title: DsRNA-mediated targeting of ribosomal transcripts RPS6 and RPL26 induces long-lasting and significant reductions in fecundity of the vector Aedes aegypti.
abstract: Ribosomal transcripts produce critical proteins that are involved in most cellular production processes. Targeting ribosomal transcripts has produced mortality in mites and ticks but the effect of ribosomal transcript knockdown has not been thoroughly examined in mosquitoes. We examine the effects of triggers targeting four ribosomal proteins (RP) transcripts. Although no significant mortality was observed after dsRNA microinjection and subsequent blood feeding, significant contrasts were observed on fecundity. Triggers targeting RPS6 and RPL26 effectively reduced gene expression but more importantly, reduced reproductive output by more than 96% and 91% at the first oviposition while triggers targeting RPL1 and RPS2 did not cause a reduction although gene expression was reduced. Significantly reduced fecundity continued through a second oviposition cycle in dsRPS6 and dsRPL26 cohorts, although the effect was not as strong. Relative gene expression levels confirmed specific transcript knockdown up to 20days post-injection in mosquitoes that did not oviposit or produced reduced clutch sizes. Dissections at 36h post-blood meal indicated defects in oocyte provisioning. The strong phenotype produced by dsRPS6 allowed us to examine the effects in various tissues as well as the dose response, trigger format, delivery method and trigger specificity in Aedes aegypti. Strong knockdown was observed in the abdomen and the ovaries. Greater than 50ng of dsRPS6 significantly reduced fecundity but not when delivered in a sugar meal or as an siRNA. Similar bioassays with mutated dsRPS6 triggers indicates that up to three mismatches per possible siRNA are still effective in reducing fecundity. These studies indicate that while active and effective triggers can be developed for vector species, the lack of an efficient delivery method is the biggest barrier to use as a potential control method.
authors.affiliation: Navy Entomology Center of Excellence, Testing & Evaluation Department, CMAVE Detachment, Naval Air Station, Jacksonville, Jacksonville, FL 32211, United States; Center for Medical, Agricultural, and Veterinary Entomology, Agricultural Research Service, United States Department of Agriculture, 1700 SW 23rd Drive, Gainesville, FL 32608, United States. Electronic address: alden.estep@ars.usda.gov.
authors.forename: A S
authors.initials: AS
authors.lastname: Estep
authors.affiliation: Center for Medical, Agricultural, and Veterinary Entomology, Agricultural Research Service, United States Department of Agriculture, 1700 SW 23rd Drive, Gainesville, FL 32608, United States.
authors.forename: N D
authors.initials: ND
authors.lastname: Sanscrainte
authors.affiliation: Center for Medical, Agricultural, and Veterinary Entomology, Agricultural Research Service, United States Department of Agriculture, 1700 SW 23rd Drive, Gainesville, FL 32608, United States.
authors.forename: J J
authors.initials: JJ
authors.lastname: Becnel
mesh_terms: D000330:Aedes / Q000235:genetics / Q000502:physiology*; D000818:Animals; D005260:Female; D005298:Fertility; D055785:Gene Knockdown Techniques; D019476:Insect Proteins / Q000235:genetics* / Q000378:metabolism; D012330:RNA, Double-Stranded / Q000235:genetics* / Q000378:metabolism; D012269:Ribosomal Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D019476:Insect Proteins; D012330:RNA, Double-Stranded; D012269:Ribosomal Proteins
keywords: Aedes aegypti; Fecundity; Gene silencing; Oviposition; Ribosome; Trigger; dsRNA; siRNA