# Anopheles larval ecology and physicochemical characterization of larval habitats in Dire Dawa: an area colonized by Anopheles stephensi in Eastern Ethiopia

## TL;DR

This study examines the larval habitats and species of Anopheles mosquitoes in Dire Dawa, Ethiopia, revealing that Anopheles stephensi is the most common species and that habitat type and urbanization influence larval density.

## Contribution

The study provides new insights into the distribution and ecology of Anopheles stephensi in urban, peri-urban, and rural settings in Ethiopia.

## Key findings

- Anopheles stephensi was the most prevalent species, found in both artificial and natural habitats.
- Uncovered water tankers (bricks) were the most productive artificial habitats in urban and peri-urban areas.
- Larval density was positively correlated with urbanization, brick availability, and proximity to houses.

## Abstract

Understanding mosquito larval ecology is essential for planning and implementations of vector control strategies. The biotic and abiotic factors affect larval occurrence , density, survival and morphogenesis of mosquitoes. Artificial containers are very suitable larval habitats for some species of Anopheles and Aedes mosquitoes in urban and peri-urban settings. Therefore, this study we identified, mapped and characterized larval habitats, estimated larval density and indices larval habitats. In addition, we determined the species composition of Anopheles mosquitoes and species evenness in urban, peri-urban and rural areas of Dire Dawa city adminstration

Larval habitats were surveyed and identified monthly for a period of 16 months from February 2023 to December 2024 in urban, peri-urban and rural areas of Dire Dawa. Mosquito larvae and pupae were collected and those larvae identified as Anopheles were fed on fish-food. Emerged adults were provided with 10% sucrose solution and kept under standard conditions in field insectary. Females Anopheles was identified morphologically and further species-specific PCR assay was employed to identify members of An.gambiae s.l. In addition, real time PCRassay was performed to identify An.stephensi and An.arabiensis. Water samples were taken from the larval habitats and the physico-chemical parameters were measured using HANNA Multi-parameter (H198194). Larval habitat diversity, larval abundance and distribution were assessed across the three ecological settings (urban, peri-urban and rural).

A total of 23, 526 larvae and 1,808 pupae of Anopheles mosquitoes were collected from 909 man-made(uncovered cemented cisterns (Brick), plastic sheets, steel drums, Tire tracks, Canal ditch, plastic tanker/Barrel) and natural habitats (River edges, ponds, animal hoof prints and swaps) in urban, peri-urban areas of Dire Dawa. The highest mean larval density (51 larvae per dip) of Anopheles mosquitoes was recorded from peri-urban sites in uncovered water tanker (brick) followed by urban site in brick (46 larvae per dip). Anopheles larvae were not found in steel drum and plastic barrels in rural sites.

A total of 2,934 adult Anopheles mosquitoes were emerged from immatures collected from all sites, of which 75% (2194/2934) were An. stephensi, and 22 % (636 / 2934) were An.arabiensis. The remaining 3.0% were An. pharoensis, An. coustani, An .amharicus, and An. pretoriensis. Anopheles stephensi, An. arabiensis and An. amharicus shared the same habitats across the three ecologies. Larval density was positively correlated with availability of brick making, proximity to houses, urban setting, presence of competitors /predators, vegetation cover, shade cover and substrate type. But larval presence was not correlated with presence/absence of intervention. Larva/pupa presence were positively correlated with pH (r=0.264, p=0.01) and water pressure (r=0.21, p<0.05).There was positive correlation among temperature, electrical conductivity (EC), total dissolved solids (TDS), salinity and dissolved oxygen (DO) and negative correlation among temperature with resistivity, pH with mvPH. Larval presence was positively correlated with water salinity and pH.

Anopheles stephensi was the predominant species found in the study area, followed by An.arabiensis, An.amharicu, An.pharoensis, An.coustani and An.pretoriensis. Uncovered water tankers (Bricks) were the most prolific artificial habitats in urban and peri-urban sites followed by plastic sheets while natural habitats such as hoof prints and river basins were the most efficient habitat types in rural and urban settings, respectively. Anopheles stephensi was found in natural habitats of Butuji and legehare rivers from urban site, and in rural sites from man-made habitat of plastic sheet.

Anopheles amharicus larvae was found in plastic sheet, an artificial habitat common in urban, peri-urban and rural areas.

We report here the occurrence of An.stephensi in rural areas, breeding in natural habitats, and co-existing with An. gambiae s.l complex.

Availability of brick making, shorter distance from living houses being in urban ecology were directly correlated with larval density. Habitat abundance and positivity of uncovered water tankers (cemented cisterns or bricks) in urban and pre-urban sites could indicate for feasibility and proper implementation of larval source management.