Global issues in setting legal limits on soil metal contamination: A case study of Chile.

The establishment of legal limits for soil contamination with trace elements is a global issue that has not yet been resolved. However, the resolution of any global problem begins at the national level. In this vein, we present the case of Chile, the world's leading copper producer, where soil contamination by trace elements in mining areas has been severe. We evaluated the magnitude of the ecological and human health risks from exposure to arsenic (As), copper (Cu), zinc (Zn), and lead (Pb) in soils of the La Ligua and Petorca basins, two important mining areas in Chile. Contrary to what might be expected in soils affected by Cu mining activities, As was identified as the most hazardous element in the studied soils, both in terms of ecological and human health risks. On the other hand, Chile does not currently have specific legislation establishing legal limits on soil contamination with trace elements. Since Chile is geochemically similar to New Zealand, Mexico, and Italy, we used the limits of these three countries as benchmarks. We determined the background concentrations of As, Cu, Zn, and Pb in the soils of the two river basins under study and found that they tend to exceed the limits established by foreign laws. We also found that the differences in background elemental concentrations in the studied soils were primarily due to the varied lithology of soil-forming rocks. This means that absolute "one-limit-fits-all" values of element concentrations may not be adequate to regulate the level of soil contamination in areas affected by mining. As a fundamental first step, it is necessary to establish background soil concentrations of trace elements in each river basin in Chile. It is clear that Chile urgently needs to move from rubber-stamping foreign laws to the development of national legislation on soil metal contamination.

Monitoring Chilecomadia valdiviana (Lepidoptera: Cossidae) Using Sex Pheromone-Baited Traps in Apple Orchards in Chile.

Chilecomadia valdiviana (Philippi) (Lepidoptera: Cossidae) is a native xylophagous pest in apple orchards in Chile. A series of experiments evaluated the efficacy of trap type, sex pheromone (Z7,Z10-16:Ald) dose, and trap location in the apple tree canopy on trap catch of male adults. Bucket traps (6 L), with and without roof and cross vane spacers, together with bucket traps (20 L) without roof and spacers, showed higher catches among the four types of traps evaluated. In a second experiment, the UNI-trap and Delta trap showed higher catches than Multipher, wing, and bucket traps (6 L). Male catches were not affected by height when tested at 0, 1.5, and 3 m in the canopy. A 300 µg dose of Z7,Z10-16:Ald showed higher catch than the control treatment. This dose allowed monitoring of male flight of C. valdiviana for at least five weeks in apple orchards in Chile. Based on relative trap costs, we propose the use of 6 L bucket traps for male mass trapping, while Delta traps can be used for monitoring of male flight. We found that male flight of C. valdiviana occurred mainly from mid-August to late November, reaching its maximum in mid-September.

Diffusible Compounds Produced by Hanseniaspora osmophila and Gluconobacter cerinus Help to Control the Causal Agents of Gray Rot and Summer Bunch Rot of Table Grapes.

Gray and summer bunch rot are important diseases of table grapes due to the high economic and environmental cost of their control with synthetic fungicides. The ability to produce antifungal compounds against the causal agents Botrytis, Aspergillus, Penicillium, and Rhizopus of two microorganisms isolated from table grapes and identified as Hanseniaspora osmophila and Gluconobacter cerinus was evaluated. In dual cultures, both biocontrol agents (together and separately) inhibited in vitro mycelial growth of these pathogens. To identify the compounds responsible for the inhibitory effect, extractions were carried out with organic solvents from biocontrol agents separately. Through dual cultures with pathogens and pure extracts, only the hexane extract from H. osmophila showed an inhibitory effect against Botrytis cinerea. To further identify these compounds, the direct bioautography technique was used. This technique made it possible to determine the band displaying antifungal activity at Rf = 0.05-0.2. The compounds present in this band were identified by GC-MS and compared to the NIST library. The most abundant compounds, not previously reported, corresponded to alkanes, ketones, alcohols, and terpenoids. H. osmophila and G. cerinus have the potential to control the causal agents of gray and summer bunch rot of table grapes.

3,7-Dimethylpentadecane: a Novel Sex Pheromone Component from Leucoptera sinuella (Lepidoptera: Lyonetiidae).

Leucoptera sinuella is a leaf-miner moth present in several regions in the world, which has been recently introduced into Chile. The larvae feed exclusively on the leaves of poplar and willow trees, and the damage caused by the feeding behavior poses a threat to the wood-producing industry. Besides, L. sinuella larvae invade nearby orchards for pupation, causing rejections in Chilean fresh fruit for export. Here we report the identification of the female-produced sex pheromone of L. sinuella as a first step towards the development of pheromone-based methods for pest management of this species. First, we analyzed hexane extracts of the abdominal glands of virgin females by gas chromatography coupled with mass spectrometry and identified the major compound in these extracts to be 3,7-dimethylpentadecane, while minor compounds in the extracts proved to be 3,7-dimethyltetradecane and 7-methylpentadecane. Structure assignments were carried out by comparison of retention times and mass spectra of the natural products with those of authentic reference samples. Second, we conducted field tests, which showed that traps baited with synthetic 3,7-dimethylpentadecane were significantly attractive to males in a dose-dependent response. Our results also showed that a mixture of 3,7-dimethylpentadecane, 3,7-dimethyltetradecane, and 7-methylpentadecane in proportions similar to those found in gland extracts was the most attractive lure.

Linoleic acid and stearic acid are biosynthetic precursors of (7Z,10Z)-7,10-hexadecadienal, the major component of the sex pheromone of Chilecomadia valdiviana (Lepidoptera: Cossidae).

The main pheromone compound of Chilecomadia valdiviana (Lepidoptera: Cossidae) has been recently identified as (7Z,10Z)-7,10-hexadecadienal. The biosynthesis of this pheromone compound showing attributes of both Type I and Type II lepidopteran pheromones was studied by the topical application of isotope-labeled fatty acids to the pheromone gland and subsequent analysis of the gland contents (pheromone compounds and fatty acyl compounds) by gas chromatography-mass spectrometry. The deuterium label of D11-linoleic acid was incorporated into the pheromone compound and its putative acyl precursor (7Z,10Z)-7,10-hexadecadienoate, demonstrating that the pheromone compound is biosynthesized from linoleic acid by chain-shortening and further functional group transformation. Furthermore, the deuterium label of D3-stearic acid was also incorporated into the pheromone compound, which indicates that the pheromone can be synthesized de novo by C. valdiviana, as is the case for Type I lepidopteran pheromone compounds.

Volatiles of Grape Inoculated with Microorganisms: Modulation of Grapevine Moth Oviposition and Field Attraction.

Semiochemicals released by plant-microbe associations are used by herbivorous insects to access and evaluate food resources and oviposition sites. Adult insects may utilize microbial-derived nutrients to prolong their lifespan, promote egg development, and offer a high nutritional substrate to their offspring. Here, we examined the behavioral role of semiochemicals from grape-microbe interactions on oviposition and field attraction of the grapevine moth Lobesia botrana (Denis & Schiffermüller). The volatile constituents released by grape inoculated with yeasts (Hanseniaspora uvarum (Niehaus), Metschnikowia pulcherrima (Pitt.) M.W. Miller, Pichia anomala, Saccharomyces cerevisiae Meyen ex E.C. Hansen, and Zygosaccharomyces rouxii (Boutroux) Yarrow), sour rot bacteria (Acetobacter aceti (Pasteur) Beijerinck and Gluconobacter oxydans (Henneberg) De Ley), and a fungal pathogen (Botrytis cinerea Pers.) all endemic of the vineyard were sampled by solid-phase microextraction and analyzed by gas-chromatography coupled with mass spectrometry. Ethanol, 3-methyl-1-butanol, and ethyl acetate were the most common volatiles released from all microbe-inoculated grapes. In addition, acetic acid was released at a substantial amount following bacteria inoculation and in a three-way inoculation with yeasts and the fungus. 2-phenylethanol, a compound reported to attract tortricid moths when used in combination with acetic acid, was found at a relatively low level in all microbial combinations as well as in the control grape. While grapes inoculated with a consortium of yeasts stimulated oviposition in comparison with uninoculated berries, the phytopathogenic fungus deterred egg-laying. Nonetheless, the highest preference to lay eggs was measured when the yeasts were co-inoculated with the fungus. The lowest preference was obtained when grapes were inoculated with sour rot bacteria and their binary co-inoculation with yeasts and the fungus. Interestingly, oviposition on berries simultaneously inoculated with all the three microbial groups was unaffected. Lures loaded with either acetic acid or 2-phenylethanol were not attractive when placed in traps as single component in vineyards, but a binary blend attracted both sexes of grapevine moth in significant numbers. Further addition of the three most common volatiles released by infected berries (ethanol, 3-methyl-1-butanol, and ethyl acetate) did not significantly increase moth catch with this binary blend. The ecological implications of the grape-microorganism and grapevine moth interaction as well as the possibility to develop a pest monitoring system based on microbial volatiles are discussed.

Use of Mixture Designs to Investigate Contribution of Minor Sex Pheromone Components to Trap Catch of the Carpenterworm Moth, Chilecomadia valdiviana.

Field experiments were carried out to study responses of male moths of the carpenterworm, Chilecomadia valdiviana (Lepidoptera: Cossidae), a pest of tree and fruit crops in Chile, to five compounds previously identified from the pheromone glands of females. Previously, attraction of males to the major component, (7Z,10Z)-7,10-hexadecadienal, was clearly demonstrated while the role of the minor components was uncertain due to the use of an experimental design that left large portions of the design space unexplored. We used mixture designs to study the potential contributions to trap catch of the four minor pheromone components produced by C. valdiviana. After systematically exploring the design space described by the five pheromone components, we concluded that the major pheromone component alone is responsible for attraction of male moths in this species. The need for appropriate experimental designs to address the problem of assessing responses to mixtures of semiochemicals in chemical ecology is described. We present an analysis of mixture designs and response surface modeling and an explanation of why this approach is superior to commonly used, but statistically inappropriate, designs.

Identification of a Novel Moth Sex Pheromone Component from Chilecomadia valdiviana.

Chilecomadia valdiviana (Philippi) (Lepidoptera: Cossidae) is an insect native to Chile. The larval stages feed on the wood of economically important fruit tree species such as apple, pear, olive, cherry, and avocado, and also on eucalyptus. This causes weakening and, in case of severe infestation, death of the tree. We report identification of the sex pheromone produced by females of this species. Hexane extracts of the abdominal glands of virgin females were analyzed by gas chromatography (GC) with electroantennographic detection, GC coupled with mass spectrometry, and GC coupled to infrared spectroscopy. The major pheromone component was identified as (7Z,10Z)-7,10-hexadecadienal (Z7,Z10-16:Ald), and minor components present in the extracts were (Z)-7-hexadecenal and (Z)-9-hexadecenal, hexadecanal, and (9Z,12Z)-9,12-octadecadienal. Structural assignments were carried out by comparison of analytical data of the natural products and their dimethyl disulfide adducts with those of authentic reference samples. In field tests, traps baited with Z7,Z10-16:Ald captured significantly more males than control traps.

Toxicity of Six Insecticides on Codling Moth (Lepidoptera: Tortricidae) and Effect on Expression of Detoxification Genes.

The codling moth, Cydia pomonella (L.), is a key worldwide fruit pest that has evolved high levels of resistance to almost all classes of conventional insecticides. Neonicotinoids, a new reduced-risk biorational insecticide class, have remained an effective control approach. In this study, the toxicity and sublethal effect of conventional and reduced-risk biorational insecticides on transcripts abundance of three detoxification genes in codling moth were determined. Bioassays on a codling moth laboratory strain suggested that acetamiprid had the highest oral toxicity against the third-instar larvae compared with the other five pesticides. Results also indicated that acetamiprid exhibits long-term efficacy against codling moth even at 120 h post feeding. Real-time quantitative polymerase chain reaction showed that the detoxification genes CYP9A61, CpGST1, and CpCE-1 were differentially induced or suppressed by deltamethrin, cypermethrin, methomyl, carbaryl, and imidacloprid, depending on the type of insecticides; in contrast, no significant difference in CYP9A61, CpGST1, and CpCE-1 expressions were observed after acetamiprid exposure, when compared with the control. These results suggest that the reduced-risk biorational insecticide acetamiprid is an effective insecticide with no induction of detoxification genes and can be integrated into the management of codling moth.

Organophosphate Resistance and its Main Mechanism in Populations of Codling Moth (Lepidoptera: Tortricidae) from Central Chile.

The codling moth, Cydia pomonella (L.), is the key pest of apple production worldwide. Insecticide resistance has been reported in all producing countries, based on five different mechanisms. Codling moth in Chile has resistance to azinphos-methyl and tebufenozide in post-diapausing larvae. However, there are no studies about the susceptibility of these populations to insecticides from other chemical groups. Therefore, the efficacy of azinphos-methyl, chlorpyrifos-ethyl, esfenvalerate, methoxyfenozide, tebufenozide, and thiacloprid on neonate and post-diapausing larvae from six field populations was investigated, and identified resistance mechanisms in this species were evaluated. Neonate larvae were susceptible to all insecticides studied, but post-diapausing larvae from four populations were resistant to chlorpyrifos, one of them was also resistant to azinphos-methyl, and another one was resistant to tebufenozide. The acetylcholinesterase insensitivity mutation was not detected, and the sodium channel knockdown resistance mutation was present in a low frequency in one population. Detoxifying enzymatic activity of glutathione S-transferases, esterases, and cytochrome P-450 monooxygenases in adults differed among populations, but chlorpyrifos resistance was associated only with a decreased esterase activity as shown by a significant negative correlation between chlorpyrifos mortality and esterase activity.