Obtaining antisera to Pseudomonas aeruginosa and Proteus antigens for performing immunoenzyme analyses in clinical practice

The data presented in this work indicate that specific antisera to P. aeruginosa and Proteus antigens can be produced by using extracts from these microorganisms, destroyed by ultrasonic treatment or by multiple freezing and thawing, for the immunization of rabbits. Blood serum samples from patients with purulent septic complications were studied for the presence of P. aeruginosa and Proteus antigens in ELISA with the use of peroxidase-labeled antibodies from antisera to P. aeruginosa and Proteus. This investigation revealed that during the first 3 days from the beginning of the clinical manifestations of the complications P. aeruginosa and Proteus antigens were detected in 86.4% and 83.4% of the patients, respectively. In the subsequent bacteriological study of wound discharge from these patients the corresponding microflora was detected.     

Isolation and study of the properties of a polyvalent corpuscular Pseudomonas aeruginosa vaccine. An evaluation of the biological properties of a polyvalent corpuscular Pseudomonas aeruginosa vaccine

P. aeruginosa killed polyvalent corpuscular vaccine, tested in a trial on 42 volunteer donors, is safe, low reactogenic and possesses pronounced immunogenicity, as the injection of the vaccine induced a rise in the titer of antibodies to P. aeruginosa up to 1:1280 in 95% of the immunized donors. To obtain specific antibodies in the blood plasma of the donors in an amount sufficient for the protective activity of the plasma becoming manifest, it is expedient to use the vaccination schedule providing for subcutaneous injections of 0.5, 0.5 and 1.0 ml at intervals of 7 days. Intense immunity thus induced in the donors lasts for 3-4 months. The hyperimmune plasma obtained from the donors has an antibody titer of at least 1:320 and shows a 90-100% protective effect in mice infected intraperitoneally with P. aeruginosa. The preliminary results of the clinical trial of anti-P. aeruginosa plasma have demonstrated its efficiency as a part of the complex treatment of patients with purulent septic complications of P. aeruginosa etiology.     

Use of a polyvalent erythrocyte diagnostic agent for determining antibodies to Pseudomonas aeruginosa in clinical practice

The use of polyvalent erythrocyte diagnosticum prepared on the basis of 5 polysaccharide antigens of P. aeruginosa slime, isolated from strains belonging to the most widespread serovars, makes it possible to check up the humoral response of donors after their immunization with P. aeruginosa polyvalent corpuscular vaccine with the aim of obtaining anti-P. aeruginosa donor plasma. Antibody titers, determined in the passive hemagglutination test with the use of the proposed diagnosticum and corresponding to a serum dilution of 1:320 and greater, age tentatively diagnostic, which may be indicative of P. aeruginosa in the development of purulent septic complications in patients. The use of the passive hemagglutination test with the newly developed polyvalent erythrocyte diagnosticum makes it possible to check up the specific response of patients having P. aeruginosa infection in the process of their treatment with anti-P. aeruginosa hyperimmune plasma used as a part of complex therapy.     

Pseudomonas aeruginosa and Proteus antigenemia and antibody formation in mono- and mixed infections in patients with suppurative inflammatory diseases

P. aeruginosa and Proteus antigenemia and antibody production have been studied in 335 patients with purulent inflammatory diseases. The study has revealed that in the association of P. aeruginosa and Proteus with staphylococci and representatives of the family Enterobacteriaceae the level of antigenemia is considerably lower than in monoinfections or in the association of these microorganisms with streptococci. In mixed infections humoral immune response develops later than in cases of monoinfection. An important role of ecological and physiological relationships between microorganisms in the course of purulent processes and in their influence on the host has been confirmed. The use of the enzyme immunoassay in the clinical practice has made it possible to determine the etiological role of P. aeruginosa and Proteus in the development of suppurative-inflammatory diseases, to select adequate immunotherapy, including the use of anti-P. aeruginosa and anti-Proteus plasmas, and to evaluate the effectiveness of treatment.     

Effectiveness of gentamicin sulfate in suppurative-inflammatory processes of varying localization]

Antibiotic sensitivity of 486 strains of grampositive and gramnegative organisms isolated from patients with purulent infections was studied in vitro. Gentamicin was shown to be highly active as compared to kanamycin and other antibiotics against the main causative agents of purulent inflammatory infections including multiresistant E. coli, Proteus, Ps. aeruginosa, Staphylococcus. High efficiency of gentamicin in therapy of peritonitis, septic conditions, purulent postoperative wounds, infections of the urinary tract, pneumonia, etc. (197 patients) was shown. Positive results were obtained in 87.4 per cent of the cases. Side effects, such as albuminuria, hyperthermic reaction, rash, pruritis were registered in 3 per cent of the patients.     

The antibacterial immunity of people under dynamic observation in an altered radiation situation]

The comparative study of the isolation rate, level, antigenic and class specificity of serum antibodies to the causative agents of purulent septic infections (Staphylococcus aureus, Pseudomonas aeruginosa, Proteus mirabilis) and acute enteric infections in healthy adults with different ABO blood groups before (836 persons) and after (1,429 persons) the catastrophe at the Chernobyl nuclear power station was made. The study revealed the fact that the genesis of antibodies directed against different microorganisms can be stimulated without additional antigenic challenge in the form of disease or immunization, which was definitely indicative of the influence of small radiation doses in Kiev on the humoral immunity of the population. The multifactor character of the dependence of antibacterial antibody formation under altered radiation conditions on the specific features of the infective agent and the intensity of its circulation among the population, individual immune responsiveness of the body and concrete radiation conditions was established.     

Antibiotic sensitivity of the main opportunistic pathogenic aerobic bacteria

Antibiotic sensitivity of 1421 strains of Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Proteus mirabilis, Proteus vulgaris, Pseudomonas aeruginosa and Klebsiella spp. was studied. Gentamicin, levomycetin (chloramphenicol) and ristomycin proved to be the antibiotics of choice in treatment of purulent inflammatory diseases caused by S. epidermidis and S. aureus. For antibiotic therapy of infections caused by gram-negative organisms gentamicin and polymixin might be recommended.     

Mode of the rational use of Pseudomonas aeruginosa bacteriophages in therapeutic and epidemic control practice

Ecological aspects of the circulation of P. aeruginosa and P. aeruginosa bacteriophages under hospital conditions were under study. The statement concerning the formation of triple parasitic systems was put forward. The influence of these systems on the formation of phage and antibiotic resistance in P. aeruginosa hospital strains was studied. Spontaneous circulation of faintly virulent phages taking part in the formation of triple parasitic systems was shown not to ensure the elimination of P. aeruginosa hospital strains in clinics. Construction of highly virulent phages adapted to local P. aeruginosa strains was the only way of ensuring the protection of patients. Theoretical and practical approaches to the use of highly active bacteriophages for controlling P. aeruginosa infection were substantiated. The realization of these approaches resulted in achieving not only a clinical, but also essential epidemic control effect in cases of purulent septic infections caused by P. aeruginosa (a decreased frequentcy of hospital infections from 40.8% to 8.93%).     

Prolonged antibacterial action of polymer coated suture materials]

The antimicrobial activity of capromed, a surgical polymer-coated sutural material containing dioxidine, quinoxidine or gentamicin was studied in vitro and in vivo. Capromed was shown to be active against the hospital strains of S. aureus, E. coli, Proteus spp., Klebsiella spp. and P. aeruginosa. The antimicrobial activity of ADH and AG threads was preserved for 3 to 4 days after implantation. The DH-2 and G-2 threads preserved their activity for 6 to 7 days. It was concluded that the duration of thread antimicrobial activity depended on the properties of the polymer coating the thread. Capromed was applied to 280 operation wounds in 275 patients. There was no wound suppuration in the group of patients after pure operations (n = 62). In the group of patients after conditionally pure operations (n = 130) suppuration was observed in 2 patients (1.3 per cent). In patients with contaminated wounds (n = 88) suppuration in 5 of them (5.7 per cent) was recorded. The total number of the purulent complications after using capromed in surgical operations amounted to 2.5 per cent. In the control group purulent complications were stated in 8.2 per cent of the cases.     

Rifampicin in the treatment of infections of non-tuberculous etiology

Clinical efficacy of rifampicin, a semisynthetic broad spectrum antibiotic was estimated in 247 patients with purulent inflammations. It was shown advisable to use rifampicin intravenously in treatment of severe bronchopulmonary pathology, disorders of the bile excretion system, osteomyelitis, severe wound infections and in prophylaxis of postoperative purulent complications in cardiovascular surgery and other cases. High rifampicin sensitivity of staphylococci and streptococci belonging to various species was revealed. Rifampicin was found to be less active against gramnegative pathogens. The isolation frequency of rifampicin sensitive strains of E. coli, Proteus spp., Klebsiella spp. and P. aeruginosa amounted to 88.4, 52.1, 58.8 and 49.3 per cent respectively.     

The effect of an immune antibacterial plasma and a donor leukocyte mass on the immune system of patients with suppurative-septic processes

The influence of immune antistaphylococcal, anti-Proteus and anti-Pseudomonas aeruginosa plasma and donor leukocyte mass on some humoral and cell-mediated immunity characteristics in patients with purulent and septic diseases has been studied. The study has shown that treatment with immune plasma leads to an increase in the amount of circulating T lymphocytes and in the concentration of IgA, IgM and IgG, to the activation of phagocytosis and to an increase in the titers of the corresponding antimicrobial antibodies simultaneously with a decrease in the content of bacterial antigens in the blood serum. Treatment with donor leukocyte mass has also been found to lead to an increase in the concentration of T lymphocytes and immunoglobulins, to enhance the functional activity of phagocytizing neutrophils and to promote the normalization of the content of leukocytes in the peripheral blood. The use of these preparations as stimulating agents in the treatment of patients having purulent septic diseases and, simultaneously, low cell-mediated immunity characteristics is recommended.     

Bacteriological monitoring of infective agents of purulent septic diseases in an emergency hospital

During the period of 1995-2001 bacteriological examination of 13,842 patients with purulent septic diseases (PSD) was carried out. The statistical processing of data revealed that equalization of the dynamic rows of intensive and extensive values characterizing the number of patients with PSD caused by etiologically important infective agents made it possible to evaluate the reliability of information. A trend to increased etiological importance of the genera Pseudomonas, Escherichia and Klebsiella was established. The number of diseases caused by Staphylococcus, Streptococcus, Enterococcus and Proteus decreased.     

Pseudomonas aeruginosa septic arthritis of knee after intra-articular ozone injection

We describe a case of septic arthritis caused by Pseudomonas aeruginosa in an immunocompetent patient following intra-articular ozone injection into the knee. To the best of our knowledge, and after considering the current literature,we believe this case is unique as no other reports of septic arthritis caused by P. aeruginosa following intra-articular ozone injection has been made.     

Experimental study of photodynamic effect on bacterial wound microflora

The in vitro study of the influence of photodynamic action (with the use of photosensitizer "Photosens", laser and non-laser irradiation) on the strains of the main representatives purulent wound microflora was carried out. Staphylococcus aureus, S. epidermidis, Pseudomonas aeruginosa, Escherichia coli, Proteus mirabilis cultures have been isolated from wound secretions and identified. The photosensitizer was shown to produce no bactericidal effect by itself. Irradiation with laser and non-laser light sources induced a sharp decrease in the number of viable cells in the cultures under study. The irradiation of the photosensitizer led to its activation, manifested in bactericidal action. The results thus obtained confirm good prospect of using photodynamic therapy for the treatment of purulent wounds.     

Human anti-scrub typhus rickettsia and rabbit anti-Proteus antibodies recognize similar epitope in the O-polysaccharide part of Proteus mirabilis OXK lipopolysaccharide

In the Weil-Felix test, sera from patients infected with Orientia tsutsugamushi reacted with lipopolysaccharide (LPS) from Proteus mirabilis OXK strains. The O-polysaccharide of P. mirabilis OXK LPS consisted of pentasaccharide repeating units, with amidically-linked lysine residues. The lysine, linked to galacturonic residues, which plays an important role in the reaction with rabbit anti-OXK antibodies, was revealed with the aid of synthetic antigens. Using ELISA, immunoglobulin M antibodies from scrub typhus patients reacted with the O-specific polysaccharide of strain OXK LPS only. This reaction was inhibited by rabbit antibodies specific to the O-antigen of strain OXK LPS. Both human and rabbit antibodies may bind to similar epitopes on the O-polysaccharide part of P. mirabilis OXK LPS.     

Immunoenzyme analysis method for detecting anti-Pseudomonas aeruginosa antibodies in persons inoculated with pyoimmunogen

The possibility of using the indirect ELISA techniques for evaluating the level of the post-vaccinal production of humoral antibodies in donors immunized with Pyoimmunogen. P. aeruginosa vaccine, has been studied. The specificity and high resolution of this test system, based on the immobilization of the antigens of the vaccinal preparation on a solid-phase carrier, have been demonstrated. A rational method for the evaluation of specific antibody titers with due regard to the spectrophotometric data indicating the results of the reaction and the degree of the dilution of the serum under test has been proposed.     

Clinical effectiveness of carbenicillin in suppurative inflammatory processes of varying localization]

The study on sensitivity of clinical strains of the causative agents of purulent infections to carbenicillin showed that 34.6% of the staphylococcal strains, 48.1% of the E. coli strains and 40.3% of the Proteus strains were sensitive to the antibiotic. The strains of Ps. aeruginosa were characterized by moderate sensitivity to carbenicillin. The MTC for most of the isolates ranged within 25-128 microgram/ml. High therapeutic efficacy of carbenicillin in treatment of cases with purulent inflammatory processes of various localization was shown. Positive results were obtained in 82.5% of the adults and 76.2% of the premature infants treated with carbenicillin. A satisfactory therapeutic effect was observed in the cases with sepsis, diffuse purulent peritonitis and abscessing pneumonia treated with carbenicillin in combination with gentamicin.     

Characterization of epitope specificity of Proteus penneri 7 lipopolysaccharide core region

To extend the knowledge on the fragments of Proteus penneri lipopolysaccharide core regions, which determine the cross-reactions with specific antibodies, serological studies were performed by use of P. penneri 7 core-specific antiserum and Proteus sp. lipopolysaccharides. Different reactivity of the tested antiserum with three groups of antigens suggested differences in their core regions' epitope specificity. Comparing the results of the serological investigations with the previously determined structures of the core regions of the tested P. penneri lipopolysaccharides allowed distinguishing two potential tri- and tetrasaccharide epitopes and a third fragment which could not be determined precisely.     

Specific In Situ Visualization of Plasma Cells Producing Antibodies against Porphyromonas gingivalis in Gingival Radicular Cyst: Application of the Enzyme-Labeled Antigen Method

The enzyme-labeled antigen method was applied to visualize plasma cells producing antibodies to Porphyromonas gingivalis, flora of the human oral cavity. Antibodies to P. gingivalis have reportedly been detected in sera of patients with periodontitis. Biotinylated bacterial antigens, Ag53, and four gingipain domains (Arg-pro, Arg-hgp, Lys-pro, and Lys-hgp) were prepared by the cell-free protein synthesis system using the wheat germ extract. In paraformaldehyde-fixed frozen sections of rat lymph nodes experimentally immunized with Ag53-positive and Ag53-negative P. gingivalis, plasma cells were labeled with biotinylated Arg-hgp and Lys-hgp. Antibodies to Ag53 were detected only in the nodes immunized with Ag53-positive bacteria. In two of eight lesions of gingival radicular cyst with inflammatory infiltration, CD138-positive plasma cells in frozen sections were signalized for Arg-hgp and Lys-hgp. An absorption study using unlabeled antigens confirmed the specificity of staining. The AlphaScreen method identified the same-type antibodies in tissue extracts but not in sera. Antibodies to Ag53, Arg-pro, and Lys-pro were undetectable. In two cases, serum antibodies to Arg-hgp and Lys-hgp were AlphaScreen positive, whereas plasma cells were scarcely observed within the lesions. These findings indicate the validity of the enzyme-labeled antigen method. This is the very first application of this novel histochemical technique to human clinical samples.     

Antibacterial activity of rifampicin against the causative agents of suppurative, septic diseases]

The antibacterial activity of rifampicin was studied in comparison with other antibiotics with respect to clinical strains isolated from cases with various purulent inflammatory processes caused by Staphylococcus, E. coli, Ps. aeruginose, Proteus. The aim of the study was to define the role of rifampicin in the treatment of the above infections. No rifampicin resistant strains were found among staphylococci belonging to the phenotype carrying the determinants of resistance to 2-8 antibiotics. Rifampicin was less active against gramnegative organisms. High heterogeneity of the microbial population of rifampicin was shown with respect to all microbial strains tested. The rate of the spontaneous mutants was high. The average rate of the mutants was 1-7.7-10-8. The studies on the dynamics of the rifampicin resistance increase in the strains of Staphylococci, E. Coli, Ps. aeruginosa and Proteus showed that the resistance increased after 1-2 passages, which means that one-stage mutation was characteristic rifampicin.