Coupled transcriptome and proteome analysis of L3 and L4 developmental stages of Anisakis simplex s. s.: insights into target genes under glucose influence.

Anisakis simplex is a cosmopolitan parasitic nematode of marine organisms with a complex life cycle. Consuming fish infected with its larvae poses a health risk, as the parasites can penetrate gastrointestinal mucosa, damage stomach and intestinal walls, and trigger allergic reactions. The resulting disease is known as anisakiasis. The European Food Safety Authority (EFSA) classifies A. simplex as a biohazard. Most of its developmental stages (L3, L4, and adults) occur under anaerobic conditions, and larvae derive energy mainly from saccharides. However, the effect of glucose on L3 and L4 larvae-stages pathogenic to humans-has not been described. This study aimed to identify genes and pathways involved in glucose (10 mg/mL) response through transcriptomic and proteomic analyses of L3 and L4 larvae. Differentially expressed genes (DEGs), long non-coding RNAs (DELs), and differentially regulated proteins (DRPs) were identified. DEGs were involved in cuticle structure, lyase activity, and metabolic processes. Comparing L3 CTR and L4 CTR (control) to glucose-treated samples revealed 1,969 DEGs; 259 overlapped between L4 GLU vs. L3 GLU, (glucose-treated) with 11 showing reversed expression. Additionally, 84 DELs were identified in L3 GLU vs. control, 40 in L4 GLU vs. control, and 163 between glucose-treated L4 and L3. Larval and glucose-specific alternative splicing events were also analyzed. Proteomic analysis revealed 35 DRPs-5 more abundant in L4, 30 in L3. The data reveal that developmental stage exerts a more substantial influence than glucose exposure on gene and protein expression profiles. However, glucose still modulates several pathways related to translation, cytoskeletal remodeling, extracellular matrix (ECM) reorganization, and energy metabolism.