A highly efficient and consistent method for harvesting large volumes of high-titre lentiviral vectors

Lentiviral vectors pseudotyped with vesicular stomatitis virus glycoprotein (VSV-G) are emerging as the vectors of choice for in vitro and in vivo gene therapy studies. However, the current method for harvesting lentivectors relies upon ultracentrifugation at 50 000 g for 2 h. At this ultra-high speed, rotors currently in use generally have small volume capacity. Therefore, preparations of large volumes of high-titre vectors are time-consuming and laborious to perform. In the present study, viral vector supernatant harvests from vector-producing cells (VPCs) were pre-treated with various amounts of poly-L-lysine (PLL) and concentrated by low speed centrifugation. Optimal conditions were established when 0.005% of PLL (w/v) was added to vector supernatant harvests, followed by incubation for 30 min and centrifugation at 10 000 g for 2 h at 4 degreesC. Direct comparison with ultracentrifugation demonstrated that the new method consistently produced larger volumes (6 ml) of high-titre viral vector at 1 x 10(8) transduction unit (TU)/ml (from about 3000 ml of supernatant) in one round of concentration. Electron microscopic analysis showed that PLL/viral vector formed complexes, which probably facilitated easy precipitation at low-speed concentration (10 000 g), a speed which does not usually precipitate viral particles efficiently. Transfection of several cell lines in vitro and transduction in vivo in the liver with the lentivector/PLL complexes demonstrated efficient gene transfer without any significant signs of toxicity. These results suggest that the new method provides a convenient means for harvesting large volumes of high-titre lentivectors, facilitate gene therapy experiments in large animal or human gene therapy trials, in which large amounts of lentiviral vectors are a prerequisite.

Optimisation-based Design of a Manipulator for Harvesting Capsicum

This paper presents a global-optimisation frame-work for the design of a manipulator for harvesting capsicum(peppers) in the field. The framework uses a simulated capsicum scenario with automatically generated robot models based on DH parameters. Each automatically generated robot model is then placed in the simulated capsicum scenario and the ability of the robot model to get to several goals (capsicum with varying orientations and positions) is rated using two criteria:the length of a collision-free path and the dexterity of the end-effector. These criteria form the basis of the objective function used to perform a global optimisation. The paper shows a preliminary analysis and results that demonstrate the potential of this method to choose suitable robot models with varying degrees of freedom.

On Visual Detection of Highly-occluded Objects for Harvesting Automation in Horticulture

Developing accurate and reliable crop detection algorithms is an important step for harvesting automation in horticulture. This paper presents a novel approach to visual detection of highly-occluded fruits. We use a conditional random field (CRF) on multi-spectral image data (colour and Near-Infrared Reflectance, NIR) to model two classes: crop and background. To describe these two classes, we explore a range of visual-texture features including local binary pattern, histogram of oriented gradients, and learn auto-encoder features. The pro-posed methods are evaluated using hand-labelled images from a dataset captured on a commercial capsicum farm. Experimental results are presented, and performance is evaluated in terms of the Area Under the Curve (AUC) of the precision-recall curves.Our current results achieve a maximum performance of 0.81AUC when combining all of the texture features in conjunction with colour information.

Improved Algorithms for Reservoir Capacity Calculation Incorporating Storage-Dependent Losses and Reliability Norm

Two algorithms that improve upon the sequent-peak procedure for reservoir capacity calculation are presented. The first incorporates storage-dependent losses (like evaporation losses) exactly as the standard linear programming formulation does. The second extends the first so as to enable designing with less than maximum reliability even when allowable shortfall in any failure year is also specified. Together, the algorithms provide a more accurate, flexible and yet fast method of calculating the storage capacity requirement in preliminary screening and optimization models.

Induced Resistance: Potential for Control of Postharvest Diseases of Horticultural Crops

Fortunately, plants have developed highly effective mechanisms with which to defend themselves when attacked by potentially disease-causing microorganisms. If not, then they would succumb to the many pathogenic fungi, bacteria, viruses, nematodes and insect pests, and disease would prevail. These natural defence systems of plants can be deliberately activated to provide some protection against the major pathogens responsible for causing severe yield losses in agricultural and horticultural crops. This is the basis of what is known as ‘induced’ or ‘acquired’ disease resistance in plants. Although the phenomenon of induced resistance has been known amongst plant pathologists for over 100 years, its inclusion into pest and disease management programmes has been a relatively recent development, ie. within the last 5 years. This review will discuss very briefly some of the characteristics of the induced resistance phenomenon, outline some of the advantages and limitations to its implementation and provide some examples within a postharvest pathology context. Finally some approaches being investigated by the fruit pathology team at DPI Indooroopilly and collaborators will be outlined.

Cooking in the ungra area: Fuel efficiency, energy losses, and opportunities for reducing firewood consumption

Cooking efficiency and related fuel economy issues have been studied in a particular rural area of India. Following a description of the cooking practices and conditions in this locale, cooking efficiency is examined. A cooking efficiency of only 6% was found. The use of aluminium rather than clay pots results in an increased efficiency. In addition, cooking efficiency correlates very well with specific fuel consumption. The latter parameter is much simpler to analyse than cooking efficiency. The energy losses during cooking are examined in the second part of this case study. The major energy losses are heating of excess air, heat carried away by the combustion products, heat transmitted to the stove body and floor, and the chemical energy in charcoal residue. The energy loss due to the evaporation of cooking water is also significant because it represents about one-third of the heat reaching the pots.

Evaluation of Charges for Power Transmission and Losses in Bilateral Power Contracts

The paper presents a method for transmission loss charge allocation in deregulated power systems based on Relative Electrical Distance (RED) concept. Based on RED between the generator and load nodes and the predefined bilateral power contracts, charge evaluation is carried out. Generally through some power exchange mechanism a set of bilateral contracts are determined that facilitate bilateral agreements between the generation and distribution entities. In this paper the possible charges incurred in meeting loads like generation charge, transmission charge and charge due to losses are evaluated. Case studies have been carried out on a few practical equivalent systems. Due to space limitation results for a sample 5 bus system are presented considering ideal load/generation power contracts and deviated load/generation power contracts. Extensive numerical testing indicates that the proposed allocation scheme produces loss allocations that are appropriate and that behave in a physically reasonable manner.

Can feral pig harvesting and co-ordinated control reduce damage to grain crops?

Commercial and recreational harvesting of pigs is often encouraged by pest managers because it is essentially a ‘free’ reduction in pest density. However, the reduction in numbers may provide minimal damage mitigation and may be inappropriately allocated in space and time. Additionally, more effective control (e.g. baiting) may not occur because of the incorrect perception that harvesting is effective or because pigs are valued for recreational use.

Losses of nitrogen in surface runoff from a plantation horticulture farm in coastal Queensland, Australia.

Surface losses of nitrogen from horticulture farms in coastal Queensland, Australia, may have the potential to eutrophy sensitive coastal marine habitats nearby. A case-study of the potential extent of such losses was investigated in a coastal macadamia plantation. Nitrogen losses were quantified in 5 consecutive runoff events during the 13-month study. Irrigation did not contribute to surface flows. Runoff was generated by storms at combined intensities and durations that were 20–40 mm/h for >9 min. These intensities and durations were within expected short-term (1 year) and long-term (up to 20 years) frequencies of rainfall in the study area. Surface flow volumes were 5.3 ± 1.1% of the episodic rainfall generated by such storms. Therefore, the largest part of each rainfall event was attributed to infiltration and drainage in this farm soil (Kandosol). The estimated annual loss of total nitrogen in runoff was 0.26 kg N/ha.year, representing a minimal loading of nitrogen in surface runoff when compared to other studies. The weighted average concentrations of total sediment nitrogen (TSN) and total dissolved nitrogen (TDN) generated in the farm runoff were 2.81 ± 0.77% N and 1.11 ± 0.27 mg N/L, respectively. These concentrations were considerably greater than ambient levels in an adjoining catchment waterway. Concentrations of TSN and TDN in the waterway were 0.11 ± 0.02% N and 0.50 ± 0.09 mg N/L, respectively. The steep concentration gradient of TSN and TDN between the farm runoff and the waterway demonstrated the occurrence of nutrient loading from the farming landscapes to the waterway. The TDN levels in the stream exceeded the current specified threshold of 0.2–0.3 mg N/L for eutrophication of such a waterway. Therefore, while the estimate of annual loading of N from runoff losses was comparatively low, it was evident that the stream catchment and associated agricultural land uses were already characterised by significant nitrogen loadings that pose eutrophication risks. The reported levels of nitrogen and the proximity of such waterways (8 km) to the coastline may have also have implications for the nearshore (oligotrophic) marine environment during periods of turbulent flow.

Increased reproductive success in older female red kangaroos and the impact of harvesting

Wildlife harvesting has a long history in Australia, including obvious examples of overexploitation. Not surprisingly, there is scepticism that commercial harvesting can be undertaken sustainably. Kangaroo harvesting has been challenged regularly at Administrative Appeals Tribunals and elsewhere over the past three decades. Initially, the concern from conservation groups was sustainability of the harvest. This has been addressed through regular, direct monitoring that now spans > 30 years and a conservative harvest regime with a low risk of overharvest in the face of uncertainty. Opposition to the harvest now continues from animal rights groups whose concerns have shifted from overall harvest sustainability to side effects such as animal welfare, and changes to community structure, genetic composition and population age structure. Many of these concerns are speculative and difficult to address, requiring expensive data. One concern is that older females are the more successful breeders and teach their daughters optimal habitat and diet selection. The lack of older animals in a harvested population may reduce the fitness of the remaining individuals; implying population viability would also be compromised. This argument can be countered by the persistence of populations under harvesting without any obvious impairment to reproduction. Nevertheless, an interesting question is how age influences reproductive output. In this study, data collected from a number of red kangaroo populations across eastern Australia indicate that the breeding success of older females is up to 7-20% higher than that of younger females. This effect is smaller than that of body condition and the environment, which can increase breeding success by up to 30% and 60% respectively. Average age of mature females in a population may be reduced from 9 to 6 years old, resulting in a potential reduction in breeding success of 3-4%. This appears to be offset in harvested populations by improved condition of females from a reduction in kangaroo density. There is an important recommendation for management. The best insurance policy against overharvest and unwanted side effects is not research, which could be never-ending. Rather, it is a harvest strategy that includes safeguards against uncertainty such as harvest reserves, conservative quotas and regular monitoring. Research is still important in fine tuning that strategy and is most usefully incorporated as adaptive management where it can address the key questions on how populations respond to harvesting.

Gaseous Nitrogen Losses Following Soil Amendment with Biosolids under Controlled Conditions.

This paper quantifies gaseous N losses due to ammonia volatilisation and denitrification under controlled conditions at 30 degrees C and 75% to 150% of Field Capacity (FC). Biosolids were mixed with two contrasting soils from subtropical Australia at a rate designed to meet crop N requirements for irrigated cotton or maize (i.e., equivalent to 180 kg N ha(-1)). In the first experiment, aerobically (AE) and anaerobically (AN) digested biosolids were mixed into a heavy Vertosol soil and then incubated for 105 days. Ammonia volatilization over 72 days accounted for less than 4% of the applied NH4-N but 24% (AN) to 29% (AE) of the total applied biosolids' N was lost through denitrification in 105 days. In the second experiment AN biosolids with and without added polyacrimide polymer were mixed with either a heavy Vertosol or a lighter Red Ferrosol and then incubated for 98 days. The N loss was higher from the Vertosol with 16-29% of total N applied versus the Red Ferrosol with 7-10% of total N applied, while addition of polymer to the biosolids increased N loss from 7 to 10% and from 16 to 29% in the Red Ferrosol and Vertosol, respectively. A major product from the denitrification process was N-2 gas, accounting for >90% of the emitted N gases from both experiments. Our findings demonstrate that denitrification could be a major pathway of gaseous N losses under warm and moist conditions.

Efficient Energy Management Policies for Networks with Energy Harvesting Sensor Nodes

We study sensor networks with energy harvesting nodes. The generated energy at a node can be stored in a buffer. A sensor node periodically senses a random field and generates a packet. These packets are stored in a queue and transmitted using the energy available at that time at the node. For such networks we develop efficient energy management policies. First, for a single node, we obtain policies that are throughput optimal, i.e., the data queue stays stable for the largest possible data rate. Next we obtain energy management policies which minimize the mean delay in the queue. We also compare performance of several easily implementable suboptimal policies. A greedy policy is identified which, in low SNR regime, is throughput optimal and also minimizes mean delay. Next using the results for a single node, we develop efficient MAC policies.

Influence of harvesting strategy on nutrient supply and production of dairy cows consuming diets based on grass and red clover silage

The main objective of this thesis was to elucidate the effects of regrowth grass silage and red clover silage on nutrient supply and milk production of dairy cows as compared with primary growth grass silages. In the first experiment (publication I), two primary growth and four regrowth grass silages were harvested at two stages of growth. These six silages were fed to 24 lactating dairy cows with two levels of concentrate allowance. Silage intake and energy corrected milk yield (ECM) responses, and the range in these response variables between the diets, were smaller when regrowth silages rather than primary growth silages were fed. Milk production of dairy cows reflected the intake of metabolizable energy (ME), and no differences in the ME utilization were found between the diets based on silages harvested from primary growth and regrowth. The ECM response to increased concentrate allowance was, on average, greater when regrowth rather than primary growth silages were fed. In the second experiment (publication II), two silages from primary growth and two from regrowth used in I were fed to rumen cannulated lactating dairy cows. Cows consumed less feed dry matter (DM), energy and protein, and produced less milk, when fed diets based on regrowth silages rather than primary growth silages. Lower milk production responses of regrowth grass silage diets were mainly due to the lower silage DM intake, and could not be accounted for by differences in energy or protein utilization. Regrowth grass silage intake was not limited due to neutral detergent fibre (NDF) digestion or rumen fill or passage kinetics. However, lower intake may be at least partly attributable to plant diseases such as leaf spot infections, dead deteriorating material or abundance of weeds, which are all higher in regrowth compared with primary growth, and increase with advancing regrowth. In the third experiment (publications III and IV), red clover silages and grass silages harvested at two stages of growth, and a mixed diet of red clover and grass silages, were fed to five rumen cannulated lactating dairy cows. In spite of the lower average ME intake for red clover diets, the ECM production remained unchanged suggesting more efficient utilisation of ME for red clover diets compared with grass diets. Intake of N, and omasal canal flows of total non-ammonia N (NAN), microbial and non-microbial NAN were higher for red clover than for grass silage diets, but were not affected by forage maturity. Delaying the harvest tended to decrease DM intake of grass silage and increase that of red clover silage. The digestion rate of potentially digestible NDF was faster for red clover diets than for grass silage diets. Delaying the harvest decreased the digestion rate for grass but increased it for red clover silage diets. The low intake of early-cut red clover silage could not be explained by silage digestibility, fermentation quality, or rumen fill but was most likely related to the nutritionally suboptimal diet composition because inclusion of moderate quality grass silage in mixed diet increased silage DM intake. Despite the higher total amino acid supply of cows fed red clover versus grass silage diets, further milk production responses on red clover diets were possibly compromised by an inadequate supply of methionine as evidenced by lower methionine concentration in the amino acid profile of omasal digesta and plasma. Increasing the maturity of ensiled red clover does not seem to affect silage DM intake as consistently as that of grasses. The efficiency of N utilization for milk protein synthesis was lower for red clover diets than for grass diets. It was negatively related to diet crude protein concentration similarly to grass silage diets.

Yield losses in grain sorghum due to rust infection

Although rust (caused by Puccinia purpurea) is a common disease in Australian grain sorghum crops, particularly late in the growing season (April onwards), its potential to reduce yield has not been quantified. Field trials were conducted in Queensland between 2003 and 2005 to evaluate the effect of sorghum rust on grain yield of two susceptible sorghum hybrids (Tx610 and Pride). Rust was managed from 28-35 days after sowing until physiological maturity by applying oxycarboxin (1 kg active ingredient/100 L of water/ha) every 10 days. When data were combined for the hybrids, yield losses ranged from 13.1% in 2005 to 3.2% in 2003 but differences in yield the between sprayed and unsprayed treatments were statistically significant (P a parts per thousand currency signaEuro parts per thousand 0.05) only in 2005. Final area under the disease progress curve (AUDPC) values reflected the yield losses in each year. The higher yield loss in 2005 can be attributed primarily to the early development of the rust epidemic and the higher inoculum levels in spreader plots at the time of planting of the trials.