Background The phenotypic heterogeneity of circulating tumor cells (CTC) in peripheral blood and disseminated tumor cells (DTC) in bone marrow is an important constraint for clinical decision making. Here, we investigated the implications of two different subpopulations of these cells in gastric cancer (GC). Methods GC patients (n=228) who underwent elective gastric resections were prospectively examined for CTC/DTC. The cells obtained from peripheral blood and bone marrow aspirates were sorted by flow cytometry and CD45- cells expressing cytokeratins (8, 18, and 19) and CD44 were identified by immunofluorescent double staining. Results Ninety-three (41%) patients had cytokeratin-positive tumor cells in either blood or bone marrow, while cells expressing CD44 were found in 22 (10%) cases. CK+CD44+ cells were significantly more common among patients with distant metastases (50 vs 19%, P=0.001), while no such correlations were demonstrated for CK+CD44- cells. Detection of CK+CD44+ cells, but not CK+CD44-, was associated with significantly shortened survival. Moreover, the Cox proportional hazards model identified CK+CD44+ cells as a negative prognostic factor with an odds ratio of 2.38 (95% CI 1.28-4.41, P=0.006). Conclusion CD44+ phenotype of cytokeratin-positive cells in blood and bone marrow is an independent prognostic factor in patients with gastric cancer.