When is a slender not a slender? The cell-cycle arrest and scarcity of slender parasites challenges the role of bloodstream trypanosomes in infection maintenance

The development of Trypanosoma brucei in its mammalian host is marked by a distinct morphological change as replicative "slender" forms differentiate into cell-cycle arrested "stumpy" forms in a quorum-sensing dependent manner. Although stumpy forms dominate chronic infections at the population level, the proportion of replicative parasites at the individual cell level and the irreversibility of arrest in the bloodstream is unclear. Here, we experimentally demonstrate that developmental cell cycle arrest is definitively irreversible in acute and chronic infections in mice. Furthermore, analysis of replicative capacity and single-cell transcriptome profiling reveals a temporal hierarchy, whereby cell-cycle arrest and appearance of a stumpy-like transcriptome precedes irreversible commitment and morphological change. Unexpectedly, we show that proliferating parasites are exceptionally scarce in the blood after infections are established. This challenges the ability of bloodstream trypanosomes to sustain infection by proliferation or antigenic variation, these parasites instead being overwhelmingly adapted for transmission.