Improved Approach for Analyzing Bromophenols in Seafood Using Stable Isotope Dilution Analysis in Combination with SPME

An analytical method for the measurement of five naturally occurring bromophenols of sensory relevance in seafood (barramundi and prawns) is presented. The method combines simultaneous distillation−extraction followed by alkaline back extraction of a hexane extract and subsequent acetylation of the bromophenols. Analysis of the bromophenol acetates was accomplished by headspace solid phase microextraction and gas chromatography−mass spectrometry using selected ion monitoring. The addition of 13C6 bromophenol stable isotope internal standards for each of the five congeners studied permitted the accurate quantitation of 2-bromophenol, 4-bromophenol, 2,6-dibromophenol, 2,4-dibromophenol, and 2,4,6-tribromophenol down to a limit of quantification of 0.05 ng/g of fish flesh. The method indicated acceptable precision and repeatability and excellent linearity over the typical concentration range of these compounds in seafood (0.5−50 ng/g). The analytical method was applied to determine the concentration of bromophenols in a range of farmed and wild barramundi and prawns and was also used to monitor bromophenol uptake in a pilot feeding trial.

Frequency and precision of aerial surveys for kangaroo management

The appropriate frequency and precision for surveys of wildlife populations represent a trade-off between survey cost and the risk of making suboptimal management decisions because of poor survey data. The commercial harvest of kangaroos is primarily regulated through annual quotas set as proportions of absolute estimates of population size. Stochastic models were used to explore the effects of varying precision, survey frequency and harvest rate on the risk of quasiextinction for an arid-zone and a more mesic-zone kangaroo population. Quasiextinction probability increases in a sigmoidal fashion as survey frequency is reduced. The risk is greater in more arid regions and is highly sensitive to harvest rate. An appropriate management regime involves regular surveys in the major harvest areas where harvest rate can be set close to the maximum sustained yield. Outside these areas, survey frequency can be reduced in relatively mesic areas and reduced in arid regions when combined with lowered harvest rates. Relative to other factors, quasiextinction risk is only affected by survey precision (standard error/mean × 100) when it is >50%, partly reflecting the safety of the strategy of harvesting a proportion of a population estimate.

Prioritising potential guilds of specialist herbivores as biological control agents for prickly acacia through simulated herbivory

Understanding plant response to herbivory facilitates the prioritisation of guilds of specialist herbivores as biological control agents based on their potential impacts. Prickly acacia (Acacia nilotica ssp. indica) is a weed of national significance in Australia and is a target for biological control. Information on the susceptibility of prickly acacia to herbivory is limited, and there is no information available on the plant organ (i.e. leaf, shoot and root in isolation or in combination) most susceptible to herbivory. We evaluated the ability of prickly acacia seedlings, to respond to different types of simulated herbivory (defoliation, shoot damage, root damage and combinations), at varying frequencies (no herbivory, single, two and three events of herbivory) to identify the type and frequency of herbivory that will be required to reduce the growth and vigour. Defoliation and shoot damage, individually, had a significant negative impact on prickly acacia seedlings. For the defoliation to be effective, more than two defoliation events were required, whereas a single bout of shoot damage was enough to cause a significant reduction in plant vigour. A combination of defoliation + shoot damage had the greatest negative impact. The study highlights the need to prioritise specialist leaf and shoot herbivores as potential biological control agents for prickly acacia.

The meleagrid herpesvirus 1 genome is partially resistant to transposition

The propagation of herpesvirus genomes as infectious bacterial artificial chromosomes (iBAC) has enabled the application of highly efficient strategies to investigate gene function across the genome. One of these strategies, transposition, has been used successfully on a number of herpesvirus iBACs to generate libraries of gene disruption mutants. Gene deletion studies aimed at determining the dispensable gene repertoire of the Meleagrid herpesvirus 1 (MeHV-1) genome to enhance the utility of this virus as a vaccine vector have been conducted in this report. A MeHV-1 iBAC was used in combination with the Tn5 and MuA transposition systems in an attempt to generate MeHV-1 gene interruption libraries. However, these studies demonstrated that Tn5 transposition events into the MeHV-1 genome occurred at unexpectedly low frequencies. Furthermore, characterization of genomic locations of the rare Tn5 transposon insertion events indicated a nonrandom distribution within the viral genome, with seven of the 24 insertions occurring within the gene encoding infected cell protein 4. Although insertion events with the MuA system occurred at higher frequency compared with the Tn5 system, fewer insertion events were generated than has previously been reported with this system. The characterization and distribution of these MeHV-1 iBAC transposed mutants is discussed at both the nucleotide and genomic level, and the properties of the MeHV-1 genome that could influence transposition frequency are discussed. © American Association of Avian Pathologists.

Rapid genome wide mapping of phosphine resistance loci by a simple regional averaging analysis in the red flour beetle, Tribolium castaneum

Background Next-generation sequencing technology is an important tool for the rapid, genome-wide identification of genetic variations. However, it is difficult to resolve the ‘signal’ of variations of interest and the ‘noise’ of stochastic sequencing and bioinformatic errors in the large datasets that are generated. We report a simple approach to identify regional linkage to a trait that requires only two pools of DNA to be sequenced from progeny of a defined genetic cross (i.e. bulk segregant analysis) at low coverage (<10×) and without parentage assignment of individual SNPs. The analysis relies on regional averaging of pooled SNP frequencies to rapidly scan polymorphisms across the genome for differential regional homozygosity, which is then displayed graphically. Results Progeny from defined genetic crosses of Tribolium castaneum (F4 and F19) segregating for the phosphine resistance trait were exposed to phosphine to select for the resistance trait while the remainders were left unexposed. Next generation sequencing was then carried out on the genomic DNA from each pool of selected and unselected insects from each generation. The reads were mapped against the annotated T. castaneum genome from NCBI (v3.0) and analysed for SNP variations. Since it is difficult to accurately call individual SNP frequencies when the depth of sequence coverage is low, variant frequencies were averaged across larger regions. Results from regional SNP frequency averaging identified two loci, tc_rph1 on chromosome 8 and tc_rph2 on chromosome 9, which together are responsible for high level resistance. Identification of the two loci was possible with only 5-7× average coverage of the genome per dataset. These loci were subsequently confirmed by direct SNP marker analysis and fine-scale mapping. Individually, homozygosity of tc_rph1 or tc_rph2 results in only weak resistance to phosphine (estimated at up to 1.5-2.5× and 3-5× respectively), whereas in combination they interact synergistically to provide a high-level resistance >200×. The tc_rph2 resistance allele resulted in a significant fitness cost relative to the wild type allele in unselected beetles over eighteen generations. Conclusion We have validated the technique of linkage mapping by low-coverage sequencing of progeny from a simple genetic cross. The approach relied on regional averaging of SNP frequencies and was used to successfully identify candidate gene loci for phosphine resistance in T. castaneum. This is a relatively simple and rapid approach to identifying genomic regions associated with traits in defined genetic crosses that does not require any specialised statistical analysis.

High frequency fire alters C:N:P stoichiometry in forest litter

Fire is a major driver of ecosystem change and can disproportionately affect the cycling of different nutrients. Thus, a stoichiometric approach to investigate the relationships between nutrient availability and microbial resource use during decomposition is likely to provide insight into the effects of fire on ecosystem functioning. We conducted a field litter bag experiment to investigate the long-term impact of repeated fire on the stoichiometry of leaf litter C, N and P pools, and nutrient-acquiring enzyme activities during decomposition in a wet sclerophyll eucalypt forest in Queensland, Australia. Fire frequency treatments have been maintained since 1972, including burning every two years (2yrB), burning every four years (4yrB) and no burning (NB). C:N ratios in freshly fallen litter were 29-42% higher and C:P ratios were 6-25% lower for 2yrB than NB during decomposition, with correspondingly lower 2yrB N:P ratios (27-32) than for NB (34-49). Trends in litter soluble and microbial N:P ratios were similar to the overall litter N:P ratios across fire treatments. Consistent with these, the ratio of activities for N-acquiring to P-acquiring enzymes in litter was higher for 2yrB than NB while 4yrB was generally intermediate between 2yrB and NB. Decomposition rates of freshly fallen litter were significantly lower for 2yrB (72±2% mass remaining at the end of experiment) than for 4yrB (59±3%) and NB (62±3%), a difference that may be related to effects of N limitation, lower moisture content, and/or litter C quality. Results for older mixed-age litter were similar to those for freshly fallen litter although treatment differences were less pronounced. Overall, these findings show that frequent fire (2yrB) decoupled N and P cycling, as manifested in litter C:N:P stoichiometry and in microbial biomass N:P ratio and enzymatic activities. These data indicate that fire induced a transient shift to N-limited ecosystem conditions during the post-fire recovery phase. This article is protected by copyright. All rights reserved.

An unusual combination in papaya ( Carica papaya): the good (glucosinolates) and the bad (cyanogenic glycosides)

Glucosinolates are a group of sulphur-containing glycosides found in the plant order Brassicales which includes the Brassica vegetables such as broccoli, cabbage and cauliflower. When brought into contact with the plant enzymes, myrosinases, the glucosinolates break down releasing glucose and other products which serve principally in plant defence against herbivores. The most important of the products from a human nutritional viewpoint, are the isothiocyanates. These potent inducers of detoxifying enzymes bestow the distinct anti-cancer properties on these plants. Unique among tropical fruits, papaya is known to contain an abundance of one particular glucosinolate, glucotropaeolin. Other compounds that play a pivotal role in the chemical defence system of many plants are the cyanogenic glycosides. Cyanogenic glycosides are activated by plant enzymes in the event of pest attack, releasing the deterrent: toxic hydrogen cyanide. Papaya, in addition to glucosinolates, also contains low levels of cyanogenic glycosides, an unusual occurrence because it was assumed that the two classes of metabolites were mutually exclusive. Studies measuring the levels of both in the edible parts of the papaya fruit and other utilised tissues are discussed and considered in the context of potential human health ramifications. All rights reserved, Elsevier.

Issues and solutions for researching weed eradication target species

Species biology drives the frequency, duration and extent of survey and control activities in weed eradication programs. Researching the key biological characters can be difficult when plants occur at limited locations and are controlled immediately by field crews who are dedicated to preventing reproduction. Within the National Four Tropical Weeds Eradication Program and the former National Siam Weed Eradication Program, key information needed by the eradication teams has been obtained through a combination of field, glasshouse and laboratory studies without jeopardising the eradication objective. Information gained on seed longevity, age to reproductive maturity, dispersal and control options has been used to direct survey and control activities. Planned and opportunistic data collections will continue to provide biological information to refine eradication activities.

Impact of nitrification inhibitor (DMPP) on soil nitrous oxide emissions from an intensive broccoli production system in sub-tropical Australia

Vegetable cropping systems are often characterised by high inputs of nitrogen fertiliser. Elevated emissions of nitrous oxide (N2O) can be expected as a consequence. In order to mitigate N2O emissions from fertilised agricultural fields, the use of nitrification inhibitors, in combination with ammonium based fertilisers, has been promoted. However, no data is currently available on the use of nitrification inhibitors in sub-tropical vegetable systems. A field experiment was conducted to investigate the effect of the nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP) on N2O emissions and yield from broccoli production in sub-tropical Australia. Soil N2O fluxes were monitored continuously (3 h sampling frequency) with fully automated, pneumatically operated measuring chambers linked to a sampling control system and a gas chromatograph. Cumulative N2O emissions over the 5 month observation period amounted to 298 g-N/ha, 324 g-N/ha, 411 g-N/ha and 463 g-N/ha in the conventional fertiliser (CONV), the DMPP treatment (DMPP), the DMMP treatment with a 10% reduced fertiliser rate (DMPP-red) and the zero fertiliser (0N), respectively. The temporal variation of N2O fluxes showed only low emissions over the broccoli cropping phase, but significantly elevated emissions were observed in all treatments following broccoli residues being incorporated into the soil. Overall 70–90% of the total emissions occurred in this 5 weeks fallow phase. There was a significant inhibition effect of DMPP on N2O emissions and soil mineral N content over the broccoli cropping phase where the application of DMPP reduced N2O emissions by 75% compared to the standard practice. However, there was no statistical difference between the treatments during the fallow phase or when the whole season was considered. This study shows that DMPP has the potential to reduce N2O emissions from intensive vegetable systems, but also highlights the importance of post-harvest emissions from incorporated vegetable residues. N2O mitigation strategies in vegetable systems need to target these post-harvest emissions and a better evaluation of the effect of nitrification inhibitors over the fallow phase is needed.

Floristic composition and pasture condition of Aristida/Bothriochloa pastures in central Queensland. I. Pasture floristics

A survey was conducted in central inland Queensland, Australia of 108 sites that were deemed to contain Aristida/Bothriochloa native pastures to quantitatively describe the pastures and attempt to delineate possible sub-types. The pastures were described in terms of their floristic composition, plant density and crown cover. There were generally ~20 (range 5–33) main pasture species at a site. A single dominant perennial grass was rare with three to six prominent species the norm. Chrysopogon fallax (golden-beard grass) was the perennial grass most consistently found in all pastures whereas Aristida calycina (dark wiregrass), Enneapogon spp. (bottlewasher grasses), Brunoniella australis (blue trumpet) and Panicum effusum (hairy panic) were all regularly present. The pastures did not readily separate into broad floristic sub-groups, but three groups that landholders could recognise from a combination of the dominant tree and soil type were identified. The three groups were Eucalyptus crebra (narrow-leaved ironbark), E. melanophloia (silver-leaved ironbark) and E. populnea (poplar box). The pastures of the three main sub-groups were then characterised by the prominent presence, singly or in combination, of Bothriochloa ewartiana (desert bluegrass), Eremochloa bimaculata (poverty grass), Bothriochloa decipiens (pitted bluegrass) or Heteropogon contortus (black speargrass). The poplar box group had the greatest diversity of prominent grasses whereas the narrow-leaved ironbark group had the least. Non-native Cenchrus ciliaris (buffel grass) and Melinis repens (red Natal grass) were generally present at low densities. Describing pastures in terms of frequency of a few species or species groups sometimes failed to capture the true nature of the pasture but plant abundance for most species, as density, herbage mass of dry matter or plant crown cover, was correlated with its recorded frequency. A quantitative description of an average pasture in fair condition is provided but it was not possible to explain why some species often occur together or fail to co-exist in Aristida/Bothriochloa pastures, for example C. ciliaris and E. bimaculata rarely co-exist whereas Tragus australianus (small burrgrass) and Enneapogon spp. are frequently recorded together. Most crown cover was provided by perennial grasses but many of these are Aristida spp. (wiregrasses) and not regarded as useful forage for livestock. No new or improved categorisation of the great variation evident in the Aristida/Bothriochloa native pasture type can be given despite the much improved detail provided of the floristic composition by this survey.

Sequence diversity and differential expression of major phenylpropanoid-flavonoid biosynthetic genes among three mango varieties

Background: Mango fruits contain a broad spectrum of phenolic compounds which impart potential health benefits; their biosynthesis is catalysed by enzymes in the phenylpropanoid-flavonoid (PF) pathway. The aim of this study was to reveal the variability in genes involved in the PF pathway in three different mango varieties Mangifera indica L., a member of the family Anacardiaceae: Kensington Pride (KP), Irwin (IW) and Nam Doc Mai (NDM) and to determine associations with gene expression and mango flavonoid profiles. Results: A close evolutionary relationship between mango genes and those from the woody species poplar of the Salicaceae family (Populus trichocarpa) and grape of the Vitaceae family (Vitis vinifera), was revealed through phylogenetic analysis of PF pathway genes. We discovered 145 SNPs in total within coding sequences with an average frequency of one SNP every 316bp. Variety IW had the highest SNP frequency (one SNP every 258bp) while KP and NDM had similar frequencies (one SNP every 369bp and 360bp, respectively). The position in the PF pathway appeared to influence the extent of genetic diversity of the encoded enzymes. The entry point enzymes phenylalanine lyase (PAL), cinnamate 4-mono-oxygenase (C4H) and chalcone synthase (CHS) had low levels of SNP diversity in their coding sequences, whereas anthocyanidin reductase (ANR) showed the highest SNP frequency followed by flavonoid 3'-hydroxylase (F3'H). Quantitative PCR revealed characteristic patterns of gene expression that differed between mango peel and flesh, and between varieties. Conclusions: The combination of mango expressed sequence tags and availability of well-established reference PF biosynthetic genes from other plant species allowed the identification of coding sequences of genes that may lead to the formation of important flavonoid compounds in mango fruits and facilitated characterisation of single nucleotide polymorphisms between varieties. We discovered an association between the extent of sequence variation and position in the pathway for up-stream genes. The high expression of PAL, C4H and CHS genes in mango peel compared to flesh is associated with high amounts of total phenolic contents in peels, which suggest that these genes have an influence on total flavonoid levels in mango fruit peel and flesh. In addition, the particularly high expression levels of ANR in KP and NDM peels compared to IW peel and the significant accumulation of its product epicatechin gallate (ECG) in those extracts reflects the rate-limiting role of ANR on ECG biosynthesis in mango. © 2015 Hoang et al.

Automating Crop Damage Assessments with Unmanned Aerial Systems and Machine Learning

Agricultural pests are responsible for millions of dollars in crop losses and management costs every year. In order to implement optimal site-specific treatments and reduce control costs, new methods to accurately monitor and assess pest damage need to be investigated. In this paper we explore the combination of unmanned aerial vehicles (UAV), remote sensing and machine learning techniques as a promising methodology to address this challenge. The deployment of UAVs as a sensor platform is a rapidly growing field of study for biosecurity and precision agriculture applications. In this experiment, a data collection campaign is performed over a sorghum crop severely damaged by white grubs (Coleoptera: Scarabaeidae). The larvae of these scarab beetles feed on the roots of plants, which in turn impairs root exploration of the soil profile. In the field, crop health status could be classified according to three levels: bare soil where plants were decimated, transition zones of reduced plant density and healthy canopy areas. In this study, we describe the UAV platform deployed to collect high-resolution RGB imagery as well as the image processing pipeline implemented to create an orthoimage. An unsupervised machine learning approach is formulated in order to create a meaningful partition of the image into each of the crop levels. The aim of this approach is to simplify the image analysis step by minimizing user input requirements and avoiding the manual data labelling necessary in supervised learning approaches. The implemented algorithm is based on the K-means clustering algorithm. In order to control high-frequency components present in the feature space, a neighbourhood-oriented parameter is introduced by applying Gaussian convolution kernels prior to K-means clustering. The results show the algorithm delivers consistent decision boundaries that classify the field into three clusters, one for each crop health level as shown in Figure 1. The methodology presented in this paper represents a venue for further esearch towards automated crop damage assessments and biosecurity surveillance.