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.     

Acellular pertussis vaccine

Protective, immunogenic, toxic, and sensitizing properties of acellular pertussis vaccine (aPV) developed according to original technology were studied, aPV had marked protective activity which lasted more than 2 years. Sera of mice immunized by aPV also possess protective properties, and they were more prominent than in sera of mice immunized by pertussis bacteria suspension (PS). Immune sera to aPV neutralized cytopathogenic effect of pertussis toxin (PT) on ovarian Chinese hamster cells in 1:250 dilution, whereas neutralizing activity of sera to PS was very low. Level of antibodies to PT was higher in rabbits immunized, according to schedules and dosage recommended for children, by aPV than by PS. High immunogenicity of aPV was proved also by levels of IgG to PT in sera of mice immunized three times by aPV in human dosage. During experiments on mice and guinea pigs aPV had mild toxicity, did not induce autoimmune process, did not have anaphylactogenic properties compared with bacterial suspension characterized by high anaphylactogenic activity. Histamine-sensitizing abilityof aPVwas 40 times lower than that of PS. Assessment of pyrogenic properties of aPV and PS performed on rabbits showed that aPV was 1,000 times less pyrogenic than PS. Obtained results demonstrate high protective and immunogenic properties of domestic acellular pertussis vaccine and its low toxic and sensitizing characteristics.     

Acetone-treated pertussis vaccine--a potent and safer new pertussis vaccine

A vaccine was prepared from the growth of Bordetella pertussis by repeated treatment with acetone. The vaccine has been designated as acetone-treated pertussis vaccine (ATPV). A total of ten batches of ATPV were prepared, five each from B. pertussis strains 134 and 509. These strains are routinely employed at this Institute for the production of conventional whole-cell pertussis vaccine (WCPV) for blending in diphtheria-pertussis-tetanus vaccine. The mouse protective and histamine sensitizing activities of ATPV and WCPV were compared. The ATPV showed 1.5- to 2-fold higher potency than the WCPV. The histamine sensitizing activity of ATPV was much reduced compared with that of the WCPV. No appreciable difference was observed in the results of a mouse weight-gain tests between the ATPV and WCPV. The details of the preparation of ATPV have been described. Because of higher potency and reduced histamine sensitizing activity, the ATPV may prove more acceptable in immunization programmes against pertussis, even in countries where WCPV is unpopular due to its suspected reactogenicity.     

Effectiveness of oral immunization of white mice with a complex S. typhimurium antigen]

The protective properties of hydroxylamine preparation obtained from a virulent strain of S. typhimurium were studied in experiments with natural infection after a single oral immunization. The new data obtained in these experiments suggest that the treatment of bacteria with hydroxylamine allows to produce the preparation which, when administered orally, has the immunizing dose only 20 times as great as its immunizing dose for subcutaneous administration. The action of gastric juice on hydroxylamine preparation, as well as the duration and specificity of immunity induced by the oral administration of this preparation were studied. The oral administration of some adjuvants was found to make it possible to considerably decrease the effective dose of the vaccine.     

Detoxified bacterial endotoxns. II. Preparation and biological properties of chemically modified crude endotoxins from Salmonella typhimurium

Martin, William J. (University of Utah, Salt Lake City), and Stanley Marcus. Detoxified bacterial endotoxins. II. Preparation and biological properties of chemically modified crude endotoxins from Salmonella typhimurium. J. Bacteriol. 91:1750-1758. 1966.-Chemical modification of a crude endotoxin prepared by the Roschka-Edwards (RE) procedure from a strain of Salmonella typhimurium yielded products which were nontoxic for mice and had reduced fever effects in rabbits. A reduction in rabbit pyrogenicity of approximately 100 times was noted with a potassium periodate-treated RE preparation when compared with the parent RE preparation. Measured in a similar fashion, pyrogenicity of a potassium methylate-treated RE preparation was reduced by a factor of 10 while pyrogenicity of a boron trifluoride RE preparation was unchanged. All of these endotoxoids, including the parent RE preparation, showed little toxicity for mice. Immunogenicity was determined in mice by comparing Boivin, RE, and endotoxoid preparations with a heat-killed, phenol-preserved (HP) vaccine prepared from the same strain of S. typhimurium. Employing a 10 ld(50) challenge, the protective immunogenicity of the respective vaccines was determined by active immunized mouse protection tests. Although two 100 mug immunizing doses of the Boivin, RE, and the respective endotoxoid preparations varied in mouse protection (potassium methylate RE > Boivin > RE > boron trifluoride RE > potassium periodate RE), it was evident that, with the exception of the potassium methylate preparation, the HP vaccine yielded greatest protection against the 10 ld(50) challenge with S. typhimurium. Further mouse protection experiments suggested that the minimal immunogenic dose of the potassium methylate RE vaccine preparation was approximately 50 mug. These data indicated an approximate fivefold difference between the minimal pyrogenic dose (10 mug) and the minimal immunogenic dose (50 mug). These findings further suggest that potassium methylate RE vaccine preparations should be considered in the search for less toxic enteric fever vaccines.     

The efficacy of killed Trypanosoma evansi vaccines in mice.

Four strains of Trypanosoma evarsi (D3, D4, D5 and D6) isolated from German shepherd dogs were inoculated into mice, and infected blood was used to prepare 9 separate killed vaccines. White mice inoculated with 1:100 diluted PBS vaccine, 0.5% carbol vaccine, or 100% Lugol vaccine showed survival rates of more than 60%. Among these 3 vaccines PBS vaccine and 0.5% carbol vaccine showed higher survival rates at 1:500 and 1:1000 dilutions, respectively. When young mice (15-20 g) were immunized with PBS vaccine, they resisted challenge with homologous strains, D3 strain in single injection, D6 strain in double injections and all strains in 5 injections. Protection however was not observed in old mice (25-30 g) give the same vaccine preparation. When mice were vaccinated with a single injection of D3 vaccine and challenged with heterologous strains, only those challenged with D4 strain at 10-5 dilution showed a survival rate of more than 60%. There was no difference in protective ability among PBS vaccine, agar adjuvant and kaolin adjuvant vaccines. Agglutinating antibody was demonstrated in mice receiving 5 injections of PBS vaccine.     

Detoxified bacterial endotoxins. I. Preparation and biological properties of an acetylated crude endotoxin from Salmonella typhimurium

Martin, William J. (University of Utah, Salt Lake City), and Stanley Marcus. Detoxified bacterial endotoxins. I. Preparation and biological properties of an acetylated crude endotoxin from Salmonella typhimurium. J. Bacteriol. 91:1453-1459. 1966.-Acetylation of a crude endotoxin prepared by the Roschka-Edwards (RE) procedure from a strain of Salmonella typhimurium yields a product with reduced pyrogenicity in rabbits as well as one which is nontoxic in mice. A reduction in pyrogenicity of approximately 100 times was noted with this acetylated crude endotoxin when compared with the parent RE preparation. A comparison was made of immunogenicity with mice of Boivin, RE, and acetylated Roschka-Edwards (Acet-RE) preparations with a heat-killed, phenol-preserved (HP) vaccine prepared from the same strain of S. typhimurium. Less than pyrogenic doses of all vaccines were not protective. The least pyrogenic preparation (Acet-RE) was immunogenically effective in about five times the minimal pyrogenic dose. The data suggest that the Acet-RE preparation should be considered further in the search for enteric fever vaccines with lowered potential for undesirable systemic responses.     

Immunogenic dialyzable factor derived from a ribosomal fraction of Salmonella typhimurium. I. Preparation of the protective dialyzable factor from the ribosomal fraction by the freeze-thaw procedure

The preparation, properties, and immunogenicity of the dialyzable factor from a ribosomal fraction of Salmonella typhimurium are described. The ribosomal fraction was purified to eliminate O-antigenic components, by affinity chromatography (Sepharose-anti-O antibody conjugates used as immunoadsorbent). The dialyzable factor was obtained in the concentrated dialysate of the purified ribosomal fraction which was alternately frozen in dry-ice acetone and thawed in an 80 C water bath, for a total of five or six cycles. When this preparation was tested for its ability to protect mice against challenge with 1,000 LD50 of the homologous bacteria, it afforded 100% protection at a dose equivalent to 5.0 micrograms of RNA. The protection conferred by this factor was mainly cell mediated but immune serum enhanced this immunity despite the fact that no antibodies were detected in it. The protective activity of this factor was sensitive to RNase digestion but resistant to proteolytic enzymes. Ion exchange chromatography of this factor with DEAE-Sephadex A-25 (in 7 M Urea-0.02 M Tris-HCl buffer, pH 7.5) resulted in a single A260 peak which was found to be immunogenic. Chemical analysis of this peak after it was concentrated and desalted revealed that this immunogenic fraction was composed mainly of mixed nucleotides. The data indicate that protective immunity conferred by a ribosomal vaccine is associated with RNA but may not require the intact RNA molecule.     

Glutaraldehyde inactivated pertussis vaccine: a less histamine sensitizing vaccine

The effects of different inactivating agents on the biological activity of the histamine sensitization factor of Bordetella pertussis toxin were examined. The agents were used for inactivation in the preparation of whole cell pertussis suspension. The histamine sensitizing activity was reduced to 36.9-13.3% by treatment with glutaraldehyde, to about 50% by treatment with formaldehyde and by the acetone-II treatment, relative to the reduction by heat treatment. Treatment with thimerosal and the acetone-I treatment did not reduce the histamine sensitizing activity as the 50% histamine sensitizing doses of the heat inactivated pertussis preparation, the thimerosal inactivated pertussis preparation and the acetone-I treated pertussis preparation were very similar. Glutaraldehyde has thus been found to be a better inactivating agent for the preparation of a safe pertussis suspension as it considerably reduced the histamine sensitizing activity of pertussis toxin.     

Comparative analysis of the major protective antigens production in Vibrio cholerae recombinant and producer strains of the classical biovar

The comparative study of 4 constructed protective antigen producing strains of the classical biovar and V. cholerae strains 569 B Inaba and M41 Ogawa, used in manufacturing the cholera chemical vaccine "cholerogen-toxoid", was carried out. The study revealed that V. cholerae plasmid strains 2414 Ogawa, 2415 Inaba and nonplasmid strains 2416 Ogawa, 2417 Inaba had a higher level of production of the main protective antrigens in comparison with producer strains. They also synthesized much more (4-5 fold) cholera toxin, toxin co-regulated adhesion pili, contained protein OmpU in their outer membrane, exceeded 2- to 3-fold in the synthesis of pathogenicity enzymes (proteases, phospholipases) and synthesized the same amounts of 01 antigen, serovars Inaba and Ogawa. The use of the newly created protective-antigen producing strains in vaccine manufacturing could facilitate the preparation of a more effective cholera chemical vaccine "cholerogen-toxoid".     

Optimal inoculation doses of meningococcal chemical polyvalent ABC vaccine for immunizing children of various ages and adolescents. I. Safety and reactogenicity of different dosages of the preparation

The investigation was carried out to study the safety and reactogenicity of meningococcal chemical polyvalent (ABC) vaccine with the aim of finding the optimum vaccination doses of this vaccine for the immunization of children of different age groups. The investigation, carried out in accordance with the methodological principles of a strictly controlled epidemiological field trial, showed the preparation to be safe, nonreactogenic and to produce no pronounced sensitizing effect.     

Relationship of the sensitizing and protective properties of pertussis antigens

When comparing delayed hypersensitivity (DN) to B. pertussis corpuscular antigens with the agglutinogenic composition of B. pertussis, as well as to its histamine-sensitizing, leukocyte-sensitizing, adjuvant and hemagglutinating activity, no correlation was detected between the specific sensitizing properties of these antigens and the serotype and studied nonspecific properties of B. pertussis. The DH level correlated with the protective activity of B. pertussis corpuscular antigens and the ultrasonic fractions of B. pertussis. The close correlation of these two phenomena suggest that DH-inducing and protective B. pertussis antigens are identical, though their action has different manifestations, depending on the method of administration of the antigen preparation. On these grounds a new method for evaluating the immunogenicity of B. pertussis antigens is proposed. This method consists in comparing the sensitizing activity of the antigen under test and that of the national standard.     

Presence of CpG DNA and the local cytokine milieu determine the efficacy of suppressive DNA vaccination in experimental autoimmune encephalomyelitis.

We here study the adjuvant properties of immunostimulatory DNA sequences (ISS) and coinjected cytokine-coding cDNA in suppressive vaccination with DNA encoding an autoantigenic peptide, myelin basic protein peptide 68-85, against Lewis rat experimental autoimmune encephalomyelitis (EAE). EAE is an autoaggressive, T1-mediated disease of the CNS. ISS are unmethylated CpG motifs found in bacterial DNA, which can induce production of type 1 cytokines in vertebrates through the innate immune system. Because ISS in the plasmid backbone are necessary for efficient DNA vaccination, we studied the effect of one such ISS, the 5'-AACGTT-3' motif, in our system. Treatment with a DNA vaccine encoding myelin basic protein peptide 68-85 and containing three ISS of 5'-AACGTT-3' sequence suppressed clinical signs of EAE, while a corresponding DNA vaccine without such ISS had no effect. We further observed reduced proliferative T cell responses in rats treated with the ISS-containing DNA vaccine, compared with controls. We also studied the possible impact of coinjection of plasmid DNA encoding rat cytokines IL-4, IL-10, GM-CSF, and TNF-alpha with the ISS-containing DNA vaccine. Coinjection of IL-4-, IL-10-, or TNF-alpha-coding cDNA inhibited the suppressive effect of the DNA vaccine on EAE, whereas GM-CSF-coding cDNA had no effect. Coinjection of cytokine-coding cDNA with the ISS-deficient DNA vaccine failed to alter clinical signs of EAE. We conclude that the presence of ISS and induction of a local T1 cytokine milieu is decisive for specific protective DNA vaccination in EAE.     

IgA1 Protease as a Vaccine Basis for Prevention of Bacterial Meningitis

Russian Journal of Bioorganic Chemistry - The review covers the study of the protective properties of IgA1 protease and the possibility of creating a vaccine preparation for the prevention of...     

Removal of lipopolysaccharide from acellular Bordetella pertussis vaccine by detergent treatment

Protective antigen was extracted from Bordetella pertussis cells with 1.0 M NaCl and precipitated with ammonium sulfate, 20-40% saturation (designated fraction 15A-1B). The protective antigen was purified further by detergent (Emulphogene BC720) treatment and adsorption to aluminum hydroxide gel (designated fraction 15A-108A). Compared with B. pertussis vaccine and fraction 15A-1B, fraction 15A-108A retained protective activity as assessed by the mouse protection test, but had reduced protein and markedly reduced endotoxin content. Fraction 15A-108A also had reduced leukocytosis-promoting, histamine sensitizing splenomegaly-inducing, and adjuvant activities. Emulphogene treatment provided a relatively simple method for removing endotoxin from a potential acellular B. pertussis vaccine.     

Studies on histoplasmosis. V protective effect of HIF-coated Histoplasma against challenge

Summary Results of testing a total of 145 hamsters in 2 separate trials (a total of 90 challenged animals) suggest the potentiality of HIF-coatedHistoplasma yeast cells as a vaccine. Animals challenged with 5×10⁵ Histoplasma yeast cells in saline 3 weeks after initial inoculation with 5 × 10⁴ Histoplasma yest cells in 1) saline or 2) crude HIF in kidney homogenate or 3) the active 280 mµ fraction thereof were protected by the 280 mµ fraction > the crude HIF > saline suspensions ofHistoplasma, based on analysis of tissue sections and cultures.It is suggested that since HIF has also been found in experimental blasomycosis it should be sought in other mycoses, perhaps in all infectious diseases, and its potential use in a vaccine preparation should be more extensively tested.Supported in part by Public Health Service Grant No. 1-SO-IFR-05359-01 to the senior author at The George Washington University.     

T cell epitope-based peptide-DNA dual vaccine induces protective immunity against Schistosoma japonicum infection in C57BL/6J mice

Schistosomiasis is a major public health problem that primarily affects developing countries. Although schistosomicidal drugs exist, the development of an efficacious vaccine would potentially be the most powerful means of controlling this disease. Previous studies have shown that vaccination with selected protective epitopes successfully induced partial protection and/or reduced female fecundity in animal models. Thus, we investigated whether the T cell epitope P5 from the host-interactive tegument of Schistosoma japonicum 22.6 (S. japonicum) could act as a protective epitope. The protective potential of P5 in a vaccine against S. japonicum was determined by using a T cell epitope based peptide-DNA dual vaccine (PDDV). In our experiments, the vaccine construct (P5-18K-PDDV) contains the peptide of the T cell epitope (P5) and plasmid DNA, encoding P5 and adjuvant GM-CSF. We show that P5-18K-PDDV induced both cell-mediated and humoral immune responses in vivo and achieved partial protection against S. japonicum infection in C57BL/6J mice. Histopathological studies reveal that P5-18K-PDDV immunized mice had substantially reduced liver pathology compared to the control groups. Together, these results suggest that P5 could be used as a vaccine immunogen for both worm killing and disease prevention against S. japonicum.     

Control of immunologically crossreactive leptospiral infection by administration of lipopolysaccharides from a nonpathogenic strain of Leptospira biflexa

In our previous paper (Matsuo, K., Isogai, E., and Araki, Y., Carbohydr. Res., 328: 517-524, 2000), antigenic polysaccharides obtained from the lipopolysaccharide (LPS) fraction of a nonpathogenic leptospira, Leptospira biflexa patoc Patoc I, are shown to be broadly crossreactable with most rabbit antisera elicited by immunization with various pathogenic leptospires. The result led us to test a protective effect of the same LPS in a hamster model system by heterologously challenging with a pathogenic leptospira, L. interrogans manilae UP-MMG. Firstly, a similarity in the antigenic epitopes of L. biflexa and L. interrogans was confirmed by the following assays. In the microscopic agglutination test (MAT), a hamster antiserum elicited by immunization with the L. biflexa-LPS preparation was shown to agglutinate cells of L. interrogans. Contrarily, in the enzyme-linked immunosorbent assay (ELISA), the L. biflexa-LPS preparation was shown to crossreact with a hamster antiserum elicited by immunization with whole cells of L. interrogans. These results suggest that the same or closely related antigens may be present on the cell surfaces of both L. biflexa patoc Patoc I and L. interrogans manilae UP-MMG. Furthermore, in a protective assay, the prior administration of a L. biflexa-LPS preparation resulted in raising a protective response in hamsters against challenge by L. interrogans without any side effect. The protective effect was strongly dependent on the dose amounts and/or administration times of L. biflexa-LPS. Thus, L. biflexa-LPS preparations can use as a potent vaccine against leptospirosis caused by various leptospires.     

Hematoporphyrin ethers--III. Cellular uptake and photosensitizing properties

1. The cellular uptake and the efficiency in sensitizing cells to photoinactivation were determined for hematoporphyrin (Hp) diphenyl ether, Hp dicyclohexyl ether and Hp dihexyl ether. 2. The phenyl diether was taken up by the cells to the same degree as was the clinically used porphyrin preparation photofrin II, while the dihexyl and notably the dicyclohexyl ether were taken up 3-4 times better. 3. Furthermore, the quantum yields for photoinactivation of cells were similar for the three diethers and twice as large as that for photofrin II. 4. Fluorescence- and absorption spectroscopy indicate that these findings are related to the fact that photofrin II is much more aggregated in the cells than are the three Hp diethers. 5. When cells loaded with the porphyrins are incubated with porphyrin-free medium containing serum a certain percentage of the cell-bound drug is removed: 14% for photofrin II, 28% for Hp diphenyl ether, 50% for Hp dicyclohexyl ether and 20% for Hp dihexyl ether. 6. With respect to cell uptake and retention of the dyes, the data did not show any uniform relationship to the polarity of the drugs, in contrast to what has been found earlier for Hp diethers of linear hydrocarbons.     

Evaluation of influenza vaccine Vaxigrip in the combined immunization of conscripts during influenza seasons of 2002-2003

The immunization properties of the influenza vaccine Vaxigrip, used in combination with vaccines against pneunococcal infection and hepatitis A (respectively, Pneumo 23 and Avaxim), were evaluated. In Central Russia in one of the units of the internal forces of the RF Ministry of Internal Affairs 3 groups totaling 755 servicemen were formed, depending on the complex of the introduced vaccines. Active medical observation and the registration of the complaints of the vaccinees at the postvaccinal period did not reveal unusual reactions and complications in none of the groups under observation. In the evaluation of the level of specific antibodies to the circulating influenza viruses prior to vaccination the low level of collective protection to influenza B virus was determined: protective antibody levels were registered only in 14-18% of the servicemen, while the corresponding data with respect of influenza viruses A(H1N1) and A(H3N2) were 45-50% and 56-63% respectively. At the same time, in seronegative persons the vaccine Vaxigrip exhibited high immunogenic activity with respect of all 3 influenza strains; seroconversion to them was determined in 84-92% of the vaccines, and the level of protective antibody titers before the beginning of the epidemic season was 86-99% in the whole of the group. The characteristics of the prophylactic effectiveness of the vaccine Vaxigrip in relation to the influenza virus infection level 1 were 4.7 (index) and 79% (coefficient). In addition, the frequency of influenza cases, clinically pronounced and confirmed by laboratory methods, in patients who had been immunized with 3 vaccines was 6.7%, which was 10.3 times less frequent than number of cases in the groups of comparison (68.2% on the average). The coefficient of epidemiological effectiveness of the prophylaxis of influenza was 90.2%. The complex use of 3 preparations did not affect the immunization properties of the vaccine Vaxigrip.