During the bloodstream stage of the Trypanosoma brucei lifecycle, the parasite exists
in two different states: the proliferative slender form and the non-proliferative,
transmissible, stumpy form. The transition from the slender to stumpy form is
stimulated by a density-dependent mechanism and is important in infection
dynamics, ordered antigenic variation and disease transmissibility. The slender to
stumpy transition and the contribution of stumpy formation to within-host dynamics
have been difficult to analyse, however, because cell-type specific markers have been
restricted to imprecise morphological criteria.
PAD1 is a recently identified stumpy-specific protein which acts as a molecular
marker for stumpy formation and a functional marker for transmission. Here, the
control of stumpy-specific gene expression via the 3’UTR has been analysed,
identifying that there are repressive elements in the 3’UTR preventing inappropriate
expression during the slender life stage. Further, both pleomorphic and monomorphic
transgenic reporter cell lines utilising the PAD1 3’UTR have been created that report
on stumpy formation in vitro and these have been used for the analysis of stumpyinducing
chemical compounds.
Finally, a sensitive and accurate qRT-PCR assay has been developed and optimised
that faithfully reports both parasitaemia and stumpy formation throughout host
infection. Using a chronic infection rodent model, stumpy levels have been
monitored on the basis of conventional morphological and cell cycle assays, as well
as by qRT-PCR for PAD1 expression. The results define the temporal order of events
that result in the generation of stumpy forms early in a parasite infection and
thereafter describe the dynamics of slender and stumpy forms in chronic infections
extending over several weeks. This quantitative data has allowed the mathematical
modelling of transmission competence in trypanosome infections, suggesting
dominance of transmission stages throughout infection
