Analysis of the synergic and sublethal effect of the pesticides imidacloprid, acetamiprid and tebuconazole on brain metabolism in the non-bee pollinator Eristalis tenax

Abstract

Abstract
Introduction: A great decline of wild pollinators has been observed, the decline is believed to be a result of many stressors, whereas pesticide usage being one of them. Wild pollinator insects are significantly important to maintain and preserve food production and biodiversity.

Background: Pesticides are used to protect and enhance crop yield, by targeting insect and fungi pests. Imidacloprid (IMI) and acetamiprid (ACE) are both neonicotinoids with same mode of action. Tebuconazole (TEB) is a fungicide, that might exhibit synergistic effect when combined with a neonicotinoid. Studies done one effects of the combinations are limited, and strictly only involving bee pollinators. Furthermore, the metabolomic effect of pesticides as well as the sample handling and sample treatment procedure is not thoroughly investigated.
Aim(s): This study aims to investigate how different pre-treatments as immobilization techniques affect the metabolomics in flies. It also aims to investigate how the metabolism of the hover fly is altered by the pesticides IMI, ACE and TEB, alone, in binary and in ternary mixtures.

Methods: Two different immobilization techniques were evaluated; carbon dioxide or ice and compared to when no anaesthesia was used. After each treatment the fly was either dissected in room temperature or in cold. The same experiment was performed with acetamiprid orally exposed and control flies. Lastly, a design of experiment approach using face centred composite design was implemented for the single, binary, and ternary exposed mixtures of the three pesticides. A simple extraction procedure was used where the brains were homogenized with a ball mill, followed by freezing, centrifuged, dried and derivatization. The sample was analysed using gas chromatography-mass spectrometry in SIM mode.

Results: The different pre-treatment showed different accumulation and restricted metabolite. When no anaesthesia was used, an accumulation of GABA and cysteine was observed indicating oxidative stress. CO2 and cold anaesthesia exhibited a shift in energy demand. When exposed to acetamiprid, the three immobilization techniques resulted in the same reaction suggesting that the anaesthesia may not affect when exposing to a pesticide. The effect of three pesticides IMI, ACE and TEB, displayed the highest effect of ACE. IMI exhibited low effects suggesting a low dose used. TEB showed an up-regulation of cell membrane metabolite, suggesting possibly the fungicide affecting the cell membrane not only in fungi but also in insects. Interaction terms exhibited a synergistic effect. Quadratic effects was significant for almost all pesticides suggesting a non-linear relationship for the regulation of the metabolites.

Conclusion: The three pre-treatment exhibited different result. Acetamiprid exposure resulted in the same outcome for all methods. Model selections were performed and demonstrated. ACE displayed the highest effect, IMI showed low effect, TEB showed an effect but on fewer metabolites. Interaction terms showed a synergistic effect and quadratic effects was displayed as significant for many metabolites.

Popular Abstract

Popular Scientific Summary
Pollinators like bees, flies, moths, and birds are responsible for 35% of the world food production. Unfortunately, in recent years a decline of pollinators has been observed. The decline is believed to be influenced by pesticides. Pesticides are chemicals used on crops and plants to protect from diseases, fungi, weeds, and harmful insects. A lot of research on pesticide effects are focused on bee-pollinators, despite the wild insect pollinators being at greater risk. Wild insect pollinators are not well managed and controlled like bees, which makes them more at danger. On top of that, almost no research has been done on the molecular level and the studies done lack the evaluation of sample handling and pre-treatment.
To analyze pesticide effect on non-bee pollinators, this study aims to analyze molecules in the brain defined as metabolites, in the non-bee pollinator the hover fly Eristalis tenax. The purpose of this study is also to examine how different immobilization techniques affect the metabolite in the brain, and if this effect is changed when the flies are exposed to pesticide. Immobilization techniques are used to handle insects because some insects possess toxins or can sting. But also, to make it easier to handle the insects, since insects could fly or move. In addition, three different pesticides; imidacloprid, acetamiprid and tebuconazole were analyzed based on their single and combined metabolomic effect. The metabolites were analyzed with gas chromatography where they get separated by a column and analyzed with a mass spectrometer.
When no immobilization technique was used, metabolites that expresses stress was affected, meaning that the flies were experiencing fear. The ice and carbon dioxide treatment showed an increased demand for energy, to compromise for the heat loss due to freezing and to compensate for oxygen reduction. When exposed to acetamiprid, the three immobilization techniques showed similar responses, which could mean that the pesticide treatment resulted in such a huge reaction that the immobilization technique did not affect. It was also shown that the flies was actively trying to reduce the toxicity of acetamiprid by producing more metabolite that are involved in the metabolism of toxins. Metabolites involved in energy production was also affected, which could be a result of more energy is needed to recover from the exposure. And lastly, acetamiprid displayed the highest effect, imidacloprid showed low effect, tebuconazole showed an effect but on fewer metabolites. Interaction terms showed a synergistic effect and quadratic effects was displayed as significant for many metabolites.

• author: Fathi, Mona
• supervisor: Simon Palmer
• organization: Department of Chemistry
• course: KEMR30 20231
• year: 2023
• type: H2 - Master's Degree (Two Years)
• subject: Chemistry
• keywords: metabolomics, insect brain tissue, GC-MS, pesticides, analytical chemistry
• language: English
• id: 9123343
• date added to LUP: 2023-06-13 11:11:43
• date last changed: 2024-02-09 08:58:48