The Phylodynamic and Spread of the Invasive Asian Malaria Vectors, Anopheles stephensi, in Sudan

Abstract

Anopheles stephensi is an invasive Asian malaria vector that initially emerged in Africa in 2012 and was reported in Sudan in 2019. We investigated the distribution and population structure of An. stephensi throughout Sudan by using sequencing and molecular tools. We confirmed the presence of An. stephensi in eight border-states, identifying both natural and human-made breeding sites. Our analysis revealed the presence of 20 haplotypes with different distributions per state. This study revealed a countrywide spread of An. stephensi in Sudan, with confirmed presence in borders states with Chad, Egypt, Eritrea, Ethiopia, Libya, Republic of Central Africa, and South Sudan. Detection of An. stephensi at points of entry with these countries, particularly Chad, Libya, and South Sudan, indicates the rapid previously undetected spread of this invasive vector. Our phylogenetic and haplotype analysis suggested local establishment and evolutionary adaptation of the vector to different ecological and environmental conditions in Sudan. Urgent engagement of the global community is essential to control and prevent further spread into Africa.

Keywords: Africa; Anopheles stephensi; International Health Regulations; climate change; haplotypes analysis; invasive disease vector; malaria epidemics; phylogenetic analysis; vector control and surveillance.

Figure 1

Sudanese States (highlighted in light blue) where active surveys were implemented to detect An. stephensi mosquitoes. The red circles indicate the number of An. stephensi mosquitoes collected.

Figure 2

Representative breeding sites that were positive for presence of aquatic stages of An. stephensi (larvae and pupae). (A) Plastic cistern, (B) plastic barrel, (C) mud pot, (D) clay pot (Zeir/Jar), (E) ground water-basin, (F) plastic jerrycan, (G) iron barrel, (H,I) leakage of broken water supply, and (J) rainwater pond trapped in a rocky valley.

Figure 3

Sequence alignment of the 20 Sudanese An. stephensi haplotypes. Substitutions were indicted with their nucleotide codes; no deletion nor insertions were present. The dots (.) indicate identical nucleotides at the specified position in comparison with the reference sequence KT899888.1 (ref). * Nucleotide substitution positions, based on the start of the complete mitochondrial cytochrome c oxidase 1 (CO1) gene, are read vertically.

Figure 4

Phylogenetic tree showing the relationship between the Sudanese An. stephensi haplotypes with 15 reference sequences. Sudanese haplotypes (Sudan Hap01–Sudan Hap20) are in bold. The reference sequences along with their accession numbers and origin of isolate were included for each. Drosophila melanogaster was used as an outgroup taxon.

Figure 5

Median-joining haplotype network of the 19 Sudanese An. stephensi haplotypes and worldwide An. stephensi sequences. Haplotypes of each region are presented in color code. Black dashes between the haplotype lines represent the number of substitutions.

Figure 6

Pie charts inserted to show the haplotypic composition of An. stephensi population across the states of Sudan. Haplotypes of each region are presented in color code.