Cyto-Genotoxic and Behavioral Effects of Flubendiamide in Allium cepa Root Cells, Drosophila melanogaster and Molecular Docking Studies

Abstract

Flubendiamide (FLB) is an insecticide that is commonly employed to control pests on a variety of vegetables and fruits, with low toxicity for non-target organisms. However, due to its widespread use, the environmental risks and food safety have become major concerns. In this study, the toxicity potential of FLB was studied in the model organisms, Allium cepa and Drosophila melanogaster. The cyto-genotoxic effects of FLB on the root growth, mitotic index (MI), chromosomal aberrations (CAs) and deoxyribonucleic acid (DNA) damage in A. cepa root meristematic cells were investigated using the root growth inhibition Allium test and Comet assays. FLB caused CAs in the form of disturbed ana-telophase, chromosome laggards, stickiness, anaphase-bridge and polyploidy depending on the concentration and the exposure time. The toxicity and genotoxicity of FLB at various doses (0.001, 0.01, 0.1 and 1 mM) on D. melanogaster were investigated from the point of view of larval weight and movement, pupal formation success, pupal position, emergence success and DNA damage, respectively. FLB exposure led to a significant reduction of the locomotor activity at the highest concentration. While DNA damage increased significantly in the FLB-treated onions depending on the concentration and time, DNA damage in the FLB-treated D. melanogaster significantly increased only at the highest dose compared to that which occurred in the control group. Moreover, to provide a mechanistic insight into the genotoxic and locomotion-disrupting effects of FLB, molecular docking simulations of this pesticide were performed against the DNA and diamondback moth (DBM) ryanodine receptor (RyR) Repeat34 domain. The docking studies revealed that FLB binds strongly to a DNA region that is rich in cytosine-guanine-adenine bases (C-G-A) in the minor groove, and it displayed a remarkable binding affinity against the DBM RyR Repeat34 domain.

Keywords: DNA damage; chromosome aberrations; environmental risk assessment; pesticides; toxicity.

Figure 1

The inhibitory effect of FLB on A. cepa root growth. r = −0.973; p = 0.01.

Figure 2

The variation of the total number of CAs in A. cepa root cells depending on FLB concentrations and time exposure. Dose dependent: For 24 h, r = 0.71; p = 0.01. For 48 h, r = 0.725; p = 0.01. For 72 h, r = 0.616; p = 0.05. For 96 h, r = 0.518; p = 0.05. Time dependent: For 625 mg/L, r = 0.635; p = 0.01. For 1250 mg/L, r = 0.646; p = 0.01. For 2500 mg/L, r = 0.586; p = 0.01.

Figure 3

Typical stages of mitosis (a–d) and ana-telophase anomalies (e–i) induced by FLB in A. cepa cells. Scale bars: 10 µm.

Figure 4

DNA damage induced by FLB in A. cepa roots. Dose dependent: For 24 h, r = 0.969; p = 0.01. For 48 h, r = 0.895; p = 0.01. For 72 h, r = 0.918; p = 0.01. For 96 h, r = 0.900; p = 0.01. Time dependent: For 625 mg/L, r = 0.888; p = 0.01. For 1250 mg/L, r = 0.962; p = 0.01. For 2500 mg/L, r = 0.953; p = 0.01.

Figure 5

Effect of FLB exposure on pupa position and formation success of D. melanogaster.

Figure 6

Effect of FLB exposure on larval crawling of D. melanogaster.

Figure 7

Effect of FLB exposure on larval weight of D. melanogaster.

Figure 8

FLB-induced DNA damage scores in D. melanogaster.

Figure 9

Top-ranked conformation of the intermolecular interaction between flubendiamide and B-DNA. (a) Cartoon view of the DNA–flubendiamide complex; (b) 3D DNA-ligand interaction diagram of the top-ranked docking conformation. Nucleotides in interaction with flubendiamide are outlined in red boxes. The green and light grey dashed lines on the right image represent hydrogen bonds, whereas the purple and light cyan dashed lines represent the hydrophobic contact and halogen (fluorine) interactions, respectively.

Figure 10

Post-docking top-ranked conformation of flubendiamide in complex with DBM RyR Repeat34 domain. The green and light grey dashed lines represent hydrogen bonds, whereas the purple and light cyan dashed lines represent hydrophobic contacts and halogen (fluorine) interactions, respectively.