Metabolic dysfunction-associated steatotic liver disease (MASLD) and opisthorchiasis, caused by Opisthorchis viverrini (O. viverrini) infection, frequently co-exist in Northeast Thailand. However, the underlying pathophysiology remains unknown. We aimed to investigate the effect of a high-fat/high-fructose (HFF) diet combined with O. viverrini infection on MASLD. Four groups each of ten male golden hamsters were established: normal controls (NC), O. viverrini-infected (OV), HFF-fed, and HFF-fed plus O. viverrini infection (HFF+OV). After four months of treatment, histopathological study indicated substantial hepatic damage in groups given the HFF diet. In particular, the HFF+OV group demonstrated marked lipid-droplet accumulation, hepatocyte ballooning, inflammatory-cell clustering, and widespread fibrosis. Biochemical tests indicated that the HFF+OV group had the highest concentrations of alanine aminotransferase and triglycerides, but cholesterol and low-density lipoprotein levels had increased in both HFF groups. Increased expression of Tgf-β1 and α-SMA, indicative of greater fibrosis, was demonstrated by picrosirius-red staining in the HFF+OV group. There was a significant increase in levels of inflammatory markers (HMGB-1, p65, and F4/80) and expression of genes related to the synthesis of fatty acids and glucose. FTIR microspectroscopy revealed distinct changes in fatty acids and proteins, associated with the more pronounced histopathology and impaired liver function in the HFF+OV group. The findings indicate that the interplay of a HFF diet and O. viverrini infection aggravates the progression of MASLD by augmenting liver damage, inflammation, fibrogenesis, and metabolic dysfunction. This study highlights the significance of incorporating both nutritional and infection factors into the management of liver disorders, especially in areas where opisthorchiasis is common.