An Ustilago maydis Mutant Partially Blocked in P45014DM Activity Is Hypersensitive to Azole Fungicides

An Ustilago maydis ergosterol biosynthesis mutant (A14) which is partially blocked in sterol 14alpha-demethylase (P45014DM) activity is described. This mutant accumulated the abnormal 14alpha-methyl sterols, eburicol, 14alpha-methylfecosterol, and obtusifoliol, along with significant amounts of ergosterol. Although the A14 mutant grew nearly as well as the wild type, it was impaired in cell extension growth, which indicated a dysfunction in apical cell wall synthesis. The mutant was also found to be hypersensitive to the azole fungicides penconazole and tebuconazole.     

Limits to Catalysis by Ribonuclease A

Bovine pancreatic ribonuclease A (RNase A) catalyzes the cleavage of the P-O(5') bond in RNA. Although this enzyme has been the object of much landmark work in bioorganic chemistry, the nature of its rate-limiting transition state and its catalytic rate enhancement had been unknown. Here, the value of k(cat)/K(m) for the cleavage of UpA by wild-type RNase A was found to be inversely related to the concentration of added glycerol. In contrast, the values of k(cat)/K(m) for the cleavage of UpA by a sluggish mutant of RNase A and the cleavage of the poor substrate UpOC(6)H(4)-p-NO(2) by wild-type RNase A were found to be independent of glycerol concentration. Yet, UpA cleavage by the wild-type and mutant enzymes was found to have the same dependence on sucrose concentration, indicating that catalysis of UpA cleavage by RNase A is limited by desolvation. The rate of UpA cleavage by RNase A is maximal at pH 6.0, where k(cat) = 1.4 × 10(3) s(-1) and k(cat)/K(m) = 2.3 × 10(6) M(-1)s(-1) at 25°C. At pH 6.0 and 25°C, the uncatalyzed rate of [5,6-(3)H]Up[3,5,8-(3)H]A cleavage was found to be k(uncat) = 5 × 10(-9) s(-1) (t(1/2) = 4 years). Thus, RNase A enhances the rate of UpA cleavage by 3 × 10(11)-fold by binding to the transition state for P-O(5') bond cleavage with a dissociation constant of <2 × 10(-15) M.     

Evidence of absence of Trypanosoma cruzi kinetoplast DNA methylation by restriction endonuclease analysis.

Eight kinetoplast DNA samples from very different T. cruzi zymodemes were digested with the isoschizomer group of enzymes (MspI-HpaII) and (MboI-Sau 3AI), able to detect DNA methylation on cytidine and adenine for the CCGG and GATC sequences, respectively. Restriction digestion analysis of each kDNA with both isoschizomer groups of enzymes did not display a different profile suggesting that maxicircles and minicircles on this trypanosomatid are not methylated.     

Culture and growth of Lessonia trabeculata (Phaeophyta,Laminariales) juvenile sporophytes in La Herradura de Guayacan Bay,northern Chile

Lessonia trabeculata is one of the major kelps found along the northern coast of Chile. In addition to its ecological and economic importance, L. trabeculata may be severely affected by environmental disturbances such as El Níño, which during 1982–1983 cleared wide areas along the coast of Peru and Chile. The main goal of this work was to mass culture L. trabeculata and to observe the growth of sporophytes obtained in the laboratory and cultured in the sea. Juvenile sporophytes obtained in the laboratory were attached between 1 and 6 m in depth. The linear growth rate, as blade elongation, was recorded weekly for seven months. No significant differences (p < 0.05) were found in sporophyte blade linear growth at different depths. The best elongation growth rate was 7.5 ± 1.6 mm d^–1 at 3 m during March. This preliminary work suggests that L. trabeculata follows an annual growth cycle similar to that of other Laminariales with a high rate of blade elongation during the summer and decreasing towards autumn. This species can be considered a potential candidate for aquaculture to increase the availability of raw material and aid in repopulation of overexploited areas.     

Effects of flavonoids on tongue squamous cell carcinoma

Squamous cell carcinoma (SCC) of the tongue is associated with tobacco use, alcohol abuse, and human papillomavirus (HPV) infections. While clinical outcomes have recently improved for HPV‐positive patients in general, 50% of patients suffering from tongue cancer die within 5 years of being diagnosed. Flavonoids are secondary plant metabolites with a wide range of biological activities including antioxidant, anti‐inflammatory, and anticancer activities. Flavonoids have generated high interest as therapeutic agents owing to their low toxicity and their effects on a large variety of cancer cell types. In this literature review, we evaluate the actions of flavonoids on SCC of the tongue demonstrated in both in vivo and in vitro models.     

Hexazonium pararosaniline as a fixative for animal tissues

Hexazonium pararosaniline is a valuable reagent that has been used in enzyme activity histochemistry for 50 years. It is an aqueous solution containing the tris-diazonium ion derived from pararosaniline, an aminotriarylmethane dye, and it contains an excess of nitrous acid that was not consumed in the diazotization reaction. Other investigators have found that immersion for 2 min in an acidic (pH 3.5) 0.0015 M hexazonium pararosaniline solution can protect cryostat sections of unfixed animal tissues from the deleterious effects of aqueous reagents such as buffered solutions used in immunohistochemistry, while preserving specific affinities for antibodies. In the present investigation hexazonium pararosaniline protected lymphoid tissue and striated muscle against the damaging effects of water or saline. The same protection was conferred on unfixed sections treated with dilute nitrous or hydrochloric acid in concentrations similar to those in hexazonium pararosaniline solutions. Model tissues (solutions, gels or films containing gelatin and/or bovine albumin) responded predictably to well known cross-linking (formaldehyde) or coagulant (mercuric chloride) fixatives. Hexazonium pararosaniline solutions prevented the dissolution of protein gels in water only after 9 or more days of contact, during which time considerable swelling occurred. It is concluded that there is no evidence for a “fixative” action of hexazonium pararosaniline. The protective effect on frozen sections of unfixed tissue is attributable probably to the low pH of the solution.     

Efficacy of a Vaccine Formula against Tuberculosis in Cattle

“Test-and-slaughter” has been successful in industrialized countries to control and eradicate tuberculosis from cattle; however, this strategy is too expensive for developing nations, where the prevalence is especially high. Vaccination with the Calmette-Guérin (BCG) strain has been shown to protect against the development of lesions in vaccinated animals: mouse, cattle and wildlife species. In this study, the immune response and the pathology of vaccinated (BCG-prime and BCG prime-CFP-boosted) and unvaccinated (controls) calves were evaluated under experimental settings. A 10⁶ CFU dose of the BCG strain was inoculated subcutaneously on the neck to two groups of ten animas each. Thirty days after vaccination, one of the vaccinated groups was boosted with an M. bovis culture filtrate protein (CFP). Three months after vaccination, the three groups of animals were challenged with 5×10⁵ CFU via intranasal by aerosol with a field strain of M. bovis. The immune response was monitored throughout the study. Protection was assessed based on immune response (IFN-g release) prechallenge, presence of visible lesions in lymph nodes and lungs at slaughter, and presence of bacilli in lymph nodes and lung samples in histological analysis. Vaccinated cattle, either with the BCG alone or with BCG and boosted with CFP showed higher IFN-g response, fewer lesions, and fewer bacilli per lesion than unvaccinated controls after challenge. Animals with low levels of IFN-g postvaccine-prechallenge showed more lesions than animals with high levels. Results from this study support the argument that vaccination could be incorporated into control programs to reduce the incidence of TB in cattle in countries with high prevalence.