Background
Current strategies used in the management of malaria are threatened by the development of insecticide and behavioural resistance in human malaria vectors [
1,
2]. Efforts to develop tools to complement those currently in use in integrated vector management (IVM) are required, particularly those targeting exophilic mosquitoes, as an increasing proportion of people are now at risk of infective bites from mosquitoes outdoors [
3‐
5]. To this end, it is essential to increase the understanding of the ecology and behaviour of malaria vectors outdoors, to identify novel targets for IVM tool development [
5,
6]. In particular, targeting gravid malaria vectors, a life stage currently lacking tools in IVM, has the potential to reduce the mosquito density, and the vectorial capacity, of a competent mosquito population [
6].
Recent research has begun to shed light on the ecology of ovipositing malaria vectors, in part, by investigating how gravid females select and discriminate among potential oviposition sites [
7‐
12]. Aside from humidity [
13], odours emanating directly from potential oviposition sites and associated vegetation are used by gravid mosquitoes for both selection of sites and stimulation of oviposition [
7‐
11,
14‐
21]. Odours emanating from either wild or domesticated grasses appear to provide accurate signals for the quality of oviposition sites for
Anopheles arabiensis and
Anopheles coluzzii, two of the primary malaria vectors in Sub-Saharan Africa [
8‐
11]. Through a classical chemical ecology approach, including behavioural and electrophysiological analyses of volatile compounds from the headspace volatiles of rice, maize and sugarcane, synthetic odour blends have been developed [
9‐
11]. The behavioural response of gravid
An. arabiensis to these blends reflects that of the natural odours [
9‐
11]. Of the three synthetic blends, the most attractive, rice, was evaluated under semi-field conditions, demonstrating a recapture rate of more than 70% [
9].
While synthetic odour lures based on natural sources may be effective, these are potentially constrained by their restricted effectiveness in direct competition with the natural odour source, and may be affected by the previous odour experience of an individual [
22]. To overcome such constraints, bioactive compounds identified in preferred odour sources may be combined into a blend, thereby avoiding the direct mimicking of a natural odour [
23,
24]. Such “super blends” have been used effectively to control plant pests from various insect orders [
25,
26], but as of yet, are not available for the control of haematophagous insects.
In this study, the principles set out by Del Socorro et al
. [
27] and Gregg et al
. [
25] were used in designing a chimeric blend based on the previously identified attractive blends of domesticated grasses. Moreover, following the ratio-specific hypothesis proposed by Bruce et al
. [
28] and Bruce and Pickett [
29], this blend was assayed for attractiveness at various ratios within that of the natural emission rates of the individual compounds under laboratory conditions. The most attractive chimeric blend was subsequently evaluated under field conditions in malaria endemic villages in Ethiopia, using different trapping methods. The findings are discussed in the context of its potential for the development of a gravid trap for malaria mosquitoes and its potential to influence the strategy and goals set by the World Health Organization through its vector ecology and management, and its sustainable development divisions [
6].
Discussion
Odour-based tools, targeting gravid
An. arabiensis, are required to complement existing intervention strategies, which mainly target the indoor feeding and resting population. Gravid
An. arabiensis are attracted to natural and synthetic odours of domesticated grasses [
9‐
11,
39]. Bioactive volatile compounds identified and shared among these domesticated grasses were used to develop a ratio-optimized chimeric odour blend, providing a workflow for the development of blends that target oviposition-site seeking mosquitoes. In a two-choice assay, gravid mosquitoes preferred the chimeric odour blend (blend M) over that of a synthetic odour of rice, previously demonstrated to be highly effective in attracting and capturing mosquitoes in laboratory and semi-field experiments [
9]. While field assessment demonstrated that the chimeric blend (blend M) may be effective in direct competition with natural odour sources, future work is required to enhance the applicability of this odour-based intervention tool.
Originally described by Del Soccorro et al
. [
27], synthetic attractant blends do not need to rely on a mimic of bioactive compounds in a single resource. In fact, blends of bioactive compounds that are shared among attractive resources may provide an enhancement of attraction, thus providing a competitive advantage compared to any existing natural source [
26,
27,
39]. Similar to Gregg et al
. [
25,
40], we used an empirical approach to develop the chimeric blend used to attract gravid
An. arabiensis, in which the ratio of the individual bioactive compounds was optimized [
28,
29]. While the inclusion of additional bioactive compounds from preferred, and even non-preferred, vegetation associated with mosquito potential breeding sites may improve the efficacy of the chimeric blend under field conditions, the results from the laboratory bioassays clearly demonstrate that the chimeric blend may be superior to that of previously identified attractants for gravid
Anopheles mosquitoes [
7,
9‐
11,
15‐
17,
21]. Moreover, this study provides proof-of-principle that chimeric blends, also referred to as super-blends, may be developed for surveillance and control of vector mosquitoes, by combining similar approaches as those used for plant-feeding insect pests. With recent progress in understanding the chemical ecology of behaviours involved in floral, host and oviposition-site selection, it is becoming clear that bioactive compounds are used by mosquitoes parsimoniously [
12,
24]. These compounds provide a basis for the future development of chimeric blends that attract mosquitoes of different species and physiological states.
The number of mosquitoes caught in the field experiments was dependent on trap type and placement with respect to ground level. Similar to that reported by Lindh et al
. [
7], the number of mosquitoes caught per trap per night was low, which can be explained, at least in part, by the low population densities at the time of study. By placing the traps in hotspots for resting mosquitoes [
31], shaded sites that were ≥ 50 m from the household, a higher number of mosquitoes were caught, than those reported by Lindh et al
. [
7], even after adjusting for differences in population density. However, trap capture was still low, possibly reflecting the difficulty in luring gravid mosquitoes from their resting sites, and the current understanding of how gravid mosquitoes move within the landscape to locate potential oviposition sites. These field experiments designed to assess the dose-dependent attraction of malaria vectors, and generally identified the lower range of doses (3–10 ng µl
−1) to be the most effective. This is consistent with our previous results from semi-field trials with the synthetic rice blend, emphasizing the superior sensitivity of the olfactory system of
Anopheles mosquitoes [
9]. The high density of active mosquitoes near households led us to evaluate a low release rate of the chimeric blend outdoors and next to the houses, which resulted in significantly higher numbers of captures in traps baited with the lure. Whether distance from the households affects the efficacy of the lure described in this study needs further analysis, as this was not directly assessed here. The data presented in this study suggest that the activity of
Anopheles mosquitoes with various physiological states varies depending on distance from the households, with gravid mosquitoes being more amenable to trapping close to rest sites ≥ 50 m from households, which is in line with previous observations [
31].
From this study and others [
7,
41,
42], it is obvious that trap type and placement with respect to ground level is critical for ensuring the optimal efficacy of odour-based lures for malaria vectors. Previous evaluations of gravid traps targeting gravid
Anopheles mosquitoes have identified the presence of water in, the direction of airflow into (up- or down-draft) and the placement of the trap with respect to ground level to be important factors when capturing these females [
7,
9,
41,
42]. In addition, trap type has been identified as a critical factor when capturing host-seeking
Anopheles mosquitoes, with various versions of the BG-Sentinel traps, including the BG-Malaria trap and the Suna trap, often demonstrated to be superior compared to other trap types, e.g., the CDC light trap [
33,
43,
44]. Using a similar approach to that of Batista et al. [
44], 3-dimensional video-graphic analysis may be used to improve the design of gravid traps for
Anopheles mosquitoes. The addition of other cues, including water vapor [
13] and visual stimuli [
41,
45], should be considered in future development of trapping systems for gravid
Anopheles.
Conclusions
The Global Vector Control Response (WHO 2017–2030) lists the innovation of new tools, particularly those targeting vectors outdoors, with the express purpose to integrate these in sustainable IVM programs as one of two foundational elements for effective and locally-adaptive vector control systems. Innovations, based on the fundamental understanding of behaviours affecting vectorial capacity, e.g., oviposition, are critical to tackle the increased population of outdoor malaria vectors. The identification of a chimeric odour blend has yielded a workflow designed for the development of odour-based lures based on the natural odour space of vector insects, here resulting in a lure for gravid malaria vectors. This lure may be used in existing trapping systems, or may serve as the basis for the development of novel systems, further designed to optimize trap capture of Anopheles mosquitoes. Chimeric lure systems can be customized to the local vector environments with minimal input, once local ecological conditions are known, and with multiple lures available, these may be used in rotation to increase sustainability by avoiding behavioural resistance to any one blend.
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