In September 2020, a preliminary survey was conducted in the study sites, and most of the larval habitats were found to be located approximately 2 km from the centre of each community (Fig. 2 ). These areas were selected for detailed study. Thorough searches of Anopheles larval habitats (e.g., man-made ponds, wells, swamps, furrows, puddles) were conducted and their locations mapped using a global positioning system (GPS: Garmin etrex® 10) unit. A total of 59 larval habitats within the study sites, identified to consistently have immature mosquitoes, were chosen: 18 in Dodowa, and 42 in Anyakpor (Fig. 2 ). These 59 positive habitats were sampled for mosquito larvae once every two weeks from October 2020 to May 2021.![]()
The larval habitats were grouped into temporary or permanent habitats. Temporary habitats were mainly rain-dependent and dried up when rain ceased for a while [43 (link)]. The permanent habitats was defined as habitats in which Anopheles larvae were found at least once, and contained water that was fed by natural underground sources throughout the sampling period [43 (link)].
Habitat stability was indicated by the availability of water in a habitat for 14 days, following previous reported studies that showed that egg-adult cycle of An. gambiae s.l. can be completed in this length of time [17 (link), 37 (link)]. To determine productivity, the habitats were visited and examined once every two weeks for the presence of aquatic stages of anopheline and culicine mosquitoes. In addition, the area (length and width) of the water surface was measured and recorded in metres with a metal ruler and grouped as small (≤ 10 m2) or large (10–100 m2).
Mosquito larval surveys were also carried out to generate stage-specific estimates of larval densities. Water was dipped up to 20 times using a standard dipper (350 mL, BioQuip Products, Inc., CA, USA). When a habitat was too small to make 20 dips, water was dipped as many times as possible. Larval abundance was calculated as the number of larvae per number of dips made in each habitat. The number of larvae and pupae in each habitat was collected and recorded, with larvae classified as early instars (L1 and L2) or late instars (L3 and L4). Larval samples collected from each habitat were pooled into sterile plastic containers and transported to the insectary of the Department of Medical Microbiology, University of Ghana Medical School, where they were bred into adults. At the insectary, the larvae were fed on Tetramin® fish meal and maintained at 27 ± 2 °C.
The study sites and locations of larval habitats
The larval habitats were grouped into temporary or permanent habitats. Temporary habitats were mainly rain-dependent and dried up when rain ceased for a while [43 (link)]. The permanent habitats was defined as habitats in which Anopheles larvae were found at least once, and contained water that was fed by natural underground sources throughout the sampling period [43 (link)].
Habitat stability was indicated by the availability of water in a habitat for 14 days, following previous reported studies that showed that egg-adult cycle of An. gambiae s.l. can be completed in this length of time [17 (link), 37 (link)]. To determine productivity, the habitats were visited and examined once every two weeks for the presence of aquatic stages of anopheline and culicine mosquitoes. In addition, the area (length and width) of the water surface was measured and recorded in metres with a metal ruler and grouped as small (≤ 10 m2) or large (10–100 m2).
Mosquito larval surveys were also carried out to generate stage-specific estimates of larval densities. Water was dipped up to 20 times using a standard dipper (350 mL, BioQuip Products, Inc., CA, USA). When a habitat was too small to make 20 dips, water was dipped as many times as possible. Larval abundance was calculated as the number of larvae per number of dips made in each habitat. The number of larvae and pupae in each habitat was collected and recorded, with larvae classified as early instars (L1 and L2) or late instars (L3 and L4). Larval samples collected from each habitat were pooled into sterile plastic containers and transported to the insectary of the Department of Medical Microbiology, University of Ghana Medical School, where they were bred into adults. At the insectary, the larvae were fed on Tetramin® fish meal and maintained at 27 ± 2 °C.
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