A transmission bottleneck for malaria? Quantification of sporozoite expelling from laboratory and natural P. falciparum infections

Abstract

It is currently unknown whether all Plasmodium falciparum infected mosquitoes are equally infectious. We assessed sporogonic development using cultured gametocytes in the Netherlands and natural infections in Burkina Faso. We quantified the number of sporozoites expelled into artificial skin in relation to intact oocysts, ruptured oocysts, and residual salivary gland sporozoites. Sporozoites were quantified by highly sensitive qPCR; intact and ruptured oocysts by fluorescence microscopy following anti-circumsporozoite antibody staining. In laboratory conditions, higher total sporozoite burden in mosquitoes was associated with a shorter duration of sporogony (p<0.001). Overall, 53% (116/216) of P. falciparum infected An. stephensi mosquitoes expelled sporozoites into artificial skin. The geometric means of expelled and residual salivary gland sporozoites were 116 (interquartile range (IQR: 33-501) and 21,016 (IQR: 9127-78,380), respectively. There was a strong correlation between ruptured oocyst number and salivary gland sporozoite load (ρ=0.74; p<0.0001) and a weaker positive correlation between salivary gland sporozoite load and the number of sporozoites expelled (ρ=0.35; p=0.0002). In Burkina Faso, An. coluzzii mosquitoes were infected by natural gametocyte carriers. Among mosquitoes that were salivary gland sporozoite positive, 97.2% (36/37) expelled sporozoites with a geometric mean of 420 expelled sporozoites (IQR: 116-2,779) and harbored a geometric mean of 35,149 residual salivary gland sporozoites (IQR: 20,310-164,900). Again, we observed a strong correlation between ruptured oocyst number and salivary gland sporozoite load (ρ=0.84; p<0.0001) and a positive correlation between salivary gland sporozoite load and the number of sporozoites expelled (ρ=0.68; p=0.0003). Whilst sporozoite expelling was regularly observed from mosquitoes with low infection burdens, our findings indicate that mosquito infection burden is associated with the number of expelled sporozoites and may need to be considered in estimations of transmission potential.