The evolution of insecticide resistance in the brown planthopper (Nilaparvata lugens Stål) of China in the period 2012-2016

Abstract

The brown planthopper, Nilaparvata lugens, is an economically important pest on rice in Asia. Chemical control is still the most efficient primary way for rice planthopper control. However, due to the intensive use of insecticides to control this pest over many years, resistance to most of the classes of chemical insecticides has been reported. In this article, we report on the status of eight insecticides resistance in Nilaparvata lugens (Stål) collected from China over the period 2012-2016. All of the field populations collected in 2016 had developed extremely high resistance to imidacloprid, thiamethoxam, and buprofezin. Synergism tests showed that piperonyl butoxide (PBO) produced a high synergism of imidacloprid, thiamethoxam, and buprofezin effects in the three field populations, YA2016, HX2016, and YC2016. Functional studies using both double-strand RNA (dsRNA)-mediated knockdown in the expression of CYP6ER1 and transgenic expression of CYP6ER1 in Drosophila melanogaster showed that CYP6ER1 confers imidacloprid, thiamethoxam and buprofezin resistance. These results will be beneficial for effective insecticide resistance management strategies to prevent or delay the development of insecticide resistance in brown planthopper populations.

Figure 1

Collection sites of Nilaparvata lugens populations during 2012–2016. The map in this figure were generated by software Adobe Photoshop CS5 version (San Jose, CA, http://www.adobe.com/products/photoshop.html) based on our own data.

Figure 2

Resistance ratios to eight insecticides of the mixed population in each year. The data represents means and upper or lower limit.

Figure 3

Resistance ratios to four nicotinic acetylcholine receptor (nAChR) competitive modulators, imidacloprid (A), thiamethoxam (B), nitenpyram (C) and sulfoxaflor (D) of Nilaparvata lugens populations during 2012–2016.

Figure 4

Resistance ratios to buprofezin (A), chlorpyrifos (B), pymetrozine (C) and flufiprole (D) of Nilaparvata lugens populations during 2012–2016.

Figure 5

Fold change in expression of CYP6ER1 and CYP6AY1 in eight resistant N. lugens strains compared with the susceptible reference SS as determined by qPCR. Error bars display 95% fiducial limits.

Figure 6

Transgenic expression of CYP6ER1 (A,C,E and G) and CYP6AY1 (B,D,F and H) in D. melanogaster and their effects on imidacloprid, thiamethoxam and buprofezin resistance. (A and B) The expressions of CYP6ER1 and CYP6AY1 were confirmed by RT-PCR in two control lines and transgenic line. Three biological replicates of Da-GAL4, flies with genetic background correspond to Da-GAL4; Three biological replicates of UAS-CYP6ER1 (A) or UAS-CYPAY1 (B), flies not expressing the CYP6ER1 or CYP6AY1; Three biological replicates of Da > CYP6ER1 (A) or Da > CYP6AY1 (B), transgenic flies expressing the CYP6ER1 or CYP6AY1. (C,D,E,F,G and H). The comparison between survival rates of two control lines and transgenic line exposed to 1 mg/ml imidacloprid (C and D), thiamethoxam (E and F) and 400 mg/ml buprofezin (G and H). The data shown are the mean ± s.e.m. (n = 3). **P < 0.01, ***P < 0.001 (Chi-squared Test), ns (no significant).

Figure 7

Knockdown in the expression of CYP6ER1 in N. lugens field-resistance strain reduced its resistance to thiamethoxam and buprofezin. (A and C) The mRNA levels of CYP6ER1 were quantified by qRT-PCR at three days after dsRNA injection in FQ2016 (A) or buprofezin-resistance (C) strains, respectively. The data shown are mean + SEM (n = 3). Statistical significance of the gene expression between two samples was calculated using Student’s t test. *P < 0.05, **P < 0.01 and ***P < 0.001. (B and D) Effects on the mortality rate after injection of dsGFP or dsCYP6ER1 and followed by application of thiamethoxam (5 mg/L) (B) or buprofezin (50 mg/L) (D).