Abstract: Epinephelus malabaricus (Perciformes, Serranidae), a rare marine commercial fish, inhabits in the warm-water reef along the southeast coast of China. Chromosome specimens of grouper E. malabaricus, sampled from kidney cells in metaphase by vivo injection of phytohemagglutinin (PHA) and treatment with colchicine, from which well-scattered, clear, and longitudinal bands were obtained by restriction endonucleases (Alu I) demonstrated by in situ nick translations. Most chromosomes presented 8-10 bands, at least 4. The number and characteristics of the bands in the homologous chromosomes were stable, and the major features of the same serial chromosomes from different mitotic metaphase figures were identical. The chromosome specimens were then painted with human X and Y chromosome-specific DNA probes. Surprisingly, some homologous segments of human X chromosome were found in the E. malabaricus genome, and all the segments were scattered in three pairs (t7, t13, t22) in homologous grouper chromosomes. The nearest distance between the centromere and the fluorescent in situ hybridization (FISH) signal from the t7, t13 and t22, was 62.3%, 43.4% and 44.4%, respectively, and the homologous segment of human X chromosome was about 4.63% of the E. malabaricus genome. No homologous segment of the human Y chromosome was found. These may provide a new research approach for the evolution from lower vertebrates to human sex chromosomes.