Reductions in malaria burden worldwide coincides with the massive scale-up of malaria treatment and prevention measures, of which vector control is the major component, particularly in SSA. The key vector control interventions in Africa including Uganda, rely heavily on utilisation of long-lasting insecticidal nets (LLINs) and insecticide-based indoor residual spraying (IRS). This study assessed the bio-efficacy of insecticides used for vector control in LLINs and IRS in Busia and Tororo, and assess effectiveness of the different control frameworks in the two districts. Samples of Anopheles mosquito larvae were collected from various breeding grounds before rearing them in an insectary. Additionally, a nine-year period malaria cases data set (2012-2020) was obtained from the health management database to depict malaria trends. Emerged adults were observed under a dissecting microscope and identified using standardized morphological keys after respective bioassays on used nets and sprayed walls. Independent two sample T-test was used to test for significant differences in the mean malaria vectors diversity, mean variations, mortalities by net and wall type. Results showed that mean mortalities of Anopheles mosquitoes to used brands of nets from Busia (9.86±11.35) and Tororo (9.64 ±11.12) varied insignificantly (t=0.119, p=0.906). A disaggregated analysis for each net used revealed that, the DAWA plus 2.0 registered a highest mean mortality of mosquitos, followed by PermaNet 3.0+PBO, Olyset and PermaNet 2.0 respectively. Fludora fusion revealed highest mortalities on plastered painted wall, followed by Brick plain then mud/wattle walls at all times. The trend is similar for Actellic, but unlike Fludora fusion, Actellic exhibited mortalities lower than the 80% threshold for all the wall types and T-test (T-test P values<0.001) indicated Actellic efficacy to be significantly lower than that of Fludora fusion on all the three wall substrates. Considering the malaria cases registered on annual basis, Mann- Kendal test revealed a drastic decline of malaria cases over the nine year period (2012 -2020) in Tororo district with Sen’s slope of -22, while for Busia district it revealed an increase of malaria cases (Sen’s slope +28). Equally, these trends reveal varied spatial patterns over the two districts. Both the districts revealed a high prevalence of malaria between May-November in respect to bimodal rainfall pattern, matching to the perennial transmission setting of Uganda. This study has shown that vector behaviour, biology and physiology need consistent monitoring and surveillance for further entomological characterisation. Keywords: Bimodal, Bio-efficacy, Sentinel areas, Oscillation, Plasmodium vectors

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