In vitro activity of <Emphasis Type="Italic">Penicillium chrysogenum</Emphasis> antifungal protein (PAF) and its combination with fluconazole against different dermatophytes

Strains of five dermatophyte species (Microsporum canis, Microsporum gypseum, Trichophyton mentagrophytes, Trichophyton rubrum and Trichophyton tonsurans) were selected for testing against Penicillium chrysogenum antifungal protein (PAF) and its combination with fluconazole (FCZ). Inhibition of microconidia germination and growth was detected with MICs of PAF ranging from 1.56 to 200 mug ml(-1) when it was used alone, or at constant concentration (100 mug ml(-1)) in combination with FCZ at from 0.25 to 32 mug ml(-1). The MICs for FCZ were found to be between 0.25 and 128 mug ml(-1). PAF caused a fungicidal effect at 200 mug ml(-1) and reduced growth at between 50 and 200 mug ml(-1). Total growth inhibition with fungistatic activity was detected at 64 mug ml(-1) of FCZ for M. gypseum, T. mentagrophytes, and T. tonsurans, and at 32 mug ml(-1) FCZ for M. canis and T. rubrum. PAF and FCZ acted synergistically and/or additively on all of the tested fungi except M. gypseum, where no interactions were detected.     

A gene-based genetic linkage map of the collared flycatcher (Ficedula albicollis) reveals extensive synteny and gene-order conservation during 100 million years of avian evolution

By taking advantage of a recently developed reference marker set for avian genome analysis we have constructed a gene-based genetic map of the collared flycatcher, an important "ecological model" for studies of life-history evolution, sexual selection, speciation, and quantitative genetics. A pedigree of 322 birds from a natural population was genotyped for 384 single nucleotide polymorphisms (SNPs) from 170 protein-coding genes and 71 microsatellites. Altogether, 147 gene markers and 64 microsatellites form 33 linkage groups with a total genetic distance of 1787 cM. Male recombination rates are, on average, 22% higher than female rates (total distance 1982 vs. 1627 cM). The ability to anchor the collared flycatcher map with the chicken genome via the gene-based SNPs revealed an extraordinary degree of both synteny and gene-order conservation during avian evolution. The great majority of chicken chromosomes correspond to a single linkage group in collared flycatchers, with only a few cases of inter- and intrachromosomal rearrangements. The rate of chromosomal diversification, fissions/fusions, and inversions combined is thus considerably lower in birds (0.05/MY) than in mammals (0.6-2.0/MY). A dearth of repeat elements, known to promote chromosomal breakage, in avian genomes may contribute to their stability. The degree of genome stability is likely to have important consequences for general evolutionary patterns and may explain, for example, the comparatively slow rate by which genetic incompatibility among lineages of birds evolves.     

Reliable and rapid discrimination of congeneric species by mtDNA SNP analysis by multiplex PCR: application on three Trachurus and two Mullus fish species as model cases

We present a reliable, time-saving, and cost-effective multiplex PCR assay for discriminating congeneric species. Three fish species of the genus Trachurus and two of the genus Mullus served as model cases. Our multiplex PCR method interrogates species-specific diagnostic mitochondrial single nucleotide polymorphisms (mtSNPs). We selected two sets of mtSNPs that are organized in two multiplex systems for the Trachurus and the Mullus species, respectively. In both systems, all individuals tested could be safely assigned to one of the three Trachurus or two Mullus species. This novel SNP typing system offers a convenient and robust DNA profiling system suitable for large-scale identification of commercial fish species, for species with cryptic larvae or during juvenile stages, as well as for wildlife forensics. Handling editor: Christian Sturmbauer     

Periodically interrupted amperometry. A way of improving analytical performance of membrane coated electrodes

Amperometry is a powerful voltammetric measuring method. Its application is specially advantageous when used in combination with a separation step or with some other sample treatment method providing selectivity. The selectivity is often achieved by coating the amperometric working electrode surface with a membrane of special character. Size exclusion membrane, immobilized enzyme containing reaction layer, protecting dialysis membrane, perm selective ion exchange film etc can be mentioned here. In conventional amperometry the measuring potential is continuously applied, therefore in case of membrane coated electrodes the electrode process depletes the diffusion layer. In this work the performance of a new periodically interrupted amperometric (PIA) measuring program has been investigated in case of glucose enzyme sensor. The measuring program allowing time for reloading the diffusion layer provided higher current and therefore improved sensitivity and lower limit of detection.     

Germline mutation in ATR in autosomal- dominant oropharyngeal cancer syndrome

ATR (ataxia telangiectasia and Rad3 related) is an essential regulator of genome integrity. It controls and coordinates DNA-replication origin firing, replication-fork stability, cell-cycle checkpoints, and DNA repair. Previously, autosomal-recessive loss-of-function mutations in ATR have been demonstrated in Seckel syndrome, a developmental disorder. Here, however, we report on a different kind of genetic disorder that is due to functionally compromised ATR activity, which translates into an autosomal-dominant inherited disease. The condition affects 24 individuals in a five-generation pedigree and comprises oropharyngeal cancer, skin telangiectases, and mild developmental anomalies of the hair, teeth, and nails. We mapped the disorder to a ～16.8 cM interval in chromosomal region 3q22-24, and by sequencing candidate genes, we found that ATR contained a heterozygous missense mutation (c.6431A>G [p.Gln2144Arg]) that segregated with the disease. The mutation occurs within the FAT (FRAP, ATM, and TRRAP) domain-which can activate p53-of ATR. The mutation did not lead to a reduction in ATR expression, but cultured fibroblasts showed lower p53 levels after activation of ATR with hydroxyurea than did normal control fibroblasts. Moreover, loss of heterozygosity for the ATR locus was noted in oropharyngeal-tumor tissue. Collectively, the clinicopathological and molecular findings point to a cancer syndrome and provide evidence implicating a germline mutation in ATR and susceptibility to malignancy in humans.     

The abc1-/coq8- respiratory-deficient mutant of Schizosaccharomyces pombe suffers from glutathione underproduction and hyperaccumulates Cd2+

The abc1 ⁻ /coq8 ⁻ gene deletion respiratory-deficient mutant NBp17 of fission yeast Schizosaccharomyces pombe displayed a phenotypic fermentation pattern with enhanced production of glycerol and acetate, and also possessed oxidative stress-sensitive phenotypes to H₂O₂, menadione, tBuOOH, Cd²⁺, and chromate in comparison with its parental respiratory-competent strain HNT. As a consequence of internal stress-inducing mutation, adaptation processes to restore the redox homeostasis of mutant NBp17 cells were detected in minimal glucose medium. Mutant NBp17 produced significantly increased amounts of O₂^•− and H₂O₂ as a result of the decreased internal glutathione concentration and the only slightly increased glutathione reductase activity. The Cr(VI) reduction capacity and hence the ^•OH production ability were decreased. The mutant cells demonstrated increased specific activities of superoxide dismutases and glutathione reductase (but not catalase) to detoxify at least partially the overproduction of reactive oxygen species. All these features may be explained by the decreased redox capacity of the mutant cells. Most notably, mutant NBp17 hyperaccumulated yellow CdS.     

VEGF and IL-18 in induced sputum of lung cancer patients

Cytokines are key players in the biological processes of malignant tumors and special interest has been focused on cytokines exerting tumor and anti-tumor properties, such as vascular endothelial growth factor (VEGF) and Interleukin-18 (IL-18). Aim of this study was to assess IL-18 and VEGF levels in induced sputum of lung cancer patients at diagnosis, and assess their possible association with the histological type of cancer, the stage and the overall patient survival. Seventy six patients with a diagnosis of lung cancer were recruited and were followed up for 48months. Thirteen healthy smokers and 16 healthy non-smokers were used as control groups. VEGF and IL-18 were measured by ELISA in sputum supernatants at the time of diagnosis. Lung cancer patients had significantly higher baseline IL-18 and VEGF levels compared to healthy controls (p<0.001). No difference was found in IL-18 and VEGF levels between the various stages in non-small cell lung cancer (NSCLC) and between limited and extended small cell lung cancer (SCLC). However, the ratio of VEGF/IL-18 was significantly higher in NSCLC compared to SCLC patients (p=0.018). In extended SCLC overall survival was inversely associated with baseline sputum VEGF levels (p=0.034) and estimated mortality risk was 1.14 (95% CI 1.006-1.283) for an increase of 100pg/ml in VEGF levels. Such association was not found regarding baseline IL-18 levels. VEGF levels in induced sputum may have a prognostic role in the survival of SCLC. The ratio VEGF/IL-18 in induced sputum differs between NSCLC and SCLC, indicating differences in angiogenesis mechanisms and/or immunological response in these two major histological types of lung cancer.     

Thermodynamic and kinetic processes during the unfolding of BSA in the presence of the mycotoxin patulin

The effects of the mycotoxin patulin on the thermodynamics and kinetics of the transition of bovine serum albumin (BSA) in aqueous solution were studied by Differential Scanning Calorimetry and Photoluminescence methods. Results show that in the presence of patulin, the free enthalpy change during the transition of BSA was decreased by an average of ～ 46 kJ/mol, the free energy change was decreased by ～ 4 kJ/mol, and the activation energy fell from ～ 1546 to ～ 840 kJ/mol. These results indicate that the bioactivity of patulin is based on the kinetic rather than on the thermodynamic properties of the transition. This is the first evidence of the direct interaction of patulin with the free thiol-containing BSA, a process which could contribute to the adverse cyto- and genotoxic effects induced by patulin.