Table 2.

Summary of crop protection applications using an encapsulation system for RNAi delivery.

Encapsulation System	Potential Crop Protection Application	Strategy	Reference
Guanylated 2-(aminoethyl) methacrylate (AEMA)/dsRNA polyplex nanoparticles.	Insecticide induces decreased feeding in Lepidopteran larvae (Spodoptera exigua), then promoting weight loss, developmental halt, and mortality.	Increases RNAi efficiency in targeting the essential gene chitin synthase B (ChSB), while preventing the degradation of dsRNA in the alkaline gut of insects and enhancing its cellular uptake in midgut cells.	[47]
poly-[N-(3-guanidinopropyl) methacrylamide] (pGPMA)/dsRNA interpolyelectrolyte nanocomplex.	Ingestion insecticide regulates gene silencing in Lepidopteran larvae (Spodoptera frugiperda), increasing mortality from starvation and growth stunting.	Increased internalization and protection of dsRNA in insect cells, decreasing the accumulation of target mRNA due to the knockdown of genes related to vital functions such as nutrient absorption (sfVATPase), intracellular transport (sfKIF), and cell division (sfCDC27).	[52]
Chitosan/dsRNA polyplex nanoparticles	Nematicide can homogeneously enter the nematode’s body (Caenorhabditis elegans) through noncanonical endocytotic pathways and attack specific genes. The combined effect decreases the development of the nematode by the action of the chitosan vehicle.	Increases RNAi efficiency of gene knockdown throughout the whole body of the nematode by introducing intact dsRNA through the Clathrin-mediated endocytosis pathway, which is different from the canonical pathway (sid-1 and sid-2) in the study model. Furthermore, chitosan was shown to effectively decrease myosin gene expression, which is critical for the growth and reproduction of the model nematode.	[22]
Chitosan/dsRNA polyplex nanoparticles	Insecticide against Lepidopteran larvae (Spodoptera frugiperda) acts on genes related to the apoptosis pathway, inducing growth impairment and larval mortality.	Improves RNAi efficiency through the protection of dsRNA from degradation by intracellular and intercellular RNases. It also reduces the accumulation of dsRNA in the endosome while favoring its transport to the cytoplasm, where the formation of siRNAs is promoted, producing knockdown of apoptosis-related genes (iap).	[45]
Layered double hydroxide (LDH) clay nanosheets/dsRNA	Develop a topical product that induces viral resistance in plants (against PMMoV and CMV) using dsRNA absorption technology in clay nanosheets (Bio-Clay).	Increased persistence of the topical treatment due to strong adhesion of dsRNA in the vehicle (LDH) and with the leaves. It also allows the controlled release of the biomolecule and confers protection against environmental degradation while favoring the internalization of dsRNA in the plant.	[44]
Lipofectamine 2000 liposomes/dsRNA.	Insecticide against Diptera of the genus Drosophila (D. melanogaster, D. sechellia, D. yakuba, and D. pseudoobscura) acting by ingestion. It attacks essential genes of development through knockdown management.	Promotion of dsRNA internalization in insects through encapsulation protection, increasing silencing efficiency by promoting more significant RNAi accumulation in larvae. Knockdown of the genes of the VATPase (gut lumen pH stabilizer associated with nutrient uptake) and gTub23C (mitosis-related g-tubulin protein, essential for microtubule organization).	[55]
Lipofectamine 2000 liposomes/dsRNA.	Specific insecticide against larvae and adults of Drosophila suzukii combining synergic effect of multiple gene knockdown. Oral administration route.	It facilitates uptake in the insect’s gut. It causes significant mortality in larvae and adults by the reduction in transcript levels of essential genes rps13 (housekeeping), alpha COP (coatomer subunit for trans-organelles transport), and vha26 (subunit of the vacuolar ATPase). The synergistic action of knockdown of the rps13 and alpha COP genes significantly increases mortality in the insect.	[53]
Liposomes/dsRNA	Oral insecticide for the control of nymphs of Euschistus heros (hemiptera: pentatomidae), which is one of the main soybean pests in the field.	Protection of dsRNA against degradation promoted by the ribonuclease action of insect saliva. Enhanced silencing activity of target genes vATPaseA (V-type proton ATPase catalytic subunit A) and act-2 (muscle actin).	[56]
Recombinant Flock House Virus FHV/dsRNA	Recombinant insecticide based on a viral vehicle transporting dsRNA silencers of essential genes in Drosophila melanogaster. For potential massive application in other species susceptible to FHV infection.	Use of the insect cell machinery to assemble infective recombinant FHV virions that carry target sequences for the production of dsRNA when replicating in cells. Thus, virions protect the sequences responsible for silencing the rps13 (housekeeping), alpha COP (coatomer subunit for trans-organelle transport), and vha26 (subunit of the vacuolar ATPase) genes while at the same time favoring dispersal in insects. It simulates natural viral infection.	[54]
Virus-Like Particles (VLP)/dsRNA	Oral insecticide for the control of ants of several genera (Solenopsis invicta (fire ants), Camponotus pennsylvanicus and Camponotus floridanus (carpenter ants), Linepithema humile (Argentine ants), Tapinoma sessile (odorous ants), Tetramorium caespitum (pavementom ants), and Monstrous ants) pharaonis (pharaoh ants); inducing the silencing of physiological genes required for the survival of the colony.	Recombinant production in E. coli, which through specific plasmids manufactures capsid proteins of bacteriophages Qß and MS2 and inducible RNAi precursor sequences. The packaging of dsRNAs in VLPs protects them from degradation by nonspecific environmental organisms and the intestinal RNases of the target organism. It also favors its absorption by lining the gut cells. The silenced genes are related to the viability of the colony, for example, the induction of sterility and individual mortality. VLP carrying dsRNA is sprayed on the ground for spot application or incorporated into the bait. Target genes include VgR (vitellogenin receptor protein), TVXl (telomerase variant XI protein), PBAN (pheromone biosynthesis activating neuropeptide), PBANR (pheromone biosynthetic activating neuropeptide receptor), WLS (wntless protein), MEGF10 (multiple epidermal growth factor-like domain proteins 10), CHCP (clatherin heavy chain protein), CDC7 (cell division cycle 7-related protein), Cep89 (centrosomal protein 89 kdal), PSMBl (beta subunit of the type-1 proteosome), A5C (actin 5C protein), ATPSD (ATP synthase delta subunit); as well others related with anamorsin, beta actin, and Csp9 proteins	[46] WO2017/136353Al for APSE RNA Containers (ARCs)
Ribonucleoprotein particle (RNP)/dsRNA	Insecticide for control of cotton boll weevil (Anthonomus grandis) adults.	Developing a protection and stability system for dsRNA avoids degradation by nucleases in the insect’s gut and favors rapid cellular incorporation. The above is based on a chimeric protein PTD-DRBD (peptide transduction domain–dsRNA binding domain) combined with dsRNA. This type of resulting protein is known as cell-penetrating peptide (CPP).	[51]