Sensitivity of Vi phages III to gamma-radiation in the presence of cisplatin

In this study we determined Vi bacteriophage III sensitivity to native cisplatin, gamma radiation ((60)Co) or to irradiated cisplatin, and checked the possibility of enhanced Vi bacteriophage III inactivation under combined exposure to cisplatin and gamma radiation. We used highly purified phage suspensions in 0.9% NaCl solution or phosphate-buffered saline. Phage suspensions were titrated using a double agar layer method. Our study implies that survival of Vi bacteriophage III shows an exponential inverse correlation with cisplatin concentration in the incubation medium and the time of phage incubation in the presence of cisplatin. The use of irradiated cisplatin reduces phage survival in comparison with suspensions containing non-irradiated cisplatin. Irradiation of phage suspension with cisplatin causes a significant increase of phage inactivation in comparison with either treatment alone. Our results suggest that presence of cisplatin in irradiated medium enhances the radiobiological effect on Vi bacteriophages III.     

Yeast as a biosensor for antioxidants: simple growth tests employing a Saccharomyces cerevisiae mutant defective in superoxide dismutase

Mutants of Saccharomyces cerevisiae devoid of Cu,Zn-superoxide dismutase are hypersensitive to a range of oxidants, hyperbaric oxygen and hyperosmotic media, show lysine and methionine auxotrophy when grown under the atmosphere of air and have a shortened replicative life span when compared to the wild-type strain. Ascorbate and other antioxidants can ameliorate these defects, which may be a basis of simple tests sensing the presence of antioxidants. In particular, tests of growth on solid medium (colony formation) in the absence of methionine and/or lysine, or in the presence of 0.8 M NaCl can be useful for detection and semiquantitative estimation of compounds of antioxidant properties. Hypoxic atmosphere was found to increase the sensitivity of detection of antioxidants. The test of abolishment of lysine auxotrophy showed a concentration dependence of the antioxidant effects of cysteine and N-acetylcysteine which, however, lost their protective action at high concentration, in contrast to glutathione which was effective also at higher concentrations.     

Prentice's approach and the meta-analytic paradigm: a reflection on the role of statistics in the evaluation of surrogate endpoints

We put a perspective on the strengths and limitations of statistical methods for the evaluation of surrogate endpoints. Whereas using several trials overcomes some of the limitations of a single-trial framework (Prentice, 1989, Statistics in Medicine 8, 431-440), arguably the evaluation of surrogate endpoints can never be done using only statistical evidence but such evidence should be seen as but one component in a decision-making process that involves, among others, a number of clinical and biological considerations. We briefly present a hierarchical framework that incorporates ideas from Prentice's work and is uniformly applicable to different types of surrogate and true clinical outcomes.     

The O-acetylation patterns in the O-antigens of Hafnia alvei strains PCM 1200 and 1203, serologically closely related to PCM 1205

Serological tests revealed immunochemical similarities between the lipopolysaccharides of Hafnia alvei strains PCM 1200, 1203 and 1205. Immunoblotting and ELISA showed cross-reactions between the strains. NMR spectroscopy showed that the O-deacetylated O-specific polysaccharides isolated from lipopolysaccharides of H. alvei strains PCM 1200 and 1203 possessed the same composition and sequence as the O-deacetylated O-specific polysaccharide of H. alvei strain PCM 1205, that is a glycerol teichoic-acid-like polymer with a repeating unit of the following structure: [carbohydrate structure: see text] NMR spectroscopic studies of the polysaccharides concluded that O-3 of the side chain beta-D-GlcpNAc is partially O-acetylated (50-80%) in both investigated strains. In strain PCM 1203 an additional O-acetyl group (50-80%) is linked to O-6 of the chain -->3)-alpha-D-GlcpNAc-(1--> residue. The structural features of the isolated O-specific polysaccharides were also the same as those of the O-specific polysaccharides on the bacterial cells directly observed by the HR-MAS NMR technique.     

Effect of different anti-Abeta antibodies on Abeta fibrillogenesis as assessed by atomic force microscopy

Extensive data suggest that the conversion of the amyloid-beta (Abeta) peptide from soluble to insoluble forms is a key factor in the pathogenesis of Alzheimer's disease (AD). In recent years, atomic force microscopy (AFM) has provided useful insights into the physicochemical processes involving Abeta morphology, and it can now be used to explore factors that either inhibit or promote fibrillogenesis. We used ex situ AFM to explore the impact of anti-Abeta antibodies directed against different domains of Abeta on fibril formation. For the AFM studies, two monoclonal antibodies (m3D6 and m266.2) were incubated in solution with Abeta(1-42) with a molar ratio of 1:10 (antibody to Abeta) over several days. Fibril formation was analyzed quantitatively by determining the number of fibrils per microm(2) and by aggregate size analysis. m3D6, which is directed against an N-terminal domain of Abeta (amino acid residues 1-5) slowed down fibril formation. However, m266.2, which is directed against the central domain of Abeta (amino acid residues 13-28) appeared to completely prevent the formation of fibrils over the course of the experiment. Inhibition of fibril formation by both antibodies was also confirmed by thioflavin-T (ThT) fluorescence experiments carried out with Abeta(1-40) incubated for five days. However, unlike AFM results, ThT did not differentiate between the samples incubated with m3D6 versus m266.2. These results indicate that AFM can be not only reliably used to study the effect of different molecules on Abeta aggregation, but that it can provide additional information such as the role of epitope specificity of antibodies as potential inhibitors of fibril formation.     

Antibiotic susceptibility and occurrence of ESBL in Pseudomonas aeruginosa strains isolated from different clinical specimens

The aim of this study was to evaluate the drug susceptibility of 132 P. aeruginosa strains isolated from patients hospitalized in SPSK University Hospital in Bialystok. The isolates were obtained from clinical specimens over an 11-month period in 2001 and 2002. All the strains were identified in automatic ATB system using API 20 NE strips, and their susceptibility to antibiotics was tested by standard disc-diffusion method and agar dilution method. The minimal inhibitory concentration (MIC) was determined for five antibiotics: piperacillin, amikacin, ceftazidime, imipenem and ciprofloxacin. The majority of strains were susceptible to ceftazidime (91.7%), piperacillin combined with tazobactam (85.6%), amikacin (80.3%), meropenem and imipenem (81.8%). Many of our strains were resistant to cefotaxime (73.5%), ticarcillin (53%) and ciprofloxacin (48.5%). Also, the trial was undertaken to detect strains producing extended-spectrum beta-lactamases (ESBL) and inducible beta-lactamases (IBL) among P. aeruginosa rods isolated from different specimens. ESBL-producing strains were detected with double disc test (DDST) and combination double disc (CD) test. Clavulanate was applied as the inhibitor of these beta-lactamases. Strains producing ESBL were not found. On the other hand, as many as 127 P. aeruginosa strains (96.2%) produced inducible beta-lactamases (IBL).     

Differentiation of mitochondrial DNA and Y chromosomes in Russian populations

The genetic composition of the Russian population was investigated by analyzing both mitochondrial DNA (mtDNA) and Y-chromosome loci polymorphisms that allow for the different components of a population gene pool to be studied, depending on the mode of DNA marker inheritance. mtDNA sequence variation was examined by using hypervariable segment I (HVSI) sequencing and restriction analysis of the haplogroup-specific sites in 325 individuals representing 5 Russian populations from the European part of Russia. The Y-chromosome variation was investigated in 338 individuals from 8 Russian populations (including 5 populations analyzed for mtDNA variation) using 12 binary markers. For both uniparental systems most of the observed haplogroups fell into major West Eurasian haplogroups (97.9% and 99.7% for mtDNA and Y-chromosome haplogroups, respectively). Multidimensional scaling analysis based on pairwise F(ST) values between mtDNA HVSI sequences in Russians compared to other European populations revealed a considerable heterogeneity of Russian populations; populations from the southern and western parts of Russia are separated from eastern and northern populations. Meanwhile, the multidimensional scaling analysis based on Y-chromosome haplogroup F(ST) values demonstrates that the Russian gene pool is close to central-eastern European populations, with a much higher similarity to the Baltic and Finno-Ugric male pools from northern European Russia. This discrepancy in the depth of penetration of mtDNA and Y-chromosome lineages characteristic for the most southwestern Russian populations into the east and north of eastern Europe appears to indicate that Russian colonization of the northeastern territories might have been accomplished mainly by males rather than by females.     

Perturbations of malate accumulation and the endogenous rhythms of gas exchange in the Crassulacean acid metabolism plant<Emphasis Type="Italic"> Kalanchoë daigremontiana</Emphasis>: testing the tonoplast-as-oscillator model

In continuous light, leaves of the Crassulacean acid metabolism (CAM) plant Kalanchoë daigremontiana Hamet et Perrier exhibit a circadian rhythm of CO₂ uptake, stomatal conductance and leaf-internal CO₂ pressure. According to a current quantitative model of CAM, the pacemaking mechanism involves periodic turgor-related tension and relaxation of the tonoplast, which determines the direction of the net flux of malate between the vacuole and the cytoplasm. Cytoplasmic malate, in turn, through its inhibitory effect on phosphoenolpyruvate carboxylase, controls the rate of CO₂ uptake. According to this mechanism, when the accumulation of malate is disrupted by removing CO₂ from the ambient air, the induction of a phase delay with respect to an unperturbed control plant is expected. First, using the mathematical model, such phase delays were observed in numerical simulations of three scenarios of CO₂ removal: (i) starting at a trough of CO₂ uptake, lasting for about half a cycle (ca. 12 h in vivo); (ii) with the identical starting phase, but lasting for 1.5 cycles (ca. 36 h); and (iii) starting while CO₂ increases, lasting for half a cycle again. Applying the same protocols to leaves of K. daigremontiana in vivo did not induce the predicted phase shifts, i.e. after the end of the CO₂ removal the perturbed rhythm adopted nearly the same phase as that of the control plant. Second, when leaves were exposed to a nitrogen atmosphere for three nights prior to onset of continuous light to prevent malate accumulation, a small, 4-h phase advance was observed instead of a delay, again contrary to the model-based expectations. Hence, vacuolar malic acid accumulation is ruled out as the central pacemaking process. This observation is in line with our earlier suggestion [T.P. Wyka, U. Lüttge (2003) J Exp Bot 54:1471–1479] that in extended continuous light, CO₂ uptake switches gradually from a CAM-like to a C3-like mechanism, with oscillations of the two CO₂ uptake systems being tightly coordinated. It appears that the circadian rhythm of gas exchange in this CAM plant emerges from one or several devices that are capable of generating temporal information in a robust manner, i.e. they are protected from even severe metabolic perturbations.Abbreviations CAM Crassulacean acid metabolism - c_ia Ratio of mesophyll CO₂ concentration to external CO₂ concentration - J_C Rate of carbon dioxide uptake - J_W Transpiration rate - g_W Stomatal conductance - LL Continuous light conditions - PEPC Phosphoenolpyruvate carboxylase - Rubisco d-Ribulose-1,5-bisphosphatecarboxylase/oxygenase - Effective quantum yield of photosystem II