CRISPR/Cas9-mediated knockout of the NlCSAD gene results in darker cuticle pigmentation and a reduction in female fecundity in Nilaparvata lugens (Hemiptera: Delphacidae)

Chen, Jingxiang; Li, Wan-Xue; Lyu, Jun; Hu, Yu-Tao; Huang, Guohong; Zhang, Wenqing

doi:10.1016/j.cbpa.2021.110921

Cited by 17 publications

(16 citation statements)

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“…Mutations in genes that control cuticle biogenesis and color have been used as phenotypic markers for assessing the success of RNAi and CRISPR/Cas9 editing strategies in the Hemiptera. Genes that influence the cuticle were successfully mutagenized using CRISPR/Cas9-mediated gene editing in N. lugens, E. heros, and A. pisum ( Le Trionnaire et al, 2019 ; Cagliari et al, 2020 ; Chen et al, 2021 ). In N. lugens, Chen et al (2021) studied mutants in the target gene cysteine sulfinic acid decarboxylase ( NlCSAD ), which influences dark melanin pigment accumulation.…”

Section: Introductionmentioning

confidence: 99%

“…Genes that influence the cuticle were successfully mutagenized using CRISPR/Cas9-mediated gene editing in N. lugens, E. heros, and A. pisum ( Le Trionnaire et al, 2019 ; Cagliari et al, 2020 ; Chen et al, 2021 ). In N. lugens, Chen et al (2021) studied mutants in the target gene cysteine sulfinic acid decarboxylase ( NlCSAD ), which influences dark melanin pigment accumulation. They compared the outcomes of silencing NlCSAD by RNAi versus generation of NlCSAD null mutations.…”

Section: Introductionmentioning

confidence: 99%

“…Therefore, sequencing of DNAs extracted from the discarded exuvia from hatched 5 th -instar G0 nymphs was used to identify putative NlCSAD mutants. Mutations in the NlCSAD gene occurred at a frequency of 9.5% and seven mutant alleles were detected ( Chen et al, 2021 ). This low frequency may explain the absence of phenotype.…”

Section: Introductionmentioning

confidence: 99%

“…To demonstrate inheritance, two mutant G0 generation females were outcrossed with wild-type males to generate G1 generation progeny, of which 36.3% displayed a darker cuticle than wild-type controls ( Chen et al, 2021 ). Mutant alleles were verified using 5th-instar exuvia and G1 adults with identical alleles (a 4-bp deletion in NlCSAD ) were mated to produce the G2 generation.…”

Section: Introductionmentioning

confidence: 99%

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Gene Editing and Genetic Control of Hemipteran Pests: Progress, Challenges and Perspectives

Pacheco

Walling

Atkinson

2022

Front. Bioeng. Biotechnol.

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The origin of the order Hemiptera can be traced to the late Permian Period more than 230 MYA, well before the origin of flowering plants 100 MY later in during the Cretaceous period. Hemipteran species consume their liquid diets using a sucking proboscis; for phytophagous hemipterans their mouthparts (stylets) are elegant structures that enable voracious feeding from plant xylem or phloem. This adaptation has resulted in some hemipteran species becoming globally significant pests of agriculture resulting in significant annual crop losses. Due to the reliance on chemical insecticides for the control of insect pests in agricultural settings, many hemipteran pests have evolved resistance to insecticides resulting in an urgent need to develop new, species-specific and environmentally friendly methods of pest control. The rapid advances in CRISPR/Cas9 technologies in model insects such as Drosophila melanogaster, Tribolium castaneum, Bombyx mori, and Aedes aegypti has spurred a new round of innovative genetic control strategies in the Diptera and Lepidoptera and an increased interest in assessing genetic control technologies for the Hemiptera. Genetic control approaches in the Hemiptera have, to date, been largely overlooked due to the problems of introducing genetic material into the germline of these insects. The high frequency of CRISPR-mediated mutagenesis in model insect species suggest that, if the delivery problem for Hemiptera could be solved, then gene editing in the Hemiptera might be quickly achieved. Significant advances in CRISPR/Cas9 editing have been realized in nine species of Hemiptera over the past 4 years. Here we review progress in the Hemiptera and discuss the challenges and opportunities for extending contemporary genetic control strategies into species in this agriculturally important insect orderr.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Gene Editing and Genetic Control of Hemipteran Pests: Progress, Challenges and Perspectives

Pacheco

Walling

Atkinson

2022

Front. Bioeng. Biotechnol.

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“…Each microinjection was performed under a Nikon C‐DSS230 stereomicroscope (Nikon, Tokyo, Japan), using a FemtoJet 4i microinjector (Eppendorf, Hamburg, Germany). The dsRNA was injected into the insect body through a site positioned between the middle leg and hind leg of BPH 31 . Each BPH was injected with 50 nL of dsRNA (5000, 2500, and 1250 ng/μL).…”

Section: Methodsmentioning

confidence: 99%

RNAi‐mediated silencing of the autophagy‐related gene NlATG3 inhibits survival and fecundity of the brown planthopper, Nilaparvata lugens

Feng

et al. 2021

Pest Management Science

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BACKGROUND The brown planthopper (BPH), Nilaparvata lugens, is a serious insect pest of rice. Autophagy and its related gene ATG3 play multiple roles in insects. However, information about the functions of ATG3 in BPH (NlATG3) is unavailable, and its potential as a target for pest control remains unclear. RESULTS RT‐qPCR results showed a relatively low expression of NlATG3 in 1st–4th‐instar nymphs, which increased through 9‐day‐old adults. The expression of NlATG3 increased continuously in 1‐day‐old through 5‐day‐old eggs, whereas it decreased thereafter. The mRNA level of NlATG3 was markedly higher in the ovary (1.16) and head (1.00) compared to the rest body parts of BPH adults. Injecting nymphs with dsNlATG3 at doses from 62.5 to 250 ng per insect had strong lethal effect upon them. For the 5th‐instar nymphs, all individuals died within 5 days after receiving the dsNlATG3, and importantly, no individual successfully molted. Transmission electron microscopy revealed the new cuticle of nymphs injected with dsNlATG3 became loose and curved, which is clearly different from that of the control. Correspondingly, the obvious vesicles in epidermal cells disappeared after dsNlATG3‐treatment. RNAi of NlATG3 significantly reduced the total number of eggs laid per female as well as the eggs' hatchability, especially in the dsNlATG3♀ × dsGFP♂ group, whose total number of eggs laid per female largely decreased by 80.4%, and whose eggs' hatchability was significantly reduced from 95.7% to zero, when compared with the control (dsGFP♀ × dsGFP♂). CONCLUSION NlATG3 is a promising target for developing RNAi‐based insect management strategies. © 2021 Society of Chemical Industry.

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CRISPR/Cas9‐mediated knockout of NlCYP6CS1 gene reveals its role in detoxification of insecticides in Nilaparvata lugens (Hemiptera: Delphacidae)

Zhang

Gao

et al. 2023

Pest Management Science

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BACKGROUND: The brown planthopper (Nilaparvata lugens) is one of the major rice insect pests in Asia. Recently, high levels of insecticide resistance have been frequently reported and cytochrome P450 monooxygenase (P450)-mediated metabolic detoxification is a common resistance mechanism in N. lugens. However, there has been no persuasive genetic method to prove the role of P450s in insecticide resistance in N. lugens.RESULTS: Here, CRISPR/Cas9 system was used to disrupt the P450 gene NlCYP6CS1 to elucidate its role in insecticide resistance in field populations of N. lugens. We successfully constructed a homozygous strain (Nl6CS1-KO) with a 5-bp deletion and 1-bp insertion mutation of NlCYP6CS1. Compared with a background resistant strain (Nl-R), the susceptibility of knockout strain Nl6CS1-KO to imidacloprid, nitenpyram, thiamethoxam, dinotefuran, and pymetrozine was increased by 2.3-, 3.4-, 7.0-, 4.2and 3.9-fold, respectively, but not significantly changed to triflumezopyrim, chlorpyrifos and buprofezin. Life table analysis demonstrated that the Nl6CS1-KO strain resembled the Nl-R strain in terms of egg and nymph developmental duration and adult lifespan, but differed from the Nl-R strain in the survival rate of eggs and nymphs, reproduction, and body weight.CONCLUSIONS: Our study demonstrates the effect of functional deletion of NlCYP6CS1 on multiple insecticide resistance in N. lugens. For the first time, we applied CRISPR/Cas9 system to reveal the mechanism of insecticide resistance in N. lugens, which may shed light on similar studies in other hemipteran insects.

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CRISPR/Cas9-mediated knockout of the NlCSAD gene results in darker cuticle pigmentation and a reduction in female fecundity in Nilaparvata lugens (Hemiptera: Delphacidae)

Cited by 17 publications

References 45 publications

Gene Editing and Genetic Control of Hemipteran Pests: Progress, Challenges and Perspectives

Gene Editing and Genetic Control of Hemipteran Pests: Progress, Challenges and Perspectives

RNAi‐mediated silencing of the autophagy‐related gene NlATG3 inhibits survival and fecundity of the brown planthopper, Nilaparvata lugens

CRISPR/Cas9‐mediated knockout of NlCYP6CS1 gene reveals its role in detoxification of insecticides in Nilaparvata lugens (Hemiptera: Delphacidae)

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