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).     

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).     

Safety and sensitizing properties of a polyvalent corpuscular Pseudomonas aeruginosa vaccine and its effect on hematological indices

The acute and chronic toxicity, influence on hematological characteristics and sensitizing properties of P. aeruginosa polyvalent corpuscular vaccine have been studied in experiments on 3 species of animals. The acute experiment has shown that the LD50 of the preparation contains not less than 7800 million cells, which is almost 160 times higher than the recommended immunizing dose (500 million cells). The safety of the preparation is confirmed by the data obtained in the histological and histochemical investigations of the tissues and organs of animals subjected to multiple immunizations with the vaccine. These investigations have revealed no pathological changes in the animals. During the study of the chronic toxicity of the preparation the hematological characteristics of the animals have been found to remain within normal limits. The vaccine has been shown to possess low sensitizing activity, which is manifested by the absence of severe reactions to allergic skin tests with different bacterial allergens (specific allergens obtained from P. aeruginosa and allergens obtained from other bacterial species), made on completion of the course of immunization with the vaccine.     

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).     

Experimental immunological effectiveness and safety of pyoimmunogen vaccine against Pseudomonas aeruginosa infection

Pyoimmunogen, a polycomponent vaccine against P. aeruginosa infection, has been obtained in laboratory and semi-industrial conditions. The microbial biomass obtained from the strains belonging to O-serotypes (immunotypes) most frequently occurring in clinical practice has been used for producing protective antigens. The preparations have been found to contain proteins (peptides) and carbohydrates in the ratio 6 : 1 to 8 : 1, as well as traces of 2-keto-3-desoxyoctanate, which is indicative of the low content of endotoxin. The immunogenicity of the preparations has been studied experimentally by the active immunization of mice. In these experiments the animals vaccinated in a single injection were found to be protected from challenge with both homologous and heterologous P. aeruginosa strains. The high level of protection from infection caused by toxigenic strain PA-103 was registered. The preparations have low toxicity: LD50 for mice exceeds 2 mg (in protein content): after the multiple administration (7-10 times) of the preparation to mice and rats the weight of the experimental animals was not significantly different from the weight of the control animals.     

The immunological response in volunteer donors immunized with a Pseudomonas vaccine]

The trial of experimental vaccine consisting of protective protein antigens of P. aeruginosa cell wall was carried out on 114 volunteers. The vaccine proved to be faintly reactogenic and induced the formation of specific humoral immunity in 98% of the volunteers who retained a high level of anti-P. aeruginosa antibodies in their blood for up to 5 months (the term of observation after the course of immunization was over.     

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.     

Production and study of the properties of a polyvalent corpuscular Pseudomonas aeruginosa vaccine. IV. The effect of immunization with Pseudomonas aeruginosa vaccine on the immune status of volunteer donors

The influence of immunization with P. aeruginosa vaccine on the immune status of volunteer donors has been studied. Immunization with P. aeruginosa vaccine in doses of 0.5-0.5-1.0 ml at intervals of 7 days has been found to lead to the 13-fold increase of the titer of specific antibodies, lasting for 3-4 months, which ensures the possibility of obtaining anti-P. aeruginosa hyperimmune plasma with standard titers. The injection of P. aeruginosa vaccine to donors leads to the activation of humoral immunity simultaneously with the increase of the absolute and relative number of rosette-forming B-lymphocytes without essential changes in the amount of rosette-forming T-lymphocytes.     

Protective effects of affinity-purified antibody and truncated vaccines against Pseudomonas aeruginosa V-antigen in neutropenic mice

Virulent P. aeruginosa strains express PcrV, one of the translocational components of the type III secretion system. PcrV has been reported to be a protective antigen against lethal P. aeruginosa infection. The PcrV region, which contributes to protective immunity against P. aeruginosa infection, was investigated by using genetically engineered, truncated PcrV proteins and affinity-purified anti-PcrV antibodies against the truncated PcrV proteins. The efficacy of active and passive immunization against PcrV was tested in mice with cyclophosphamide-induced immunosuppression by intraabdominal challenge of P. aeruginosa . Active immunization with either full-length PcrV1-294 or PcrV139-294 significantly improved the survival of mice infected with P. aeruginosa , while PcrV139-258, PcrV139-234, PcrV197-294, and PcrV261-294 were not protective. These results suggest that an effective PcrV vaccine needs to contain not only the Mab166 epitope (PcrV144-257) but also the carboxyl terminal tail of PcrV. In the case of passive immunization, administration of affinity-purified anti-PcrV IgG against either PcrV1-294 or PcrV139-258 showed significantly higher efficacy against lethal P. aeruginosa infection than did original anti-PcrV IgG and Mab166. The increased efficacy of affinity-purified anti-PcrV IgG implies that more potent anti-PcrV strategies are possible. The results of this study are crucial to the development of an effective PcrV vaccine for active immunization and to an appropriate blocking anti-PcrV antibody against P. aeruginosa infection in humans.     

Effect of a corpuscular polyvalent Pseudomonas aeruginosa vaccine on the proliferation and differentiation of hematopoietic stem cells and erythropoiesis

P. aeruginosa corpuscular polyvalent vaccine stimulates hematopoiesis in sublethally stimulated mice. The stimulating effect is dose-dependent. The most effective method of immunization is the intravenous injection of the vaccine. The degree to which the stimulation of hematopoiesis is manifested varies in different strains of mice. The stimulation of hematopoiesis is not linked with an increase in the content of erythropoietin. P. aeruginosa polyvalent corpuscular vaccine and monovaccine, prepared from P. aeruginosa strain 1313 and incorporated into the polyvalent vaccine, protect lethally irradiated mice in the postradiation survival test.     

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 properties of anatoxin obtained from a homogeneous preparation of Pseudomonas aeruginosa exotoxin A

The protective properties of formulated toxoid obtained from the highly purified preparation of P. aeruginosa exotoxin A have been studied in the test of the active immunization of mice. The study has revealed that the preparation when introduced in 1 or 2 injections in a dose of 15 micrograms, shows faint protective potency with respect to P. aeruginosa strains differing in virulence. Immunization with this toxoid in 3 and 4 injections has been found to ensure 60-100% and 50-60% protection of mice infected with P. aeruginosa toxigenic and proteolytic strains respectively. Immunization with toxoid has been found to induce the appearance of short-term antibacterial immunity which loses its capacity to protect the immunized animals, challenged with both toxigenic and proteolytic P. aeruginosa strains, as early as on day 28. The immunization of mice with toxoid in 4 injections has been shown to induce the development of antitoxic immunity capable of neutralizing up to 150 LD50 of purified exotoxin A.     

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.     

Immunological activity and toxicity of a peroral vaccine made from mutant Re Salmonella minnesota

The possibility of the oral use of heated corpuscular vaccine prepared from S. minnesota mutant R 595 (chemotype Re) for protection against Pseudomonas aeruginosa has been studied. Oral immunization in 3 doses, each containing 10(9) cells of the vaccine strain, has been shown to protect mice from death after the intravenous injection of P. aeruginosa culture in a dose of 5 LD50 and induce a rise in the titers of antibodies to Re-glycolipid (Re-hemagglutinins). After multiple oral administration Re-vaccine shows low acute and chronic toxicity and induces local and systemic immunological transformation.     

Protective effect of the aerosol immunization of mice with the polycomponent vaccine Immunovac VP-4 after the challenge of the animals with Klebsiella pneumoniae virulent strain

Used four schemes of the administration of the preparation with different time of the exposition of the animals in an aerosol chamber were tested with their subsequent intraperitoneal challenge with K. pneumoniae virulent strain K16. Irrespective of the number of immunization courses, the administration of the preparation made at intervals of 1 day, or daily, did not ensure any protective effect, but only led to an insignificant increase in their survival time in comparison with nonimmunized animals. After intervals between immunizations were increased to 3 days the protective effect of aerosol immumization was obtained (the survival rate was 65-80 % and considerably differed from that of the controls). The protective effect of aerosol immunization thus obtained was comparable with the effectiveness immunization made in a single subcutaneous injection. Aerosol immunization resulted in low antibody titers to the antigens contained in the vaccine, while after a single subcutaneous injection high antibody titers to Klebsiella and Proteus antigens were detected. The antigen-stimulated blast transformation of spleen lymphocytes in mice subjected to aerosol immunizations in 5 exposures was high. After subcutaneous immunization significant changes in such characteristics were detected on day 15. The data thus obtained were indicative of good prospects in the development Immunovac VP-4 as the medicinal form intended for use in aerosols.     

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.     

Antigenic properties of Pseudomonas aeruginosa anatoxin and the protective action of antitoxic anti-Pseudomonas aeruginosa serum

The antigenic properties of P. aeruginosa toxoid, prepared with the use of casein culture medium, were not inferior to those of the toxoid obtained in Martin broth. In experiments on white mice antisera obtained by the immunization of rabbits with the toxoid prepared on the basis of casein culture medium showed sharply pronounced protective properties against P. aeruginosa homologous and heterologous strains, as well as toxigenic reference strain PA-103.     

Morphological study of the organs and tissues of mice infected after a burn with a Pseudomonas aeruginosa strain and treated with a Pseudomonas aeruginosa polyvalent corpuscular vaccine and tobramycin

The effect of tobramycin and polyvalent corpuscular Ps. aeruginosa vaccine on the infectious process in mice with grade III burns inoculated with Ps. aeruginosa 1312 was studied. The highest percentage of the survival (100 per cent) among the animals was observed, when the vaccine was applied locally every day for 7 days. With the use of tobramycin administered intramuscularly for 2 times 95 per cent of the animals survived. When the vaccine was administered subcutaneously, 96.6 per cent of the animals survived. Morphological investigation of the organs and tissues of the mice showed that the vaccine applications to the infected burns promoted rapid elimination of microorganisms in the wounds and prevented development of sepsis due to Ps. aeruginosa. At the same time early epithelization of the wounds was observed (by the 4th-7th day). Intramuscular injections of tobramycin and subcutaneous injection of vaccine also prevented development of sepsis due to Ps. aeruginosa and protected the animals from death. Still no epithelization of the wounds by that period was observed. Microscopic examinations revealed necrosis of the epiderma and derma elements and microbial swarms on the skin surface.     

Polish preparations for preventing and treating infections caused by Pseudomonas aeruginosa

Three preparations, viz. sheep serum for local application ("Sanoserum"), anti-Pseudomonas sheep immunoglobulin for passive immunization ("Immunoglobulin Pseudomonas") and Pseudomonas vaccine for active immunization ("Pseudovac"), have been obtained at the Central Laboratory of Sera and Vaccines in Warsaw, Poland. The laboratory study and clinical trial of these preparations have been carried out. The preparations have proved to be effective as supplement to the conventional methods for the early diagnosis and treatment of P. aeruginosa infection in burn patients.     

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.