Chronic nanoplastic exposure progressively disrupts immune-related traits across generations in Drosophila melanogaster

ABSTRACTF Nanoplastics (NP) are an emerging type of environmental contaminant. However, their long-term biological effects, particularly on terrestrial organisms, remain poorly understood. This study investigated the impact of chronic dietary exposure to NP on the reproductive output, immune-related traits, and antibacterial performance across ten successive generations of Drosophila melanogaster . The flies were continuously exposed to four concentrations of NP (0, 1, 100, and 500 µg NP g⁻¹ food). Reproductive parameters (e.g., oviposition) were monitored alongside immune endpoints, including hemocyte populations dynamics, antimicrobial peptide gene expression (Cecropin A1, Drosomycin, and Bomanin), and functional antibacterial responses. Reproductive output remained stable across generations, indicating short-term resilience to NP exposure. However, immune-related traits showed progressive, generation-dependent changes. F10 restructured hemocyte populations, increasing plasmatocytes, and reducing crystal cells, and lamellocytes. Gene expression analyses revealed AMP-specific modulation: Cecropin A1 (CecA1) and Bomanin (S4) were consistently suppressed, while Drosomycin (Drs) exhibited a non-monotonic response. Importantly, antibacterial performance decreased from the F5 generation onward, with a dose-dependent increase in colony-forming unit counts (up to threefold at 500 µg NP g⁻¹ food) in response to challenges with both Bacillus thuringiensis and Serratia marcescens . This study provides the first evidence of multigenerational immune disruption in a terrestrial invertebrate chronically exposed to NP, without any adverse effects on oviposition. Our results emphasize the potential for cumulative immunotoxicity in environments contaminated with NP, highlighting the need for long-term ecological risk assessments.