Elsevier

Aquatic Toxicology

Volume 248, July 2022, 106181
Aquatic Toxicology

Exposure of Anopheles gambiae larvae to a sub-lethal dose of an agrochemical mixture induces tolerance to adulticides used in vector control management

https://doi.org/10.1016/j.aquatox.2022.106181Get rights and content

Highlights

  • Exposure to agrochemicals can affect the tolerance of mosquitoes larvae to insecticides.

  • The potential carry-over of such effect to the adult stage has been far less documented.

  • Exposure of An. gambiae larvae to agrochemicals increase tolerance to Fludora® fusion in adults.

  • Fludora® fusion efficacy for vector control may be locally affected by the ecological context.

Abstract

The heavy use of pesticides in agricultural areas often leads to the contamination of nearby mosquito larvae breeding sites. Exposure to complex mixtures of agrochemicals can affect the insecticide sensitivity of mosquito larvae. Our study objective was to determine whether agrochemical residues in Anopheline larval breeding sites can affect the tolerance of adults to commonly used adulticides. We focussed on Fludora® Fusion, a vector control insecticide formulation combining two insecticides (deltamethrin and clothianidin) with different modes of action. An. gambiae larvae were exposed to a sub-lethal dose of a mixture of agrochemical pesticides used in a highly active agricultural area on the Ivory Coast. Comparative bioassays with Fludora Fusion mixture and its two insecticide components (deltamethrin and clothianidin) were carried out between adult mosquitoes exposed or not to the agrochemicals at the larval stage. A transcriptomic analysis using RNA sequencing was then performed on larvae and adults to study the molecular mechanisms underlying the phenotypic changes observed. Bioassays revealed a significantly increased tolerance of adult females to clothianidin (2.5-fold) and Fludora Fusion mixture (2.2-fold) following larval exposure to agrochemicals. Significantly increased tolerance to deltamethrin was not observed suggesting that insecticide exposure affects the adult efficacy of the Fludora Fusion mixture mainly through mechanisms acting on clothianidin. Transcriptomic analysis revealed the potential of agrochemicals to induce various resistance mechanisms including cuticle proteins, detoxification action and altered insecticide sequestration. These results suggest that although the Fludora Fusion mixture is effective for adult vector control, its efficacy may be locally affected by the ecological context. The present study also suggests that, although the complex interactions between the use of agrochemicals and vector control insecticides are difficult to decipher in the field, they still must be considered in the context of insecticide resistance management programmes.

Introduction

In 2020, malaria caused 627,000 deaths. These mainly occurred in Africa (96%) and more than 80% were children less than 5 years old (WHO, 2021). Despite significant advances in malaria vaccines and treatments, control of Anopheles mosquitoes in Africa remains an important malaria control strategy that decreases malaria prevalence and mortality in endemic regions (Bhatt et al., 2015; Coleman et al., 2017; Lim et al., 2011; Tizifa et al., 2018; WHO, 2021). Vector control relies on the use of insecticide treated nets (ITN) or indoor residual insecticide sprays (IRS). Pyrethroid insecticides are most commonly used because of their strong effects on the target mosquitoes and their low toxicity to the environment and mammals (Zaim et al., 2000).

However, after decades of use, pyrethroid efficacy has been reduced by resistance mechanisms selected in Anopheles populations (Chouaibou et al., 2017; Fane et al., 2012; Nwane et al., 2013; Reid and McKenzie, 2016). Pyrethroid resistance is mainly associated with target site mutations affecting the voltage gated sodium channel (VGSC, Knock Down Resistance ‘kdr’ mutations) and increased insecticide metabolism mediated by detoxification enzymes (metabolic resistance). Kdr mutations are widely distributed in African Anopheles populations and encompass kdr West (L1014F) and kdr East (L1014S) mutations (Martinez-Torres et al., 1998; Ranson et al., 2000). Metabolic resistance occurs through increased activities of detoxification enzymes, resulting in increased insecticide metabolism (Li et al., 2007). The main detoxification enzyme families involved include cytochrome P450 monooxygenases (P450, CYP for genes), carboxyl/cholinesterases (CCE), glutathione-S-transferase (GST), UDP-glycosyl-transferases (UDPGT) and sulfotransferases (SULT) (David et al., 2013; Liu, 2015; Moyes et al., 2017; Riveron et al., 2018). In addition to target-site modifications and metabolic resistance, additional mechanisms involving cuticle modifications, altered insecticide transport and sequestration, sensory appendage protein (SAP) and chemosensory proteins (CSP) have been reported or suggested (Ingham et al., 2020, 2018; Liu, 2015; Yahouedo et al., 2017).

Introduction of new insecticides with different modes of action has been suggested as a component of insecticide resistance management strategies (Darriet and Chandre, 2013; WHO, 2012). Among the novel insecticides proposed for managing pyrethroid resistance, neonicotinoids have been presented as a good alternative because they target nicotinic acetylcholine receptors (nAChR) which represent a new biochemical target in public health insects (Agossa et al., 2018; Ngufor et al., 2017). A new insecticide formulation combining the neonicotinoid clothianidin and the pyrethroid deltamethrin (8:1 w/w) under the brand name Fludora® Fusion was developed by Bayer for indoor residual spraying (IRS) as a tool for insecticide resistance management (IRM). Field trials using Fludora® Fusion demonstrated its high efficacy against various Malaria vectors, including pyrethroid-resistant populations (Fongnikin et al., 2020; Kamaraju et al., 2021; Ngufor et al., 2017). Fludora® Fusion may also help slow the selection of pyrethroid and neonicotinoid resistance alleles (Zoh et al., 2021). However, environmental factors that may affect the sensitivity of mosquitoes to Fludora® Fusion remain to be investigated.

Agriculture yields in Africa are associated with the use of agrochemicals. Heavy use of agrochemicals, including herbicides and insecticides, in intensive agricultural areas often leads to contamination of nearby mosquito larvae breeding sites (Chouaibou et al., 2016). Exposure to complex agrochemical mixtures can lead to reduced sensitivity to insecticides due to ‘gene x environment’ interactions (Liu, 2015; Riveron et al., 2018). These interactions can affect the transient expression of a range of proteins though the induced expression of detoxification enzymes. Cuticle proteins and transporters may also play roles in the increased resistance phenotypes observed. For example, Riaz et al. (2009) showed that Aedes aegypti larvae exposed to a sublethal dose of the herbicide glyphosate or the PAH benzo[a]pyrene showed increased tolerance to the insecticides imidacloprid and permethrin associated with increased transcription of several P450 genes. Similarly, exposure of Ae. aegypti larvae to a sublethal dose of the fungicide copper sulfate led to increased tolerance to permethrin and was associated with over-transcription of several P450 genes (Poupardin et al., 2008, 2010). Transcriptomic data confirmed the association between this increased insecticide tolerance and the overexpression of genes encoding detoxification enzymes, cuticle proteins and xenobiotic transporters (David et al., 2010; Poupardin et al., 2010). Exposure of Anopheles gambiae larvae to phytochemical leaf extracts of the curry tree revealed significant induction of transcripts associated with cuticular proteins and detoxification enzymes (Mang'era et al., 2021).

The objective of this study was to determine whether agrochemicals found in Anopheline larval breeding sites can affect the tolerance of adult mosquitoes to commonly used insecticides such as Fludora® Fusion. In this study, An. gambiae larvae were exposed to a sub-lethal dose of a mixture of agrochemicals representative of those used in an agriculture region of the Ivory Coast. Comparative bioassays were conducted between adult mosquitoes exposed, or nonexposed, to the pesticide mixture at the larval stage. A transcriptomic analysis using RNA sequencing (RNA-seq) was conducted to study the molecular mechanisms underlying the observed insecticide sensitivity changes.

Section snippets

Mosquitoes

An. gambiae sensu stricto larvae were collected near Tiassalé (south of Ivory Coast). This is a rice field area with classic vector control activities (mainly mosquito nets impregnated with pyrethroids) and heavy use of pesticides for crop protection, including herbicides, fungicides and insecticides from the pyrethroid, organophosphate, carbamate and neonicotinoid families (Chouaibou et al., 2016). Malaria vectors in this area are resistant to multiple insecticides, including pyrethroids, DDT,

Bioassays

Exposure of Tiassalé-S larvae to a sublethal dose of the agrochemical mixture differentially affected the tolerance of adult females to the Fludora Fusion mixture and its two insecticide components (Fig. 1). Comparative bioassays on adult females showed a 2.5-fold and a 2.2-fold increased tolerance to clothianidin and the Fludora Fusion mixture respectively compared to the nonexposed control (Fisher p-value <0.001). Though a slight increased tolerance to deltamethrin was observed in exposed

Discussion

Mosquito larval development often occurs in areas exposed to natural xenobiotics and anthropogenic pollutants. Xenobiotics, including residual agrochemicals, can affect subsequent larval tolerance to chemical insecticides used for mosquito control (Poupardin et al., 2010; Riaz et al., 2009; Suwanchaichinda and Brattsten, 2001). The xenobiotics most affective on larval insecticide sensitivity often act as detoxification enzyme inducers, supporting the key role of the detoxification system in

Conclusion

The WHO has encouraged development of novel vector control products that can be used for managing pyrethroid resistance (Turner et al., 2016; WHO, 2012). Among them, pyrethroid-neonicotinoid combinations such as the IRS formulation Fludora® Fusion show good potential for use against African malaria vectors (Fongnikin et al., 2020; Fuseini et al., 2019; Oxborough et al., 2019). Such combinations also show a good potential for delaying the selection of insecticide resistance alleles (Zoh et al.,

Data availability statement

The RNA-seq sequence data from this study have been deposited at NCBI Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/geo/info/ submission.html) under the accession number GSE189345.

CRediT authorship contribution statement

Marius Gonse Zoh: Methodology, Investigation, Data curation. Jordan Tutagata: . Behi K. Fodjo: Methodology, Investigation. Chouaïbou S. Mouhamadou: Methodology. Christabelle G. Sadia: Methodology, Investigation. Justin McBeath: Conceptualization. Frederic Schmitt: Conceptualization. Sebastian Horstmann: Conceptualization. Jean-Philippe David: Conceptualization, Methodology, Investigation, Data curation, Writing – original draft, Writing – review & editing, Supervision. Stéphane Reynaud:

Declaration of Competing Interest

The funding organization (Bayer) developed and commercialized Fludora® Fusion used in this study and proposed the original test hypothesis. The funder had no role in the study design, data collection, analysis and interpretation of results. JMB, FS and SH were employed by Bayer. MGZ PhD was funded by Bayer and hosted at the Laboratoire d'Ecologie Alpine (LECA). The position of JT was funded by Bayer and hosted at LECA. All other authors declare no conflict of interest.

Acknowledgements

Editorial services, including language editing and correction, were provided by XpertScientific Editing and Consulting Services.

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