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Staphylococcus aureus is a leading cause of significant morbidity and mortality and an enormous economic burden to public health worldwide. Infections caused by methicillin-resistant S. aureus (MRSA) pose a major threat as MRSA strains are becoming increasingly prevalent and multi-drug resistant. To this date, vaccines targeting surface-bound antigens demonstrated promising results in preclinical testing but have failed in clinical trials. S. aureus pathogenesis is in large part driven by immune destructive and immune modulating toxins and thus represent promising vaccine targets. Hence, the objective of this study was to evaluate the safety and immunogenicity of a staphylococcal 4-component vaccine targeting secreted bi-component pore-forming toxins (BCPFTs) and superantigens (SAgs) in non-human primates (NHPs). The 4-component vaccine proved to be safe, even when repeated vaccinations were given at a dose that is 5 to 10- fold higher than the proposed human dose. Vaccinated rhesus macaques did not exhibit clinical signs, weight loss, or changes in hematology or serum chemistry parameters related to the administration of the vaccine. No acute, vaccine-related elevation of serum cytokine levels was observed after vaccine administration, confirming the toxoid components lacked superantigenicity. Immunized animals demonstrated high level of toxin-specific total and neutralizing antibodies toward target antigens of the 4-component vaccine as well as cross-neutralizing activity toward staphylococcal BCPFTs and SAgs that are not direct targets of the vaccine. Cross-neutralization was also observed toward the heterologous streptococcal pyogenic exotoxin B. Ex vivo stimulation of PBMCs with individual vaccine components demonstrated an overall increase in several T cell cytokines measured in supernatants. Immunophenotyping of CD4 T cells ex vivo showed an increase in Ag-specific polyfunctional CD4 T cells in response to antigen stimulation. Taken together, we demonstrate that the 4-component vaccine is well-tolerated and immunogenic in NHPs generating both humoral and cellular immune responses. Targeting secreted toxin antigens could be the next-generation vaccine approach for staphylococcal vaccines if also proven to provide efficacy in humans., Competing Interests: MA and HK have stocks and RA and FH have stock options in Integrated Biotherapeutics Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Venkatasubramaniam, Liao, Cho, Adhikari, Kort, Holtsberg, Elsass, Kobs, Rudge, Kauffman, Lora, Barber, Aman and Karauzum.)

Staphylococcus aureus is a major human pathogen, and the emergence of antibiotic-resistant strains is making all types of S. aureus infections more challenging to treat. With a pressing need to develop alternative control strategies to use alongside or in place of conventional antibiotics, one approach is the targeting of established virulence factors. However, attempts at this have had little success to date, suggesting that we need to better understand how this pathogen causes disease if effective targets are to be identified. To address this, using a functional genomics approach, we have identified a small membrane-bound protein that we have called MspA. Inactivation of this protein results in the loss of the ability of S. aureus to secrete cytolytic toxins, protect itself from several aspects of the human innate immune system, and control its iron homeostasis. These changes appear to be mediated through a change in the stability of the bacterial membrane as a consequence of iron toxicity. These pleiotropic effects on the ability of the pathogen to interact with its host result in significant impairment in the ability of S. aureus to cause infection in both a subcutaneous and sepsis model of infection. Given the scale of the effect the inactivation of MspA causes, it represents a unique and promising target for the development of a novel therapeutic approach., (Copyright © 2020 Duggan et al.)

Superantigens (SAgs) play a major role in the pathogenesis of Staphylococcus aureus and are associated with several diseases, including food poisoning, bacterial arthritis, and toxic shock syndrome. Monoclonal antibodies to these SAgs, primarily TSST-1, SEB and SEA have been shown to provide protection in animal studies and to reduce clinical severity in bacteremic patients. Here we quantify the pre-existing antibodies against SAgs in many human plasma and IVIG samples and demonstrate that in a major portion of the population these antibody titers are suboptimal and IVIG therapy only incrementally elevates the anti-SAg titers. Our in vitro neutralization studies show that a combination of antibodies against SEA, SEB,and TSST-1 can provide broad neutralization of staphylococcal SAgs. We report a single fusion protein (TBA 225 ) consisting of the toxoid versions of TSST-1, SEB and SEA and demonstrate its immunogenicity and protective efficacy in a mouse model of toxic shock. Antibodies raised against this fusion vaccine provide broad neutralization of purified SAgs and culture supernatants of multiple clinically relevant S. aureus strains. Our data strongly supports the use of this fusion protein as a component of an anti-virulence based multivalent toxoid vaccine against S. aureus disease.

Staphylococcus aureus causes significant illnesses throughout the world, including toxic shock syndrome (TSS), pneumonia, and infective endocarditis. Major contributors to S. aureus illnesses are secreted virulence factors it produces, including superantigens and cytolysins. This study investigates the use of superantigens and cytolysins as staphylococcal vaccine candidates. Importantly, 20% of humans and 50% of rabbits in our TSS model cannot generate antibody responses to native superantigens. We generated three TSST-1 mutants; G31S/S32P, H135A, and Q136A. All rabbits administered these TSST-1 toxoids generated strong antibody responses (titers>10,000) that neutralized native TSST-1 in TSS models, both in vitro and in vivo. These TSST-1 mutants lacked detectable residual toxicity. Additionally, the TSST-1 mutants exhibited intrinsic adjuvant activity, increasing antibody responses to a second staphylococcal antigen (β-toxin). This effect may be due to TSST-1 mutants binding to the immune co-stimulatory molecule CD40. The superantigens TSST-1 and SEC and the cytolysin α-toxin are known to contribute to staphylococcal pneumonia. Immunization of rabbits against these secreted toxins provided complete protection from highly lethal challenge with a USA200 S. aureus strain producing all three exotoxins; USA200 strains are common causes of staphylococcal infections. The same three exotoxins plus the cytolysins β-toxin and γ-toxin contribute to infective endocarditis and sepsis caused by USA200 strains. Immunization against these five exotoxins protected rabbits from infective endocarditis and lethal sepsis. These data suggest that immunization against toxoid proteins of S. aureus exotoxins protects from serious illnesses, and concurrently superantigen toxoid mutants provide endogenous adjuvant activity., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background: Camelids and sharks possess a unique subclass of antibodies comprised of only heavy chains. The antigen binding fragments of these unique antibodies can be cloned and expressed as single domain antibodies (sdAbs). The ability of these small antigen-binding molecules to refold after heating to achieve their original structure, as well as their diminutive size, makes them attractive candidates for diagnostic assays., Results: Here we describe the isolation of an sdAb against Staphyloccocus aureus enterotoxin B (SEB). The clone, A3, was found to have high affinity (Kd = 75 pM) and good specificity for SEB, showing no cross reactivity to related molecules such as Staphylococcal enterotoxin A (SEA), Staphylococcal enterotoxin D (SED), and Shiga toxin. Most remarkably, this anti-SEB sdAb had an extremely high Tm of 85°C and an ability to refold after heating to 95°C. The sharp Tm determined by circular dichroism, was found to contrast with the gradual decrease observed in intrinsic fluorescence. We demonstrated the utility of this sdAb as a capture and detector molecule in Luminex based assays providing limits of detection (LODs) of at least 64 pg/mL., Conclusion: The anti-SEB sdAb A3 was found to have a high affinity and an extraordinarily high Tm and could still refold to recover activity after heat denaturation. This combination of heat resilience and strong, specific binding make this sdAb a good candidate for use in antibody-based toxin detection technologies.

Immunomodulators licopid (synthetic analogue of muramylpeptide) and purified staphylococcal toxoid (PST) in some variants of experiments on mice caused adverse immunologic effects: enhancement of virus-induced immunosupression, shift from latent immunosupression (revealed only by low, but not standard, doses of test-antigen) to manifested one. Described adverse effects are not a contraindication for use of the studied drugs in practice. Their adverse effects were revealed only after single inoculation, whereas in clinical conditions PST and licopid are used by courses with duration of 5-7 and 10 days respectively. Our experiments show that inoculation of suppressive doses of the preparations repeated by 2-4 times can prevent the shift from latent to manifested immunosupression. Enhancement of immunosupression was not observed in case of combined administration of suppressive doses of PST and adjuvant dose of licopid.

Influence of immunomodulator of bacterial origin - purified staphylococcal toxoid (PST) - on the synthesisof proinlammatory (IL-1beta, IL-6, TNFalpha, IFN-gamma) and anti-inflammatory (IL- 10) cytokines, as well as cytokines directing the immune response to Th1 (IL-12) or Th2 (IL-4) type was studied in mice. Serum cytokines levels as well as levels of cytokines produced by splenocytes spontaneously or after stimulation by phytohemagglutinin were measured 4 and 24 hours after inoculation of PST. It was shown that PST in wide spectrum of doses (15; 1.5; 0.15 BU per mouse) was able to enhance or suppress synthesis of cytokines. Effect was nonlinear and its direction was depended from cytokine, time interval passed before obtaining the sample and dose of PST. For example, 15 BU of PST enhanced whereas 0.15 BU of PST suppressed the IL-6 production 4 hours after inoculation. Decrease of IL-6 level in serum 24 hours after inoculation of PST was detected. Synthesis of several serum interleukins (IL-2, IL-10) did not changed 4 and 24 hours after inoculation irrespective from dose of PST. It was demonstrated that modulation of humoral immune response in vivo induced by PST did not associated with modulation of cytokine profile. For example, increase of number of cells secreting antibodies to sheep erythrocytes was registered both during increased synthesis of cytokines (4 hours, IL-1beta, IL-6, IL-12) and during period of its depression (IL-6, TNF-alpha, IFN-gamma), as well as during stable production of cytokines (IL-1beta, IL-6, IFN-gamma).

With adaptive transfer method it has been shown that immunomodulator purified staphylococcal toxoid (PST) changed (stimulate or suppress) antigen-presenting function (APF) of mice peritoneal macrophages (MF) in vivo. This phenomenon was registered during assessment of ability of peritoneal MF to present heterologous (with regard to PST) antigen--sheep erythrocytes (SE). Modulation vector depended from time interval between PST and SE inoculations. Inoculation of PST 1.5 h before SE resulted in stimulation of APF. When SE were inoculated to donor mice 24 h after PST, suppression of APF was developed. Suppression of APF was observed along with suppression of proinflammatory cytokines (IL-1beta, IL-6, TNF-alpha) as well as B-lymphocytes growth factor (IL-4). When cytokine profile was assessed 4 h after PST injection, the suppression of synthesis of these cytokines was not observed. Production of IL-12 increased in 9-12 times in 4 and 24 h after PST injection.

Effect of immunomodulators for microbial origin on innate immunity and antitumor system was continued to study. Immunomodificator Immunovac VP-4, purified staphylococcal toxoid and glucosaminyl muramyl dipeptide (GMDP) equally enhanced cytotoxicity of mononuclear leukocytes of peripheral blood of healthy donors. Index of cytotoxicity was 2.78, 2.77 and 2.70 respectively. Reduced metastatic progression of Lewis lung carcinoma in mice was observed after Immunovac VP-4 and GMDP administration. Effectiveness was seen when preparations administered according to schedules including their administration before implantation of the tumor. If preparations were administered number of metastases reduced in 4.4-5.6 times and size of metastases reduced in 7-10 times. Interplay between antitumor activity of studied immunomodulators and cytotoxic activity of NK-cells, which are base effectors of antitumor immune response, are discussed.

The partially degraded lipopolysaccharide of Burkholderia cepacia (LPSdegr) and the ornithine-containing lipids were purified from some bacteria. The substances were developed as complex lipid adjuvants, because they have weak toxicity and are able to activate the immune systems of the living body. After various toxoid antigens such as pertussis toxoid, diphtheria toxoid and tetanus toxoid were mixed with the complex lipid adjuvants, the mixtures were administered to mice subcutaneously. Antitoxoid IgG antibody titers in the serum were measured several times over 3 months. The efficacy of the LPSdegr as adjuvant was almost as high as that of the ornithine-containing lipids, and it was almost equal to that of the aluminum hydroxide adjuvant (Alum), which is generally used as a vaccine adjuvant.

The action of immunomodulators, purified staphylococcal toxoid (PST) and lycopid, on secondary immunodeficiency state developing during infection caused by Coxsackie virus B3 was studied. This defect was manifested by delayed hypersensitivity to sheep red blood cells (SRBC) and the suppression of neutralizing antibodies to poliomyelitis virus. Depending on the scheme of the experiment, PST normalized the defects of immune response to SRBC or poliovirus, increased suppression or showed no activity. Lycopid corrected the defects of humoral response to SRBC. The combination of PST and lycopid was found to produce no increase of suppression. The suggestion was made on the expediency of combination of two (and probably more) immunomodulators for increasing the efficiency of correction of secondary immunodeficiency.

On the 50 volunteers from 20 to 40 years old gomeostasis indexes of inoculating by staphylococci cleaning adsorbing anatoxini donors, before have been revactinated against tetanus, were studied. It was established, that attract of plasmaferes donors to consistent immunization by tetani and staphylococci anatoxini don't call negative alterations in their gomeostasis indexes. Results of study demonstrate, that it's possibly to attract for immunization by staphylococci anatoxini donors, before have been revactinated against tetanus.

The Russian experience with the preparation and clinical application of an antitoxic antistaphylococcal hyperimmune plasma and immunoglobulin is described. The immunotherapies were developed in the late 1960s and put into widespread use in the Soviet Union for the prophylaxis and treatment of sepsis, pneumonia, and other conditions caused by an epidemic of antibiotic-resistant Staphylococcus aureus.

In this study, the adjuvant effect of the sustained release biodegradable nanospheres (100-150 nm in diameter) has been compared with alum. Nanospheres were formulated using a biodegradable polylactic polyglycolic acid copolymer (PLGA, 50:50) containing Staphylococcal Enterotoxin B (SEB) toxoid as a model vaccine antigen. Systemic immune response of the nanospheres containing toxoid was studied in rabbits by subcutaneous immunization. The data demonstrated that approximately 30% of the toxoid activity was lost following its encapsulation into nanospheres. Under in vitro conditions, nanospheres demonstrated sustained release of the toxoid. However, only 20% of the antigenic toxoid was released over the first 2 weeks of the release study. Immunization of animals with equal doses of toxoid, either using nanospheres or alum induced a comparable systemic immune response (IgG, IgM and IgA). The immune response reached a maximum level at 7 weeks post-immunization, which then gradually declined with time. The booster dose of toxoid at 19 weeks, either using alum or nanospheres induced similar immune response in both the groups, but was greater than the primary immune response. The studies, thus, suggest that biodegradable nanospheres could be used as a vaccine adjuvant.

Mice vaccinated with a combination of two Staphylococcus aureus antigens consisting of a recombinant collagen-binding protein (CnBP) and alpha-toxoid (alpha-toxoid) were significantly protected from intramammary challenge infection with S. aureus. The average number of bacteria recovered from the glands of mice vaccinated with the combination of CnBP/alpha-toxoid was significantly lower compared to the average number of bacteria recovered from the glands of mice vaccinated with only CnBP or alpha-toxoid or controls (P< or =0.01). Histopathological examination of mammary glands of mice vaccinated with CnBP together with alpha-toxoid showed no pathological changes, whereas glands of mice vaccinated with CnBP or alpha-toxoid alone developed severe mastitis and showed both focal and disseminated necrosis.

82 blood donors have been observed, 63 of them were immunized. Blood group ABO(H), secreting group Se--se and Staphylococcus antibody contents (anti-alpha-staphylolysins) were determined in all the donors. It was found out that the donors-secretors with A(II) blood group exhibited the antibody-production increasing. It is supposed that the secreting of group-specific substance A, that has structural elements similar those of staphylococcus into saliva promotes antibody production increase against staphylococcus. The mechanism of such specific stimulation remains to be unknown and requires further studying.

Blood serum content was studied of specific antistaphylococcal antibodies (staphylolysins) in 576 donors immunized with staphylococcal anatoxin with the purpose of obtaining an antistaphylococcal plasma and antistaphylococcal immunoglobulin to be used in clinical settings. 292 donors had been immunized and examined prior to 1986, 284--after 1986 (before 1994). Comparison of the immune responses in the above periods of time permitted finding out that 13.03% of immunized donors responded to the antigenic stimulus by such paradoxical reaction as disappearance of specific antibodies; the number of persons-active respondents has gotten reduced from 17.12% to 5.98% as has the number of individuals having the baseline level of staphylolysins (1-2 ME/ml). The changes were at their greatest in donors with group A (II) blood.

Purified staphylococcal toxoid (PST) was shown to alter the spontaneous and mitogen-induced proliferation of mouse spleen cells. In vitro, PST inhibited spontaneous proliferation, as well as proliferation induced by the optimal dose of Con A (2 micrograms/ml) and the optimal and suboptimal doses of lipopolysaccharide (LPS) (100 and 50 micrograms/ml). At the same time the dose of 1.5 binding units (BU) of PST, inhibiting spontaneous proliferation in vitro, induced strong proliferative response in combination with the suboptimal dose of Con A (1 microgram/ml). Our experiments demonstrated that the spontaneous proliferation of mouse spleen cells, immunized with the toxoid, remained unchanged (0, 15 or 15 BU/mouse) or increased (1.5 BU/mouse) the response of spleen cells of immune animals to ConA and LPS also changed in comparison with the control, depending on the conditions of the experiment. After the use of the combination of 2 micrograms of Con A and 1.5 BU of PST or 100 (50) micrograms of LPS and 1.5 BU of PST the inhibition of proliferative response was observed. The summation of the signals of the suboptimal dose of Con A (1 microgram) 1.5 BU of PST was demonstrated.

Staphylococcal enterotoxin B (SEB), a primary cause of food poisoning, is also a superantigen that can cause toxic shock after traumatic or surgical staphylococcal wound [correction of would] infections or viral influenza-associated staphylococcal superinfections or when aerosolized for use as a potential biologic warfare threat agent. Intranasal or intramuscular (i.m.) immunization with formalinized SEB toxoid formulated with meningococcal outer membrane protein proteosomes has previously been shown to be immunogenic and protective against lethal respiratory or parenteral SEB challenge in murine models of SEB intoxication. Here, it is demonstrated that immunization of nonhuman primates with the proteosome-SEB toxoid vaccine is safe, immunogenic, and protective against lethal aerosol challenge with 15 50% lethal doses of SEB. Monkeys (10 per group) were primed i.m. and given booster injections by either the i.m. or intratracheal route without adverse side effects. Anamnestic anti-SEB serum immunoglobulin G (IgG) responses were elicited in all monkeys, but strong IgA responses in sera and bronchial secretions were elicited both pre- and post-SEB challenge only in monkeys given booster injections intratracheally. The proteosome-SEB toxoid vaccine was efficacious by both routes in protecting 100% of monkeys against severe symptomatology and death from aerosolized-SEB intoxication. These data confirm the safety, immunogenicity, and efficacy in monkeys of parenteral and respiratory vaccination with the proteosome-SEB toxoid, thereby supporting clinical trials of this vaccine in humans. The safety and enhancement of both bronchial and systemic IgA and IgG responses by the proteosome vaccine delivered by a respiratory route are also encouraging for the development of mucosally delivered proteosome vaccines to protect against SEB and other toxic or infectious respiratory pathogens.

Regularities in the formation of autoantibodies to human IgA, IgM, IgG and their fragments, such as F(ab')2, Fab, Fc, in donors immunized with influenza vaccine (44 subjects) and staphylococcal toxoid (15 subjects) were studied with regard to the dynamics of specific immune response. After immunization with staphylococcal toxoid autoimmune reactions were registered only in the precipitation tests during 3-5 weeks of observation. In donors immunized with influenza vaccine the induction of isotypically heterogeneous autoimmune reactions was established. These reactions were represented mainly by IgA and IgG autoantibodies to immunoglobulins of different classes and their fragments. The duration of autoimmune reactions registered in this group of donors was specially noteworthy. The levels of autoantibodies to most of the antigens under study remained significantly elevated for 6 months. After immunization with influenza vaccine the presence of direct correlation between the level of specific immune response and the level of autoantibody formation was established.

Microspheres composed of biocompatible, biodegradable poly(DL-lactide-co-glycolide) (DL-PLG) and staphylococcal enterotoxin B (SEB) toxoid were evaluated as a vaccine delivery system when subcutaneously injected into mice. As measured by circulating immunoglobulin G (IgG) antitoxin titers, the delivery of SEB toxoid via DL-PLG microspheres, 1 to 10 microns in diameter, induced an immune response which was approximately 500 times that seen with nonencapsulated toxoid. The kinetics, magnitude, and duration of the antitoxin response induced with microencapsulated toxoid were similar to those obtained when an equal toxoid dose was administered as an emulsion with complete Freund adjuvant. However, the microspheres did not induce the inflammation and granulomata formation seen with complete Freund adjuvant. The adjuvant activity of the microspheres was not dependent on the superantigenicity of SEB toxin and was equally effective at potentiating circulating IgG antitrinitrophenyl levels in response to microencapsulated trinitrophenyl-keyhole limpet hemocyanin. Empty DL-PLG microspheres were not mitogenic, and SEB toxoid injected as a mixture with empty DL-PLG microspheres was no more effective as an immunogen than toxoid alone. Antigen-containing microspheres 1 to 10 microns in diameter exhibited stronger adjuvant activity than those greater than 10 microns, which correlated with the delivery of the 1- to 10-microns, but not the greater than 10-microns, microspheres into the draining lymph nodes within macrophages. The antibody response induced through immunization with microencapsulated SEB toxoid was protective against the weight loss and splenic V beta 8+ T-cell expansion induced by intravenous toxin administration. These results show that DL-PLG microsphere vaccine delivery systems, which are composed of pharmaceutically acceptable components, possess a strong adjuvant activity for their encapsulated antigens.

The intensity and direction of the modulation of humoral immune response, induced by the action of purified staphylococcal toxoid (PST), depend on the dose of the preparation, the dose of the test antigen, the scheme of the experiment, the time of the registration of its results and the genotype of animals. PST induces the modulation of the synthesis of both IgM and IgG antibodies. The injection of small divided doses of PST enhances immune response to heterologous antigen, while a single inoculation of the total dose produces no such effect.

Animal and human blood serum contains great amount of blocked (or "silent") immunoglobulins which, being activated by heating to 60 degrees C, pH decrease to 2.0-2.5 or treatment with 5M KSCN acquire a capacity to interact with different antigens. This interaction may be equally prevented or weakened by both identical and serologically non-related antigen, i.e. activated immunoglobulins are polyspecific. Polyspecific immunoglobulins show less affinity in comparison with monospecific antibodies, their interaction with antigens depends considerably on temperature.

The protective potency of purified staphylococcal toxoid by the survival rate of immunized mice challenged with the culture of Staphylococcus aureus strain L-1726 and the antigenic properties of the toxoid, determined by the level of antitoxin in the blood of mice and by the intensity of cell-mediated immunity in the spleen-cell migration inhibition test, were studied. The experiments were made on CBA and C57BL/6 mice. Purified staphylococcal toxoid was shown to possess antigenic and protective properties in a wide range of doses between 0.15 and 15 binding units per mouse. The protective effect of the toxoid in CBA mice was manifested in the presence of circulating antibodies and cell-mediated reaction or only in the presence of the toxoid. In C57BL/6 mice the protective effect of the toxoid was less pronounced and appeared in combination with the induction of cell-mediated immunity in the presence of an extremely low antibody level (0.062 I.U.).

It has been shown that during immune system activation pronounced alterations are observed in the metabolic process course in lymphoid cells that, probably, reflect the changes in T- and B-lymphocyte functional activity, and the kinetics of intercellular relationships. The ratio of LDH1 and LDH5 fractions most differing in electrophoretic mobility has been recommended as a sign of B-system immunity activation.

This paper presents results of testing pregnant women for staphylococcal antigen sensitization using immunothermistography (ITG). The ITG is based on registration of environmental heat conduction change with a microthermistor resistor during antigen-antibody reaction. The study group comprised 75 pregnant women immunized by staphylococcal anaphylotoxin and nonimmunized pregnant and nonpregnant women. Blood alfa-antistaphylolysin levels showed a close direct correlation with ITG findings. Combined use of these methods identified a population of pregnant women requiring immunization.

The data were obtained on the presence of considerable aberrations of the isoenzymic spectrum of lactate dehydrogenase in different lymphocyte populations and subpopulations. The isoenzymic shifts reflect the changes occurring in the metabolic status of the cells and are sensitive tests detecting functional deficiency of lymphocytes.

Delayed hypersensitivity skin tests were carried out with 5 antigens - PPD, staphylococcus, streptokinase/streptodornase, candida and trichophytin-in 400 hospital patients without any known causes for diminished delayed hypersensitivity. The degree of reactivity to each antigen was: PPD. 69.50%; candida 49.75%; streptokinase/streptodornase 44.50%; trichophytin 42.00%; staphylococcus 14.25%. Reactivity to either PPD or candida occurred in 86.50% of all cases. Positive response to streptokinase/streptodornase was present in 7%; which brings the total cases with reactions to one or more of the 3 antigens to 93.50%. Those who responded to trichophytin or staphylococcus were 3% only, bringing the total response of all cases to at least one antigen, to 96.5%.

The influence of combined immunization with inactivated chromatographic vaccine and adsorbed staphylococcal toxoid on the activity of the immune system of volunteer donors has been studied. With the T- to B-cell ratio in the peripheral blood remaining stable, the third injection of staphylococcal toxoid induces a certain suppression of the capacity of lymphocytes for blast transformation in the presence of mitogen and specific antigens. The intensity of the accumulation of anti-alpha-staphylolysin is subject to considerable individual variations and depends on its level prior to immunization. The level of influenza antihemagglutinins remains most stable when the interval between the injection of inactivated chromatographic influenza vaccine and the beginning of immunization with adsorbed staphylococcal toxoid is 3-6 months.

It was shown that in the immunization of rabbits with sorbed staphylococcus toxoid and human albumin, at the height of the immune response there occurred an intensification of biosynthesis and destabilization of lysosomal membranes of blood leukocytes, with a simultaneous development of microcirculation disturbances in the lungs. Development of pneumonia against the background of immunization led to a greater destabilization of the lysosomic membranes of leukocytes and to the intensification in the lungs of infiltrative and destructive processes, in comparison with the course of inflammation in the nonimmunized animals. Peculiarities of the antigens used caused some specificity in the course of pneumonia. A relationship between the changes of blood leukocytes and disturbances in the lungs in immunization is supposed.

The isohemagglutination test with the introduction of the heat-stable fraction of microbial broth culture into the reacting system was used as a model of the antigen-antibody interaction. In the "A-anti-A" system the inhibition of isohemagglutination in the presence of staphylococcal allergen was observed. This inhibition was caused by the binding of the allergen with anti-A-antibodies, which was indicative of the antigenic similarity between staphylococcus and group-specific factor A. The influence of the similarity thus established on immune response was studied by the determination of antitoxic titers in 292 donors immunized with staphylococcal toxoid and in 86 patients with chronic hematogenic fistulous osteomyelitis with due regard of the ABO phenotype of the examinees. Subjects with group A(II) blood were found to have lesser antibody titers than those with blood of other groups and to be predisposed to the progressive course of staphylococcus-induced diseases.

The content of antistaphylococcal antitoxins--antialphastaphylolysins was examined in the blood serum of three groups of blood donor volunteers. It is shown that volunteer blood donors immunized according to two schemes with prophylactic purposes may be a source of antistaphylococcal plasma. Additionally antistaphylococcal plasma may be obtained from non-immune donors showing high titers of antialphastaphylolysins revealed by the screening method.

Ag-staining of the nucleolar organizer regions of acrocentric chromosomes of T-lymphocytes did not change during the immune response in children with porotitis and in those being in contact with parotitis-suffering children, as well as in young adults previously immunized by staphylococcal anatoxin. This character displayed individual peculiarities. No differences in these age groups were detected. A positive correlation was found between the size of Ag-band and the ability of chromosomes to make associations. Ag-staining and participation of G-chromosomes in associations was higher than those markers in D-chromosomes.

In experiments of the passive protection of mice the protective properties of sera obtained from humans before and after their immunization with Proteus vaccine used as a monopreparation or in combination with staphylococcal toxoid and/or pyoimmunogen were studied. When introduced in a single subcutaneous injection, Proteus vaccine prepared from soluble antigenic complexes ensured an increase in the protective properties of sera. The second injection of the vaccine essentially enhanced the protective potency of the sera of the immunized donors. The therapeutic injection of Proteus vaccine ensured the essential increase of the protective properties of the sera. This increase could be experimentally detected within at least 25-30 days from the beginning of immunization. The immunization of volunteers with Proteus vaccine in combination with pyoimmunogen and adsorbed staphylococcal toxoid ensured the maximum increase of the protective properties of their sera.