Identifying gaps in protection from malaria vector biting in rural Cambodia using an entomological assessment and human behaviour observations

Background: Forest-exposed populations remain the last significant, and most difficult to access, high-risk populations for malaria in Cambodia. Despite the availability of long-lasting insecticidal nets (LLINs) and/or hammock nets (LLIHNs), continued malaria transmission indicates gaps in protection. This study aimed to identify these gaps among forest-exposed individuals in Plasmodium falciparum hotspots in two provinces in Cambodia, using entomological assessments and human behaviour observations (HBOs). Methods: Anopheles bionomic traits were characterized using Human Landing Catches (HLCs) in a village setting in Mondulkiri province, and in both village and forest settings in Kampong Speu province, Cambodia. Mosquitoes were collected from 17h00 to 07h00 over 540 collection nights. Human behaviour observations (HBOs) focused on monitoring activities near HLC sites and recording the use of LLINs/LLIHNs or Project BITE’s bite prevention tools: a volatile pyrethroid spatial repellent (VPSR), topical repellent (TR), and insecticide-treated clothing (ITC). Data on mosquito landing pressure and human behaviours were integrated to generate the HBO-adjusted Human Landing Rate (HBO-adjusted HLR). Results: A total of 5,985 Anopheles mosquitoes were collected, with 608 (10%) identified molecularly to species-level. Seventeen Anopheles species were identified, including a likely novel species from the Leucosphyrus Subgroup, which was the predominant species characterized. The HBO-adjusted HLR was found to be greatest during the early evening hours, when people were outdoors awake, followed by when people were sleeping indoors without a net. Relatively few people were observed using, or correctly using, the new bite prevention tools intended for protection in the forest. Conclusion: This study demonstrates the importance of understanding spatial and temporal human exposure to mosquito bites, in the presence of proven vector control tools (LLINs, LLIHNs) and newly introduced bite prevention tools (VPSRs, ITCs, and TRs). To help achieve malaria elimination, human behaviour data on intervention use and behaviour patterns should be evaluated and integrated with entomological data towards identifying and quantifying protection conferred by current interventions, as well as remaining gaps in protection. This information supports the selection of appropriate interventions, which supplement rather than replace existing tools, to target existing gaps in protection. © The Author(s) 2025.