Investigations on the efficacy of monovalent Pseudomonas aeruginosa vaccine for oral administration in Pseudomonas aeruginosa experimental infection

Therapeutic efficacy of Pseudomonas aeruginosa vaccine for oral use (10(10) killed germs/ml), prepared from strain 4922, belonging to serotype XV, by Meitert-Meitert scheme, on 4 experimental models in mice (pneumonia, infected burn, septicaemia and urinary tract infection) was studied in comparison with monovalent Ps. aeruginosa vaccine serotype XV (10(9) killed germs/ml) for subcutaneous use and also with associated administration of the two vaccine variants. Mice immunization by using vaccine for oral use was performed by 0.5 ml vaccine per day, for 10 days and vaccine for subcutaneous use was administrated in a volume of 0.5 ml x 2, at 3 days interval. Mice immunization by using the two vaccine types, in association was concomitantly performed and in the same quantity as for separate immunization. In experimental pneumonia, Ps. aeruginosa vaccine for oral use protected mice in 35% of cases, those with infected burns were protected in 33.3% of cases, those with septicemia--in 96.6% of cases and those with urinary tract infection in 50% of cases. As compared to Ps. aeruginosa vaccine for subcutaneous use, the results obtained by vaccine for oral use are less favourable but associated administration of both vaccine variants led to superior results. Thus, in experimental pneumonia, it was obtained a surviving rate of 65% for animals immunized with both vaccine types, in comparison with 50% for animals immunized with vaccine for subcutaneous use only, and in Ps. aeruginosa infected burn, it was obtained a recovering rate of 79.1% for the animals immunized by using both vaccines, in comparison with 70.8% surviving for animals immunized with vaccine for subcutaneous use. In experimental septicaemia and urinary tract infection, combined use of both vaccine variants determined animals surviving and recovering in percents similar to those obtained by separate administration of vaccine for subcutaneous use (in septicemia--100% protection; in urinary tract infection--75% protection).     

Comparative study of the antibacterial activity of aminoglycoside antibiotics and their combinations with carbenicillin against Pseudomonas aeruginosa

Antibacterial activity of 7 aminoglycoside antibiotics and combinations of tobramycin or gentamicin with carbenicillin was studied with respect to 33 clinical strains of Ps. aeruginosa. Tobramycin, sisomicin, gentamicin and amicacin showed high levels of antibacterial activity. Tobramycin and sisomicin were 3-4 and 2 times more effective than gentamicin. 100 per cent of the Ps. aeruginosa isolates was sensitive to tobramycin and amicacin. The number of the isolates sensitive to sisomicin and gentamicin amounted to 97 and 94 per cent respectively. The respective numbers for streptomycin and kanamycin were 32 and 11 per cent. No monomycin sensitive isolates were detected. Combination of tobramycin or gentamicin with carbenicillin increased the antibacterial activity of the aminoglycoside antibiotics by 2-16 times and that of carbenicillin by 2-32 times. The synergistic effect of gentamicin or tobramycin with carbenicilin was observed with respect to 50 and 58 per cent of the isolates respectively. No antagonistic effect was detected on the combined use of the antibiotics. The majority of the isolates (96 per cent) were sensitive to combinations of carbenicillin in a concentration of 50 micrograms/ml with tobramycin or gentamicin in concentrations of 0.15 or 0.3 micrograms/ml respectively.     

Results of using polyvalent Pseudomonas aeruginosa vaccine in children with burns by various medical centers

Polyvalent Pseudomonas aeruginosa vaccine, prepared at the Institute of Hematology from 10 hospital strains isolated from burn wounds, was administered to 32 children with extensive and deep burns. The vaccine was well tolerated. The vaccine produced a high degree of the immunity against Pseudomonas aeruginosa infection. Agglutinin serum titre increased significantly. Vaccination either prevented or inhibited the infection of burn wounds with Pseudomonas aeruginosa in all immunized children. The symptoms of Pseudomonas aeruginosa infection usually disappeared following one or two vaccinations. Bacteriemia caused by P. aeruginosa was not observed in 31 out of 32 children. In the remaining child transient bacteriemia was noted. No septicemia caused by P. aeruginosa was seen. Due to the high efficiency of the polyvalent P. aeruginosa vaccine all burned children with burns exceeding 10% of the total body surface should by vaccinated to prevent the life-threatening infections with Pseudomonas aeruginosa.     

Experimental basis for the combined use of tobramycin and immune preparations for treating acute Pseudomonas aeruginosa infections

The possibility of using tobramycin (Tb) in combination with P. aeruginosa polyvalent corpuscular vaccine (PaPCV) or pyocyanosis hyperimmune plasma (PHP) for the treatment of P. aeruginosa sepsis was experimentally studied. The combined use of Tb and PHP, administered in amounts corresponding to ED50 of each preparation used separately, ensured the survival of 90% of the infected mice, and the injected of PaPCV with ED50 of the antibiotic ensured the survival of 73% of the experimental animals. The combined use of Tb and the two immuno-preparations (PaPCV and PHP) for the treatment of P. aeruginosa infection proved to be more effective than their separate administrations.     

Dynamics of the contamination of mice during the treatment of burn sepsis due to Pseudomonas aeruginosa using tobramycin alone or combined with active and passive immunization

An experimental study was made with a view to finding out the possible bacteriological advantages of the combined use of tobramycin (Tb) and P. aeruginosa corpuscular polyvalent vaccine (PaCPV) or P. aeruginosa hyperimmune plasma (PaHIP) in burn sepsis caused by P. aeruginosa. The use of the median therapeutic dose of Tb (2.5 mg/kg body weight per day), alone or in combination with immunopreparations, ensured the survival rate of the animals equal to 100%. The contamination of the body with P. aeruginosa after treatment with Tb and PaHIP or PaCPV was lower than after the administration of Tb alone, this phenomenon becoming manifest starting from day 5 of observation in the first case and from day 10 in the second case. The combined use of Tb and immunopreparations (PaCPV or PaHIP) in acute P. aeruginosa infection proved to be more effective than treatment with Tb alone.     

Effectiveness of a polyvalent corpuscular Pseudomonas aeruginosa vaccine, antibiotics and their combinations in experimental chronic Pseudomonas aeruginosa infection

The data on the development of the experimental model of P. aeruginosa chronic infection in mice, produced by their intraperitoneal inoculation with the infective agent, and on the study of the properties of this model are presented. The model has been used in the experimental study of the preventive action of P. aeruginosa polyvalent corpuscular vaccine. The comparative study, carried out with the use of the proposed model, has been made with a view to evaluating the effectiveness of different methods for the treatment of P. aeruginosa chronic septic infection by means of antibiotics (polymixin B and tobramycin), P. aeruginosa polyvalent corpuscular vaccine and their combination. The combined use of this vaccine with antibiotics (polymixin B or tobramycin) has proved to give the most pronounced curative effect with respect to P. aeruginosa chronic infection.     

Experimental vaccinal prophylaxis of Pseudomonas aeruginosa burn sepsis

After the injection of P. aeruginosa live culture under the burned skin of mice sepsis develops within the first 24 hours, finally leading to the death of the animals. The microorganisms can be isolated from the blood, liver, kidneys and mesenterial lymph nodes till day 3 and from the spleen till day 5. After the intraperitoneal injection of P. aeruginosa live culture into mice, sepsis also develops within 24 hours, and the culture can be isolated from the blood and parenchymatous organs till day 3. The LD50 of the culture is equal to 5.1 X 10(6) microbial cells when introduced intraperitoneally and to 30 microbial cells in experimental burn sepsis. Experimental burn sepsis clearly demonstrates the effectiveness of Pseudomonas acellular protein vaccine: its index of effectiveness exceeds 3,000.     

Effectiveness of tobramycin and immunologic preparations in experimental Pseudomonas infection

Certain indices of immunity were studied in mice with burn sepsis due to P. aeruginosa during their treatment with tobramycin (Tb) alone or in combination with immunological drugs. The most significant stimulation of the phagocytic function of peritoneal macrophages was observed when Tb was used in combination with polyvalent corpuscular vaccine of P. aeruginosa. When Tb was used alone or in combination with hyperimmune plasma of P. aeruginosa there was observed close correlation between the phagocytic index and the levels of cyclic adenosine-3',5'-monophosphate in them. Therapy of P. aeruginosa infection with the antibiotic and immunological drugs resulted in much higher levels of agglutinine antibodies in blood serum of the mice than the therapy with Tb alone.     

Antibiotic resistance of Pseudomonas aeruginosa strains isolated from patients with infected burns

Sensitivity to 15 drugs of 248 P. aeruginosa strains isolated from patients with infected burns was studied by the method of agar dilution. All of the strains were resistant to polymyxin M, ceporin, erythromycin and oleandomycin. Most of the strains were resistant to streptomycin, monomycin, ampicillin and rifadin. Moderate resistance of the strains to carbenicillin, morphocycline, vibramycin, kanamycin, tetraolean and tetracycline was observed: the maximum concentrations of these antibiotics (128 microgram/ml) inhibited the growth of 85, 69, 63, 51.8, 43.6 and 41.2 per cent of the strains respectively. Gentamicin proved to be most active against the strains of P. aeruginosa and inhibited 87 per cent of the strains when used in the therapeutic doses. The study provided recomendation of the drugs for parenteral and local use in treatment of burns infected with P. aeruginosa.     

Possible dependence of the results of phage typing Ps. aeruginosa strains on the presence of R plasmids

The plasmid composition of 209 strains of Ps. aeruginosa was determined. The strains were isolated from patients, animals and environment in different geographical areas. The number of the plasmid-containing strains averaged 26.8 per cent. The molecular weights of the plasmids varied from less than 10 to more than 150 MD. 41 conjugative plasmids were transmitted to the recipients of Ps. aeruginosa RAO 303. 66 per cent of them had a restrictive effect on the development of phages used in genetic studies, epidemiological phage typing (Lindberg Collection), and medical practice. This resulted in the changing of the phage type of the host strain. Similar results were obtained in the studies with 10 standard R plasmids representing different incompatibility groups. No relation between the spectrum of the phage restriction, group specificity and the other properties of the plasmids was observed. About 50 per cent of the plasmids markedly lowered the sensitivity level of Ps. aeruginosa RAO 303 to the therapeutic pyocyanic phage. The systems of restriction and modification of DNA coded with plasmids were not detected. A possible changing of the phage type of Ps. aeruginosa strains under the effect of R plasmids should be considered in epidemiological assays and respective treatment measures.     

Comparative study of the antibacterial activity of a number of new antibiotics and their combinations in relation to Pseudomonas aeruginosa]

Tobramycin and sisomycin proved to have the highest antibacterial activity against 156 clinical strains of Ps. aeruginosa and were 4--8 times more effective than monomycin, kanamycin, neomycin and to a lesser extent gentamicin. The combination of mecillinam and sisomycin had a synergistic effect with respect to 26 out of 50 strains of Ps. aeruginosa and the combination of mecillinam and tobramycin had a synergistic effect on 18 strains. An antagonistic effect was observed with the use of the above combinations in 3 cases. The effect of the combinations depended on sensitivity of Ps. aeruginosa cultures to the aminoglycoside antibiotic included into the compositions.     

Antibiotic sensitivity and pathogenetic factors of hospital strains of Pseudomonas aeruginosa isolated from patients treated in a resuscitation unit]

Strains of Pseudomonas aeruginosa were isolated from patients treated in the Centre of Thermal Affections in 1985-1989. It was shown that 72.9, 59.3, 33.8 and 54.2 per cent of the isolates were sensitive to cefotaxime, tobramycin, gentamicin and polymyxin, respectively. The study of pathogenicity factors of the isolates revealed that 83 per cent of the strains produced thermolabile enterotoxin, 79.6 per cent of the strains had adhesive activity and 71.1 per cent of the strains produced hemolysin. The study detected combinations of various pathogenicity factors. 42.3 per cent of the isolates had both adhesive and enterotoxigenic properties. Adhesiveness and hemolytic activity were shown by 13.5 per cent of the strains. 16.9 per cent of the strains produced both enterotoxin and hemolysin. Adhesive activity, enterotoxigenicity and hemolysin production were observed in 6.7 per cent of the strains. It was noted that the strains of P. aeruginosa resistant to polymyxin mainly produced enterotoxin (18.6 per cent) and those resistant to cefotaxime had adhesive activity (34.0 per cent).     

Anti-idiotype-induced, lipopolysaccharide-specific antibody response to Pseudomonas aeruginosa. II. Isotype and functional activity of the anti-idiotype-induced antibodies

We recently developed a murine anti-idiotypic mAb that functioned as a molecular mimic of the O-specific polysaccharide side chain (Ps) of Pseudomonas aeruginosa LPS in vitro, and which induced Ps-specific antibodies in syngeneic BALB/c ByJ mice. In the current studies, we demonstrate that these anti-Id-induced, Ps-specific antibodies fix complement to the bacterial cell surface, and protect neutropenic mice from fatal P. aeruginosa sepsis. The isotypic distribution of the anti-Id-induced antibodies, however, resembles the restricted pattern (IgM and IgG3) seen after administration of purified Ps to mice. The immunogenicity and number of isotypes of Ps-specific antibody produced could be enhanced by conjugating the anti-Id to keyhole limpet hemocyanin. Finally, the anti-Id administered before immunization with purified Ps, primed BALB/c ByJ mice for production of other Ps-specific antibody isotypes (IgG1). These studies show that this anti-Id induces functional anti-Ps antibodies in syngeneic mice, and when used in conjugate form or as a priming agent before Ps immunization, yields an antibody response consistent with "T cell dependence." These immunization strategies may be useful for the induction of polysaccharide-specific antibodies in man.     

Paradoxical synergistic effects of tumour necrosis factor and interleukin 1 in murine gut-derived sepsis with Pseudomonas aeruginosa.

The authors evaluated the synergistic effect of tumour necrosis factor (TNF) and interleukin 1 (IL-1) in gut-derived sepsis in mice. After colonization of Pseudomonas aeruginosa strain D4 in the gastrointestinal tract, cyclophosphamide was administered to induce bacterial translocation of the P. aeruginosa and thereby to cause gut-derived sepsis. In this model, treatment either with 8 microg/kg of recombinant human TNF-alpha (rhTNF-alpha) or 2 microg/kg of recombinant human interleukin 1alpha (rhIL-1alpha) solely did not affect the mortality, whereas combined administration of the same doses of rhTNF-alpha and rhIL-1alpha significantly increased the mortality rate in comparison with saline-treated mice. Bacterial counts in liver and blood were significantly higher in rhTNF-alpha and rhIL-1alpha treated mice than in saline-treated mice. Endogenous TNF-alpha and IL-1beta productions were stimulated after combined treatment with rhTNF-alpha and rhIL-1alpha. On the contrary to these adverse effects, combined treatment with 500 microg/kg of rhTNF-alpha and 50 microg/kg of rhIL-1alpha on the day before the administration of cyclophosphamide significantly reduced the mortality from septic infection. We conclude that TNF and IL-1 synergistically affect the mortality of mice after gut-derived sepsis due to P. aeruginosa in mice and the timing of treatment with these cytokines causes both extremes in their effects.     

Production and experimental testing of the polyvalency of corpuscular vaccine for the prevention of Pseudomonas aeruginosa infections II. Experimental testing of the immunogenicity of polyvalent corpuscular Pseudomonas aeruginosa vaccine

Newly developed P. aeruginosa vaccine has been shown to be safe and apyrogenic for experimental animals. Immunization with the vaccine in a single injection of 0.5 ml has been found to ensure the protection of 80--98% of mice from lethal infection caused by virulent vaccine strains, with the exception of P. aeruginosa strain No. 1311, for 9 weeks. Immunity to P. aeruginosa strain No. 1311 develops only by day 56 after vaccination. No sharp correlation between the specific agglutinin level and the degree of protective effect induced by the immunization of animals with the polyvalent vaccine has been established. The vaccine has been shown to possess high immunogenicity in respect to clinical P. aeruginosa strains belonging to different serotypes (homo- and heterological vaccine strains).     

Immunotyping of freshly isolated strains of Pseudomonas aeruginosa

During 1972-1982 the bacteriological study of 1391 patients with thermal burns was carried out. As the result of clinico-bacteriological studies, the occurrence of P. aeruginosa was found to increase from 39.3% to 70.5% during this period. The immunotyping of P. aeruginosa cultures isolated in 3 burn-treatment centers showed that strains belonging to immunotypes 2, 3, 7 and 3/7 were most frequently isolated from burn wounds. These strains were found to be the cause of hospital infections in burn-treatment hospitals. In connection with the data thus obtained immunological preparations intended for the prophylaxis and treatment of P. aeruginosa infection should include P. aeruginosa strains, immunotypes 2, 3, 7 and 3/7.     

Clinical study to assess the immunogenicity and safety of a recombinant Pseudomonas aeruginosa OprF-OprI vaccine in burn patients

In a recent clinical trial we evaluated the safety and immunogenicity of a recombinant OprF-OprI vaccine consisting of the mature outer membrane protein I (OprI) and amino acids 190-342 of OprF of Pseudomonas aeruginosa in burn patients and compared the elicited antibodies with antibodies against tetanus as response to a simultaneous immunization given on the day of admission. Safety and immunogenicity of the vaccine had been tested before in healthy human volunteers as published in 1999. In this first clinical trial we immunized eight burn patients suffering from second or third degree burns involving between 35% and 55% of the body surface three times with 100 microg of the OprF-OprI vaccine. The vaccine was found to be very well tolerated. The patients did not show any serious side effects - and in particular no activation of the mediator cascade was observed. None of the subjects showed systemic P. aeruginosa infections during or after the treatment of their burns. The serological tests (ELISA) for detection of antibodies against P. aeruginosa and tetanus toxoid showed seroconversion for seven patients after inoculation. The data indicate that OprF-OprI can be a useful vaccine in the therapeutic management of burn injuries.     

Protective effect of a corpuscular polyvalent Pseudomonas aeruginosa vaccine in generalized chronic Pseudomonas aeruginosa infection in mice with cyclophosphamide-induced leukopenia

The prophylactic effect of immunization with P. aeruginosa polyvalent corpuscular vaccine has been shown on the model of P. aeruginosa generalized chronic infection in mice with leukopenia induced by the intraperitoneal injection of cyclophosphamids. This effect is manifested by the increased resistance of the animals to sublethal doses of P. aeruginosa strain, as well as by more intense general and specific immunological responses in the infected animals (the increase of specific antibody titers, the number of leukocytes in the blood serum and the phagocytic activity of the cells of peritoneal exudate).     

Antibiotic sensitivity of Proteus, Pseudomonas pyocyanea and staphyloccoci isolated from scleroma and ozena patients]

Antibiotic sensitivity of 292 strains of Proteus, 60 strains of Ps, aeruginosa, 309 strains of S. aureus and 88 strains of S. epidermidis isolated from the upper respiratory tract of patients with scleroma and ozena was studied. The cultures of Pr. mirabilis were sensitive to aminoglucosides (54.9-96.2 per cent) and Pr. morganii were sensitive to levomycetin (81.5 per cent) and neomycin (92.6 per cnet). Sensitivity of Pr. vulgaris and Pr. morganii was reliably higher (p less than 0.001) than that of Pr. mirabilis. The strains of Pr. morganii were less sensitive to monomycin (P less than 0.001) and streptomycin (p less than 0.01) as compared to the cultures of other Proteus species tested. The strains of Ps. aeruginosa were sensitive only to gentamicin (90 per cent) and neomycin (81.1 per cent). Most of the strains of S. aureus (85.4-100 per cent) were sensitive to oleadomycin, erythromycin, olemorphocycline, tetraolean, oxacillin, methicillin ceporin, lincomycin, ristomycin, kanamycin, monomycin and gentamicin. Benzylpenicillin (90.8 per cent of the sensitive strains), ampicillin (67.1 per cent), tetracycline (66.7 per cent), levomycetin (68.6 per cent) and streptomycin (38.1 per cent) were less effective. Antibacterial therapy in cases with scleroma and ozena should be directed not only against causative agents of the diseases but also against the microbes developing due to disbacteriosis. Combination of parenteral and local use of the antibiotics in the treatment of chronic clebsiellesis decreased the isolation rate of Proteus and Ps. aeruginosa in the patients.     

Antibiotic resistance study of clinical strains of Pseudomonas aeruginosa]

The study of 40 clinical strains of Ps. aeruginosa isolated from the wound surfaces of the patients showed that all the isolates were resistant to one or several antibiotics. The number of the strains resistant to 5, 4, 3, 2 or 1 drug was 5, 22.5, 25. 30 or 17.5 per cent respectively. Fifteen strains carried resistance plasmids capable of conjugative transfer. Eleven out of 21 plasmids controlled resistance to chloramphenicol, 7 plasmids controlled resistance to streptomycin and sulfanylamides, 1 plasmid controlled resistance to streptomycin and chloramphenicol. The presence of two types of the plasmids controlling resistance to chloramphenicol and streptomycin + sulfanylamides respectively was found. All the plasmids proved to be capable of conjugative transfer between the strains of Ps. aeruginosa ML (PAO). The frequency of the plasmid conjugative transfer in such crosses ranged from 10(-6) to 10(-3). Most of the plasmids belonged to the incompatibility groups P-2 and P-7. One plasmid belonged to the incompatibility group P-5. It should be noted that about a half of the plasmids (11 out of 21) belonged to the incompatibility group P-7 which up to the present time was conditional, since was represented by a single plasmid Rms 148.