RESUMEN
Estimates of lower thermal limits are widely used to infer sensitivity to climate variability, local adaptation and adaptive acclimation responses in ectotherms. These inferences build on the ecological relevance of the tolerance estimates and assume that estimates can be extrapolated to relevant conditions. Methodological effects for upper thermal limits have been extensively investigated, with different ramping rates and acclimation regimes giving rise to varying, and even disparate, conclusions. However, methodological effects have received much less attention for lower thermal limits. In this study, we explicitly test whether methodology could affect estimates of lower thermal limits in interaction with acclimation temperature and thermal variability, by acclimating adult Drosophila melanogaster to different constant and fluctuating temperature regimes and generating reaction norms at different ramping rates. We find that ramping rates have no significant effect on the lower thermal limits. Constant temperature acclimation resulted in non-linear reaction norms, while the introduction of thermal variability during adult life result in linear reaction norms. Thus, applying ecologically relevant conditions (here thermal variability) potentially impacts the results and conclusions of insect low temperature tolerance and acclimation capacity.