Decapod iridescent virus 1 (DIV1), the first isolated species of the genus Decapodiridovirus (family Iridoviridae), is a major pathogen threatening farmed decapods, causing large-scale disease outbreaks and high mortality. However, the mechanisms underlying its cellular entry remain poorly understood. In this study, we investigated DIV1 internalization using hematopoietic tissue cells of Cherax quadricarinatus. Pharmacological inhibition assays showed that (i) clathrin-dependent endocytosis inhibitors (pitstop 2, chlorpromazine) failed to suppress DIV1 infection, excluding clathrin-mediated entry; (ii) cellular cholesterol disrupted with methyl-β-cyclodextrin or nystatin significantly inhibited infection, and this effect was reversed by cholesterol replenishment; and (iii) caveolar inhibitors (phorbol 12-myristate 13-acetate, genistein, wortmannin) and organelle pH neutralizers (NH4Cl, bafilomycin A1) effectively blocked DIV1 invasion. Fluorescence co-localization and pull-down assays confirmed that DIV1 associates with caveolin-1 during internalization. Additionally, DIV1 entry relies on dynamin and the microtubule cytoskeleton, and post-internalization trafficking of the virus targeted the Golgi apparatus. Collectively, DIV1 enters hematopoietic C. quadricarinatus cells via caveola-mediated endocytosis, a process dependent on cholesterol, dynamin, and microtubule cytoskeleton, with subsequent trafficking to the Golgi apparatus.IMPORTANCEElucidating viral entry mechanisms into target cells is fundamental for developing effective antiviral strategies. This study reveals that DIV1, a major threat to decapod aquaculture, utilizes caveola-mediated endocytosis to invade hematopoietic tissue cells of Cherax quadricarinatus. This entry process is dependent on membrane cholesterol, dynamin activity, microtubule-based trafficking, and a pH-responsive Golgi environment. These findings fill a critical knowledge gap in Betairidovirinae entry biology, distinguish DIV1 from other iridovirids and decapod pathogens (e.g., white spot syndrome virus), and identify key host-derived targets. This work thereby enables the development of targeted antiviral interventions to safeguard decapod aquaculture.
Zheng et al. (Mon,) studied this question.