Pesticide residues in food - acute dietary exposure

Consumer risk assessment is a crucial step in the regulatory approval of pesticide use on food crops. Recently, an additional hurdle has been added to the formal consumer risk assessment process with the introduction of short-term intake or exposure assessment and a comparable short-term toxicity reference, the acute reference dose. Exposure to residues during one meal or over one day is important for short-term or acute intake. Exposure in the short term can be substantially higher than average because the consumption of a food on a single occasion can be very large compared with typical long-term or mean consumption and the food may have a much larger residue than average. Furthermore, the residue level in a single unit of a fruit or vegetable may be higher by a factor (defined as the variability factor, which we have shown to be typically ×3 for the 97.5th percentile unit) than the average residue in the lot. Available marketplace data and supervised residue trial data are examined in an investigation of the variability of residues in units of fruit and vegetables. A method is described for estimating the 97.5th percentile value from sets of unit residue data. Variability appears to be generally independent of the pesticide, the crop, crop unit size and the residue level. The deposition of pesticide on the individual unit during application is probably the most significant factor. The diets used in the calculations ideally come from individual and household surveys with enough consumers of each specific food to determine large portion sizes. The diets should distinguish the different forms of a food consumed, eg canned, frozen or fresh, because the residue levels associated with the different forms may be quite different. Dietary intakes may be calculated by a deterministic method or a probabilistic method. In the deterministic method the intake is estimated with the assumptions of large portion consumption of a ‘high residue’ food (high residue in the sense that the pesticide was used at the highest recommended label rate, the crop was harvested at the smallest interval after treatment and the residue in the edible portion was the highest found in any of the supervised trials in line with these use conditions). The deterministic calculation also includes a variability factor for those foods consumed as units (eg apples, carrots) to allow for the elevated residue in some single units which may not be seen in composited samples. In the probabilistic method the distribution of dietary consumption and the distribution of possible residues are combined in repeated probabilistic calculations to yield a distribution of possible residue intakes. Additional information such as percentage commodity treated and combination of residues from multiple commodities may be incorporated into probabilistic calculations. The IUPAC Advisory Committee on Crop Protection Chemistry has made 11 recommendations relating to acute dietary exposure.

Discrepancy Between Antennal and Behavioral Responses for Enantiomers of a-Pinene: Electrophysiology and Behavior of Helicoverpa Armigera (Lepidoptera)

The ability of adult cotton bollworm, Helicoverpa armigera (Hubner), to distinguish and respond to enantiomers of α-pinene was investigated with electrophysiological and behavioral methods. Electroantennogram recordings using mixtures of the enantiomers at saturating dose levels, and single unit electrophysiology, indicated that the two forms were detected by the same receptor neurons. The relative size of the electroantennogram response was higher for the (−) compared to the (+) form, indicating greater affinity for the (−) form at the level of the dendrites. Behavioral assays investigated the ability of moths to discriminate between, and respond to the (+) and (−) forms of α pinene. Moths with no odor conditioning showed an innate preference for (+)-α-pinene. This preference displayed by naıve moths was not significantly different from the preferences of moths conditioned on (+)-α-pinene. However, we found a significant difference in preference between moths conditioned on the (−) enantiomer compared to naıve moths and moths conditioned on (+)-α-pinene, showing that learning plays an important role in the behavioral response. Moths are less able to distinguish between enantiomers of α-pinene than different odors (e.g., phenylacetaldehyde versus (−)-α-pinene) in learning experiments. The relevance of receptor discrimination of enantiomers and learning ability of the moths in host plant choice is discussed.

Quantifying northward movement rates of eastern king prawns along eastern Australia

Movement rates of eastern king prawns, Melicertus plebejus (Hess), were estimated from historical and recent conventional tag-recapture information collected across eastern Australia. Data from three studies and 2,656 tag recaptures were used. Recaptured males and females both moved east–north-east in central Queensland and north–north-east in southern Queensland and New South Wales. Over a period of one year, the estimated transition matrix reflected the species strong northerly movement and the more complex longitudinal movement, showing a very high probability of eastern movement in central Queensland and almost negligible eastern or western movement in northern New South Wales. The high exchange probability between New South Wales and Queensland waters indicated that spatial assessment models with movement rates between state jurisdictions would improve the management of this single-unit stock.

Quantifying northward movement rates of eastern king prawns along eastern Australia

Movement rates of eastern king prawns, Melicertus plebejus (Hess), were estimated from historical and recent conventional tag-recapture information collected across eastern Australia. Data from three studies and 2,656 tag recaptures were used. Recaptured males and females both moved east-north-east in central Queensland and north-north-east in southern Queensland and New South Wales. Over a period of one year, the estimated transition matrix reflected the species strong northerly movement and the more complex longitudinal movement, showing a very high probability of eastern movement in central Queensland and almost negligible eastern or western movement in northern New South Wales. The high exchange probability between New South Wales and Queensland waters indicated that spatial assessment models with movement rates between state jurisdictions would improve the management of this single-unit stock.

Genetic variability of Tomato spotted wilt virus in Australia and validation of real time RT-PCR for its detection in single and bulked leaf samples

The potential for large-scale use of a sensitive real time reverse transcription polymerase chain reaction (RT-PCR) assay was evaluated for the detection of Tomato spotted wilt virus (TSWV) in single and bulked leaf samples by comparing its sensitivity with that of DAS-ELISA. Using total RNA extracted with RNeasy® or leaf soak methods, real time RT-PCR detected TSWV in all infected samples collected from 16 horticultural crop species (including flowers, herbs and vegetables), two arable crop species, and four weed species by both assays. In samples in which DAS-ELISA had previously detected TSWV, real time RT-PCR was effective at detecting it in leaf tissues of all 22 plant species tested at a wide range of concentrations. Bulk samples required more robust and extensive extraction methods with real time RT-PCR, but it generally detected one infected sample in 1000 uninfected ones. By contrast, ELISA was less sensitive when used to test bulked samples, once detecting up to 1 infected in 800 samples with pepper but never detecting more than 1 infected in 200 samples in tomato and lettuce. It was also less reliable than real time RT-PCR when used to test samples from parts of the leaf where the virus concentration was low. The genetic variability among Australian isolates of TSWV was small. Direct sequencing of a 587 bp region of the nucleoprotein gene (S RNA) of 29 isolates from diverse crops and geographical locations yielded a maximum of only 4.3% nucleotide sequence difference. Phylogenetic analysis revealed no obvious groupings of isolates according to geographic origin or host species. TSWV isolates, that break TSWV resistance genes in tomato or pepper did not differ significantly in the N gene region studied, indicating that a different region of the virus genome is responsible for this trait.

Development and Accreditation of an Applied Climate Education Unit for Sustainable Land Use in Australia

In recent years, there have been significant developments in climate science relevant to agriculture and natural resource management. Assessing impacts of climate variability and use of seasonal climate forecasts have become increasingly important elements in the management "toolkit" for many Australian farmers. Consideration of climate change further increases the need for improved management strategies. While climate risk extension activities have kept pace with advances in climate science, a national review of the Vocational Education and Training system in Australia in relation to "weather and climate" showed that these topics were "poorly represented" at the management level in the Australian Qualifications Framework, and needed increased emphasis. Consequently, a new Unit of Competency concerning management of climatic risk was developed and accredited to address this deficiency. The objective of the unit was to build knowledge and skills for better management of climate variability via the elements of surveying climatic and enterprise data; analysing climatic risks and opportunities; and developing climatic risk management strategies. This paper describes establishment of a new unit for vocational education that is designed to harness recent developments in applied climate science for better management of Australia's highly variable climate. The main benefits of the new unit of competency, "Developing climatic risk management strategies,"were seen as improving decisions in climate and agriculture, and reducing climate risk exposure to enhance sustainable agriculture. The educational unit is now within the scope of agricultural colleges, universities, and registered training organisations as an accredited unit.

Pasteurella multocida Expresses Two Lipopolysaccharide Glycoforms Simultaneously, but Only a Single Form Is Required for Virulence: Identification of Two Acceptor-Specific Heptosyl I Transferases

Lipopolysaccharide (LPS) is a critical virulence determinant in Pasteurella multocida and a major antigen responsible for host protective immunity. In other mucosal pathogens, variation in LPS or lipooligosaccharide structure typically occurs in the outer core oligosaccharide regions due to phase variation. P. multocida elaborates a conserved oligosaccharide extension attached to two different, simultaneously expressed inner core structures, one containing a single phosphorylated 3-deoxy-D-manno-octulosonic acid (Kdo) residue and the other containing two Kdo residues. We demonstrate that two heptosyltransferases, HptA and HptB, add the first heptose molecule to the Kdo1 residue and that each exclusively recognizes different acceptor molecules. HptA is specific for the glycoform containing a single, phosphorylated Kdo residue (glycoform A), while HptB is specific for the glycoform containing two Kdo residues (glycoform B). In addition, KdkA was identified as a Kdo kinase, required for phosphorylation of the first Kdo molecule. Importantly, virulence data obtained from infected chickens showed that while wild-type P. multocida expresses both LPS glycoforms in vivo, bacterial mutants that produced only glycoform B were fully virulent, demonstrating for the first time that expression of a single LPS form is sufficient for P. multocida survival in vivo. We conclude that the ability of P. multocida to elaborate alternative inner core LPS structures is due to the simultaneous expression of two different heptosyltransferases that add the first heptose residue to the nascent LPS molecule and to the expression of both a bifunctional Kdo transferase and a Kdo kinase, which results in the initial assembly of two inner core structures.

Genetic control of propagation traits in a single Corymbia torelliana x Corymbia variegata family

Genetic control of vegetative propagation traits was described for a second-generation, outbred, intersectional hybrid family (N = 208) derived from two species, Corymbia torelliana (F. Muell.) K.D. Hill & L.A.S. Johnson and Corymbia variegata (F. Muell.) K.D. Hill & L.A.S. Johnson, which contrast for propagation characteristics and in their capacity to develop lignotubers. Large phenotypic variances were evident for rooting and most other propagation traits, with significant proportions attributable to differences between clones (broad-sense heritabilities 0.2-0.5). Bare root assessment of rooting rate and root quality parameters tended to have the highest heritabilities, whereas rooting percentage based on root emergence from pots and shoot production were intermediate. Root biomass and root initiation had the lowest heritabilities. Strong favourable genetic correlations were found between rooting percentage and root quality traits such as root biomass, volume, and length. Lignotuber development on a seedling was associated with low rooting and a tendency to poor root quality in cuttings and was in accord with the persistence of species parent types due to gametic phase disequilibrium. On average, nodal cuttings rooted more frequently and with higher quality root systems, but significant cutting type x genotype interaction indicated that for some clones, higher rooting rates were obtained from tips. Low germination, survival of seedlings, and rooting rates suggested strong hybrid breakdown in this family.

Characterisation of chicken Campylobacter jejuni isolates using resolution optimised single nucleotide polymorphisms and binary gene markers

The principal objective of this study was to determine if Campylobacter jejuni genotyping methods based upon resolution optimised sets of single nucleotide polymorphisms (SNPs) and binary genetic markers were capable of identifying epidemiologically linked clusters of chicken-derived isolates. Eighty-eight C. jejuni isolates of known flaA RFLP type were included in the study. They encompassed three groups of ten isolates that were obtained at the same time and place and possessed the same flaA type. These were regarded as being epidemiologically linked. Twenty-six unlinked C. jejuni flaA type I isolates were included to test the ability of SNP and binary typing to resolve isolates that were not resolved by flaA RFLP. The remaining isolates were of different flaA types. All isolates were typed by real-time PCR interrogation of the resolution optimised sets of SNPs and binary markers. According to each typing method, the three epidemiologically linked clusters were three different clones that were well resolved from the other isolates. The 26 unlinked C. jejuni flaA type I isolates were resolved into 14 SNP-binary types, indicating that flaA typing can be unreliable for revealing epidemiological linkage. Comparison of the data with data from a fully typed set of isolates associated with human infection revealed that abundant lineages in the chicken isolates that were also found in the human isolates belonged to clonal complex (CC) -21 and CC-353, with the usually rare C-353 member ST-524 being especially abundant in the chicken collection. The chicken isolates selected to be diverse according to flaA were also diverse according to SNP and binary typing. It was observed that CC-48 was absent in the chicken isolates, despite being very common in Australian human infection isolates, indicating that this may be a major cause of human disease that is not chicken associated.

The use of high resolution melting (HRM) to map single nucleotide polymorphism markers linked to a covered smut resistance gene in barley

Using an established genetic map, a single gene conditioning covered smut resistance, Ruh.7H, was mapped to the telomere region of chromosome 7HS in an Alexis/Sloop doubled haploid barley population. The closest marker to Ruh.7H, abg704 was 7.5 cM away. Thirteen loci on the distal end of 7HS with potential to contain single nucleotide polymorphisms (SNPs) were identified by applying a comparative genomics approach using rice sequence data. Of these, one locus produced polymorphic co-dominant bands of different size while two further loci contained SNPs that were identified using the recently developed high resolution melting (HRM) technique. Two of these markers flanked Ruh.7H with the proximal marker located 3.8 cM and the distal marker 2.7 cM away. This is the first report on the application of the HRM technique to SNP detection and to rapid scoring of known cleaved amplified polymorphic sequence (CAPS) markers in plants. This simple, precise post-PCR technique should find widespread use in the fine-mapping of genetic regions of interest in complex cereal and other plant genomes.

Using automated supplementation systems to meet growth targets for grazing sheep

Remote drafting technology now available for sheep allows targeted supplementation of individuals within a grazing flock. This paper reports results of three experiments. Experiment 1 examined the weight change of Merino wethers allowed access to either lupin grain or whole cottonseed 0, 1, 2 or 7 days/week for 6 weeks. Experiment 2 examined the weight change of Merino wethers allowed access to either lupins or a sorghum + cottonseed meal (CSM) supplement 0, 2, 4 or 7 days/week for 8 weeks. Experiment 3 investigated the relationship between five allocations of trough space at the supplement self-feeders (5–50 cm/sheep) and the weight change of Merino wethers allowed access to lupins 1 day/week for 8 weeks. In all experiments, the Merino wethers had free access as a single group to drinking water and low quality hay in a large group pen and were allowed access to supplement once per day on their scheduled days of access. No water was available in the areas containing supplement, but one-way flow gates allowed animals to return to the group pen in their own time. There was a linear response in growth rate to increased frequency of access to lupins in Experiments 1 and 2, with each additional day of access increasing liveweight gain by 26 and 21 g/day, respectively. Similarly, the response to the sorghum + CSM supplement was linear, although significantly lower (P < 0.05), at 12 g/day. Providing access to whole cottonseed resulted in no significant change in growth rate compared with the control animals. In Experiment 3, decreasing trough space from 50 to 5 cm/sheep had no effect on sheep liveweight change. It was concluded that the relationships developed here, for growth response to increased frequency of access to lupins or a sorghum + CSM supplement, could be used to indicate the most appropriate frequency of access to supplement, through a remote drafting unit, to achieve sheep weight change targets. Also, that a trough space of 5 cm/sheep appears adequate in this supplementation system.

Measuring and predicting canopy nitrogen nutrition in wheat using a spectral index—The canopy chlorophyll content index (CCCI).

Varying the spatial distribution of applied nitrogen (N) fertilizer to match demand in crops has been shown to increase profits in Australia. Better matching the timing of N inputs to plant requirements has been shown to improve nitrogen use efficiency and crop yields and could reduce nitrous oxide emissions from broad acre grains. Farmers in the wheat production area of south eastern Australia are increasingly splitting N application with the second timing applied at stem elongation (Zadoks 30). Spectral indices have shown the ability to detect crop canopy N status but a robust method using a consistent calibration that functions across seasons has been lacking. One spectral index, the canopy chlorophyll content index (CCCI) designed to detect canopy N using three wavebands along the "red edge" of the spectrum was combined with the canopy nitrogen index (CNI), which was developed to normalize for crop biomass and correct for the N dilution effect of crop canopies. The CCCI-CNI index approach was applied to a 3-year study to develop a single calibration derived from a wheat crop sown in research plots near Horsham, Victoria, Australia. The index was able to predict canopy N (g m-2) from Zadoks 14-37 with an r2 of 0.97 and RMSE of 0.65 g N m-2 when dry weight biomass by area was also considered. We suggest that measures of N estimated from remote methods use N per unit area as the metric and that reference directly to canopy %N is not an appropriate method for estimating plant concentration without first accounting for the N dilution effect. This approach provides a link to crop development rather than creating a purely numerical relationship. The sole biophysical input, biomass, is challenging to quantify robustly via spectral methods. Combining remote sensing with crop modelling could provide a robust method for estimating biomass and therefore a method to estimate canopy N remotely. Future research will explore this and the use of active and passive sensor technologies for use in precision farming for targeted N management.

Using harvest statistics to monitor temporal variation in kangaroo density and harvest rate

Management of the commercial harvest of kangaroos relies on quotas set annually as a proportion of regular estimates of population size. Surveys to generate these estimates are expensive and, in the larger states, logistically difficult; a cheaper alternative is desirable. Rainfall is a disappointingly poor predictor of kangaroo rate of increase in many areas, but harvest statistics (sex ratio, carcass weight, skin size and animals shot per unit time) potentially offer cost-effective indirect monitoring of population abundance (and therefore trend) and status (i.e. under-or overharvest). Furthermore, because harvest data are collected continuously and throughout the harvested areas, they offer the promise of more intensive and more representative coverage of harvest areas than aerial surveys do. To be useful, harvest statistics would need to have a close and known relationship with either population size or harvest rate. We assessed this using longterm (11-22 years) data for three kangaroo species (Macropus rufus, M. giganteus and M. fuliginosus) and common wallaroos (M. robustus) across South Australia, New South Wales and Queensland. Regional variation in kangaroo body size, population composition, shooter efficiency and selectivity required separate analyses in different regions. Two approaches were taken. First, monthly harvest statistics were modelled as a function of a number of explanatory variables, including kangaroo density, harvest rate and rainfall. Second, density and harvest rate were modelled as a function of harvest statistics. Both approaches incorporated a correlated error structure. Many but not all regions had relationships with sufficient precision to be useful for indirect monitoring. However, there was no single relationship that could be applied across an entire state or across species. Combined with rainfall-driven population models and applied at a regional level, these relationships could be used to reduce the frequency of aerial surveys without compromising decisions about harvest management.

Testing the effectiveness of the SKIM foam fractionator to reduce nutrient levels in prawn farm effluent

To experimentally investigate the effect of the “SKIM” mechanical foam fractionator on suspended material and the nutrient levels in prawn farm effluent, a series of standardised short-term treatments were applied to various effluent types in a static 10,000-litre water body. Prawn pond effluents were characterised by watercolour and dominance of phytoplankton species. Three effluent types were tested, namely 1) particulate-rich effluent with little apparent phytoplankton, 2) green mircoalgal bloom predominately made up of single celled phytoplankton, and 3) brown microalgal bloom with higher prevalence of diatoms. The effluent types were similar (P>0.05) in non-volatile particulate material, and nitrate/nitrite but varied from each other in the following ways: 1) The particulate-rich effluents were lower (P<0.05) in volatile solids (compared to brown blooms), total Kjeldahl nitrogen, dissolved organic nitrogen, dissolved organic phosphorus and chlorophyll a (compared to both green and brown blooms). 2) The brown blooms were higher (P<0.05) in ammonia (compared to green blooms), total nitrogen and total phosphorus (compared to both green and particulate-rich effluent), but were lower (P<0.05) in inorganic phosphorus (compared to both green and particulate-rich effluent). 3) The green blooms were higher (P<0.05) in dissolved (both organic and inorganic) phosphorus (compared to both brown and particulate-rich effluents). Although the effluent types varied significantly in these aspects the effect of the Skim treatment was similar for all parameters measured except total phosphorus. Bloom type and Skim-treatment period significantly (P<0.05) affected total Kjeldahl phosphorus concentrations. For all effluent types there was a continuous significant reduction (P<0.05) in total Kjeldahl phosphorus during the initial 6-hour treatment period. Levels of total suspended solids and volatile suspended solids in all effluent types were significantly (P<0.05) reduced in the first 2 hours but not thereafter. Non-volatile suspended solids were also significantly (P<0.05) reduced in the first 2 hours (30 to 40 % reduction) and a further 40% reduction occurred in the particulate-rich effluent over the next 2 hours. Mean values for total ammonia, dissolved organic nitrogen, total Kjeldahl nitrogen, total nitrogen, chlorophyll a and dissolved organic or inorganic phosphorus levels were not significantly (P>0.05) affected by the Skim unit in any bloom type during the initial 6 hours of testing. Nevertheless, non-significant nitrogen reductions did occur. Foam production by the Skim unit varied with different blooms, resulting in different concentrate volumes and different end points for separate experiments. Concentrate volumes were generally high for the particulate-rich and green blooms (175 – 370 litres) and low for the brown blooms (25 – 80 litres). This was due to the low tendency of the brown bloom to produce foam. This generated higher nutrient concentrations in the associated condensed foam, but may have limited the treatment efficiency. The results suggest that in this application, the Skim unit did not remove micro-algae as effectively as was anticipated. However, it was effective at removing other suspended solids. Considering these attributes and the other uses of this machinery documented by the manufactures, the unit’s oxygenation mixing capacities coupled with inorganic solids removal may provide a suitable mechanism for construction of a continuously mixed bioreactor that utilises the filtration and profit making abilities of bivalves.

Phenotypic and genotypic variation within populations of kikuyu (Pennisetum clandestinum) in Australia.

Experiments at 2 sites in subtropical eastern Australia investigated the variation in agronomic attributes, quality and genetic structure existing within: naturally-occurring populations of kikuyu ( Pennisetum clandestinum) from within Australia; selections produced from the treatment of Whittet seed with mutagenic chemicals; and available cultivars. Runners were collected from coastal areas extending from Western Australia to the Atherton Tableland in north Queensland. One experiment evaluated 10 mutagenic selections and 4 cultivars in a lattice design and the other evaluated 12 ecotypes and 3 cultivars in a randomised block design. The experimental unit was single plants, which were sown on a 1.5 m grid into a weed-free seed-bed (Mutdapilly) or a killed kikuyu stand (Wollongbar), both of which were kept clear of weeds and other kikuyu plants for the duration of the experiments. Foliage height, forage production and runner yield were assessed. Leaf material was analysed for concentrations of crude protein (CP), acid detergent fibre (ADF) and neutral detergent fibre (NDF) and for in vitro dry matter digestibility (IVDDM) in autumn, winter and spring. DNA was extracted from each plant in the ecotype comparison and subjected to a modified DAF (DNA amplification fingerprinting) analysis to determine the level of genetic relatedness. In the first experiment, none of the mutagenic lines derived from Whittet yielded significantly more or was more digestible than commercial Whittet material, although some selections were superior to the other commercial kikuyu cultivars, Noonan and Crofts, and 'common' kikuyu. However, there were significant differences in plant height and runner expansion. In the second experiment, significant differences in plant height, foliage yield, runner development, and leaf CP, ADF, NDF and IVDDM concentrations were demonstrated between the ecotypes, mutagenic selections and cultivars. There was a 4- to 6-fold difference in plant yield and a 6- to 10-fold difference in runner production between the ecotypes at the 2 sites. Quality of the leaf ranged from 200 to 270 g/kg (CP), from 700 to 770 g/kg (IVDDM), from 170 to 250 g/kg (ADF) and from 470 to 550 g/kg (NDF). Improvements in quality and agronomic attributes were not mutually exclusive. Genetic fingerprint analysis of the kikuyu lines indicated that they formed 2 broad groupings. Most of the regional ecotypes were grouped with 'common' kikuyu as represented by the material collected from Wollongbar, and the Beechmont, Atherton Tableland and Gympie ecotypes were grouped with the registered cultivars Whittet, Noonan and Crofts. Two lines produced by mutagenesis from Whittet remained closely linked to Whittet. These results suggest that there was variation between populations of kikuyu in yield, quality and genetic diversity but that mutagenesis by treating seed with sodium azide and diethylene sulphide did not achieve a significant change in the digestibility of leaf over cv. Whittet.