Substrate specificities of Fusarium biosynthetic enzymes explain the genetic basis of a mixed chemotype producing both deoxynivalenol and nivalenol-type trichothecenes.

Fusarium species are traditionally grouped into type A and type B trichothecene producers based on structural differences in the mycotoxin they synthesize. The type B trichothecene-producing Fusarium graminearum strains are further divided into 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), and nivalenol (NIV) chemotypes. The former two chemotypes, collectively termed a deoxynivalenol (DON) chemotype, evolved from a NIV chemotype by inactivation of FgTri13, which encodes trichothecene C-4 hydroxylase, during the evolutionary process. Despite stable overexpression of FgTri13, however, both 3-acetylnivalenol (3-ANIV) and 3-ADON accumulate equally in shake flask culture of a transgenic 3-ADON chemotype. In this study, we investigated why the "3-ANIV chemotype" could not be obtained using this strategy. When analysis was extended to the transgenic NIV chemotype, in which FgTri7 C-4 acetylase gene was disrupted and FgTri8 deacetylase gene was replaced with the 3-ADON chemotype's orthologue, C-4 unoxygenated 3-ADON, as well as C-4 oxygenated 3-ANIV, accumulated as the end product. A feeding experiment with an ΔFgtri5ΔFgtri3 double gene disruptant, a trichothecene non-producing mutant unable to acetylate C-15 of the trichothecene ring, revealed the importance of the 15-O-acetyl group for efficient C-4 hydroxylation of DON-type trichothecenes. This implies that traditional DON and NIV chemotype diversification is not solely explained by FgTri13, but is also explained by the function of the FgTri8 trichothecene deacetylase gene. None of the crude cell extracts from existing chemotypes showed highly specific C-15 deacetylation activity against 3,15-diacetylnivalenol (3,15-diANIV) without deacetylating C-15 of the C-4 unoxygenated earlier intermediate, 3,15-diacetyldeoxynivalenol. Thus, an unnatural Fusarium trichothecene, 3-ANIV, could only be synthesized as part of a mixture with 3-ADON, unless the esterase encoded by FgTri8 evolves to act on the 15-O-acetyl of 3,15-diANIV with high specificity. We also explain why the transgenic "15-ANIV chemotype", which can be generated through functional inactivation of FgTri7, uses an engineered pathway via 3,15-diANIV, but not 15-ADON, to generate 15-ANIV. Tri genes appear to evolve continuously, and altered functions of trichothecene pathway enzymes result in the generation of new trichothecenes, such as NX-2 and NX-3, which have been recently discovered in field isolates of F. graminearum. As recombination of FgTri8 between existing F. graminearum isolates could give rise to a strain that produces mixtures of DON and NIV-type trichothecenes, it may also be noteworthy to monitor the emergence of a field isolate that invalidates traditional chemotype classification.

[Trial of signal-to-noise ratio adjustment by repetition time in magnetic resonance imaging].

The purpose of this study was to find an equation between signal-to-noise ratio (SNR) and repetition time (TR) in order to adjust SNR by changing TR in magnetic resonance imaging (MRI) examinations. Using a phantom for SNR measurement, according to NEMA MS 1-1988, measurement of SNR was performed by spin-echo pulse sequences for various TR values. An equation of SNR and scanning parameters including TR were obtained from these results. In order to determine the range of TR where the images showed contrast suitable for diagnosis, the contrast-to-noise ratio (CNR) was measured for various TR values. CNR measurement was performed by scanning a brain phantom, and CNR was defined as the contrast of white matter and gray matter divided by noise. Scanning of a resolution phantom was carried out with various scanning parameters, and the usefulness of the equation obtained was determined by whether or not pins in the phantom were visible. The reliability of the equation was confirmed from this verification. Results showed that TR can be used for the adjustment of SNR using the equation obtained in this study.

[Performance evaluation of X-ray CT using visible scintillation light].

We proposed a new method of performance evaluation for X-ray CT using visible scintillation light and examined its usefulness in this study. When we scanned a plastic scintillator disk in a gantry opening of the X-ray CT, we could observe visible scintillation light. The rotation of the light-emitting area of the disk corresponded to that of the X-ray tube. We were able to record the scintillation light by digital video camera. By analyzing the area of visible scintillation light, the rotation speed of the X-ray tube, angular spread of the X-ray beam, uniformity of the incident X-rays, and change in X-ray energy were measured. No other method is available to obtain the above parameters of X-ray CT during a single CT scan. In the measurements of the uniformity of incident X-rays and change of X-ray energy, our method showed good accuracy in detecting the attenuation caused by the couch between the X-ray tube and the plastic scintillator disc. The proposed method is inexpensive and easy-to-use. We conclude that the method is a useful tool for performance evaluation as well as a maintenance tool for X-ray CT.

Total synthesis of spicamycin.

The first total synthesis of one of the spicamycin congeners, SPM VIII (3), is described. A preliminary model study for construction of the characteristic N-glycoside linkage in spicamycin using tetra-O-benzyl-beta-D-mannopyranosylamine (13) and halopurines 5 revealed that Pd-catalyzed conditions successfully provided the coupling products 14 and 15 in good yields. It was also shown that thermal anomerization of the N-glycosides easily occurred, which resulted in the predominant formation of the beta-anomer as the thermodynamically favored compound, and the activation energy of anomerization of 15 was estimated to be ca. 30 kcal/mol. The novel aminoheptose unit of spicamycin 6 was prepared stereoselectively by carbon elongation of an acyclic aldehyde, prepared by ring cleavage reaction of a highly functionalized cyclohexane derived from naturally abundant myo-inositol. The Pd-catalyzed coupling reaction of the beta-heptopyranosylamine 6 with protected 6-chloropurine 5d, followed by deprotection, provided spicamycin amino nucleoside 2, whose condensation with dodecanoylglycine completed the total synthesis of 3. This study confirmed the proposed unique structure of a novel nucleoside antibiotic.