Imidacloprid Extraction from Citrus Leaves and Analysis by Liquid Chromatography-Mass Spectrometry (HPLC-MS/MS).

A procedure was developed to extract Imidacloprid (IMD) from newly-flushed and fully-expanded citrus leaves. The extraction was conducted in a bullet blender, using a small sample mass (0.5 g of fresh tissue), stainless-steel beads (24 g), and methanol as extractant (10 mL). The extracts did not require further clean-up before analysis by HPLC-MS/MS. The method was validated with control samples from IMD-untreated Hamlin orange trees. The method limit of detection and limit of quantitation were 0.04 and 0.12 µg g(-1), respectively. IMD recoveries from fortified leaf tissue were between 92 % and 102 %, with relative standard deviations of <8 %. The method was further evaluated by extracting leaves from Hamlin orange trees treated with IMD. The treated trees showed maximum concentrations of 10.8 and 21.8 µg g(-1), observed at 20 days after applying two soil-drenching rates (0.51 and 1.02 kg IMD ha(-1)), respectively. This extraction technique will generate useful data on IMD plant uptake, foliar concentration, and correlations with Asian citrus psyllid (ACP) mortality or control. The method could be used to generate baseline data to improve IMD soil-drenching applications as the main management practice to control the ACP.

Two aquaporins, PIP1;1 and PIP2;1, mediate the uptake of neonicotinoid pesticides in plants.

Neonicotinoids (NEOs), a large class of organic compounds, are a type of commonly used pesticide for crop protection. Their uptake and accumulation in plants are prerequisites for their intra- and intercellular movements, transformation, and function. Understanding the molecular mechanisms that underpin NEO uptake by plants is crucial for effective application, which remains elusive. Here, we demonstrate that NEOs enter plant cells primarily through the transmembrane symplastic pathway and accumulate mainly in the cytosol. Two plasma membrane intrinsic proteins discovered in Brassica rapa, BraPIP1;1 and BraPIP2;1, were found to encode aquaporins (AQPs) that are highly permeable to NEOs in different plant species and facilitate NEO subcellular diffusion and accumulation. Their conserved transport function was further demonstrated in Xenopus laevis oocyte and yeast assays. BraPIP1;1 and BraPIP2;1 gene knockouts and interaction assays suggested that their proteins can form functional heterotetramers. Assessment of the potential of mean force indicated a negative correlation between NEO uptake and the energy barrier of BraPIP1;1 channels. This study shows that AQPs transport organic compounds with greater osmolarity than previously thought, providing new insight into the molecular mechanisms of organic compound uptake and facilitating innovations in systemic pesticides.

Accelerated removal of five pesticide residues in three vegetables with ozone microbubbles.

A simple and effective approach to remove pesticide residues on vegetables is necessary for food safety. Ozone microbubbles treatment as an eco-friendly washing technique was investigated for three vegetables (celery, pakchoi and cowpea) collected from the field applied with five pesticides. The removal rates of five pesticide residues on cowpea by ozone microbubbles treatment were 15 %-47 % higher than that by ozone macrobubbles. Moreover, compared with the other four systemic pesticides, emamectin benzoate had a preferable removal rate (65 %-94 %) as a non-systemic pesticide with high water solubility. Through Mass Spectrometry (MS), the double-bonded structure of emamectin benzoate made it more possible to be removed chemically, carrying a degradation rate of 88 % at 25 min in water. Additionally, cowpea showed low removal (28 %-65 %) owing to its stomata and rough surface with SEM. Conclusively, this study demonstrated the potential of the ozone microbubble treatment on pesticide residues removal to enhance food safety.

The use of phosphonates in agriculture. Chemical, biological properties and legislative issues.

The Phosphorus (III) derivatives, named Phosphonates, include congeners with properties as fungicides that are effective in controlling Oomycetes. Examples are organic compounds like Fosetyl-Al [Aluminium tris-(ethylphosphonate)] and salts formed with the anion of phosphonic acid [(OH)2HPO] and Potassium, Sodium and Ammonium cations. According to IUPAC, the correct nomenclature for these compounds is "phosphonates", but in common language and scientific literature they are often named "phosphites", creating ambiguity. The European legislation restricts the use of phosphonates, with the ban for application in organic agriculture. However, phosphonate residues were detected in some organic products due to their addition to fertilizers allowed in organic agriculture. The legitimacy of this addition is controversial, as it is not evident if phosphonates have also a nutritional role in addition to their fungicidal properties. The new European Directive EU 1009/2019 resolves the problem by banning the phosphonates addition to fertilizers and placing a limit of 0.5% by mass for unintentional addition. However, an official method is not available for phosphonates determination in fertilizers and approval by the European Committee for Standardization (CEN) is necessary in a short time. This review presents an overview about the chemical, biological, analytical and legislative aspects of phosphonates and aims at providing: clarity on the correct nomenclature to avoid misunderstandings; the evaluation of phosphonates properties with the absence of a nutritional role, justifying the ban on adding to fertilizers; a summary of analytical techniques that could be considered by CEN to complete the analytical background for the agricultural use of phosphonates.

Are Non-Target Honey Bees (Hymenoptera: Apidae) Exposed to Dinotefuran From Spotted Lanternfly (Hemiptera: Fulgoridae) Trap Trees?

The spotted lanternfly, Lycorma delicatula, is an introduced plant hopper that causes significant damage to host plants in the United States. Because of its affinity for tree of heaven, Ailanthus altissima, control efforts have focused on the use of the systemic insecticide, dinotefuran, in designated trap trees. There is concern about exposure to this pesticide by non-target species, especially honey bees, Apis mellifera, via lanternfly honeydew. Therefore, honey bee colonies were established in areas of high densities of trap trees and samples of honey, bees, and beeswax were collected in May, July, and October of 2017 for analysis. Samples were extracted by the QuEChERS method and analyzed using high-performance liquid chromatography with tandem mass spectrometry to determine the presence and quantity of dinotefuran. Additionally, honeydew from lanternflies was analyzed for dinotefuran and informal observations of trap tree visitors were made. None of the worker bee, wax, or honey samples indicated detectable levels of dinotefuran; however, honeydew samples collected did contain dinotefuran above the detection limit with amounts ranging from 3 to 100 ng per sample. The lack of dinotefuran in honey bee products matches the general absence of honey bees at trap trees in informal observations.

Imidacloprid Soil Drenches Affect Weight and Functional Response of Spined Soldier Bug (Hemiptera: Pentatomidae).

There are ongoing concerns of potential direct and indirect lethal and sublethal effects of insecticides on nontarget arthropod populations. The risk to natural enemies from systemic insecticides is mainly through exposure to the active ingredient by ingestion, and such risk may be elevated for omnivores that feed on treated plants, as well as herbivores that also feed on those same treated plants. Podisus maculiventris (Say), an important natural enemy in many agricultural systems, can be potentially exposed to the neonicotinoid imidacloprid when ingesting contaminated prey and feeding on plants subjected to soil-drench applications of this compound. In the current study, we examined the potential impact of imidacloprid soil drenches on some functional and morphological endpoints. Cabbage plants were treated with soil drenches of imidacloprid that corresponded to half and full recommended labels rates against whiteflies and aphids. Fourth instar diamondback moth, Plutella xylostella (Linnaeus) (Lepidoptera: Plutellidae), larvae on plants were used as prey in our experiments; P. xylostella is not a target of imidacloprid applications but may co-occur with other pests in systems where the insecticide is applied. We found that exposure to imidacloprid-treated plants did not cause significant mortality neither to P. maculiventris nor to P. xylostella, but both treatment concentrations impaired the predation, with consequences for predator weight gain during the assessment period. Our results corroborate those from other studies and demonstrate that effects from systemic insecticides can transcend trophic levels to affect natural enemies indirectly, such as through exposure from feeding on pests not targeted by the insecticide.

A new ecotoxicological test method for genetically modified plants and other stressors in soil with the black fungus gnat Bradysia impatiens (Diptera): current status of test development and dietary effects of azadirachtin on larval development and emergence rate.

BACKGROUND: Few suitable and standardized test methods are currently available to test the effects of genetically modified plants (GMP) on non-target organisms. To fill this gap and improve ecotoxicological testing for GMP, we developed a new soil ecotoxicological test method using sciarid larvae as test organisms. RESULTS: Bradysia impatiens was identified as a candidate species. Species of the genus Bradysia occur in high numbers in European agroecosystems and B. impatiens can be reared in the laboratory in continuous culture. A functional basic test design was successfully developed. Newly hatched larvae were used as the initial life stage to cover most of the life cycle of the species during the test. Azadirachtin was identified as a suitable reference substance. In several tests, the effects of this substance on development time and emergence rate varied for different temperatures and test substrates. The toxicity was higher at 25 °C compared to 20 °C and in tropical artificial soil compared to coconut fiber substrate. CONCLUSIONS AND OUTLOOK: Results suggest that the developed test system is suitable to enter a full standardization process, e.g., via the Organisation for Economic Co-operation and Development. Such a standardization would not only assist the risk assessment of GMP, but could include other stressors such as systemic pesticides or veterinary pharmaceuticals reaching the soil, e.g., via spreading manure. The use of sciarid flies as test organisms supports recommendations of EFSA, which stressed the ecological role of flies and encouraged including Diptera into test batteries.

An Evaluation of the Honey Bee (Hymenoptera: Apidae) Safety Profile of a New Systemic Insecticide, Flupyradifurone, Under Field Conditions in Florida.

Flupyradifurone (Sivanto) is a novel systemic insecticide from the butenolide class developed by Bayer. Based on available data (USEPA 2014), this insecticide appears to have a favorable safety profile for honey bee colonies. As a result, the label permits the product to be applied during prebloom and bloom in various crops, including citrus, except when mixed with azole fungicides during the blooming period. We placed 24 honey bee (Apis mellifera L.) colonies adjacent to eight flowering buckwheat (Fagopyrum esculentum Moench) fields that either had been sprayed with the maximum label rate of flupyradifurone or with water only (control fields), with three colonies placed adjacent to each field. We conducted colony strength assessments during which the number of adult bees, eggs, uncapped brood cells, capped brood cells, food storage cells, and weights of honey supers and brood chambers were determined prior to, during, and after the flowering period. We also analyzed bee-collected pollen and nectar for flupyradifurone residues. Overall, there were no differences in any colony strength parameter for colonies placed at control and flupyradifurone-treated buckwheat fields. Residue analyses showed that pollen (x = 565.8 ppb) and nectar (x = 259.4 ppb) gathered by bees on fields treated with flupyradifurone contained significantly higher flupyradifurone residues than did bee bread and unprocessed nectar collected by bees from control fields (75% of samples