RNA Interference Suppression of v-ATPase B and Juvenile Hormone Binding Protein Genes Through Topically Applied dsRNA on Tomato Leaves: Developing Biopesticides to Control the South American Pinworm, Tuta absoluta (Lepidoptera: Gelechiidae).

Govindaraju Ramkumar,Kandaswamy Kalaivani,Aml Ghaith,Ramasamy Asokan,Wayne Brian Hunter,Sengottayan Senthil-Nathan,Mohamed Soliman Elshikh,Sengodan Karthi,Patcharin Krutmuang,B Kariyanna,Ahmed Abdel-Megeed,N R Prasannakumar,Mona S Alwahibi

doi:10.3389/fphys.2021.742871

Abstract

The South American pinworm Tuta absoluta (Meyrick) (Family: Gelechiidae) is one of the most devastating lepidopteran pests in the developing countries of South America, Africa, and Asia. This pest is classified as the most serious threat for tomato production worldwide. In the present study, we analyzed RNAi-mediated control through exogenously applied dsRNA delivery on tomato. The dsRNA treatments were made to target the juvenile hormone binding protein and the v-ATPase B. Both mRNA targets were cloned, validated by sequencing, and used to produce each dsRNA. After treatments the relative transcript expression was analyzed using qRTPCR to assess to efficacy of RNAi. A leaf-dip assay was used to provide late 2nd instar larvae three feeding access periods: 24, 48, and 72 h, to evaluate the effect of gene silencing of each target. Larvae were fed tomato leaves coated with five different RNAi concentrations (10, 20, 30, 40, and 50 micrograms/centimeter-squared), that suppressed two genes (juvenile hormone protein, JHBP, and vacuolar-type adenosine triphosphatase enzyme, v-ATPase). Treatments with dsRNA showed a significant increase in mortality at 24, 48, and 72 h after ingestion (P < 0.01, α = 0.05), along with reduced leaf damage, and increased feeding deterrence. The results suggest that these two RNAi products may provide a suitable treatment for control of this and other lepidopteran pests.

Highlights

Tuta absoluta Meyrick (Lepidoptera: Gelechiidae) is an oligophagous pest infesting many Solanaceous crops (Global Distribution Map: CABI, Campos et al, 2017; Biondi et al, 2018; Rwomushana et al, 2019)
Significant larval mortality was observed in larvae feeding at the increasing concentrations of each double-stranded RNA (dsRNA) (Figure 1, JH binding protein (JHBP); Figure 2, v-ATPase B and LC50 values of 3.426 and 4.121 μg/μl; 5.126)
Results showed that the expression of both v-ATPase B and JHBP transcripts decreased progressively with increasing concentrations of dsRNA represented in Relative Fold Change per treatment (Figures 4, 5)

Summary

Introduction

Tuta absoluta Meyrick (Lepidoptera: Gelechiidae) is an oligophagous pest infesting many Solanaceous crops (Global Distribution Map: CABI, Campos et al, 2017; Biondi et al, 2018; Rwomushana et al, 2019). Synthetic pesticides are commonly used for pest control worldwide (Guedes et al, 2019) Application of these chemicals against T. absoluta has been reported with little success, mainly because the pest has developed resistance (Siqueira et al, 2000; Senthil-Nathan, 2020). Research shows that RNAi biopesticides can be exogenously applied in liquid sprays (Dalakouras et al, 2016; Koch et al, 2016; McLoughlin et al, 2018; Dubrovina and Kiselev, 2019; Dubrovina et al, 2019, 2020; Jalaluddin et al, 2019; Mezzetti et al, 2020) or bound with a carrier-like clay or nanotubes in sprays (Worrall et al, 2019; Fletcher et al, 2020) or applied in water or clay pellets as a soil treatment (Ghosh et al, 2018) or as plant-expressed silencing, which has met with significant success in field crops (Younis et al, 2014; Bramlett et al, 2020; Das and Sherif, 2020; Veillet et al, 2020)

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Discussion

Conclusion