Identification of Escherichia coli nitrate reductase as an antigen for a monoclonal antibody with previously unknown specificity

The immunoaffinity chromatography of total membrane proteins from Escherichia coli helped determine the specificity of the monoclonal antibody 3A6 that was obtained upon immunization of mice with nicotinamide nucleotide transhydrogenase preparations and reacted with an unknown E. coli antigen. Proteins with apparent molecular masses of 150, 45, and 20 kDa were isolated and identified by N-terminal sequencing as the subunits of nitrate reductase. This conclusion was confirmed by immunoblotting with the 3A6 antibody of the proteins from the E. coli cells grown upon induction of nitrate reductase. It was shown that the 3A6 antibody specifically recognizes the alpha subunit of nitrate reductase, and the formation of the enzyme-antibody complex does not result in a loss of the enzyme catalytic activity.     

Multicentre study of comparative efficacy of meropenem and combined regimens for empirical antibacterial therapy of severe nosocomial infections: results of clinical and pharmacoeconomic analysis]

Adequacy and effectiveness of empirical antibacterial therapy of severe nosocomial infections with meropenem vs. combined regimens of antibacterial therapy were investigated and the ratio of the cost and effectiveness of the compared regimens was evaluated. A prospective, randomized, open, comparative study of two initiative regimens of empirical antibacterial therapy of severe nosocomial infections was performed: meropenem in a daily dose of 1.5-3 g and the standard regimen with the use of betalactams and fluoroquinolones in combination with aminoglycosides and/or metronidazole. Patients with recorded diagnosis of nosocomial pneumonia (including the ventilator-associated one) or abdominal infection with the signs of severe sepsis and severity of APACHE II > 14 were enrolled. The patients were stratified into 2 groups subject to the disease severity, i.e. APACHE II 15-20 and APACHE II 21-25. One hundred thirty five out of 166 patients with recorded nosocomial infection were included into the final estimate of the therapy adequacy and effectiveness (Protocol Analysis): 62 patients were treated with meropenem and in the treatment of 73 patients the standard antibacterial therapy was used. In the group of the patients treated with meropenem there were stated significantly higher clinical effectiveness (recovery in 80.6% of the patients vs. the control of 46.6%, p < 0.01) and pathogen eradication (89.6 and 48.1% respectively, p < 0.01). The difference in the clinical and bacteriological effectiveness of meropenem and the standard therapy was more evident in the subgroups of more severe patients (APACHE > 20). With the use of meropenem the probability of recovery from nosocomial infection was significantly higher (RR 1.73-1.94, p < 0.001) vs. the control. Meropenem provided significantly higher eradication of the pathogens: P. aeruginosa (88 and 40% respectively, p = 0.007), E. coli (100 and 46.7%, p = 0.003), Acinetobacter spp. (90.9 and 40%, p = 0.02). The antibacterial therapy with the use of meropenem was assessed as adequate in 51 out of 56 patients (91.1%), that was 3 times as frequent as with the use of the standard antibacterial therapy (33.9%). The cost-effectiveness coefficient with the use of meropenem was 2.2 times lower vs. the control. Therefore, the empirical therapy of severe nosocomial infections with meropenem proved to be more adequate and from the economic viewpoint more advantageous vs. the standard combined regimens of antibacterial therapy, that was evident from significantly higher clinical and bacteriological efficacy of the treatment and decrease of the terms of the patients hospitalization in intensive care units (on the average by 5 days).     

The First Finding of the Actinia Synandwakia hozawai in the Sea of Okhotsk

The finding of the actinia Synandwakia hozawai in the coastal waters of northern Sakhalin (Sea of Okhotsk) suggests a wider range of this species, which was previously only known to inhabit the coastal waters of eastern Japan. Data are presented on the morphology of the S. hozawai specimen from the Sea of Okhotsk and the types of its nematocysts.     

Sequence analysis and expression profiling of 14-3-3 genes from the extremophile <Emphasis Type="Italic">Thelungiella salsuginea</Emphasis>, ecotype Yakutsk

Members of the 14-3-3 protein family are known to be important regulators of plant primary metabolism, hormonal signal transduction, and ion homeostasis. We identified nine isoforms of 14-3-3 genes of Thellungiella salsuginea, an extremophile relative of Arabidopsis thaliana. All the identified isoforms were designated according to their Arabidopsis orthologs: Chi, Omega, Psi, Phi, Upsilon, Lambda, Mu, Epsilon, and Omicron. Comparison of the deduced amino acid sequences reveals high degree of identity between the members of this protein family. Isoforms, designated as Ts14-3-3 Chi, Omicron, and Mu, display noticeable differences in their C-terminal domain as compared to their Arabidopsis homologs. Phylogenetic analysis demonstrated that the identified isoforms split into two groups, epsilon and non-epsilon, according to the common classification of the 14-3-3 family genes. The Thellungiella 14-3-3 isoforms are differentially expressed in various plant tissues, and real-time RT-PCR revealed that most of the isoforms are highly expressed even under normal growth conditions. In response to abiotic stress, low temperatures and high concentrations of salts, 14-3-3 genes exhibited different expression patterns. Our data suggest that, due to the high expression levels of the 14-3-3 genes, Thellungiella plants are likely pre-adapted to the stress conditions. Differences between the C-terminal domains of some Thellungiella 14-3-3 proteins and their Arabidopsis homologs may result in differences in target protein specificity.     

Functional properties of smooth muscle cells in ascending aortic aneurysm

Thoracic aortic aneurysm (TAA) develops as a result of complex sequential events that dynamically alter the structure and composition of the aortic vascular extracellular matrix (ECM). The main cellular elements that alter the composition of aortic wall are smooth muscle cells (SMCs). The purpose of the present work was to study alterations of smooth muscle cell functions derived from the patients with TAA and from healthy donors. Since it is believed that TAA associates with bicuspid aortic valve (BAV) and with tricuspid aortic valve (TAV) differed in their pathogenesis, we have compared SMCs and tissue samples from BAV and TAV patients and healthy donors. The comparison was done by several parameters: SMC growth, migration and apoptotic dynamics, metalloproteinase MMP2 and MMP9 activity (zymography), and elastin, collagen, and fibrillin content (Western blot) in both tissue samples and cultured SMCs. Proliferation of BAV and TAV SMCs was decreased and migration ability in scratch tests was increased in TAV-derived SMCs compared to donor cells. BAV-cells migration ability was not changed compared to donor SMCs. Elastin content was decreased in TAA SMCs, whereas the content of fibrillin and collagen was not altered. At the same time, the elastin and collagen protein level was significantly higher in tissue samples of TAA patients than in donorderived samples. SMC proliferation and migration is differently affected in TAV and BAV-associated TAA that supports the idea on different nature of these two TAA groups. Our data also show that SMC functional properties are altered in TAA patients and these alterations could play a significant role in the disease pathogenesis.     

An oligonucleotide microarray for detection and discrimination of orthopoxviruses based on oligonucleotide sequences of two viral genes

An oligonucleotide microarray for detection and identification of orthopoxviruses was developed. Genus specific and orthopoxvirus species-specific regions of the genes encoding chemokine binding and alpha/beta-interferon binding proteins were used as a target. The developed microarray allows the variola, monkeypox, cowpox, vaccinia, camel-pox and ectromelia (mousepox) viruses to be distinguished with a high degree of reliability.