Rapid and efficient screening of a Representational Difference Analysis library using reverse Southern hybridisation: Identification of genetic differences between Haemophilus parasuis isolates

Representational Difference Analysis (RDA) is an established technique used for isolation of specific genetic differences between or within bacterial species. This method was used to investigate the genetic basis of serovar-specificity and the relationship between serovar and virulence in Haemophilus parasuis. An RDA clone library of 96 isolates was constructed using H. parasuis strains H425(P) (serovar 12) and HS1967 (serovar 4). To screen such a large clone library to determine which clones are strain-specific would typically involved separately labelling each clone for use in Southern hybridisation against genomic DNA from each of the strains. In this study, a novel application of reverse Southern hybridisation was used to screen the RDA library: genomic DNA from each strain was labelled and used to probe the library to identify strain-specific clones. This novel approach represents a significant improvement in methodology that is rapid and efficient.

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.

A free-enzyme catalyst for the bioremediation of environmental atrazine contamination.

Herbicide contamination from agriculture is a major issue worldwide, and has been identified as a threat to freshwater and marine environments in the Great Barrier Reef World Heritage Area in Australia. The triazine herbicides are of particular concern because of potential adverse effects, both on photosynthetic organisms and upon vertebrate development. To date a number of bioremediation strategies have been proposed for triazine herbicides, but are unlikely to be implemented due to their reliance upon the release of genetically modified organisms. We propose an alternative strategy using a free-enzyme bioremediant, which is unconstrained by the issues surrounding the use of live organisms. Here we report an initial field trial with an enzyme-based product, demonstrating that the technology is technically capable of remediating water bodies contaminated with the most common triazine herbicide, atrazine.

Using bacteria in banana for efficient use of fertilizer

This project aims to develop sustainable banana production practices by improving efficiency of fertilizer use. We will investigate how we can reintroduce endophytic beneficial bacteria that can be established inside the banana host to provide lasting and durable benefits to growth. In the current research we have been able to isolate bacteria from inside banana and preliminary characterisation indicates isolates with a range of attributes for improved efficiency of fertilizer uptake. Experimentation will include evaluation in vitro and pot trials and field trials using bacteria most likely to increase plant access to nutrients as well to compare nutrient impacts from conventional verses slow release fertilizer.

Reef Catchments Catalyst

To interogate spatial data sets including satellite imagery, EM surveys and ground samples to identify the efficiencies of current management practices within Australian cane regions.

Efficient organogenesis of an Australian passionfruit hybrid (Passiflora edulis x Passiflora edulis var. flavicarpa) suitable for gene delivery

An efficient regeneration protocol based on organogenesis from cotyledon explants and suitable for gene delivery has been developed for an Australian passionfruit hybrid. Multiple shoots were regenerated from 30-day-old cotyledon explants on Murashige and Skoog (MS) medium containing 6-benzylvaminopurine (BAP) and coconut water. Media pulsing experiments were conducted to investigate the effect on organogenesis of exposure time of the explants to MS containing 10 mu M BAP and 10% (v/v) coconut water, i.e. passionfruit regeneration medium (PRM). Continuous exposure of these explants to PRM maximised the number of shoots produced to 12.1 per explant. However, periods on hormone-free medium improved the appearance of the shoots and increased the number of explants with shoots from 75 to 84.6%. Further, shoots exposed for 7 days to half-strength MS supplemented with 10 mu M NAA (1-napthalene acetic acid) produced twice as many plantlets than those on half-strength MS alone. Transient GUS histochemical assays indicated delivery of the uidA gene via Agrobacterium tumefaciens.

Identifying risk-efficient strategies using stochastic frontier analysis and simulation: An application to irrigated cropping in Australia

In irrigated cropping, as with any other industry, profit and risk are inter-dependent. An increase in profit would normally coincide with an increase in risk, and this means that risk can be traded for profit. It is desirable to manage a farm so that it achieves the maximum possible profit for the desired level of risk. This paper identifies risk-efficient cropping strategies that allocate land and water between crop enterprises for a case study of an irrigated farm in Southern Queensland, Australia. This is achieved by applying stochastic frontier analysis to the output of a simulation experiment. The simulation experiment involved changes to the levels of business risk by systematically varying the crop sowing rules in a bioeconomic model of the case study farm. This model utilises the multi-field capability of the process based Agricultural Production System Simulator (APSIM) and is parameterised using data collected from interviews with a collaborating farmer. We found sowing rules that increased the farm area sown to cotton caused the greatest increase in risk-efficiency. Increasing maize area also improved risk-efficiency but to a lesser extent than cotton. Sowing rules that increased the areas sown to wheat reduced the risk-efficiency of the farm business. Sowing rules were identified that had the potential to improve the expected farm profit by ca. $50,000 Annually, without significantly increasing risk. The concept of the shadow price of risk is discussed and an expression is derived from the estimated frontier equation that quantifies the trade-off between profit and risk.

The need for speed: Timely prevention of the dispersal of noxious weeds in relief fodder using efficient sampling procedures

Invasive and noxious weeds are well known as a pervasive problem, imposing significant economic burdens on all areas of agriculture. Whilst there are multiple possible pathways of weed dispersal in this industry, of particular interest to this discussion is the unintended dispersal of weed seeds within fodder. During periods of drought or following natural disasters such as wild fire or flood, there arises the urgent need for 'relief' fodder to ensure survival and recovery of livestock. In emergency situations, relief fodder may be sourced from widely dispersed geographic regions, and some of these regions may be invaded by an extensive variety of weeds that are both exotic and detrimental to the intended destination for the fodder. Pasture hay is a common source of relief fodder and it typically consists of a mixture of grassy and broadleaf species that may include noxious weeds. When required urgently, pasture hay for relief fodder can be cut, baled, and transported over long distances in a short period of time, with little opportunity for prebaling inspection. It appears that, at the present time, there has been little effort towards rapid testing of bales, post-baling, for the presence of noxious weeds, as a measure to prevent dispersal of seeds. Published studies have relied on the analysis of relatively small numbers of bales, tested to destruction, in order to reveal seed species for identification and enumeration. The development of faster, more reliable, and non-destructive sampling methods is essential to increase the fodder industry's capacity to prevent the dispersal of noxious weeds to previously unaffected locales.

Efficient eucalypt cell wall deconstruction and conversion for sustainable lignocellulosic biofuels

In order to meet the world’s growing energy demand and reduce the impact of greenhouse gas emissions resulting from fossil fuel combustion, renewable plant-based feedstocks for biofuel production must be considered. The first-generation biofuels, derived from starches of edible feedstocks, such as corn, create competition between food and fuel resources, both for the crop itself and the land on which it is grown. As such, biofuel synthesized from non-edible plant biomass (lignocellulose) generated on marginal agricultural land will help to alleviate this competition. Eucalypts, the broadly defined taxa encompassing over 900 species of Eucalyptus, Corymbia, and Angophora are the most widely planted hardwood tree in the world, harvested mainly for timber, pulp and paper, and biomaterial products. More recently, due to their exceptional growth rate and amenability to grow under a wide range of environmental conditions, eucalypts are a leading option for the development of a sustainable lignocellulosic biofuels. However, efficient conversion of woody biomass into fermentable monomeric sugars is largely dependent on pretreatment of the cell wall, whose formation and complexity lend itself toward natural recalcitrance against its efficient deconstruction. A greater understanding of this complexity within the context of various pretreatments will allow the design of new and effective deconstruction processes for bioenergy production. In this review, we present the various pretreatment options for eucalypts, including research into understanding structure and formation of the eucalypt cell wall.