Genome-wide detection of Wolbachia in natural Aedes aegypti populations using ddRAD-Seq

Atikah Fitria Muharromah; Jerica Isabel L Reyes; Ngure Kagia; Kozo Watanabe

doi:10.3389/fcimb.2023.1252656

Genome-wide detection of Wolbachia in natural Aedes aegypti populations using ddRAD-Seq

Front Cell Infect Microbiol. 2023 Dec 14:13:1252656. doi: 10.3389/fcimb.2023.1252656. eCollection 2023.

Authors

Atikah Fitria Muharromah^{1

2}, Jerica Isabel L Reyes¹, Ngure Kagia¹, Kozo Watanabe¹

Affiliations

¹ Molecular Ecology and Health Laboratory, Center for Marine Environmental Studies (CMES), Ehime University, Matsuyama, Japan.
² Entomology Laboratory, Department of Tropical Biology, Faculty of Biology, Universitas Gadjah Mada, Yogyakarta, Indonesia.

Abstract

Background: Wolbachia, an endosymbiotic bacterium, is globally used to control arboviruses because of its ability to block arboviral replication and manipulate the reproduction of Wolbachia host, Aedes aegypti. Polymerase chain reaction (PCR)-based Wolbachia detection has been recently reported from natural Ae. aegypti populations. However, due to the technical limitations of PCR, such as primer incompatibility, PCR-based assays are not sufficiently reliable or accurate. In this study, we examined double digestion restriction site-associated DNA sequencing (ddRAD-Seq) efficiency and limitations in Wolbachia detection and quantification in field-collected Ae. aegypti natural populations in Metro Manila, the Philippines, compared with PCR-based assays.

Methods: A total of 217 individuals Ae. aegypti were collected from Metropolitan Manila, Philippines. We separated it into 14 populations consisting of 7 female and male populations. We constructed a library for pool ddRAD-Seq per population and also screened for Wolbachia by PCR assays using wsp and 16S rRNA. Wolbachia density per population were measured using RPS17 as the housekeeping gene.

Results: From 146,239,637 sequence reads obtained, 26,299 and 43,778 reads were mapped across the entire Wolbachia genome (with the wAlbA and wAlbB strains, respectively), suggesting that ddRAD-Seq complements PCR assays and supports more reliable Wolbachia detection from a genome-wide perspective. The number of reads mapped to the Wolbachia genome per population positively correlated with the number of Wolbachia-infected individuals per population based on PCR assays and the relative density of Wolbachia in the Ae. aegypti populations based on qPCR, suggesting ddRAD-Seq-based semi-quantification of Wolbachia by ddRAD-Seq. Male Ae. aegypti exhibited more reads mapped to the Wolbachia genome than females, suggesting higher Wolbachia prevalence rates in their case. We detected 150 single nucleotide polymorphism loci across the Wolbachia genome, allowing for more accurate the detection of four strains: wPip, wRi, TRS of Brugia malayi, and wMel.

Conclusions: Taken together, our results demonstrate the feasibility of ddRAD-Seq-based Wolbachia detection from field-collected Ae. aegypti mosquitoes.

Keywords: Aedes aegypti; Philippines; Wolbachia; ddRAD-seq; genome-wide.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Aedes* / microbiology
Animals
Female
Humans
Male
Philippines
RNA, Ribosomal, 16S / genetics
Ribosomal Proteins
Wolbachia* / genetics

Substances

RNA, Ribosomal, 16S
Ribosomal Proteins

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was funded by grants from JSPS Core-to-Core Program B. Asia-Africa science platforms (JPJSCCB20190008) and the Sumitomo Electric Industries Group Corporate Social Responsibility (CSR) Foundation, Endowed Chair program.