Identification and functional analysis of a novel cytochrome P450 gene CYP9A105 associated with pyrethroid detoxification in Spondoptera exigua Hubner

Authors: WANG, RUI LONG - South China Agricultural University; LIU, SHI WEI - South China Agricultural University; Baerson, Scott; QIN, ZHONG - South China Agricultural University; MA, ZHI HUI - South China Agricultural University; SU, YI JUAN - South China Agricultural University; ZHANG, JIA EN - South China Agricultural University

Submitted to: International Journal of Molecular Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/2/2018
Publication Date: 3/5/2018
Citation: Wang, R., Liu, S., Baerson, S.R., Qin, Z., Ma, Z., Su, Y., Zhang, J. 2018. Identification and functional analysis of a novel cytochrome P450 gene CYP9A105 associated with pyrethroid detoxification in Spondoptera exigua Hubner. International Journal of Molecular Sciences. 19:737.

Interpretive Summary: Insects frequently become resistant to growth inhibitory compounds such as allelochemicals and commercial pesticides they encounter in the environment by acquiring the capacity to inactivate them, for example through the acquisition of genes encoding specialized metabolic enzymes. Cytochrome P450 proteins are enzymes often involved in the metabolism of toxins encountered by herbivorous insects during feeding. This work focuses on the cytochrome P450 genes of Spodoptera exigua, commonly referred to as the beet armyworm. This organism poses a significant threat to commercial agriculture in many parts of the world and was therefore chosen as the model organism for these studies. We show in this work that three Spodoptera exigua cytochrome P450 genes are likely involved in the metabolism of pyrethroid toxins by first severely inhibiting their expression in vivo. The resulting insects become much more sensitive to inhibition from several commonly-encountered pyrethroid toxins. Additionally, all three genes (designated CYP9A105,CYP9A106 and CYP9A107), exhibit increased expression in untreated individuals exposed to these same toxins. Taken together, the results from this work strongly suggest roles for CYP9A105,CYP9A106 and CYP9A107 in detoxifying pyrethroid compounds encountered by Spodoptera exigua while feeding.

Technical Abstract: In insects, cytochrome P450 monooxygenases (P450s or CYPs) are known to be involved in the detoxification and metabolism of insecticides, leading to increased resistance in insect populations. The beet armyworm, Spodoptera exigua, is one of the most agriculturally important polyphagous pest species, and has developed resistance to various insecticides. In this study, three CYP3 clan P450 genes (CYP9A105, CYP9A106 and CYP9A107) were identified and characterized from S. exigua. The cDNAs of CYP9A105, CYP9A106 and CYP9A107 encoded 530, 531 and 531 amino acid proteins, respectively. Quantitative real-time PCR analyses showed that CYP9A105, CYP9A106 and CYP9A107 were expressed at all developmental stages, with maximal expression observed in 5th instar stage larvae, and in dissected 5th instar larvae the highest transcript levels were found in midguts and fat bodies. The expression of CYP9A105, CYP9A106 and CYP9A107 in midguts was upregulated by treatments with the insecticides a-cypermethrin, deltamethrin and fenvalerate at both LC15 concentrations (0.10, 0.20 and 5.0 mg/L, respectively) and LC50 concentrations (0.25, 0.40 and 10.00 mg/L, respectively). RNA interference (RNAi) mediated silencing of CYP9A105, CYP9A106 or CYP9A107 led to increased mortalities of insecticide-treated 4th instar S. exigua larvae. The mortalities of S. exigua exposed to deltamethrin, a-cypermethrin and fenvalerate increased from 45.9% to 73.7%, 56.3% to 79.6% and 42.6% to 69.6%, respectively, after RNAi-mediated silencing of CYP9A105. Following RNAi-mediated silencing of CYP9A106, mortalities of S. exigua exposed to deltamethrin, a-cypermethrin and fenvalerate increased from 45.9% to 68.9%, 56.3% to 77.4% and 42.6% to 79.3%, respectively. A similar result was obtained following RNAi-mediated silencing of CYP9A107, where mortalities of S. exigua exposed to deltamethrin, a-cypermethrin and fenvalerate increased from 45.9% to 81.9%, 56.3% to 74.1% and 42.6% to 76.3%, respectively. Our results strongly suggest that these three CYP9A genes play important roles in pyrethroid insecticide resistance in S. exigua.