A wide range of factors can be attributed to the syndrome of fatty liver observed in some cultured fish species. The objective of the study was therefore to quantify different hepatocyte ultrastructural features as potentially influenced by twelve nutritional and husbandry factors, in order to discriminate the most influent factors in Eurasian perch (Perca fluviatilis), a typical carnivorous temperate fish species. Twenty-four groups of juveniles (initial weight 57.6 (sd 14.4) g) were intensively reared for 116 d and fed sixteen different isoproteic diets. The distribution of the experimental treatments was based on a multivariate fractional factorial design (L24212) with either high (+1) or low (-1) level of each of the following factors: diet (lipid and protein sources, lipid content, astaxanthin enrichment), feeding level, daily and weekly distribution frequency, fish density, initial weight heterogeneity, temperature, photoperiod, and light spectrum. Liver lipid droplets, glycogen, mitochondria and rough endoplasmic reticulum (RER) were semi-quantified and analysed by a soft imaging system using transmission electronic microscopy photographs. Important variability of hepatocyte ultrastructural features was observed. The present study confirms that the rearing temperature, through its influences in the general metabolic activity, seems to be the main factor modifying mainly lipid droplet accumulation and RER development. However, factors that could be pooled under the designation of factors leading to food accessibility and lipid and protein quality intensify or compensate the effect of temperature. © The Authors 2008.