Reflectance-based determination of age and species of blowfly puparia

Forensic entomology is primarily concerned with the estimation of time since death and involves determination of the age of immature insects colonising decomposing remains. Accurate age determination of puparia is usually accomplished by dissection, which means destructive sampling of evidence. As part of improving abilities to correctly identify species and developmental age, it is highly desirable to have available non-destructive methods. In this study, we acquired external hyperspectral imaging (HSI) data (77 spectral bands, 389-892 nm) from the dorsal and ventral sides of individual puparia of two species of blowfly (Diptera: Calliphoridae), Calliphora dubia Macquart 1855 and Chrysomya rufifacies Macquart 1842. Puparia were dissected to determine the presence/absence of eight internal morphological development characteristics (legs, wings, labella, abdominal segments, antennae, thoracic bristles, orbital/facial bristles and eye colour and arista). Based on linear discriminant analysis and independent validation of HSI data, reflectance features from puparia could be used to successfully (1) distinguish the two species (classification accuracy = 92.5 %), (2) differentiate dorsal and ventral sides of puparia (classification accuracy C. dubia = 81.5 %; Ch. rufifacies = 89.2 %) and (3) predict the presence of these morphological characteristics and therefore the developmental stage of puparia (average classification accuracy using dorsal imaging: C. dubia = 90.3 %; Ch. rufifacies = 94.0 %). The analytical approach presented here provides proof of concept for a direct puparial age relationship (i.e. days since the onset of pupation) between external puparial reflectance features and internal morphological development. Furthermore, this approach establishes the potential for further refinement by using a non-invasive technique to determine the age and developmental stage of blowflies of forensic importance.