Life‐Stage–Specific Hematological Profiles in Yellow Mealworm Beetles ( <i>Tenebrio molitor</i> ): Reference Intervals and Hemocyte Characterization

BACKGROUND: The edible insect industry, driven by the nutritional and environmental benefits of mealworms, faces increasing challenges from pathogens affecting Tenebrio molitor farming systems. Hemocyte-based health assessments could provide valuable diagnostic insights, yet standardized hemolymph processing protocols and hematological reference intervals (RIs) are currently lacking for this species. OBJECTIVES: This study aimed to develop practical protocols for hemolymph sampling, refine hemocyte classification, and establish hematological RIs for both larval and adult stages of T. molitor. METHODS: An optimized tearing method, involving capillary collection of hemolymph from a proleg transected at the femoral-tibial joint, was used to sample healthy individuals. Total and differential hemocyte counts were determined using a hemocytometer and Wright-Giemsa-stained smears, respectively, and from distinct insect populations (n = 41-43 samples per group). Hemolymph was analyzed in pools due to the small size of T. molitor. Transmission electron microscopy was conducted on a multinucleated giant hemocyte (MGH) isolated from a histologic slide. RIs were calculated following American Society for Veterinary Clinical Pathology (ASVCP) guidelines, and life-stage differences were statistically assessed. RESULTS: Granular cells and plasmatocytes predominated, whereas prohemocytes and oenocytoids were rare. MGHs, newly identified in T. molitor, were characterized as a fifth hemocyte population. Significant life-stage differences were observed, warranting separate RIs for larvae and adults. The established RIs were relatively wide, reflecting natural fluctuations in invertebrate hemolymph volume and the unsexed, age-diverse reference population. CONCLUSIONS: This report of hematological RIs for T. molitor lays the groundwork for implementing hemocyte-based health surveillance in yellow mealworm beetle farming.