Insect pests impose a serious threat to agricultural productivity. Initially, for pest management, several breeding approaches were applied which have now been gradually replaced by genome editing (GE) strategies as they are more efficient and less laborious. Due to its specificity and easy handling, CRISPR/Cas9-based genome editing has been applied to a wide range of organisms for various research purposes. For pest control, diverse approaches have been applied utilizing CRISPR/Cas9-like systems, thereby making the pests susceptible to various insecticides, compromising the reproductive fitness of the pest, hindering the metamorphosis of the pest, and there have been many other benefits.
Insect Species | Target Gene | Accession Number | Genetic Trait | Mutation Type |
Delivery of CRISPR Components | Findings | References |
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Drosophila melanogaster | yellow, rosy | NM_057444.3, NM_079613.3 | Pigmentation and Mating | Knockout, Knockin | Plasmid | This was the first report using the CRISPR/Cas9 system to mediate efficient genome engineering in Drosophila. | Gratz et al., 2013 |
yellow, white | NM_057444.3, NM_079613.3 |
Pigmentation and Mating | Knockout | Cas9 mRNA and sgRNA | sgRNA concentration-dependant knockout was shown for yellow gene, and highly efficient and varied genome editing efficiencies were shown by different sgRNAs. | Bassett et al., 2013 | |
yellow | NM_057444.3 | Pigmentation and Mating | Knockout | Cas9 mRNA and gRNA | This report used the approach of targeting multiple genes with different sgRNAs, and it attained a remarkably effective targeted mutagenesis. | Yu et al., 2013 | |
Ast, Eh, capa, Ccap, Crz, npf, Mip, mir-219, mir-315, white | NM_001300582.1, NM_079662.3, NM_079828.3, NM_001275917.2, NM_079626.3, NM_080493.3, NM_140714.4, NR_048289.1, NR_048297.1, X76202.1 |
Knockout | Plasmid | To obtain a Cas9–sgRNA complex for achieving targeted mutagenesis, two transgene vectors harboring expression cassettes for Cas9 and sgRNA were delivered. | Kondo and Ueda, 2013 | ||
rosy, DSH3PX1 | NM_079613.3, NM_140091.4 | Pigmentation and Mating | Knockout, Knockin | Plasmid | Executed efficient and complex genomic manipulations using CRISPR/Cas9-mediated HDR. | Gratz et al., 2014 | |
ebony, yellow, wingless, wnt | NM_079707.4, NM_057444.3, NM_078778.5 | Segmentation | Knockout, Knockin | Plasmid | Different promoters were used to drive sgRNA expression, and based on promoter properties, different patterns of expression were observed. | Port et al., 2014 | |
EGFP, mRFP | Chromogenic fluorophores | Knockout | Plasmid | Induction of mutations by injection of an sgRNA into Vasa-Cas9 transgenic fly embryos. | Sebo et al., 2014 | ||
white, piwi | NM_057439.2, NM_001298896.1 | Pigmentation and Expression of group of small RNA | Knockout, Knockin | Plasmid | Used Cas9 nickase and sgRNA pairs to prevent off-target effects during the generation of indel mutants. | Ren et al., 2014 [110] |
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ms(3)k81, white, yellow | NM_143253.2, NM_057439.2, NM_057444.3 |
Pigmentation | Knockout, Knockin | Plasmid | CRISPR mediated genome editing was shown in Drosophila. | Xue et al., 2014a | |
yellow, notch, bam, nos,ms(3)k81, cid |
NM_057444.3, NM_001258581.2, NM_057452.4, NM_057310.4, NM_143253.2, NM_079006.4 | Physiology | Knockout | Plasmid | A CRISPR/Cas9-mediated conditional mutagenesis system combined with tissue-specific expression of Cas9 was used to temporally and spatially inhibit gene expression. | Xue et al., 2014b | |
salm | NM_164966.3 | Zinc Finger Transcriptional Repressor | Knockin | mRNA, transgene | For flexible modification of fly genome, a two-step method was proposed. | Zhang X. et al., 2014 | |
ebony, yellow, vermilion | NM_079707.4, NM_057444.3, NM_078558.3 | Pigmentation | Knockout, Knockin | Plasmid, transgene | Donor template and sgRNA plasmids were injected into Cas9 transgenic embryos in Drosophila. | Ren et al., 2014b | |
ebony, yellow, white | NM_079707.4, NM_057444.3, NM_057439.2 | Pigmentation | Knockout, Knockin | Plasmid, transgene | A bicistronic Cas9/sgRNA vector was constructed which enhanced the efficiency of gene targeting. | Gokcezade et al., 2014 | |
ebony, yellow, wg, wls, Lis1, Se | NM_079707.4, NM_057444.3, NM_078778.5, NM_140188.4, NM_057812.5, NM_139978.4, NM_057812.5 | Pigmentation and Physiology | Knockout, Knockin | Plasmid, transgene | Non-transgenic individuals exhibited lessefficient knockin than transgenic individuals did. | Port et al., 2015 [111] |
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yellow | NM_057444.3 | Pigmentation | Knockin | Transgene | Heterozygous recessive mutation was converted to homozygous loss of function mutations utilizing mutagenic chain reaction (MCR) technology in Drosophila. | Gantz and Bier, 2015 | |
Dα6 | NM_164874.3 | Insecticide resistance | Knockin | Plasmid | The G275E mutation of the nAChR Dα6 subunit is directly related to Spinosad resistance. | Zimmer et al., 2016 | |
LUBEL | NM_001273232.2 | Growth and Development | Knockout | Plasmid | Flies with LUBEL mutations exhibited reduced survival and defective climbing in response to heat. | Asaoka et al., 2016 | |
Scsα | NM_079181.4 | Growth and Development | Knockout | Plasmid | Mutant flies could not produce sufficient energy to promote normal growth. | Quan et al., 2017 | |
clamp | NM_136293.4 | Sex Specific | Knockout | Plasmid | The expression of a sex-specific gene was regulated by an essential transcription factor. | Urban et al., 2016 | |
chameau, CG4221, CG5961 | NM_135273.5,NM_141949.4 | Knockin | mRNA | HDR-mediated genome modifications efficiency was tested, and a problem associated with “ends-in” recombination was resolved. | Yu et al., 2014 | ||
fdl | NM_165908.2 | Knockout | Plasmid | Capability of CRISPR/Cas9 system for analysing or manipulating protein glycosylation pathways. | Mabashi-Asazuma et al., 2015 [112] |
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mod(mdg4) | NM_163878.2 | Knockout | Plasmid | Validation of a functional gene involved in trans-splicing that influenced the development in flies. | Gao et al., 2015 [113] |
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act5C, lig4, mus308 | NM_167053.2, NM_132679.3, L76559.1 | Knockout, Knockin | Plasmid, transgene | Offered a comprehensive technique for genome editing in Drosophila S2 cells. | Kunzelmann et al., 2016 [114] |
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yellow, white, tan | NM_057444.3, NM_057439.2, NM_132315.1 |
Pigmentation | Knockin | Plasmid, transgene | Proposed a new process of attaining single or multiple allelic substitutions. | Lamb et al., 2016 | |
wntless | NM_140188.4 | Growth and development | Knockout | Plasmid | A complex of tRNA–sgRNA was proposed to amplify the cleavage efficiency of the Cpf1 and Cas9 nucleases. | Port and Bullock, 2016 | |
TpnC | NM_078895.4 | Growth and development | Knockout | Plasmid | Confirmed that the myofibril assembly is related to TpnC gene. | Chechenova et al., 2017 | |
Alk | NM_144343.3 | Growth and development | Knockout | Plasmid | Revealed that transcription factors can affect Alk gene expression by establishing mutations in Alk enhancer regions. | Mendoza-Garcia et al., 2017 | |
Drosophila suzukii | white (w-) | NM_057439.2 | Pigmentation | Knockout | Plasmid | Absence of mating and copulation failure was reported. The mutation also caused pigmentation deficiency in testis sheath, which could be a probable reason for copulation failure. | Yan et al., 2020 |
white, Sxl | NM_057439.2, XM_017083263.2 | Sex determination | Knockout | Plasmid | Sxl gene was proved as excellent gene to suppress the population growth of this destructive pest. | Li and Scott, 2016 | |
DsRed (red fluorescence protein) |
knockin | Plasmid, transgene | The enhancer/promoter of the spermatogenesis-specific beta-2-tubulin (β2t) gene was used for expression of fluorescent proteins or effector molecules in testes of pests, and this providing basis for reproductive biology studies sexing and monitoring. | Ahmed, H. M. et al., 2019 | |||
Drosophila subobscura | yellow, white | XM_034814491.1, XM_034808177.1 |
Pigmentation | Knockout | mRNA | Gene functions were analyzed in a non-model Drosophila species. | Tanaka et al., 2016 [115] |
Anastrepha ludens | Astra-2 | EU024509.1 | Sex determination | Knockout | RNP complex | The mutation caused sterility, thus, the target gene was proposed for helping in pest control. | Li et al., 2019 |
Bactrocera dorsalis | White and transformer | AY055817.1, KP342062.1 | Sex determination and reproduction | Knockout | RNP complex | CRISPR/Cas9 mediated mutation of white and transformer genes caused various phenotypic effects. | Zhao et al., 2018 |
Ceratitis capitata |
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Homology directed repair | knockin | RNP complex and a single-stranded oligo donor | Conversion of eGFP-to-BFP was demonstrated for establishing an efficient HDR through CRISPR-based genome editing. | Aumann, R. A. et al., 2018 | |
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X89933.1, XM_020858622.2 | Segmentation | Knockout | RNP complex | A simple and highly efficient RNP complex-based genome editing approach was reported with the details of designing and preparation. | Meccariello, A. et al., 2017 | |
Helicoverpa armigera | NPC1b | MK555324.1 | Growth and dietary uptake of Cholesterol | Knockout | RNP complex | NPC1b is vital for the growth and dietary cholesterol uptake. Thus, a novel pest-management technique can be developed using NPC1b as an insecticidal target. | Zheng, J. C. et al., 2020 |
HaCad | JX23382.1 | cell–cell adhesion | Knockout | sgRNAs and Cas9 mRNA | sgRNAs and Cas9 mRNA were injected into the fresh eggs, and a high editing efficiency of the HaCad locus was achieved. | Wang, J. et al., 2016 | |
HaABCA2 | KP259911.1 | Regulation of enzymes | Knockout | Cas9 mRNA and sgRNA | The knockout of HaABCA2 confirmed the role of HaABCA2 in mediating toxicity of both Cry2Aa and Cry2Ab against H. armigera. | Wang, J. et al., 2017 | |
odorant receptor 16 (OR16) |
KF768670.1 | Olfaction | Knockout | Cas9 mRNA + sgRNA and RNP complex | The results represent the basis for novel olfactory-based strategies of pest population control. |
Chang, H. et al., 2017 | |
white, ok, brown, and scarlet | XM_021344759.2, KU754490.1, KU754480.1, KU754478.1 | Pigmentation | Knockout | Cas9 mRNA | The report represented differential distribution of eye pigments in the mutants; this finding may be helpful in elucidation of biosynthetic pathway. | Khan, S. A.,et al., 2017 | |
cluster of nine P450 genes | KM016735.1, R095600.1, KM016739.1, KM016740.1, DQ256407.1, KM016743.1, KM016741 | Regulation of detoxifying enzymes | Knockout | Cas9 protein and multiple sgRNAs | The report identified the key players in the insecticide metabolism. | Wang, H. et al., 2018 | |
Plutella xylostella | Pxabd-A | XM_011570968.3 | Body segmentation | Knockout | Cas9 mRNA | CRISPR/Cas9 was used to target genes in P. xylostella for the first time which provided new ideas for pest control. | Huang et al., 2016 |
Pxdsx | XM_048630440.1 | Sex determination | Knockout | Microinjection of RNP complex | The results showed CRISPR/Cas9 system led to altered expression of sex biased genes. | Wang et al., 2019 | |
PxCHS1 | AB271784.1 | Development | Knockout | Plasmid | Description of the resistance management strategies for insect pests, it and explained the MoA behind the resistance using CRISPR/Cas9 system. | Douris et al., 2016 | |
LW | Locomotion | Knockout | RNP complex | The results showed weaker phototaxis and reduced locomotion, thus making it a helpful method for pest control | Chen et al., 2021 [116] |
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Spodoptera frugiperda | Sfabd-A | MH541836.1 | Body segmentation | Knockout | RNP complex | The results showed that gene function validation and the understanding of resistance mechanism can be performed using CRISPR/Cas9 system which can lead to the development of novel pest management approaches. | Wu et al., 2018 |
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XM_035582273.2 XM_050696092.1 XM_050696079.1 |
Growth and development | Knockout | Cas9 protein and multiple sgRNAs | The developed mutants were helpful to understand the crucial pathways of S. frugiperda and the strategy can also applied for other invasive pests. | Zhu, G. H. et al., 2020 [117] |
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Spodoptera litura | Slabd-A | GCA_002706865.1 | Body segmentation | Knockout | Cas9 mRNA and sgRNA | The direct injection of Cas9-coding mRNA and Slabd-A-specific sgRNA into the embryos of the S. litura led to the induction of the typical abd-A deficient phenotypes showing irregular segmentation and unusual pigmentation at the larval stage. | Bi, H. L. et al., 2016 |
SlitBLOS2 | XM_022977403.1 | Molecular marker | Knockout | Cas9 mRNA and sgRNA | The study demonstrated that SlitBLOS2 has a role in the coloration of the integuments, and thus, it provided a marker gene for functional studies and pest control strategies. | Zhu, G. H. et al., 2017 | |
Spodoptera littoralis | SlitOrco | Olfaction | Knockout | mRNA | The Orco gene was investigated in the insect Spodoptera littoralis. The results were helpful in making a pest control strategy and in gene function analysis. | Koutroumpa et al., 2016 | |
Spodoptera exigua | Seα6 | MN714701.1 | Knockout | RNP complex | The study demonstrated that knocked-out Seα6 was highly resistant to insecticides. | Zuo et al., 2020 | |
Dendrolimus punctatus | DpWnt-1 | KU640201.1 | Development and segmentation | Knockout | mRNA | Proved the necessity of DpWnt-1 signaling in appendage development and anterior segmentation. | Liu H. et al., 2017 |
Cydia pomonella | CpomOR1 | FJ385021.1 | Olfaction | Knockout | Cas9 mRNA and sgRNA | The report demonstrated mutation in the CpomOR1 gene via CRISPR/Cas9 affected the egg production and viability in the insect. | Garczynski, S. F. et al., 2017 |
Ostrinia furnacalis | OfAgo1 | Growth and development | knockout | sgRNA and Cas9 mRNA | Mutation in OfAgo1 gene through CRISPR/Cas9 technology caused cuticle disruption. | You et al., 2019 | |
Agrotis ipsilon | AiTH | Growth and development | Knockout | sgRNA and Cas9 mRNA | The AiTH gene knockout by CRISPR/Cas9 caused narrowing in the egg shell. | Yang et al., 2018 | |
Hyphantria cunea | Hcdsx | Reproduction | Knockout | sgRNA and Cas9 mRNA | Knocked-out Hcdsx gene by CRISPR/Cas9 caused sex-specific sterility, thus making it a pest control method. | Li et al., 2020 | |
Mythimna separata | NPC1b | MZ209049.1 | Intestinal absorption and sterol trafficking | Knockout | RNP complex | Knockout of NPC1b can hamper nutrient absorption. | Tang et al. 2022 |
Nilaparvata Lugens | Nl-cn and Nl-w |
MH105806.1 | Pigmentation | Knockout | Cas9 mRNA and sgRNA | Two genes for eye pigmentation were targeted using CRISPR/Cas9, and the results paved path for gene-function interrogation. | Xue. et al., 2018 |
Diaphorina citri | ACP-TRX-2 | XM_026831570.1 | Physiology | Knockout | BAPC-assisted delivery of CRISPR components | The method incorporated BAPC-assisted delivery of CRISPR/Cas9 into nymphs and adults, thus resulting an innovative breakthrough in gene editing, it and has shown a significant improvement over efforts using injection of eggs. | Hunter et al., 2018 |
Diaphorina citriHomalodisca vitripennis, Bemisia argentifolii | Thioredoxin and Vermillion | XM_046819472.1 | Physiology andEye color | Knockout | BAPC, plasmid, dsRNA | The BAPC-assisted delivery system developed gene editing methods across the all hemipteran pests by permitting the use of nymphs and adults. BAPC-assisted CRISPR delivery transformed the approaches to protect food crops from different pathogens and insect vectors. | Hunteret al., 2019 |
Bemisia tabaci | white | XM_019053144.1 | Pigmentation | Knockout | SgRNA + Cas9 protein fused with overy targeting peptide ligand (BtKV) | The method has significantly expanded the capability of CRISPR techniques for whitefly research. | Heu et al., 2020 |
Euschistus heros | abnormal wing disc (awd), tyrosine hydroxylase (th) and yellow (yel) | NP_001119625.1, XP_008182999.1, XP_001948479.1 | Body segmentation and pattern | Knockdown and knockout | dsRNA, RNP complex | Use of RNAi and CRISPR/Cas9 techniques for managing insect pests. | Cagliari et al., 2020 |
Tribolium castaneum | Tribolium E-cadherin | XM_961215.3 | Dorsal closure defect | Knockout | Plasmid | Tribolium E-cadherin gene was targeted for knockout study. | Gilles et al., 2015 |
Leptinotarsa decemlineata | vestigial gene (vest) | XM_023168389.1 | Growth and development | Knockout | RNP complex | Functionally characterized vest gene and CRISPR/Cas9 protocol was established for mutagenesis. | Gui, S. et al., 2020 |
Locusta migratoria | Orco | JN989549.1 | Olfaction | Knockout | mRNA | Functional genetic studies of locusts by generation of loss-of-function mutation for managing insect pests. | Li Y. et al., 2016 |
Tetranychus urticae | PSST | KX806605.1 | Knockout | Plasmid | Substitution in the H92R amino acid of the PSST homolog was related to pyridaben resistance and the mutation into the Drosophila PSST homolog using CRISPR/Cas9 genome-editing tools. | Bajda et al., 2017 | |
phytoene desaturase | MF167355.1 | Knockout | RNP complex | Induction of two mutagenetic events using CRISPR/Cas9 providing basis for functional studies. | Dermauw, W. et al., 2020 |
This entry is adapted from the peer-reviewed paper 10.3390/agriculture12111896