Calliphora vicina (Diptera: Calliphoridae) pupae: a timeline of external morphological development and a new age and PMI estimation tool.

The minimum postmortem interval (PMI(min)) is commonly estimated using calliphorid larvae, for which there are established age estimation methods based on morphological and development data. Despite the increased duration and sedentary nature of the pupal stage of the blowfly, morphological age estimation methods are poorly documented and infrequently used for PMI determination. The aim of this study was to develop a timeline of metamorphosis, focusing on the development of external morphology (within the puparium), to provide a means of age and PMI estimation for Calliphora vicina (Rob-Desvoidy) pupae. Under controlled conditions, 1,494 pupae were reared and sampled at regular time intervals. After puparium removal, observations of 23 external metamorphic developments were correlated to age in accumulated degree hours (ADH). Two age estimation methods were developed based on (1) the combination of possible age ranges observed for each characteristic and (2) regression analyses to generate age estimation equations employing all 23 characteristics observed and a subset of ten characteristics most significantly correlated with age. Blind sample analysis indicated that, using the combination of both methods, pupal age could be estimated to within ±500 ADH with 95% reliability.