Comparison of the effect of patients?�? own sera versus na??ve sera on infectivity of Plasmodium vivax to Anopheles arabiensis from naturally infected gametocyte carriers using membrane feeding assay in Ethiopia

Abstract

Despite the achievements obtained so far, malaria continued to be the burden of public health which may indicate the need to update and strengthen the malaria control strategies. One potential candidate that the strategy toolbox can benefit from is malaria transmission blocking/reducing vaccines. In mosquito membrane feeding experiments, where mosquitoes were artificially fed on the blood of infected humans, they could not develop oocysts showing that some individuals produced antibodies against the sexual stage surface antigens of the parasite in blood which can be taken by mosquitoes during blood meal. This study focused on comparing the effect of these sexual stage immunity in the patients’ own serum with naïve serum as transmission reducing immunity from naturally P. vivax infected gametocyte carriers using serum replacement direct membrane feeding assay. The effect of patients’ own sera on infectivity of P. vivax and mixed infection (P. vivax and P. falciparum) to laboratory reared Anopheles arabiensis from naturally infected patients around Arbaminch town, Ethiopia, was assessed by comparing with naïve serum using serum replacement membrane feeding assay. Feeding assays used 4–6 days-old female Anopheles arabiensis mosquitoes after starvation for 12 hrs. Oocysts development was assessed microscopically seven days post-feeding. Asexual parasites and gametocytes were quantified in donor blood by microscopy and quantitative PCR (qPCR / RT-qPCR). Eighteen individual participants confirmed of malaria positive included in feeding experiments of which 11/18 (61.1%) were P. vivax mono infection, 1/18 (5.5%) was P. falciparum mono infection and 3/18 (16.7%) were mixed infection whereas 3/18 (16.7%) of them later confirmed by nested and qPCR to be negative. A total of 3600 Anopheles arabiensis used for feeding from which >80% successfully fed. For each individual feeding, midgut of 30-32 Anopheles mosquitoes were dissected to detect and count the oocysts number per midgut. The proportion of infected mosquitoes for patients’ own and naive serum was not significantly associated with gametocyte density determined by microscope (ρ(11) = -0.0275; P = 0.946) and stage based transcript RT-qPCR (ρ(11) = 0.477; P = 0.138). Likewise, it is not also associated with parasite density assessed by microscopy (ρ (11) = - 0.0459; P = 0.452) and qPCR (ρ(11) = -0092; P = 0.989). From total feeding done, 83.3% (10/12) of P. vivax single species and 33%, (1/3) of mixed species patients infected at least one mosquito whereas no mosquitoes infected by P. falciparum. The proportion of infected mosquitoes increases Page | VIII when naive serum (median of proportion of infected mosquitoes, 99%; IQR, 67-100) than the patient's own serum (median 80%; IQR, 43-87; P=0.0023, Wilcoxon signed-rank test) was used. The overall median oocysts intensity per infected midgut was significantly lower in patient’s own sera (median, 10; IQR, 3-30; range, 1-187; P<0.001; Signed-rank test) than naive serum (median, 38; IQR, 6-74; range, 1-224). Transmission reduction characteristics of individual patient serum was measured by a reduction in the oocysts number and classified as 18.2% non-blocking sera and 81.8% reducing sera. There is an indication that there are antibodies from naturally infected individuals in malaria exposed populations in Ethiopia that act as a transmission reducing immunity in our study. This need further study on such antibodies for their prevalence, robustness, kinetics and blocking, or reducing, or enhancing potential against selected antigens in different endemic settings