Intervention reducing malaria parasite load in vector mosquitoes: No impact on Plasmodium falciparum extrinsic incubation period and the survival of Anopheles gambiae.

In the fight against malaria, transmission blocking interventions (TBIs) such as transmission blocking vaccines or drugs, are promising approaches to complement conventional tools. They aim to prevent the infection of vectors and thereby reduce the subsequent exposure of a human population to infectious mosquitoes. The effectiveness of these approaches has been shown to depend on the initial intensity of infection in mosquitoes, often measured as the mean number of oocysts resulting from an infectious blood meal in absence of intervention. In mosquitoes exposed to a high intensity of infection, current TBI candidates are expected to be ineffective at completely blocking infection but will decrease parasite load and therefore, potentially also affect key parameters of vector transmission. The present study investigated the consequences of changes in oocyst intensity on subsequent parasite development and mosquito survival. To address this, we experimentally produced different intensities of infection for Anopheles gambiae females from Burkina Faso by diluting gametocytes from three natural Plasmodium falciparum local isolates and used a newly developed non-destructive method based on the exploitation of mosquito sugar feeding to track parasite and mosquito life history traits throughout sporogonic development. Our results indicate the extrinsic incubation period (EIP) of P. falciparum and mosquito survival did not vary with parasite density but differed significantly between parasite isolates with estimated EIP50 of 16 (95% CI: 15-18), 14 (95% CI: 12-16) and 12 (95% CI: 12-13) days and median longevity of 25 (95% CI: 22-29), 15 (95% CI: 13-15) and 18 (95% CI: 17-19) days for the three isolates respectively. Our results here do not identify unintended consequences of the decrease of parasite loads in mosquitoes on the parasite incubation period or on mosquito survival, two key parameters of vectorial capacity, and hence support the use of transmission blocking strategies to control malaria

Effect of infectious blood dilution on the prevalence and intensity of infection in mosquitoes.

(a) Oocyst prevalence 7 dpbm (number of mosquitoes with at least one oocyst in their midguts out of the total number of mosquitoes dissected) as a function of gametocytemia (the estimated number of infectious gametocytes per microliter of blood in each of the dilution groups: 24, 45, 48, 72, 91, 98, 136, 261 and 784 gametocytes/μl of blood, note that to avoid overlapping of the x-axis labels, the concentrations of 48 and 91 gam/μl are not indicated). Each colored circle represents a dissected mosquito (red: parasite isolate A with an initial gametocytemia of 72, green: parasite isolate B with initial gametocytemia of 136, and blue: parasite isolate C with an initial gametocytemia of 784). The colored lines represent the best-fit logistic growth curves for each parasite isolate. Note that the x-axis is on a log10 scale. (b) Oocyst intensity 7 dpbm (number of oocysts in infected mosquitoes) as a function of gametocytemia. Each colored circle represents a P. falciparum oocyst-positive midgut. The colored lines represent the linear relationship for each parasite isolate, while the black line (± se) for all data regardless of isolate origin. Note that the x- and y- axes are on a log10 scale. (c) Sporozoite prevalence (number of mosquitoes with heads/thoraxes detected positive to P. falciparum by qPCR at the time of death of the individual out of the total number of tested heads/thoraxes) as a function of gametocytemia. Each colored circle represents a tested head/thorax. The x-axis is on a log10 scale. (d) Amount of parasite DNA in mosquito heads/thoraxes expressed as the inverse of the qPCR cycle threshold (1/Ct, the higher the inverse of threshold cycle, the higher the intensity of infection). For each mosquito, 1/Ct value is the average over 4 to 6 technical replicates. The x- and y-axes are on a log10 scale. Each colored circle represents a P. falciparum positive head/thorax using qPCR.</p

Relationship between parasite density and EIP.

(a) EIP (the time between the infectious blood meal and the detection of P. falciparum in mosquito saliva collected from cotton balls) as a function of gametocytemia (the number of gametocytes per microliter of blood in each of the dilution groups: 24, 45, 48, 72, 91, 98, 136, 261 and 784 gametocytes/μl of blood, note that to avoid overlapping of the x-axis labels, the concentrations of 48 and 91 gam/μl are not indicated). Each colored circle represents an infected mosquito from which EIP was measured (red: parasite isolate A with an initial gametocytemia of 72, green: parasite isolate B with initial gametocytemia of 136, and blue: parasite isolate C with an initial gametocytemia of 784). The x-axis is on a log10 scale. (b) EIP as a function of 1/Ct in mosquito infected heads/thoraxes extracts (the higher the 1/Ct value, the higher the infection intensity). For each cotton ball, 1/Ct value is the average over 3 technical replicates. The colored lines in panels (a) and (b) represent the linear relationship for each parasite isolate. (c) Kaplan–Meier curves representing the temporal dynamics of sporozoite detection in cotton balls used to collect saliva from individual mosquitoes fed on each parasite isolate.</p

Relationship between parasite density and mosquito longevity.

(a) Kaplan–Meier curves representing survival in days post blood meal for each infection status (red: mosquitoes exposed to infectious blood and having developed Plasmodium, black: mosquitoes exposed to infectious blood and which remained uninfected). (b) Mosquito longevity in days as a function of gametocytemia (the number of gametocytes per microliter of blood in each of the dilution groups: 24, 45, 48, 72, 91, 98, 136, 261 and 784 gametocytes/μl of blood, note that to avoid overlapping of the x-axis label, the concentrations of 48 and 91 gam/μl are not indicated). Each colored circle represents a mosquito exposed to the infectious blood (red: infected mosquitoes, black: mosquitoes that remained uninfected). The x-axis is on a log10 scale. (c) Mosquito longevity in days as a function of gametocytemia (the number of gametocytes per microliter of blood in each of the dilution groups: 24, 45, 48, 72, 91, 98, 136, 261 and 784 gametocytes/μl of blood, note that to avoid overlapping of the x-axis label, the concentrations of 48 and 91 gam/μl are not indicated). Each colored circle represents a mosquito exposed to one of the three parasite isolates (red: parasite isolate A with an initial gametocytemia of 72, green: parasite isolate B with initial gametocytemia of 136, and blue: parasite isolate C with an initial gametocytemia of 784). The x-axis is on a log10 scale. (d) mosquito longevity in days as a function of 1/Ct in mosquito infected heads/thoraxes extracts (the higher the 1/Ct value, the higher the infection intensity). For each cotton ball, 1/Ct value is the average over 3 technical replicates. Each colored circle represents an infected mosquito. The colored lines in panels (b) and (c) represent the linear relationship for each parasite isolate. (e) Kaplan–Meier curves representing survival in days post blood meal for each parasite isolate.</p