The aminoglycoside hypersensitive mutant of Klebsiella pneumoniae (Kp-126) was isolated and used for specific detection of new aminoglycoside-type antibiotics. From the fermentation broth of a butirosin-producing strain, 3,3'-neotrehalosadiamine (BMY-28251) was detected as a minor component specifically active against Kp-126. This communication describes the isolation and identification of BMY-28251 from Bacillus circulans YQW-B6. The strain was fermented in medium FR8, and the broth filtrate was processed via Amberlite IRC-50 adsorption, NH4OH elution, and concentration to obtain a total active fraction. Further purification using Amberlite CG-50 column chromatography yielded component X (BMY-28251), which showed high activity against Kp-126 but weak activity against Bacillus subtilis PCI 219. Component X gave positive ninhydrin and anthrone reactions (negative Tollens reaction), had [α]27D +33° (c 1.0, H2O), end UV absorption, a carbohydrate-type IR spectrum, and NMR signals including a doublet anomeric proton (5.35 ppm, J=3.5 Hz). By TLC comparison with an authentic sample and physicochemical analysis, component X was identified as 3,3'-diamino-3,3'-dideoxy-α,β-trehalose (BMY-28251), which was also found in Bacillus pumilus K169-B91 using Kp-126.