As 16S rRNA sequence analysis has proven inadequate for the differentiation of Bacillus cereus from closely related species, we employed the gyrase B gene (gyrB) as a molecular diagnostic marker. The gyrB genes ofB. cereus JCM 2152T,Bacillus thuringiensis IAM 12077T,Bacillus mycoides ATCC 6462T, andBacillus anthracis Pasteur #2H were cloned and sequenced. Oligonucleotide PCR primer sets were designed from withingyrB sequences of the respective bacteria for the specific amplification and differentiation of B. cereus, B. thuringiensis, andB. anthracis. The results from the amplification of gyrB sequences correlated well with results obtained with the 16S rDNA-based hybridization study but not with the results of their phenotypic characterization. Some of the reference strains of both B. cereus (three serovars) and B. thuringiensis (two serovars) were not positive in PCR amplification assays with gyrB primers. However, complete sequencing of 1.2-kb gyrB fragments of these reference strains showed that these serovars had, in fact, lower homology than their originally designated species. We developed and tested a procedure for the specific detection of the target organism in boiled rice that entailed 15 h of preenrichment followed by PCR amplification of the B. cereus-specific fragment. This method enabled us to detect an initial inoculum of 0.24 CFU of B. cereus cells per g of boiled rice food homogenate without extracting DNA. However, a simple two-step filtration step is required to remove PCR inhibitory substances.

A cationic fluorescent nanogel thermometer based on thermo-responsive N-isopropylacrylamide and environment-sensitive benzothiadiazole was developed with a new azo compound bearing imidazolium rings as the first cationic radical initiator. This cationic fluorescent nanogel thermometer showed an excellent ability to enter live mammalian cells in a short incubation period (10 min), a high sensitivity to temperature variations in live cells (temperature resolution of 0.02-0.84 °C in the range 20-40 °C), and remarkable non-cytotoxicity, which permitted ordinary cell proliferation and even differentiation of primary cultured cells.

To determine the clinical relevance of in vitro drug chemoresistance in childhood acute myeloid leukemia, we used an MTT assay to test leukemic cells from 132 newly diagnosed children. Patients were diagnosed according to the French-AmericanBritish (FAB) classification as follows: M0 (n = 12), M1 (n = 16), M2 (n = 53), M4 (n = 17), M5 (n = 19) and M7 (n = 15). The results revealed that, compared to leukemic cells from completeresponders (n = 107), those from non-responders who failed induction therapy (n = 17) were 1.4 to 5.0 times more resistant in vitro to cytarabine (P = 0.005), melphalan (P = 0.003), etoposide (P = 0.011), L-asparaginase (P = 0.017), aclarubicin (P = 0.026) and dexamethasone (P = 0.039). For seven other drugs tested, the median lethal dose of 70% and leukemic cell survival of non-responders were higher than those of complete-responders, but the difference was not statistically significant. We sought correlations between FAB subtypes and in vitro drug resistance. Leukemias of the FAB M4 and M5 subtype were more sensitive to L-asparaginase (P = 0.01, P = 0.0036) than those of the FAB M2 subtype. FAB M5 leukemia was more sensitive to etoposide than were the FAB M2, M4 and M7 subtypes (P = 0.001, P = 0.034, P = 0.023, respectively). By contrast, FAB M5 leukemia was significantly more resistant to prednisolone and dexamethasone than were the FAB M0, M1, M2, M4 and M7 subtypes. We sought correlations between in vitro drug resistance and long-term clinical outcome, but found no associations in this case. These results suggest that in vitro resistance to cytarabine, melphalan, etoposide, L-asparaginase, aclarubicin and dexamethasone might represent factors that can predict response to the early course of therapy. Selecting an appropriate anti-cancer drug according to the FAB classification together with drug sensitivity testing may contribute to improved prognoses in childhood acute myeloid leukemia. Leukemia (2001Leukemia ( ) 15, 1892Leukemia ( -1897

Six antitumor antibiotics of a new structure class, indoxamycins A-F (1-6), were isolated from a saline culture group of marine-derived actinomyces whose strains showed approximately 96% sequence homology of 16S rDNA with the family streptomycetaceae. The structures of these indoxamycins, which are unusual polyketides composed of six consecutive chiral centers, were assigned by combined spectral and chemical methods. In feeding experiments using a stable isotope label, indoxamycin A was assembled from propionate units initially forming the "aglycon" pentamethyl indeno furan. The discovery of these unprecedented compounds from marine-derived actinomycetes, a low gene homology genus, offers a significant opportunity for drug discovery.

Summary. To explore the potential efficacy of l-asparaginase treatment in acute myeloid leukaemia (AML) patients, we studied the in vitro resistance of French-AmericanBritish (FAB) subtypes of childhood AML to l-asparaginase using a methyl-thiazol-tetrazolium assay. We tested leukaemic cells obtained from 177 common acute lymphoblastic leukaemia (cALL) and 228 AML children at diagnosis. The median 70% lethal dose of l-asparaginase (LD70 asp ) (U/ml) was 0AE46 in the cALL and 6AE70 in the AML samples. The median LD70 asp among each FAB subtype of AML was 0AE76 (M0), 0AE46 (M1), 10AE00 (M2), 10AE00 (M3), 1AE18 (M4), 1AE35 (M5) and 10AE00 (M7). Type M3 samples had the highest LD70 asp . The LD70 asp of the M2 samples was significantly higher than that of the M1, M4 and M5 samples. When the LD70 asp values were classified as low (0AE016-0AE159), intermediate (0AE16-1AE59) or high (1AE6-10AE00), the frequency of low, intermediate or high LD70 asp among the M1 samples were similar to those among the cALL samples. In conclusion, cells from AML types M1, M4 and M5 were relatively sensitive to l-asparaginase, and M1 cells were as sensitive as those of cALL, suggesting that l-asparaginase treatment may be effective for these subtypes of AML.