X-Ray-Based Irradiation of Larvae and Pupae of the Navel Orangeworm (Lepidoptera: Pyralidae).

The suitability of adult male the navel orangeworm, Amyelois transitella (Walker) for Sterile Insect Technique (SIT) has been reported for both high energy gamma (>1 MeV) and low energy x-ray (90 keV) sterilization. However, research regarding sterilization of NOW larvae and pupae by gamma irradiation indicated nonsuitability due to high mortality. Here, NOW larvae and pupae were irradiated to doses up to 50 Gy with 90 keV x-rays, then paired with nonirradiated colony mates. Sterility of surviving insects was determined by the presence or absence of hatched neonates. While presence of offspring does not guarantee viability, the absence does guarantee sterility (as is appropriate for SIT) and was thus the measure used here. Early stage larvae experienced 77% mortality at a dose of 30 Gy, versus 20% for nonirradiated control. At 40 Gy, mortality reached 98%. Of surviving early stage larvae at 30 Gy, 29% of moth pairs produced offspring. For late stage larvae, no offspring were produced at 40 Gy, but mortality was 73%. For pupae, mortality reached 53% at 30 Gy with 13% still producing neonates, while mortality reached 98% at 40 Gy. These results are consistent with reported results for gamma irradiation of NOW larvae where sterility was observed somewhere between the 30 Gy and 60 Gy data points, but mortality was high. This further confirms the lack of suitability of NOW irradiated in the larval stage, whether by gamma or x-ray, and supports the hypothesis that x-ray and gamma treatments are biologically equivalent at equal doses.

Silo-Stored Pistachios at Varying Humidity Levels Produce Distinct Volatile Biomarkers.

Fungal-contaminated tissues are known to produce volatile profiles that are different from uncontaminated tissues. Fungi require certain water activity levels before growth can occur. For nonxerophilic fungi, a water activity of 0.85 is typical for growth, and for extreme xerophilic fungi, the water activity can be as low as 0.64. Recent investigations with stored pistachios (kernels in shell, no hull tissue) at varying relative humidities showed differences among the collected volatile profiles at the tested humidities (ambient, 63, 75, and 84%). Water activities of the kernel and shell were also measured. Results showed significant changes in volatile profiles as a function of water activity of the corresponding pistachio tissue with measured water activity levels at or below that of what is considered extreme xerophilic activities. Because fungal growth, including mycotoxigenic fungi, is dependent upon water activity, the detected volatile profiles could be used for early detection of fungal presence. Multivariate analysis of the volatile data demonstrated significant differences among the volatile profiles at the tested relative humidity levels, and several volatiles were identified as biomarkers of increased humidity and likely fungal development.

Differentiation of Volatile Profiles from Stockpiled Almonds at Varying Relative Humidity Levels Using Benchtop and Portable GC-MS.

Contamination by aflatoxin, a toxic metabolite produced by Aspergillus fungi ubiquitous in California almond and pistachio orchards, results in millions of dollars of lost product annually. Current detection of aflatoxin relies on destructive, expensive, and time-intensive laboratory-based methods. To explore an alternative method for the detection of general fungal growth, volatile emission profiles of almonds at varying humidities were sampled using both static SPME and dynamic needle-trap SPE followed by benchtop and portable GC-MS analysis. Despite the portable SPE/GC-MS system detecting fewer volatiles than the benchtop system, both systems resolved humidity treatments and identified potential fungal biomarkers at extremely low water activity levels. This ability to resolve humidity levels suggests that volatile profiles from germinating fungal spores could be used to create an early warning, nondestructive, portable detection system of fungal growth.

Rehabilitation needs of the survivors of the 2013 Ya'an earthquake in China.

OBJECTIVE: To determine the physical, functional and psychosocial rehabilitation needs of the injured survivors of the 2013 earthquake in Ya'an, China. DESIGN: Cross-sectional survey. METHODS: A total of 143 injured survivors (80 males, 63 females, mean age 41.4 years (standard deviation (SD) 20.2)) were recruited for the study. A questionnaire was designed to collect their demographic and environmental information. Manual muscle testing and active range of motion (AROM) were used to assess physical function. The Modified Barthel Index was used to assess activities of daily living (ADL) and the Post-Traumatic Stress Disorder - Self Rating Scale was used to assess psychological status. The rehabilitation needs of the injured survivors were identified based on the assessment results. RESULTS: Physical problems of the back and lower limbs were more severe than those of the upper limbs, as measured by MMT and AROM. Various degrees of dependence in ADL were measured in 74.2% of subjects, and psychological distress was present in more than 50% of subjects. Rehabilitation training prescribed by local therapists included functional training (74.8%), exercises (49.0%), provision of splints (44.8%), etc. Psychological interventions were re-commended to most of the patients. There were high levels of need for assistive devices, and home and community modifications. CONCLUSION: There is a high level of need for early rehabilitation in hospital, followed by continued home and community rehabilitation services, particularly for injured survivors who are returning to villages with very limited resources.

In-field Volatile Analysis Employing a Hand-held Portable GC-MS: Emission Profiles Differentiate Damaged and Undamaged Yellow Starthistle Flower Heads.

INTRODUCTION: Understanding the complex chemical signalling of plants and insects is an important component of chemical ecology. Accordingly, the collection and analysis of chemical cues from plants in their natural environment is integral to elucidation of plant-insect communications. Remote plant locations and the need for a large number of replicates make in situ headspace analyses a daunting logistical challenge. A hand-held, portable GC-MS system was used to discriminate between damaged and undamaged Centaurea solstitialis (yellow starthistle) flower heads in both a potted-plant and natural setting. OBJECTIVE: To determine if a portable GC-MS system was capable of distinguishing between undamaged and mechanically damaged plant treatments, and plant environments. METHODOLOGY: A portable GC-MS utilising needle trap adsorbent technology was used to collect and analyse in situ headspace volatiles of varying yellow starthistle treatments. Principal component analysis (PCA) was used to distinguish treatments and identify biomarker volatiles. Analysis of variance (ANOVA) was used to determine differences between treatment volatile amounts. RESULTS: The portable GC-MS system detected 31 volatiles from the four treatments. Each GC-MS run was completed in less than 3 min. PCA showed four distinct clusters representing the four treatments - damaged and undamaged potted plant, and damaged and undamaged natural plant. Damage-specific volatiles were identified. CONCLUSION: The portable GC-MS system distinguished the treatments based on their detected volatile profiles. Additional statistical analysis identified five possible biomarker volatiles for the treatments, among them cyclosativene and copaene, which indicated damaged flower heads.

The major volatile compound 2-phenylethanol from the biocontrol yeast, Pichia anomala, inhibits growth and expression of aflatoxin biosynthetic genes of Aspergillus flavus.

Aspergillus flavus is a ubiquitous saprophyte that is able to produce the most potent natural carcinogenic compound known as aflatoxin B1 (AFB1). This toxin frequently contaminates crops including corn, cotton, peanuts, and tree nuts causing substantial economic loss worldwide. Consequently, more than 100 countries have strict regulations limiting AFB1 in foodstuffs and feedstuffs. Plants and microbes are able to produce volatile compounds that act as a defense mechanism against other organisms. Pichia anomala strain WRL-076 is a biocontrol yeast currently being tested to reduce AF contamination of tree nuts in California. We used the SPME-GC/MS analysis and identified the major volatile compound produced by this strain to be 2-phenylethanol (2-PE). It inhibited spore germination and AF production of A. flavus. Inhibition of AF formation by 2-PE was correlated with significant down regulation of clustering AF biosynthesis genes as evidenced by several to greater than 10,000-fold decrease in gene expression. In a time-course analysis we found that 2-PE also altered the expression patterns of chromatin modifying genes, MYST1, MYST2, MYST3, gcn5, hdaA and rpdA. The biocontrol capacity of P. anomala can be attributed to the production of 2-PE, which affects spore germination, growth, toxin production, and gene expression in A. flavus.

Headspace volatiles from 52 oak species advertise induction, species identity, and evolution, but not defense.

Leaf volatiles convey information about a plant to other organisms in their proximity. Despite increasing interest in understanding the relevance of volatile emissions for particular ecological interactions, there has been relatively little effort to assess generally what information volatile profiles transmit. We surveyed the volatile profiles of wounded and unwounded leaves of 52 oak (Quercus) species. We used phylogenetic comparison and multivariate techniques to assess in what circumstances oak individuals advertised their species identity, evolutionary history, direct defenses, or damage. We found that both species identity and evolutionary history were advertised when leaves were wounded, but species could not be differentiated by odor when leaves were not wounded. Various fatty-acid derivative compounds showed the strongest phylogenetic signal suggesting that they may best disclose taxonomic affiliations in oaks. We tested whether oak volatile composition or diversity advertised high defensive investment, but we found no evidence for this. Wounded leaves disclose much about an oak species' identity and taxonomic affiliation, but unwounded leaves do not. This is consistent with the idea that volatile information is targeted toward natural enemy recruitment.

Generation of the volatile spiroketals conophthorin and chalcogran by fungal spores on polyunsaturated fatty acids common to almonds and pistachios.

The spiroketal (E)-conophthorin has recently been reported as a semiochemical of the navel orangeworm moth, a major insect pest of California pistachios and almonds. Conophthorin and the isomeric spiroketal chalcogran are most commonly known as semiochemicals of several scolytid beetles. Conophthorin is both an insect- and plant-produced semiochemical widely recognized as a nonhost plant volatile from the bark of several angiosperm species. Chalcogran is the principal aggregation pheromone component of the six-spined spruce bark beetle. Recent research has shown conophthorin is produced by almonds undergoing hull-split, and both spiroketals are produced by mechanically damaged almonds. To better understand the origin of these spiroketals, the volatile emissions of orchard fungal spores on fatty acids common to both pistachios and almonds were evaluated. The volatile emission for the first 13 days of spores placed on a fatty acid was monitored. The spores investigated were Aspergillus flavus (atoxigenic), A. flavus (toxigenic), Aspergillus niger, Aspergillus parasiticus, Penicillium glabrum, and Rhizopus stolonifer. The fatty acids used as growth media were palmitic, oleic, linoleic, and linolenic. Spores on linoleic acid produced both spiroketals, those on linolenic acid produced only chalcogran, and those on palmitic and oleic acid did not produce either spiroketal. This is the first report of the spiroketals conophthorin and chalcogran from a fungal source.

Hull split and damaged almond volatiles attract male and female navel orangeworm moths.

A blend of volatiles derived from the emissions of almonds at hull split and mechanically damaged almonds was compared to almond meal, the current monitoring standard for the insect pest navel orangeworm (NOW). Field trapping studies were performed to determine the blend's ability to attract adult NOW. The blend comprised racemic 1-octen-3-ol, ethyl benzoate, methyl salicylate, acetophenone, and racemic (E)-conophthorin. Ethyl acetate was used as a solvent with a blend component concentration of 100 mg/mL. The blend attracted both sexes of NOW when tested in five 2-week intervals spanning the first three flights of NOW in commercial almond orchards in the southern Central Valley of California. The blend demonstrated consistently higher capture rates for female NOW throughout the evaluation period, but unlike almond meal it significantly attracted males. Reported is a survey of the major and minor volatiles emitted from almonds at hull split, the key period of vulnerability to NOW infestation. Also reported is the attractancy of a formulated test blend based on the host plant volatile emissions, electroantennographic screening experiments, and field trapping studies. The results of this test blend highlight progress toward a host-plant-based attractant for NOW, a major insect pest of California tree nuts that presently lacks an adequate monitoring tool.

Electroantennographic bioassay as a screening tool for host plant volatiles.

Plant volatiles play an important role in plant-insect interactions. Herbivorous insects use plant volatiles, known as kairomones, to locate their host plant. When a host plant is an important agronomic commodity feeding damage by insect pests can inflict serious economic losses to growers. Accordingly, kairomones can be used as attractants to lure or confuse these insects and, thus, offer an environmentally friendly alternative to pesticides for insect control. Unfortunately, plants can emit a vast number volatiles with varying compositions and ratios of emissions dependent upon the phenology of the commodity or the time of day. This makes identification of biologically active components or blends of volatile components an arduous process. To help identify the bioactive components of host plant volatile emissions we employ the laboratory-based screening bioassay electroantennography (EAG). EAG is an effective tool to evaluate and record electrophysiologically the olfactory responses of an insect via their antennal receptors. The EAG screening process can help reduce the number of volatiles tested to identify promising bioactive components. However, EAG bioassays only provide information about activation of receptors. It does not provide information about the type of insect behavior the compound elicits; which could be as an attractant, repellent or other type of behavioral response. Volatiles eliciting a significant response by EAG, relative to an appropriate positive control, are typically taken on to further testing of behavioral responses of the insect pest. The experimental design presented will detail the methodology employed to screen almond-based host plant volatiles by measurement of the electrophysiological antennal responses of an adult insect pest navel orangeworm (Amyelois transitella) to single components and simple blends of components via EAG bioassay. The method utilizes two excised antennae placed across a "fork" electrode holder. The protocol demonstrated here presents a rapid, high-throughput standardized method for screening volatiles. Each volatile is at a set, constant amount as to standardize the stimulus level and thus allow antennal responses to be indicative of the relative chemoreceptivity. The negative control helps eliminate the electrophysiological response to both residual solvent and mechanical force of the puff. The positive control (in this instance acetophenone) is a single compound that has elicited a consistent response from male and female navel orangeworm (NOW) moth. An additional semiochemical standard that provides consistent response and is used for bioassay studies with the male NOW moth is (Z,Z)-11,13-hexdecadienal, an aldehyde component from the female-produced sex pheromone.

Volatile analysis of ground almonds contaminated with naturally occurring fungi.

Aflatoxigenic aspergilli inflict major economic damage to the tree nut industry of California, with the highest negative impact to almonds. Aspergilli and fungi in general are known to emit volatiles in varying quantity and composition dependent upon their growth media. The goal of the study was to determine the volatile emission of whole and blanched almonds that had been picked out and labeled as inedible by processors. The aflatoxin content and number of colony forming units of each sample were also determined. A total of 23 compounds were consistently detected and identified. Several volatiles from the blanched almonds demonstrated significant increases when compared to the emissions of whole almonds. Several of these volatiles are considered fatty acid decomposition products and included hexanal, heptanal, octanal, nonanal, 3-octen-2-one, tetramethylpyrazine, and decanal. The almond samples investigated were characteristic of a typical postharvest environment and illustrative of potential contamination within a stockpile or transport container. Volatiles indicative of fatty acid decomposition were predominant in the samples that underwent some form of blanching. The emission amounts of hexanal, heptanal, octanal, and hexanoic acid increased 3-fold in samples contaminated with aflatoxin; however, due to variability between samples they could not be considered as indicator volatiles for aflatoxin content. The emission profile of volatiles from almond kernels contaminated with naturally occurring aspergilli and associated fungi is heretofore unreported.

In situ seasonal study of the volatile production of almonds (Prunus dulcis) var. 'Nonpareil' and relationship to navel orangeworm.

Nonpareil almonds, Prunus dulcis , account for the largest percentage of almond varieties grown in the Central Valley of California. Several studies have investigated the various nonvolatile and volatile components of various plant parts; however, the volatile organic compound (VOC) emission of almonds from a single cultivar has not been studied over the course of a growing season. This aspect is particularly relevant to research concerning the navel orangeworm (NOW), a major insect pest of almonds and other tree nuts. Despite the continued presence of NOW, the identification of particular VOCs and their relationship to NOW have not been addressed. The VOC emission of Nonpareil almonds was collected in situ over the course of a growing season by solid-phase microextraction (SPME). The VOCs (Z)-hex-3-enyl acetate, (Z)-hex-3-enyl butyrate, undecan-2-ol, beta-bourbonene, and tetradecane were present for the majority of the days investigated. Several VOCs exhibited positive electroantennographic signals from male and/or female NOW moths.