The most sensitive screening technique for genes that predispose patients for particular cancers is direct sequencing. However, sequencing of complex genes is technically demanding, costly and time-consuming. We have tested alternate screening techniques to find a fast sensitive method for detecting alterations of DNA in the large BRCA1 gene prior to sequencing. Sequencing of this gene is particularly arduous because it lacks clearly defined mutation sites. The single-strand conformation polymorphism (SSCP) technique is one of the most frequently used pre-screening methods but its sensitivity and efficiency is not completely satisfying. We have compared the SSCP assay with a newly developed technique called denaturing high performance liquid chromatography (DHPLC) to screen the BRCAl gene. We studied 23 patients at high risk for early onset breast and ovarian cancer and four controls. In these patients, a total of 113 fragments with sequence variations in the BRCA1 gene could be identified. The DHPLC technique resolved 100% of the DNA alterations that were observed in cycle sequencing. In contrast, mutation analysis by SSCP accounted for 94% of the detected variations. In addition, DHPLC screening allowed us to discriminate between different alterations in a single fragment, because of the characteristic elution profiles of the DNA molecules. Polymorphisms that were present in our samples could be predicted by means of DHPLC testing independently of sequence analysis. We conclude that DHPLC is a highly potent screening method for genetic analyses. It is highly sensitive, efficient and economical and can be automated.

Breast Neoplasms↗

PubMedDate not supplied

Synthesis of fluorochromes and pigments in Malassezia furfur by use of tryptophan as the single nitrogen source.

A new minimal medium consisting only of L-tryptophan (L-Trp) and a lipid source induced formation of brown pigmentation only in the species Malassezia furfur, which diffuses into the agar. Strains of the species M. sympodialis and M. pachydermatis failed to grow on this medium. On mDixon medium, however, after replacement of peptone by L-Trp, growth of all three Malassezia species was achieved. Under these conditions pigment production was observed with all M. furfur strains tested, although the results for M. pachydermatis strains were inconsistent. M. sympodialis strains showed no pigment production. On the minimal medium pigmentogenesis was induced in M. furfur by only 0.01 g% tryptophan; the pH optimum was pH 5. In all M. furfur strains, alternative amino nitrogen sources given concurrently with Trp suppressed pigmentogenesis. Furthermore, there were differences in the optimal temperature among the individual M. furfur strains. CBS 7019, CBS 6000 and CBS 6001 failed to produce pigment at 37 degrees C. The extract of the culture exhibited remarkable fluorescence, and several indole derivatives with a broad spectrum of colours were detected. This finding may have an impact on the clinical appearance of pityriasis versicolor, a very common skin disease caused by lipophilic yeasts of the genus Malassezia. We hypothesize that in pityriasis versicolor metabolic adaptation of Malassezia yeasts to altered nitrogen conditions on superficial skin might be of patho-physiological importance. Tryptophan as an inducer of pigmentogenesis probably accumulates during excessive sweating, a well-known manifestation of pityriasis versicolor.

Fluorescent Dyes↗

PubMedDate not supplied

Limited joint mobility in the diabetic foot: relationship to neuropathic ulceration.

Limited joint mobility in the hand is a common manifestation of diabetes with the reported prevalence in insulin-dependent diabetes varying between 8 and 43%. Sixty-two subjects were studied in three groups (controls, diabetic patients without foot problems, and diabetic patients with neuropathic ulceration) to determine whether similar changes occur in the joints of the foot and to examine any possible relationship with neuropathic ulceration. There was a significant impairment of mobility in the range of motion of the sub-talar joint in diabetic patients with ulcers when compared with controls (p = 0.0001) or with the other diabetic patients (p = 0.004). There was a significant correlation between sub-talar range of motion and mobility in other joints of the foot such as at the hallux (r = 0.59, p less than 0.001), or with mobility of the 5th finger (r = 0.41, p less than 0.01). There was also a significant association between the clinical presence of limited joint mobility in the hand, Dupuytren's contracture, and mobility of the sub-talar joint (p less than 0.05). Furthermore, impairment of mobility of the sub-talar joint was greatest on the affected side in those diabetic patients with neuropathic ulceration (p = 0.029). We conclude that the syndrome of limited joint mobility also affects the joints of the feet of diabetic patients and may predispose to ulceration in susceptible neuropathic feet.

Ankle Joint↗

PubMedDate not supplied

Metabolites of 2-aminophenol from fruit bodies of Lepiota americana (Agaricales).

From the acetone extract of the North American toadstool Lepiota americana 2-aminophenoxazin-3-one (1) and a novel amino-1,4-benzoquinone derivative, lepiotaquinone (2), were isolated. The structure of 2 was confirmed by its preparation from 2-aminophenol and amino-1,4-benzoquinone.

Agaricales↗