[Photosensitizing properties of 3,3'-diethylthiacarbocyanine in biological media].

The objective of the study is elucidation of perspectives of 3,3'-diathylcarbocyaine application as a photosensitizer for curing viral infections by photodynamic therapy. Lipid-containing bacteriophage PM-2 of Pseudoalteromonas espejiana was used as a model. The testing was carried out at a special installation modeling photodynamic exposure conditions towards a non-fractionated phage lysate. 3,3'-DECC demonstrated a rapid photo-bleaching when added tothe phage lysate but not to water. The initial rate of PM-2 phage photoinactivation was proportional to the square concentration of the dye in the range of 0.5-9 µmol/L. This confirms a hypothesis that the dimer is the principal photochemically active form of the dye. An improved ability to form dimers was found in the dye in the phage lysate (10-folds better than in the water). The dye formed a stable adduct with the bacteriophage material. This adduct had an extinction maximum at λ(max) = 594 nm and demonstrated the properties of a polymer (sedimentation under a low-speed centrifugation).

[Application of the complex of DNA with the congo red anionic diazo dye for detection of nuclease-producing colonies of marine bacteria].

This study was aimed at the development of a method for detection of colonies of nuclease-secreting marine bacteria. The BAL nuclease-producing marine bacterium Pseudoalteromonas espejiana BAL-31 was used as the test object. A new method was developed involving the congo red (CR) anionic dye. The P. espejiana culture was plated on nutrient agar with CR and denatured DNA. In such media. CR was found to form complexes with DNA. After two days of incubation at 30 degrees C, halos were found around the P. espejiana colonies. No halos appeared when DNA was not introduced, when BAL nuclease was inactivated, or when the medium was inoculated with Escherichia coli. It was concluded that the halos around the colonies indicated nuclease excretion. The halos were shown to result from the coagulation of CR released after digestion of the CR-DNA complex by the nuclease. This method for detection of nuclease-producing colonies can probably be used for all marine bacteria and possibly for halophilic bacteria as well.

[A novel method of transfection of marine bacteria Pseudoalteromonas espejiana with deoxyribonucleic acid of bacteriophage PM2].

DNA of bacteriophage PM2 is a convenient test object for studying DNA-damaging genotoxic agents. The extent of DNA damage can be estimated by the ability of damaged DNA for transfection of host cells, marine bacterium Pseudoalteromonas espejiana (Pae), str. BAL-31. The efficiency of transfection of Pae lines maintained for long periods without freezing was found to be very low upon the use of a widely accepted transfection method developed by van der Schans et al. (1971). Such cultures grown in a medium with 10 mM Ca2+ standard for Pae contained cell aggregates and exopolymer material. Pae was found to be capable of growing in a medium without the calcium supplement in the presence of chelator EGTA (low-calcium medium, LCM). After growth in LCM, cells did not aggregate, cultures lacked the activity of nuclease BAL, and transfection efficiency of cells grown in LCM drastically increased. Based on these results, a novel procedure of transfection with an efficiency of 2 x 10(4)-2 x 10(5) infectious centers per microgram of PM2 DNA was developed.

[Spontaneous mutants of Alteromonas espejiana, resistant to kanamycin and bleomycin]. / Spontannye mutanty Alteromonas espejiana, rezistentnye k kanamitsinu i bleomitsinu.

An attempt was made to induce insertions in marine bacterium Alteromonas espejiana Bal-31 (Ae) using the TnphoA transposon. The Ae mutants selected on a kanamycin-containing medium after conjugation of the Ae with the transposon donor, Escherichia coli SM10(pRt291), were resistant not only to kanamycin (Kn), but also to bleomycin (Bm), and were sensitive to tetracycline. Although the mutants were phenotypically similar to insertion mutants, the mutations appeared to be spontaneous. The sensitivity of spontaneous Kmr Ae mutants selected at various Km concentrations to Bm was investigated. The mutants selected at low Km concentrations were resistant to Bm, whereas those selected at high Km concentrations were sensitive to Bm. The possible mechanisms underlying the dual resistance to Bm and aminoglycosides in bacteria are discussed.

[Loss of adsorptive ability--the reason for inactivation of the RM2 bacteriophage during storage]. / Poteria sposobnosti k adsorbtsii--prichina inaktivatsii bakteriofaga RM2 pri khranenii.

The kinetics of bacteriophage PM2 inactivation at storage was compared with the kinetics of bacteriophage adsorption on Alteromonas espejiana BAL-31 host cells. Adsorption ability and infectivity are lost with the same rate at temperatures 4-28 degrees C suggesting the loss of adsorption ability to result in bacteriophage inactivation. At higher temperatures infectivity is lost more rapidly than the ability of adsorption. The single hit kinetics of adsorption ability loss suggests the simple model of independent inactivation of 12 antireceptors located at the tops of icosaedric capsid to be erroneous. At bacteriophage inactivation the major port of protein I, a fragment of antireceptors, is preserved in the capsid composition.

[Inactivation of bacteriophage PM2 during storage]. / Inaktivatsiia bakteriofaga PM2 pri khranenii.

The kinetics of bacteriophage inactivation in the medium that is optimal for its storage has been studied at temperatures from 4 to 55 degrees C. The plot of Arrhenius dependence of the constant of inactivation rate consists of the two linear parts with the energies of activation Ea = 25 kcal/mol for 4-37 degrees C and Ea = 91 kcal/mol for 37-55 degrees C. The DNA of inactivated bacteriophage remained mostly in superspiralized form and completely preserved its biological activity as tested by transfection in spheroplasts. The analysis of inactivation kinetics suggests ageing of virions cultivated at 4 degrees C. The addition of watersoluble antioxidant amoxipin did not change the inactivation kinetics. The addition of antioxidant ionol with twin-80 increased the inactivation that was paralleled by the bacteriophage DNA degradation.

[Significance of van Merten's index in the diagnosis of ventilation-perfusion disorders and causes of respiratory insufficiency]. / O znachenii indeksa van Mertena v diagnostike ventiliatsionno-perfuzionnykh narushenii i prichin dykhatel'noi nedostatochnosti.

The ventilation-perfusion correlations were studied in 30 normal persons and in 60 patients with chronic heart and pulmonary diseases with the aid of van Marten's index (RCO2). It is shown that capnographic determination of the index is of paramount importance in the diagnosis of ventilation-perfusion disorders and of the causes of respiratory failure since the index rises only in patients with the clinical signs of obstructive ventilatory insufficiency.

[Electrophoresis of DNA fragments renatured and cross-linked with psoralen: kinetic analysis of DNA photocross-linking]. / Elektroforez sshitykh psoralenom i renaturirovannykh fragmentov DNK: kineticheskii analiz fotosshivaniia DNK.

The kinetics of photoreactions between DNA and psoralen was analysed. A method is described for determining the dependence of the average number of cross-links per base-pair (n) on the fluence of UV-irradiation. This calibration curve allows to specify the conditions of irradiation necessary to obtain any desired value of n.

[Use of psoralen for detecting the conjugated damage to 2 chains in irradiated phage PM2 DNA]. / Ispol'zovanie psoralen dlia obnaruzheniia sopriazhennykh povrezhdenii dvukh tsepei v obluchennoi DNK faga PM2.

Using psoralen for the photochemical cross-linking of DNA chains the authors have demonstrated the formation of conjugated lesions, of both opposite and non-opposite types, in X-irradiated superhelical DNA of PM2 phage. It is suggested that these lesions result from hitting a DNA molecule by a spur.

DNA replication and head assembly in bacteriophage T4.

Phage DNA was accumulated in cells of E. coli B, infected with the phage T4DtsLB3 (gene 42), without the synthesis of late proteins (in the presence of chloramphenicol). Then (stage II), chloramphenicol was removed and further replication of the phage DNA suppressed with hydroxyurea and by simultaneously raising the temperature to 40 degrees. The media M9 or M9 with 1% amino acid were used; the times of addition of chloramphenicol and the hydroxyurea concentration were also varied. It was also shown that in medium M9, at stage II, chiefly early proteins were synthesized. In the medium containing amino acids, at stage II the following was observed: 1) DNA synthesis was entirely suppressed and a degradation of DNA occurred; 2) both early and late proteins were synthesized, with a predominance of the latter; 3) an assembly of the elements of the phage tails and capsids occurred without the neck and flagellum, and a small number of phage particles were also found; 4) the capsids, isolated in a sucrose density gradient after lysis with chloroform, contained the proteins Palt, P20, P23, P24, several unidentified proteins, and did not contain Pwac, P23, and P22, 5) the yield of viable phage varied from 0.05 to 15% per cell. Thus, the entire morphogenesis of T4 phage can occur without accompanying replication of phage DNA.