Approximately 30% of plant nuclear genes appear to encode proteins targeted to the plastids or endoplasmic reticulum (ER). The signals that direct proteins into these compartments are diverse in sequence, but, on the basis of a limited number of tests in heterologous systems, they appear to be functionally conserved across species. To further test the generality of this conclusion, we tested the ability of two plastid transit peptides and an ER signal peptide to target green fluorescent protein (GFP) in 12 crops, including three monocots (barley, sugarcane, wheat) and nine dicots (Arabidopsis, broccoli, cabbage, carrot, cauliflower, lettuce, radish, tobacco, turnip). In all species, transient assays following microprojectile bombardment or vacuum infiltration using Agrobacterium showed that the plastid transit peptides from tomato DCL (defective chloroplast and leaves) and tobacco RbcS [ribulose bisphosphate carboxylase (Rubisco) small subunit] genes were effective in targeting GFP to the leaf plastids. GFP engineered as a fusion to the N-terminal ER signal peptide from Arabidopsis basic chitinase and a C-terminal HDEL signal for protein retention in the ER was accumulated in the ER of all species. The results in tobacco were confirmed in stably transformed cells. These signal sequences should be useful to direct proteins to the plastid stroma or ER lumen in diverse plant species of biotechnological interest for the accumulation of particular recombinant proteins or for the modification of particular metabolic streams.

Conserved water-mediated H-bonding dynamics of catalytic Asn 175 in plant thiol protease

The role of invariant water molecules in the activity of plant cysteine protease is ubiquitous in nature. On analysing the 11 different Protein DataBank (PDB) structures of plant thiol proteases, the two invariant water molecules W I and W2 (W220 and W222 in the template 1PPN structure) were observed to form H-bonds with the Ob atom of Asn 175. Extensive energy minimization and molecular dynamics simulation studies up to 2 ns on all the PDB and solvated structures clearly revealed the involvement of the H-bonding association of the two water molecules in fixing the orientation of the asparagine residue of the catalytic triad. From this study, it is suggested that H-bonding of the water molecule at the W1 invariant site better stabilizes the Asn residue at the active site of the catalytic triad.

The original native pasture ecosystems of the eastern and western Darling Downs - Can they be restored?

The original pasture ecosystems of southern inland Queensland ranged from treeless grasslands on cracking clays through grassy woodlands of varying density on a great range of soil types to those competing at the dynamic edges of forests and scrubs. Fire, both wild and aboriginal-managed, was a major factor, along with rainfall extremes, in shaping the pastures and tree:grass balance. Seedling recruitment was driven by rainfall extremes, availability of germinable seed and growing space, with seed availability and space being linked to the timing and intensity of recent fires and rain. The impact of insects, diseases, severe wind and hailstorms on recruitment should not be underestimated. The more fertile soils had denser grass growth, greater fire frequency and thinner tree cover than infertile soils, except where trees were so dense that grass growth was almost eliminated. The pastures were dominated by perennial tussock grasses of mid-height but included a wide array of minor herbaceous species whose abundance was linked to soil type and recent seasonal conditions. Many were strongly perennial with Asteraceae, Fabaceae, Malvaceae, Cyperaceae and Goodeniaceae most common in an environment, which can experience effective rainfall at any time of year. The former grassland communities that are now productive farming lands are not easily returned to their original composition. However, conservation of remnant examples of original pasture types is very achievable provided tree density is controlled, prescribed burning and grazing are used and rigorous control of invasive, exotic species is undertaken. This should be done with a clear understanding that significant short-and medium-term fluctuations in botanical composition are normal.

Development of a small-plant bioassay to assess banana grown from tissue culture for consistent infection by Fusarium oxysporum f. sp. cubense

Two reliable small-plant bioassays were developed using tissue-cultured banana, resulting in consistent symptom expression and infection by Fusarium oxysporum f. sp. cubense (Foc). One bioassay was based on providing a constant watertable within a closed pot and the second used free-draining pots. Culture medium for spore generation influenced infectivity of Foc. Inoculation of potted banana by drenching potting mix with a conidial suspension, consisting mostly of microconidia, few macroconidia and no chlamydospores, generated from one-quarter-strength potato dextrose agar + streptomycin sulfate, resulted in inconsistent infection. When a conidial suspension that consisted of all three spore types, microconidia, macroconidia and chlamydospores, prepared from spores generated on carnation leaf agar was used, all plants became infected, indicating that the spore type present in conidial suspensions may contribute to inconsistency of infection. Inconsistency of infection was not due to loss of virulence of the pathogen in culture. Millet grain precolonised by Foc as a source of inoculum resulted in consistent infection between replicate plants. Sorghum was not a suitable grain for preparation of inoculum as it was observed to discolour roots and has the potential to stunt root growth, possibly due to the release of phytotoxins. For the modified closed-pot system, a pasteurised potting mix consisting of equal parts of bedding sand, perlite and vermiculite plus 1 g/L Triabon slow release fertiliser was suitable for plant growth and promoted capillary movement of water through the potting mix profile. A suitable potting mix for the free-draining pot system was also developed.

Non-destructive assessment of Arundo donax (Poaceae) leaf quality

Leaf carbon (C) content, leaf nitrogen (N) content, and C:N ratio are especially useful for understanding plant-herbivore interactions and may be important in developing control methods for the invasive riparian plant Arundo donax L. We measured C content, N content, C:N ratio, and chlorophyll index (SPAD 502 reading) for 768 leaves from A. donax collected over a five year period at several locations in California, Nevada, and Texas. Leaf N was more variable than leaf C, and thus we developed a linear regression equation for estimating A. donax leaf N from the leaf chlorophyll index (SPAD reading). When applied to two independent data sets, the equation (leaf N content % = -0.63 + 0.08 x SPAD) produced realistic estimates that matched seasonal and spatial trends reported from a natural A. donax population. Used in conjunction with the handheld SPAD 502 meter, the equation provides a rapid, non-destructive method for estimating A. donax leaf quality.

Melaleuca densispicata in Currawinya National Park, South-West Queensland: Ecology and preliminary implications for management of a rare plant species

Melaleuca densispicata Byrnes is an uncommon species with a limited distribution, comprising disjunct populations in inland southern Queensland and northern New South Wales, Australia. It is a dense, woody shrub, 2–4 m in height, which exhibits a marked 'clumping' growth habit. It has thick, papery bark and displays many white flowers during spring or early summer. Although it has long been known to exist, M. densispicata was only formally described in 1984, and very little is currently known about its ecology or specific management requirements. There are only seven known subpopulations of the species across its range. A major population at the western limit of its distribution occurs on Currawinya National Park (28°52'S, 144°30'E). Here, it is locally abundant and listed as a noteworthy plant species under the Management Plan (Queensland Parks & Wildlife Service 2001). This study aimed to identify patterns in the distribution of M. densispicata in Currawinya National Park, describe its ecological niche and role, and provide management recommendations for the species within the study area. Recent anecdotal observations of recruitment failure in south-western Queensland (Peter McRae, QPWS, October 2004, pers. comm.; Dick O'Connell, local grazier, July 2005 pers. comm.) caused additional emphasis to be placed on the examination of recruitment and recruitment factors.

Beyond chemical dependency for managing plant-parasitic nematodes: Examples from the banana, pineapple and vegetable industries of tropical and subtropical Australia

Plant-parasitic nematodes are important pests of horticultural crops grown in tropical and subtropical regions of Australia. Burrowing nematode (Radopholus similis) is a major impediment to banana production and root-knot nematodes (predominantly Meloidogyne javanica and M. incognita) cause problems on pineapple and a range of annual vegetables, including tomato, capsicum, zucchini, watermelon, rockmelon, potato and sweet potato. In the early 1990s, nematode control in these industries was largely achieved with chemicals, with methyl bromide widely used on some subtropical vegetable crops, ethylene dibromide applied routinely to pineapples and non-volatile nematicides such as fenamiphos applied up to four times a year in banana plantations. This paper discusses the research and extension work done over the last 15 years to introduce an integrated pest management approach to nematode control in tropical and subtropical horticulture. It then discusses various components of current integrated pest management programs, including crop rotation, nematode monitoring, clean planting material, organic amendments, farming systems to enhance biological suppression of nematodes and judicious use of nematicides. Finally, options for improving current management practices are considered.

The environmental fate of diuron under a conventional production regime in a sugarcane farm during the plant cane phase

BACKGROUND: Field studies of diuron and its metabolites 3-(3,4-dichlorophenyl)-1-methylurea (DCPMU), 3,4-dichlorophenylurea (DCPU) and 3,4-dichloroaniline (DCA) were conducted in a farm soil and in stream sediments in coastal Queensland, Australia. RESULTS: During a 38 week period after a 1.6 kg ha^-1 diuron application, 70-100% of detected compounds were within 0-15 cm of the farm soil, and 3-10% reached the 30-45 cm depth. First-order t1/2 degradation averaged 49 ± 0.9 days for the 0-15, 0-30 and 0-45 cm soil depths. Farm runoff was collected in the first 13-50 min of episodes lasting 55-90 min. Average concentrations of diuron, DCPU and DCPMU in runoff were 93, 30 and 83-825 µg L^-1 respectively. Their total loading in all runoff was >0.6% of applied diuron. Diuron and DCPMU concentrations in stream sediments were between 3-22 and 4-31 µg kg^-1 soil respectively. The DCPMU/diuron sediment ratio was >1. CONCLUSION: Retention of diuron and its metabolites in farm topsoil indicated their negligible potential for groundwater contamination. Minimal amounts of diuron and DCMPU escaped in farm runoff. This may entail a significant loading into the wider environment at annual amounts of application. The concentrations and ratio of diuron and DCPMU in stream sediments indicated that they had prolonged residence times and potential for accumulation in sediments. The higher ecotoxicity of DCPMU compared with diuron and the combined presence of both compounds in stream sediments suggest that together they would have a greater impact on sensitive aquatic species than as currently apportioned by assessments that are based upon diuron alone.

Host-plant resistance and biopesticides: Ingredients for successful integrated pest management (IPM) in Australian sorghum production

There are two major pests of sorghum in Australia, the sorghum midge, Stenodiplosis sorghicola (Coquillett), and the corn earworm, Helicoverpa armigera (Hübner). During the past 10 years the management of these pests has undergone a revolution, due principally to the development of sorghum hybrids with resistance to sorghum midge. Also contributing has been the adoption of a nucleopolyhedrovirus for the management of corn earworm. The practical application of these developments has led to a massive reduction in the use of synthetic insecticides for the management of major pests of sorghum in Australia. These changes have produced immediate economic, environmental and social benefits. Other flow-on benefits include providing flexibility in planting times, the maintenance of beneficial arthropods and utilisation of sorghum as a beneficial arthropod nursery, a reduction in midge populations and a reduction in insecticide resistance development in corn earworm. Future developments in sorghum pest management are discussed.

Host plant resistance in grain crops and prospects for invertebrate pest management in Australia: An overview

An integrated pest management (IPM) approach that relies on an array of tactics is adopted commonly in response to problems with pesticide-based production in many agricultural systems. Host plant resistance is often used as a fundamental component of an IPM system because of the generally compatible, complementary role that pest-resistant crops play with other tactics. Recent research and development in the resistance of legumes and cereals to aphids, sorghum midge resistance, and the resistance of canola varieties to mite and insect pests have shown the prospects of host plant resistance for developing IPM strategies against invertebrate pests in Australian grain crops. Furthermore, continuing advances in biotechnology provide the opportunity of using transgenic plants to enhance host plant resistance in grains.

Development of hazard site surveillance programs for forest invasive species: A case study from Brisbane, Australia

Hazard site surveillance is a system for post-border detection of new pest incursions, targeting sites that are considered potentially at high risk of such introductions. Globalisation, increased volumes of containerised freight and competition for space at domestic ports means that goods are increasingly being first opened at premises some distance from the port of entry, thus dispersing risk away from the main inspection point. Hazard site surveillance acts as a backstop to border control to ensure that new incursions are detected sufficiently early to allow the full range of management options, including eradication and containment, to be considered. This is particularly important for some of the more cryptic forest pests whose presence in a forest often is not discovered until populations are already high and the pest is well established. General requirements for a hazard site surveillance program are discussed using a program developed in Brisbane, Australia, in 2006 as a case study. Some early results from the Brisbane program are presented. In total 67 species and 5757 individuals of wood-boring beetles have been trapped and identified during the program to date. Scolytines are the most abundant taxa, making up 83% of the catch. No new exotics have been trapped but 19 of the species and 60% of all specimens caught are exotics that are already established in Australia.

Letter to the editor - Name that plant scientifically, please!

The report of Brunfelsia poisoning of dogs in Sydney by Drs Singh, Cowan and Child is very welcome, but it raises one important scientific point that has been further highlighted by the letter from Dr Seavers. Neither in the case report nor in Dr Seavers' letter is the species or cultivar of the plants identified. There are about 40 known Brunfelsia species, a number of which are cultivated in tropical to temperate climate gardens in Australia. It is noteworthy that Dr Seavers speculates about the Brunfelsia plants involved in dog poisonings in Australia, suggesting that 'dwarf' plants may be more poisonous ...