Estimating Post-Feeding Developmental Time of Sarcophaga peregrina (Diptera: Sarcophagidae) Larvae at 25 °C Using ATR-FTIR Spectroscopy and Differential Gene Expression Analysis

Estimating the post-feeding developmental time of necrophagous larvae remains challenging because body size and behavior provide limited resolution after larvae cease feeding. This study integrated behavioral observation, attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), chemometric regression modeling, transcriptomic analysis, and reverse transcription quantitative polymerase chain reaction (RT-qPCR) validation to characterize post-feeding development in Sarcophaga peregrina (Robineau-Desvoidy) under 25 °C. The post-feeding stage began at 84.83 ± 1.34 h after larviposition and lasted 38.21 ± 2.29 h. ATR-FTIR spectra from 0, 10, 20, 30, and 40 h post-feeding larvae showed temporal changes in the 1800–900 cm−1 fingerprint region, mainly involving bands associated with proteins, lipids, polysaccharides, and chitin. Principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and variable importance in projection (VIP) analyses identified discriminative spectral regions, while regression models further evaluated time-estimation performance. Among the models, partial least squares regression (PLS-R) showed the most balanced predictive ability, with the coefficient of determination in prediction (R2P) = 0.920 and the root mean square error of prediction (RMSEP) = 4.00 h in the prediction set. Transcriptomic analysis identified 4641 differentially expressed genes, and RT-qPCR confirmed temporal expression patterns of selected candidate genes. These molecular changes were mainly associated with cuticle/chitin remodeling, protein metabolism, and energy metabolism. Overall, ATR-FTIR combined with chemometric modeling provides a complementary biochemical approach for estimating post-feeding developmental time in S. peregrina under controlled 25 °C conditions, although validation at other temperatures is required before broader application.