Acute Smurf mortality and inter-phase dependence in Drosophila and mice identified through comprehensive modelling and statistical analysis of two-phase ageing

Ageing is traditionally conceived as a continuous process of progressive physiological decline. Recent evidence across multiple species challenges this view, suggesting ageing may proceed through distinct phases. Here we present a rigorous statistical framework to test and refine the two-phase ageing model using longitudinal survival data from Drosophila melanogaster. We analyzed 1,159 individually tracked female flies from the Smurf assay, which identifies a transition from a non-Smurf state to a Smurf state characterized by increased intestinal permeability that precedes death. Using non-parametric hazard rate estimation followed by mechanistic modelling, we reveal three key findings. First, the Smurf transition rate follows a Gompertz-Makeham law, increasing exponentially with age. Second, contrary to previous constant-rate assumptions, newly transitioned Smurf flies exhibit remarkably high mortality - approximately 40% die within 24 hours - followed by an exponential decline in death rate. Third, we identified a subtle but statistically significant negative dependence between time spent non-Smurf and subsequent Smurf lifespan, though this relationship varies: flies transitioning before 200 hours show minimal dependence and higher early mortality. Our best-fit model captures the bimodal nature of mortality curves using simple, biologically interpretable functions. Validation using data from two mouse strains confirms the broader applicability of this framework. These results establish a quantitative foundation for the two-phase ageing paradigm and highlight a critical period of vulnerability immediately following the physiological transition to frailty.