Dataset Information

First Report on CRISPR/Cas9-Based Genome Editing in the Destructive Invasive Pest Tuta Absoluta (Meyrick) (Lepidoptera: Gelechiidae).

ABSTRACT: The tomato leaf miner Tuta absoluta (Meyrick) is one of the world's most destructive pests of tomato, and because of its severe economic impacts, as well as the development of pesticide resistance, the species has been intensively studied, especially in regard to the identification of targets for T. absoluta control. However, functional genomic studies of T. absoluta have been constrained by a lack of effective genetic tools. Therefore, the aim of the present study was to develop a CRISPR/Cas9 zygote microinjection protocol for generating heritable mutations in T. absoluta, using the ommochrome synthesis gene cinnabar as an easily evaluated target gene. The injection of fertilised eggs with Cas9 protein and four sgRNAs, which targeted cinnabar exon 3, resulted in a mutagenesis rate of 31.9% for eggs reaching adulthood, and cinnabar mutagenesis resulted in either red or mosaic eye colour phenotypes. As such, this study is the first to report a complete and detailed CRISPR/Cas9 workflow for the efficient genome editing of the globally important invasive pest T. absoluta. The application of this robust genome-editing tool to T. absoluta will greatly facilitate the discovery of suitable RNAi control targets and the subsequent development of novel control strategies.

SUBMITTER: Ji SX

PROVIDER: S-EPMC9160428 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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First Report on CRISPR/Cas9-Based Genome Editing in the Destructive Invasive Pest Tuta Absoluta (Meyrick) (Lepidoptera: Gelechiidae).

Ji Shun-Xia SX Bi Si-Yan SY Wang Xiao-Di XD Wu Qiang Q Tang Yan-Hong YH Zhang Gui-Fen GF Wan Fang-Hao FH Lü Zhi-Chuang ZC Liu Wan-Xue WX

Frontiers in genetics 20220519

The tomato leaf miner Tuta absoluta (Meyrick) is one of the world's most destructive pests of tomato, and because of its severe economic impacts, as well as the development of pesticide resistance, the species has been intensively studied, especially in regard to the identification of targets for T. absoluta control. However, functional genomic studies of T. absoluta have been constrained by a lack of effective genetic tools. Therefore, the aim of the present study was to de ...[more]

PMID: 35664294

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Project description:Tuta absoluta (the tomato pinworm) is an invasive insect pest with a highly damaging effect on tomatoes causing between 80 and 100% yield losses if left uncontrolled. Resistance to chemical pesticides have been reported in some T. absoluta populations. Insect microbiome plays an important role in the behavior, physiology, and survivability of their host. In a bid to explore and develop an alternative control method, the associated microbiome of this insect was studied. In this study, we unraveled the bacterial biota of T. absoluta larvae and adults by sequencing and analyzing the 16S rRNA V3-V4 gene regions using Illumina NovaSeq PE250. Out of 2,092,015 amplicon sequence variants (ASVs) recovered from 30 samples (15 larvae and 15 adults), 1,268,810 and 823,205 ASVs were obtained from the larvae and adults, respectively. A total of 433 bacterial genera were shared between the adults and larval samples while 264 and 139 genera were unique to the larvae and adults, respectively. Amplicon metagenomic analyses of the sequences showed the dominance of the phylum Proteobacteria in the adult samples while Firmicutes and Proteobacteria dominated in the larval samples. Linear discriminant analysis effect size (LEfSe) comparison revealed the genera Pseudomonas, Delftia and Ralstonia to be differentially enriched in the adult samples while Enterococcus, Enterobacter, Lactococcus, Klebsiella and Wiessella were differentially abundant in the larvae. The diversity indices showed that the bacterial communities were not different between the insect samples collected from different geographical regions. However, the bacterial communities significantly differed based on the sample type between larvae and adults. A co-occurrence network of significantly correlated taxa revealed a strong interaction between the microbial communities. The functional analysis of the microbiome using FAPROTAX showed that denitrification, arsenite oxidation, methylotrophy and methanotrophy as the active functional groups of the adult and larvae microbiomes. Our results have revealed the core taxonomic, functional, and interacting microbiota of T. absoluta and these indicate that the larvae and adults harbor a similar but transitory set of bacteria. The results provide a novel insight and a basis for exploring microbiome-based biocontrol strategy for this invasive insect pest as well as the ecological significance of some of the identified microbiota is discussed.

Project description:BackgroundTuta absoluta Meyrick 1917 (Lepidoptera: Gelechiidae) is an invasive, pesticide resistant, and a major treat of tomato production in the world. It needs effective management options that naturally infect the insect without causing any identified side effects. Entomopathogenic fungi (EPF) are the most important options. However, geographic origin and climatic condition apparently creates genetic variation among EPF strains that influence on their pathogenicity. Thus, screening of effective EPF strains from the local source is vital to develop environmental friendly pest control tactic for T. absoluta.ResultsIn this study, 27 indigenous Beauveria were isolated from the various types of soil and 12 of the isolates were screened based on their biological efficiency index (BEI). These isolates scored 65.7-95.7% and 68.3-95% of mortality against second and third instar larvae of T. absoluta at concentration of 1 × 107spores·ml-1 in 7 days post inoculation, respectively. Out of these, five (18.5%) isolates scored above 90% mortality on both instar larvae with LT50 value of 3.33 to 5.33 days at the lowest (104 spores·ml-1) and 1.93 to 3.17 days at highest (108 spores·ml-1) spore concentrations and has LC50 value of 1.5 × 103 to 1.1× 105 spores·ml-1. Moreover, isolates exhibited the promising mortality better (1.5 × 106 to 3.5 × 107 spores·ml-1), sporulated over the larval cadavers, well grown at optimal temperature, and produced chitinolytic enzymes. Molecular analysis showed that isolates have nearly monophyletic characters and grouped under species of Beauveria bassiana.ConclusionDifferent types of soil in Ethiopia are an important source of B. bassiana, and these isolates showed promising pathogenicity against T. absoluta, which is crucial for ecofriendly biopesticide development. Although isolates were nearly monophyletic in phylogenetic study, five of them were highly effective in the laboratory bioassays against T. absoluta; however, further field evaluation is required for mass production.

Project description:Tomato (Solanum esculentum) is one of the vegetable crops grown by both smallholder and commercial farmers in the Kingdom of Eswatini. Tomato leaf miner, Tuta absoluta M. is a major insect pest of tomatoes resulting in reduced tomato yields throughout the country. The study investigated the virulence of two sub-tropical EPN species on T. absoluta larvae. Steinernema yirgalemense and S. jeffreyense at different concentrations (0, 20, 40, 60 IJs/insect) were screened for efficacy (i.e. mortality rate) against larvae of T. absoluta in laboratory bioassays. Results obtained showed that S. yirgalemense and S. jeffreyense were able to kill T. absoluta larvae without significant differences between the two EPN species in 24-well bioassay trays. Significantly higher (p < 0.05) mortality was observed when 60 IJs/insect was used. The combination of S. yirgalemense at 60 IJs/insect (100%) and S. jeffreyense at 60 IJs/insect (98.3%) resulted in significantly higher (p < 0.05) mortality, compared with the other four combinations of EPN and concentration tested. In the leaf bioassays, S. yirgalemense (58.8%) resulted in significantly higher (p < 0.05) mean mortality compared to S. jeffreyense (46.4%). Steinernema yirgalemense at 60 IJs/insect resulted in significantly higher mean mortality compared to the other EPN and concentration combinations in the leaf bioassay. The results indicated that both EPNs tested were effective against T. absoluta larvae. Steinernema yirgalemense at 60 IJs/insect can effectively find T. absoluta larvae inside leaf mines, but large-scale field trials are recommended to demonstrate the potential use of the biocontrol agent within integrated pest management programs.Tomato (Solanum esculentum) is one of the vegetable crops grown by both smallholder and commercial farmers in the Kingdom of Eswatini. Tomato leaf miner, Tuta absoluta M. is a major insect pest of tomatoes resulting in reduced tomato yields throughout the country. The study investigated the virulence of two sub-tropical EPN species on T. absoluta larvae. Steinernema yirgalemense and S. jeffreyense at different concentrations (0, 20, 40, 60 IJs/insect) were screened for efficacy (i.e. mortality rate) against larvae of T. absoluta in laboratory bioassays. Results obtained showed that S. yirgalemense and S. jeffreyense were able to kill T. absoluta larvae without significant differences between the two EPN species in 24-well bioassay trays. Significantly higher (p < 0.05) mortality was observed when 60 IJs/insect was used. The combination of S. yirgalemense at 60 IJs/insect (100%) and S. jeffreyense at 60 IJs/insect (98.3%) resulted in significantly higher (p < 0.05) mortality, compared with the other four combinations of EPN and concentration tested. In the leaf bioassays, S. yirgalemense (58.8%) resulted in significantly higher (p < 0.05) mean mortality compared to S. jeffreyense (46.4%). Steinernema yirgalemense at 60 IJs/insect resulted in significantly higher mean mortality compared to the other EPN and concentration combinations in the leaf bioassay. The results indicated that both EPNs tested were effective against T. absoluta larvae. Steinernema yirgalemense at 60 IJs/insect can effectively find T. absoluta larvae inside leaf mines, but large-scale field trials are recommended to demonstrate the potential use of the biocontrol agent within integrated pest management programs.

Project description:(1) Background: The tomato leafminer Tuta absoluta (Lepidoptera: Gelechiidae) is a key tomato insect pest. At present, it is considered to be a serious threat in various countries in Europe, North Africa, and Middle East. The extensive use and the developed resistance of T. absoluta to spinosad causes some concern, which leads to the need for alternative products. (2) Materials and Methods: Several laboratory experiments were conducted to investigate the ovicidal properties of a zeolite particle film on T. absoluta. The toxicity of three different zeolites and six zeolite formulations to T. absoluta eggs and larvae was determined using different exposure methods. (3) Results: In general, the formulated zeolites yielded higher egg and larvae mortality values, especially when the zeolite particle film was residually applied. Notable differences in mortality rates from exposure to zeolites compared to other products, such as kaolin, its formulated product Surround, and the insecticide spinosad, were observed. Kaolin and Surround exhibited little or no effect for both application methods, while the hatch rate was reduced by 95% when spinosad was applied topically. Spinosad yielded egg and larvae mortality rates of 100% for both application methods. Additionally, increased oviposition activity was observed in adults exposed to the wettable powder (WP) formulations. These WP formulations increased egg deposition, while Surround and spinosad elicited a negative oviposition response. (4) Conclusions: It can be derived that the tested products, zeolites BEA (Beta polymorph A), FAU (Faujasite), LTA (Linde type A), and their formulations, had no real insecticidal activity against the eggs of T. absoluta. Nevertheless, egg exposure to zeolites seemed to affect the development process by weakening the first instar larvae and increasing their mortality. Subsequently, based on the choice test, no significant difference was observed between the number of eggs laid on the treated leaves and control leaves.

Dataset Information

First Report on CRISPR/Cas9-Based Genome Editing in the Destructive Invasive Pest Tuta Absoluta (Meyrick) (Lepidoptera: Gelechiidae).

Publications

First Report on CRISPR/Cas9-Based Genome Editing in the Destructive Invasive Pest <i>Tuta Absoluta</i> (Meyrick) (Lepidoptera: Gelechiidae).

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