Sequential blood meals augment vector infectiousness by promoting Leishmania replication and triggering amplification of metacyclics via a novel retroleptomonad developmental stage

Sand flies, similar to most vectors, take multiple blood meals during their lifetime(1-4). The effect of subsequent blood meals on pathogens developing in the vector, and their impact on disease transmission have never been examined. Here, we show that ingestion of a second uninfected blood meal by Leishmania-infected sand flies triggers dedifferentiation of metacyclic promastigotes, considered a terminally differentiated stage inside the vector(5), to a leptomonad-like stage, the retroleptomonad promastigote. Reverse metacyclogenesis occurs after every subsequent blood meal where retroleptomonad promastigotes rapidly multiply and differentiate to metacyclic promastigotes enhancing sand fly infectiousness. Importantly, a subsequent blood meal amplifies the few Leishmania parasites acquired by feeding on infected hosts by 125 folds, and increases lesion frequency by 4 folds, in twice-fed compared to single-fed flies. These findings place readily available blood sources as a critical element in transmission and propagation of vector-borne pathogens.