Thirteen DDBs were isolated from every enrichment culture using the R2A agar

or 100-fold-diluted NA plates. Gram staining revealed that nine strains were Gram-positive and four were Gram-negative. The bacterial 16S rRNA genes were analysed and the results are summarized in Table 1. Phylogenetic analysis was performed by constructing neighbour-joining trees. As shown in Fig. 2a, the Gram-positive strains (SS1, SS2, SS3, SS4, LS1, LS2, YMN1, YUL1, PFS1) were closely related to the genus Nocardioides in the family Nocardioidaceae, forming four clusters. Levels of 16S rRNA gene sequence similarity ranged from 92% to 100%. The Gram-negative strains (SS5, RS1, NKK1, NKJ1) were closely related to the genus Devosia in the family Hyphomicrobiaceae, forming two clusters, and their 16S rRNA gene sequence similarities ranged from 95% to 100%. HM781-36B research buy The initial DON degradation rates using the washed cells of the strains preincubated LY294002 mw with DMM, 1/3LB and 1/3R2A were examined (Table 1). All of the strains preincubated with DMM showed DON-degrading activities, and degraded 100 μg mL−1 of DON

to below the detection limit (0.5 μg mL−1) after the 24 h of incubation. Among the strains, SS5 and RS1 showed high rates of DON degradation, which were more than three times those of the other strains. Although strains NKK1 and NKJ1 were closely related to strains SS5 and RS1, the degradation rates were lower. Strains SS5, RS1 and NKJ1 expressed DON-degrading activities regardless of the preincubation media used. Preincubation with 1/3LB enhanced the DON-degrading activities of strains SS5 and RS1, but repressed that of NKK1. These results provided insight into the diversity of DON-degradation phenotypes within closely related strains. Meanwhile,

all of the Gram-positive strains exhibited high DON-degrading activities by preincubation with DMM, although they exhibited Metalloexopeptidase very low activities by preincubation with 1/3R2A or 1/3LB. That the buffer with autoclaved cells did not decrease the concentration of DON and that the buffer filtrates during DON degradation also did not (data not shown) indicate that the decrease of DON is attributed to the enzymatic reactions catalysed in the living cells. Figure 3a and b show the time course of DON degradation, and HPLC elution profiles of DON and its metabolites in washed cells of representative strains LS1, SS5 and these autoclaved strains. The profiles of the two strains showed at least three peaks in addition to the DON peak (6.5 min); one peak corresponded to the peak in the authentic standards of 3-epi-DON (4.5 min), indicating that both strains produced 3-epi-DON. The HPLC elution profiles also revealed unidentified peaks at 3.0 and 6.9 min in the RS1 sample, and at 1.6 and 4.8 min in the LS1 sample. These peaks were not detected when DDBs were autoclaved or were incubated without DON (Fig. 3c), indicating that these peaks were the products derived from DON.