Wai S. Gee

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.

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 blends 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.

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.

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.

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.

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.

Volatile emissions from the flea beetle Altica litigata (Coleoptera: Chrysomelidae) associated with invasive Ludwigia hexapetala.

The flea beetle Altica litigata (Chrysomelidae) is an insect herbivore to plants within the families Lythraceae and Onagraceae, including ornamentals such as crape myrtle, Lagerstroemia spp. This insect is important both as a pest species and as a naturally occurring biological control agent due to its aggregate feeding behavior, which typically results in severe defoliation of the host plant. Despite the negative economic impact to ornamentals and contrary benefits as a biological control agent, there are few reports on the semiochemical communication of this family of insects. Uruguayan primrose-willow (Ludwigia hexapetala) is an invasive aquatic weed in California and serves as a host to A. litigata. To better characterize this association, the volatile emissions of A. litigata were collected while the flea beetles were: in containers by themselves, in containers with L. hexapetala leaves, in situ on L. hexapetala leaves in a growth chamber, and in situ on L. hexapetala leaves in the field. For comparison, the volatile emissions of A. litigata associated with two subspecies of creeping water primrose (L. peploides) were also evaluated. Two himachalene-type sesquiterpenes, showing the same carbon skeleton as compounds previously reported from Aphthona flava and Epitrix fuscula, were detected as volatiles from A. litigata.

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.