In Vitro Studies Of The Effect Of Anopheles Gambiae Midgut Bacteria On The Development Of Plasmodium Falciparum

Abstract

During blood feeding, female Anopheles mosquitoes may ingest Plasmodium gametocytes which undergo transformation in the gut and develop into sporozoites that are infectious to humans. Bacteria inhabit the mosquito gut, and the number and diversity of these bacteria change following blood feeding. The presence of some bacteria species results in the reduced intensity of developing Plasmodium parasites. Little attention has been given to understanding this direct mechanism of bacteria on Plasmodium parasites, and the effects of bacteria on malaria parasite developmental genes are not completely understood. This limits the scope of how gut bacteria, for example Enterobacter and Serratia, which have been found with anti-Plasmodium effects can be further explored for alternative disease control strategies. Therefore, this study investigated the lethal effect of cell-free secreted bio-products of E. cloacae and S. marcescens on a key Plasmodium parasite developmental gene (Gamete release gene, GAMER) for its potential as a target for malaria transmission-blocking. Plasmodium falciparum 3D7 and Dd2 cultures at 1% parasitaemia were independently exposed to spent Luria-Bertani (LB) medium from varying concentrations of Enterobacter cloacae and Serratia marcescens. The parasite killing effect of the bacteria were assessed with SYBR green fluorescent assay after 48 hours of co-culture. Spent media with final bacteria concentration between 10e+10-10e+20 reduced parasitaemia (P<0.001) compared to parasite culture without bacteria treatment. Using real-time (quantitative) PCR, it was found that the expression of GAMER was down regulated by 2 folds after 1 hour of screening P. falciparum 3D7 with cell-free spent medium of E. cloacae cultured for 8 hours in LB broth (Ec-8). However, the expression of GAMER was unaffected after 6 and 12 hours of screening P. falciparum 3D7 with Ec-8. These data provide information for further studies on gene and protein targets for transmission blocking interventions.