Detection and quantification of Anopheles gambiae sensu lato mosquito larvae in experimental aquatic habitats using environmental DNA (eDNA).

Abstract

Background: Growing insecticide resistance and changes in biting and resting behavior of malaria vectors threaten efficacy of insecticide treated nets and indoor residual spraying. Larval source management (LSM) is a promising approach that can target mosquitoes irrespective of their behavior as adults. However, the use of traditional monitoring methods for immature stages of Anopheles mosquitoes is a major challenge to LSM due to the variability in their breeding habitats. We evaluate the use of an environmental DNA (eDNA) analysis technique in monitoring Anopheles gambiae sensu lato larvae in experimental aquatic habitats. Methods: eDNA was simultaneously sampled and extracted from different volumes of water, number of larvae, and occupation time. Larval presence was detected using PCR and eDNA concentration in samples from 1 L habitats quantified using an IGS and cyt b TaqMan assays. The limit of detection of the two assays was tested and larval density correlated with eDNA positivity. Results: 74% of replicates in the 50 mL habitats were PCR positive with at least 6h required to get a signal from a single larva (0.02 larvae/mL). All 12 replicates where 1 L of water was used were positive with stronger PCR bands than replicates with the same larval density in 50 mL for 24 h. There was a correlation between larval densities and eDNA detection in both assays: IGS, r = 0.503, p = 0.047; and cyt b, r = 0.558, p = 0.025. There was stochasticity in eDNA detection rates, using both PCR and qPCR across all the dilutions. Conclusion: This study has demonstrated the potential use of eDNA analysis for detection and quantification of An. gambiae s.s. mosquito larvae in aquatic habitats. The stochasticity observed in eDNA detection suggest that this technique is best for monitoring aquatic habitats with many larvae at low densities.

Technical Workflow Development for Integrating Drone Surveys and Entomological Sampling to Characterise Aquatic Larval Habitats of Anopheles funestus in Agricultural Landscapes in Côte d’Ivoire

“…funestus in water bodies where their larvae were not collected or identified. e previous detection of eDNA from artificial Anopheles larval habitats has demonstrated the ability to detect 0.002 larvae/ml, and further validation of our protocol with field samples is ongoing [50]. e SOPs provided and instructions on these methods should serve as good guidelines for future researchers.…”

Investigation of mosquito larval habitats and insecticide resistance in an area with a high incidence of mosquito-borne diseases in Jining, Shandong Province

“…Therefore, aquatic habitats are important for mosquito breeding. Larval source management (LSM) involves the management of water bodies that are potential larval habitats to prevent the development of immature mosquitoes into adults [18,19], and studies have shown that mosquito LSM has an important potential impact on mosquito-borne disease transmission both in urban and rural areas [20][21][22]. For example, in urban areas of Dar es Salaam, Tanzania, the use of larvicides reduced the risk of malaria infection in 5-year-old children [23].…”

Quantitative PCR assay for the detection of Aedes vigilax in mosquito trap collections containing large numbers of morphologically similar species and phylogenetic analysis of specimens collected in Victoria, Australia

“…Further development and validation of this assay could allow the identification of the different life stages of Ae. vigilax and potentially enable detection of this mosquito species larvae in water samples, as has been outlined by previous studies [ 68 ]. Aedes vigilax collected in Victoria were seen to have a high sequence identity to those of clade III, with no separation seen between individuals from the two capture locations in Victoria, indicating mixing between these populations.…”