[Some immunological changes in children with bacterial infections treated with bacteriophages].

The aim of the study was to reveal the possible immunological changes in children with bacterial infections treated with commercial bacteriophage preparations administered per os. In case of medical indications (for treatment or diagnostic) blood sampling was carried out. In serum the antibodies against bacteriophage preparations - phage cocktail components (phages against Staphylococcus, Streptococcus, Proteus vulgaris, Escherichia coli, Pseudomonas aeruginosa) were investigated. The neutralisation reaction was used. There were processed samples from 65 children with following diagnoses: sepsis, bacterial pneumonia, urinary tract infection, bacterial infections of upper respiratory ways, bacterial diarrhea. In samples taken in the first days of treatment antibodies were revealed in infants up to one month (I group) in 0/29 cases - 0%, in infants aged from one month till one year (II group)- 1/25 - 4.0%, in children aged from 1 till 15 years (III group) - 3/9 - 33.3%; data after 14-20 days from the beginning of treatment - I group - 0/9 - 0%, II group - 4/15 - 26.7%, III group - 5/5 - 100%; data after 30-60 days from the beginning of the treatment - I group - 1/5 - 20.0%, II group - 6/10 - 60.0%, III group - 3/3 - 100%. Bacteriophages neitralisation degree varied between 50,7% and 97.3%. Any regularity regarding different components of used phage preparations was not established. In case of inclusion of commercial phage preparations administered per os in the treatment of bacterial infections in children, the anti-phage neutralizing antibodies are produced by the macroorganism. This fact limits the duration of phage therapy and its usage in the treatment of future bacterial infections in treated patients. Production of anti-phage antibodies in young infants is substantially less expressed and this indicates to purposefulness and presumably higher efficacy of bacteriophage therapy in this age period.

[What happens when the child gets bacteriophage per os?].

The aim of the work was to define the distribution of phages administered per os to children for medical reasons, and the immune response. 102 children aged from 5 days to 15 years with different diseases of bacterial etiology (pneumonia, sepsis, urinary infection, pharyngitis/sinusitis, enteral infection) were monitored. Pyobacteriophage was being included into the complex therapy. The drug was administered per os. In 6/7 of blood, 48/55 urine and 64/75 stool samples taken on the 3-5th day of treatment different components of pyobacteriophage were revealed. The titers varied from 103 to 105 pfu/ml. No age differences were seen. In two weeks after the onset of the phagotherapy the antibodies to phages were tested in the blood serum using the neutralization reaction method. The blood samples were taken from 31 patients. In 14 of them the antibodies neutralizing 52.5-97.3% of the phage activity were seen. A significant age-related peculiarity was determined: in newborns and infants the antibodies were not revealed or their activity was low. Obtained results confirm the reasonability to use of peroral phagotherapy in gastro-intestinal infections. At the same time it was ascertained that the phages taken per os can permeate into the internal environment of the organism and thus the peroral phagotherapy can be used to treat systemic infections and urinary tract infections as well. Absence or low production of the antiphage antibodies in newborns and infants suggests high efficacy of the phagotherapy in this age group.

Study of therapeutic potential of the experimental pseudomonas bacteriophage preparation.

Wide expansion of the infections caused by multi-antibiotic resistant strains of P. aeruginosa revived the idea of phage therapy with pseudomonas phage preparations for treatment and prevention of the bacterial infection diseases. The purpose of this study was examination of an experimental series of the therapeutic-prophylactic pseudomonas bacteriophage preparation, with wide spectra of lytical activity and high therapeutic potential. Newly isolated phage clones of P. aeruginosa were studied by the basic tests (such as host range, lysis stability, physiological and immunogenic properties of the phages, host dependent restriction/modification phenomena and automatic reproduction ability of the phages on the UV inactivated strains) determining their virulent nature. An experimental series of the therapeutic-prophylactic pseudomonas phage preparation were developed from the genuine virulent phage clones CF1/1; CF1/7; P.a.N1, P.a.N2 and P.a.N4. The phage preparation successfully passed in vitro (efficacy, sterility, stability) and in vivo (safety, definition of therapeutic potential) controls on experimental animals (white mice). The host range of the experimental pseudomonas phage preparation equals 99.5% of 206 strains of P. aeruginosa. Preclinical testing of the experimental pseudomonas phage preparation on white mice revealed that the therapeutic efficacy of the phage preparation was higher (80-100%) than that of the antibiotic-ciprinol (50-80%). Noteworthy, 100% therapeutic efficacy was observed after combined application of the antibiotic and the phage preparation.

Evolution of T4-related phages.

Much progress has been made in understanding T-even phage biology in the last 50 years. We now know the entire sequence of T4, encoding nearly 300 genes, only 69 of which have been shown to be essential under standard laboratory conditions; no specific function is yet known for about 140 of them. The origin of most phage genes is unclear, and only 42 genes in T4 have significant similarity to anything currently included in GenBank. Comparative analysis of related phages is now being used to gain insight into both the evolutionary origins and interrelationships of these phage genes, and the functions of their protein products. The genomes of phages isolated from Tbilisi hospitals, Long Island sewage plants, the Denver zoo, and Khabarovsk show basic similarity. However, these phages show substantial insertions and deletions in a number of regions relative to each other, and closer investigation of specific sequences often reveals much more complex relationships. There are only a few cases in T4-related phages in which there is evidence for evolution through DNA duplication. These include the fibrous products of genes 12, 34, and 37; head proteins gp23 and gp24; and the Alt enzyme and its downstream neighbors. T4 also contains 13 apparent relatives of group I and group II intron homing endonucleases. Distal portions of the tail fibers of various T-even phages contain segments closely related to tail-fiber regions of other DNA coliphages, such as Mu, P1, P2, and lambda. Horizontal gene transfer clearly emerges as a major factor in the evolution of at least the tail-fiber regions, where site-specific recombination probably is involved in the exchange of host-range determinants.

Host-controlled modification and restriction as a criterion of evaluating the therapeutical potential of Pseudomonas phage.

The recently isolated phages phi ST3 and phi ST1 were compared as to their lysis behaviour in about 100 different P. aeruginosa strains. The growth of phi ST3 varies greatly in different host strains. We demonstrated one case of "non-classical", host-dependent modification and restriction. Here the capability to adsorb, and consequently to reproduce in a given host strain differs, depending on which modification the phage acquired in its former host. The DNA-containing phage phi ST1 displays stable lysis properties in the majority of the host strains. This makes phi ST1 a candidate for therapeutic phage preparations. One of the reasons for stable lysis properties is the apparent selection against recognition sites of restriction enzymes in its genome.

[Serological characterization of bacteriophage T3 carrying different "non-classical" modifications]. / Serologicheskaia kharakteristika bakteriofagov T3 s razlichnymi neklassicheskimi modifikatsiiami.

The antigenic properties of bacteriophage T3 and a mutant T3/R7 which undergoes "non-classical" modification and restriction are compared. The "non-classical" modification of T3/R7 consists of a host-dependent, reversible change in the adsorption capacity of phage on different host strains. We have shown that this modification is connected with changes in the antigenic properties of phage components involved in phage absorption to the host cell. This means that, in contrast to the "classical" host-control-led modification and restriction of DNA, the "non-classical" modification and restriction of phage is based on protein modification.

[Investigation of serological features and molecular heterogeneity of virion proteins of morphologically related phages]. / Issledovanie serologischeskikh svoistv i molekuliarnoi geterogennosti belkov virionov morfologicheski rodstvennykh bakteriofagov.

Some interaction process peculiarites of morphologically related phate T4, DDVI and FI1 with the corresponding antiphageous sera in cross reactions of neutralisation have been investigated. Close serological relationship of phage T4 and DDVI was demonstrated. Inactivation of phage FI1 in these serological reactions showed anomalous nature and did not submit to the rules of the first step reaction. According to investigations of the protein component of virions of these phages by electrophoresis in polyacrylamide gel, phages T4 and DDVI contain twelve, FI1 and DDVI four, and T4 and FI1 two protein fractions common in molecular weight. The data obtained indicates that close serological relationship between phages T4 and DDVI is determined by the presence of common protein antigens in the absorptional apparatus of virions. Proteins of phage FI1, common with the proteins of T4 and DDVI, are obviously located in other structural components, and this determines the specific aggregation of virions in cross reactions of neutralization.

[Electron microscope study of antigen constituents of the phage DDVI and its h-mutant]. / Zlektronno-mikroskopicheskoe izuchenie antigennykh komponentov bakteriofaga DDVIh+ i ego h-mutanta.

A comparative study of the fine antigen structure of phage DDVI and its h mutant using neutralization reaction and the electron microscopy of the phage-antibody complex has shown that the head and the tail of these phages have common protein constituents. The bulk of the antigens located in tail's fibers and in a base plates is also identical. However, the application of the cross-adsorbed sera has shown that the phage DDVI and the h mutant differ by only one specific antigen located in the distal part of tail's fibers.

Electron-microscopic study of the serological affinity between the antigenic components of phages T4 and DDVI.

In an investigation of the antigenic fine structure of phages T4 and DDVI with the use of the neutralization reaction and electron-microscopic observation of the phage-antibody complexes, it has been possible to establish that the head of phage T4 consists of proteins which have antigenic determinants of two types: The first type is identical to the antigens of the head of phage DDVI, and the second type is apparently absent in phage DDVI. The phage DDVI head contains mostly determinants which are common to the phage T4 head, since it was not possible to detect antigenically specific components in the phage DDVI head. The tail sheaths of phage T4 and DDVI appear to be identical in the antigenic respect. A difference has been observed in the fibers and the base plates of the phages investigated. The presence of the following three types of antigens has been established: 1) common to phages T2, T4, and DDVI, 2) common to phages T4 and DDVI, and 3) specific for each phage investigated.