Characterization of Anopheles gambiae D7 salivary proteins as markers of human-mosquito bite contact

Abstract

Background: Malaria is transmitted when infected Anopheles mosquitoes take a blood meal. During this process, the mosquitoes inject a cocktail of bioactive proteins that elicit antibody responses in humans and could be used as biomarkers of exposure to mosquito bites. This study evaluated the utility of IgG responses to members of the Anopheles gambiae D7 protein family as serological markers of human-vector contact.

Methods: The D7L2, D7r1, D7r2, D7r3, D7r4 and SG6 salivary proteins from An. gambiae were expressed as recombinant antigens in Escherichia coli. Antibody responses to the salivary proteins were compared in Europeans with no prior exposure to malaria and lifelong residents of Junju in Kenya and Kitgum in Uganda where the intensity of malaria transmission is moderate and high, respectively. In addition, to evaluate the feasibility of using anti-D7 IgG responses as a tool to evaluate the impact of vector control interventions, we compared responses between individuals using insecticide-treated bednets to those who did not in Junju, Kenya where bednet data were available.

Results: We show that both the long and short forms of the D7 salivary gland antigens elicit a strong antibody response in humans. IgG responses against the D7 antigens reflected the transmission intensities of the three study areas, with the highest to lowest responses observed in Kitgum (northern Uganda), Junju (Kenya) and malaria-naïve Europeans, respectively. Specifically, the long form D7L2 induced an IgG antibody response that increased with age and that was lower in individuals who slept under a bednet, indicating its potential as a serological tool for estimating human-vector contact and monitoring the effectiveness of vector control interventions.

Conclusions: This study reveals that D7L2 salivary antigen has great potential as a biomarker of exposure to mosquito bites and as a tool for assessing the efficacy of vector control strategies such as bednet use.

Keywords: Anopheles gambiae; Biomarker of exposure; Infectious bites; Malaria; Plasmodium falciparum.

Fig. 1

SDS-PAGE image of the recombinant mosquito salivary antigens

Fig. 2

IgG responses to gSG6 and D7 salivary antigens. Plasma IgG levels to D7 proteins were compared between malaria-naïve European adults (n = 35), a cohort of children from Junju in Kilifi (n = 313) and a cohort Kitgum in northern Uganda (n = 147). The P-value was determined by the Kruskal–Wallis test (***P < 0.001)

Fig. 3

IgG responses to gSG6 and D7 salivary antigens stratified by age among individuals resident in Kilifi, Kenya (0–3 years [n = 62], 3–6 years [n = 85], > 7 years [n = 166]). The P-values were determined by the Kruskal–Wallis test (ns, not significant; **P < 0.01; *P < 0.05)

Fig. 4

IgG responses to recombinant D7L2 compared between individuals who used insecticide-treated bednets (n = 158) and those who did not (n = 155). The P-value was determined by the Wilcoxon rank-sum test (ns, not significant; ** P < 0.01)