Fusarium head blight (FHB) severity was evaluated in a set of recombinant inbred (RI) lines from a cross between the two-rowed barley varieties, Harbin 2-row (resistant) and Turkey 6 (susceptible). Harbin 2-row was one of the most resistant varieties to FHB among ca. 5,000 barley germplasm accessions stored at Okayama University. The FHB severity of the RI lines and their parents was evaluated by the ‘cut-spike test’ in the 2000–2001 and 2003–2004 seasons. Spikes with disease symptoms were scored based on eleven grades from resistant (0) to susceptible (10). A high-density linkage map, including 328 AFLP, 45 SSR, one RFLP-STS, one AFLP-STS, four resistance gene analog (RGA) and four morphological markers, was constructed to detect QTLs for FHB severity. Composite interval mapping enabled to detect one QTL on chromosome 2H in the 2000–2001 season, and one QTL on chromosome 4H and another QTL on chromosome 6H in the 2003–2004 season. At these three QTLs, the alleles from Harbin 2-row contributed to a lower severity. A QTL on chromosome 2H was located in the vicinity of the cleistogamy locus (cly1 or Cly2) that determines open/closed flowering type. The comparison with previously identified QTLs for FHB severity by the present authors indicated that Harbin 2-row might harbor the same resistance allele in the vicinity of the cly1/Cly2 locus as Russia 6, which was used as a resistant parent in the previous study. The QTL in the vicinity of the cly1/Cly2 locus is one of the promising sources for FHB resistance breeding in barley.