Detection of antibodies to the teichoic acids of the Staphylococcus aureus cell wall in the sera of osteomyelitis patients

The comparative analysis of serum samples from patients with hematogenic and post-traumatic osteomyelitis for the presence of antibodies to the teichoic acids of the cell wall of S. aureus has been carried out by means of ELISA. The investigation has shown that the test system developed on the basis of teichoic acids can be used for confirming the diagnosis of osteomyelitis. An attempt to use this system for the dynamic observation of the disease has been made.     

Antibodies to Staphylococcus aureus teichoic acids in the pathogenesis of chronic osteomyelitis

The titers of antibodies to S. aureus cell-wall teichoic acids have been determined in 97 orthopedic and traumatic patients with purulent diseases, differing by the activity of the process, by means of the enzyme immunoassay. These antibodies appeared in the patients' blood in active osteomyelitic process of staphylococcal etiology.     

Chemical characterization of a new surface antigenic polysaccharide from a mutant of Staphylococcus aureus

A mutant of Staphylococcus aureus H was isolated by virtue of its inability to agglutinate with antibodies against teichoic acid of S. aureus. Immunological studies revealed that the mutant, S. aureus T, possessed a new surface antigen in addition to having the antigenic determinant of the wild-type strain, the ribitol teichoic acid. The presence of this additional surface component rendered strain T resistant to staphylococcal typing phages, presumably by masking the phage-receptor sites. The polymer was separated from teichoic acid by chromatography on diethylaminoethyl cellulose and was shown to be composed of two amino sugars, N-acetyl-d-fucosamine and N-acetyl-d-mannosamin uronic acid.     

Serological diagnosis of suppurative-septic diseases of staphylococcal etiology

The parallel examination of osteomyelitis patients by means of the passive hemagglutination (PHA) test with the use of antigenic erythrocyte diagnostic agents, prepared on the basis of hydrochloric acid extract from Staphylococcus aureus strain 209P and teichoic acid extract from S. aureus strain Wood 46 has revealed that these diagnostic agents are practically equal in their diagnostic effectiveness. In the examination of endocarditis patients measurement of the total antibody activity in the PHA test with diagnosticum on the basis of strain 209P has proved to be a more sensitive method, whereas in osteomyelitis patients the total IgG activity has been more accurately measured by ELISA. The treatment of sera with 2-mercaptoethanol essentially decreased the diagnostic effectiveness of the PHA test. The study has shown the diagnostic value of not only IgG but also of IgM antibody measurements.     

Humoral response to Staphylococcus aureus antigens evaluated by the western blotting method]

Cellular antigens extracted from the cells of four Staphylococcus aureus strains from different kinds of infections (sepsis, osteomyelitis, furunculosis) were analysed by the western blotting technique. Antibiotic sensitivity pattern of the strains was compared. One isolate was found to be MRSA strain. Sera samples from patients of whom strains were isolated and four sera from blood donors (as a control) were used in the investigation. IgG levels for purified staphylococcal antigens (lipase, alpha-toxin and teichoic acid) were estimated. Interaction between extracted bacterial antigens and serum antibodies of IgG class were analysed in homologous and heterologous systems. The most strong immunological reaction of the investigated sera with staphylococcal antigens was observed in the case of homologous system. Serum from sepsis patient was found to be the most reactive serum with all staphylococcal antigens mixtures.     

Diagnosis of staphylococcal diseases using an immunoenzyme reaction

To develop an immunoenzymatic system, the preparations of teichoic acid (TA) isolated from the Staphylococcus aureus strain Wood 46 were used. The TA-based ELISA technique made it possible to diagnose infectious endocarditis of staphylococcal etiology. Further investigations are necessary to ascertain the possibility of using this system for diagnosing other staphylococcal diseases.     

Possibility of detecting human embryonal leukemia antigen]

A possibility of detecting embryonic leukemic antigen on human leukemic blast cells in an acute human leukemia cytotoxicity test with the sera and 7S and 19S serum immunoglobulins of the placental blood was studied. The presence on the blast cells of patients suffering from acute leukemia of an antigen detectable by antibodies of placental blood (of parturients) was demonstrated; this antigen was absent on the leukocytes of healthy donors.     

Identification of autoantibodies associated with systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the presence of antinuclear antibodies. We performed serological analysis of cDNA expression library (SEREX) to identify autoantibodies associated with SLE. The screening of three different cDNA expression libraries with pooled sera of patients with SLE yielded 11 independent clones that reacted with pooled sera of patients with SLE. In this screening, autoantibodies to poly(ADP-ribose) polymerase (PARP), U1snRNP, and galectin-3 were prevalent in the sera of patients with SLE (26/68, 25/68, 12/63, respectively). The frequency of autoantibody to PARP was significantly higher in SLE than that of healthy donors (0/76) (38.2% vs 0%, p<0.00001). The autoantibody to PARP was infrequently detected in the serum of patients with RA (1/50). However, autoantibody to PARP was not found in the sera of patients with other rheumatic diseases including Sjogren's syndrome (0/19), systemic sclerosis (0/18), and polymyositis/myositis (0/37). The frequency of autoantibody to human galectin-3 (12/63) was significantly higher in SLE than that of healthy donors (0/56) (19% vs 0%, p=0.0006). Autoantibody to galectin-3 was not found in the sera of patients with rheumatoid arthritis (0/50), Sjogren's syndrome (0/18), and systemic sclerosis (0/19). Interestingly, autoantibody to galectin-3 was also prevalent in the sera of patients with polymyositis/dermatomyositis (16/37, 43.2%). Further functional characterization of these autoantibodies would be necessary to determine their value as diagnostic markers or to define clinical subsets of patients with SLE. Statistical analysis revealed that the presence of autoantibody to PARP was inversely related with pleurisy, and the presence of autoantibody to galectin-3 related with renal disease.     

Antibodies to meningococcal antibodies in the sera of patients and donors

The comparative study of sera taken from healthy persons (pooled sera of 100 donors, 6 individual serum specimens) and sera taken from patients with meningococcal meningitis (pooled sera of 10 patients with meningococcal infection, group A, and 6 individual serum specimens from patients with meningococcal infection, groups A, B, C) was carried out by the method of immunoblotting. All proteins from healthy donors were found to contain antibodies to meningococcal iron-regulated protein (IRP) of 85 kD, designated as TbpB. In 30% of donor sera the presence of antibodies to meningococcal IRP of 34 kD (FbpA) was registered. Moreover, donor sera were found to contain antibodies to meningococcal IRP of 45 kD. The sera taken from convalescents were found to have the increased content of antibodies to IRP of 70 and 85 kD and somewhat lesser content of antibodies to proteins of 98, 44 and 34 kD. As regards other (non iron-regulated) proteins, in the process of convalescence the most intensive antibody production was observed with respect to minor protein with a molecular weight of 50 kD, as well as proteins of class 5, characterized by molecular weights of 30 kD and less.     

Demonstration of EBNA (Epstein-Barr virus nuclear antigen) antibodies of different immunoglobulin classes.

Anit-EBNA IgM, a previously unknown antibody, was detected by the antihuman globulin anticomplement immunofluorescence (ACIF) method in serum samples from acute infectious mononucleosis (IM) of Epstein-Barr virus (EBV) origin. The antibody disappears from the serum in some weeks during convalescence. It was absent in anti-EBV=positive sera of healthy donors and in serum samples taken from patients with IM caused by cytomegalo-virus. The antibody appears simultaneously with anti-EBV IgmM and, reaching a lower titre than the latter, its titre curve runs parallel with the anti-EBV IgM curve. Since in acute EBV infections, anti-EBNA IgM always appeared, its presence may serve as an additional evidence of the acuteness of EBV infection. In EBV-seropositive healthy subjects, the bulk of antibodies belongs to the IgG class, non-complement-fixing IgA antibodies occur only sporadically.     

Autoantibodies against aggrecan in systemic rheumatic diseases

OBJECTIVE: This study was undertaken to investigate the presence of autoantibodies against the main cartilage proteoglycan, aggrecan, in systemic rheumatic disease sera, and to identify substructure(s) responsible for the autoimmune response. METHODS: Sera were obtained from 86 patients with various systemic rheumatic diseases, 14 with osteoarthritis (OA), 18 with cancer and 40 healthy individuals. The presence of autoantibodies against aggrecan was examined by a solid phase assay and by Western blotting, using proteoglycan aggregates treated with proteolytic enzymes. The positive bands were subjected to nanohigh performance liquid chromatography (nanoHPLC)-MS, in order to identify the aggrecan substructures involved in the autoimmune response. RESULTS: Autoantibodies against aggrecan were identified in all systemic rheumatic disease sera at a high titre, almost three times that observed in healthy controls. OA and cancer sera produced a reaction equal to that of the healthy. Western blotting analysis of aggrecan proteolytic fragments revealed the presence of a triple band, reacting with the patients' sera, of about 37 kDa, which also reacted with a polyclonal antibody against hyaluronan-binding region. NanoHPLC-MS analysis suggested that this band belonged to the G2 domain of aggrecan. CONCLUSION: At least a part of the autoimmune reaction to aggrecan, displayed by the systemic disease sera, involves the G2 domain. The significant difference observed between these sera and those from other diseases, especially cancer, may suggest a possible discriminatory role of anti-aggrecan antibodies. This may help in the differential diagnosis in complicated clinical cases. However, for this to be confirmed, studies in larger cohorts of patients should be performed.     

Detection of antibodies to preparations of Staphylococcus aureus peptidoglycan, teichoic acids and polysaccharide in patients' sera

Enzyme immunoassay systems on the basis of S. aureus teichoic acids, peptidoglycan and polysaccharide-containing preparation are incapable of reliably diagnosing the staphylococcal nature of postoperative complications in oncological patients.     

The level of IgM, IgG and IgA antibodies and total antibodies to the low-molecular, cell-wall protein of Streptococcus group A without type specificity in the serum of patients with a streptococcal infection

The levels of IgM, IgG and IgA antibodies, as well as total antibodies, to group A streptococcal low-molecular cell-wall protein without type specificity were studied in the sera of patients with primary erysipelas, rheumatism in the active and inactive phases, seronegative rheumatoid arthritis, as well as in the sera of healthy donors. The average level of antibodies to low-molecular protein in the sera of all groups of patients was significantly higher than the sera of healthy donors. The analysis of the distribution of antibodies in accordance with their isotypes revealed the specific features of response, characteristic of each group of patients. For rheumatism patients, the positive correlation between response to low-molecular protein and response to group-specific polysaccharide A was established. This correlation was most pronounced in patients with rheumatism in the inactive phase.     

Identification of a novel antigen from Staphylococcus epidermidis

A genomic DNA library of Staphylococcus epidermidis NCTC 11047 was constructed, using the Lambda Zap Express cloning vector, and screened with serum collected from a patient with S. epidermidis endocarditis. Sequence analysis of a 30 kDa cloned protein, termed staphylococcal secretory antigen, SsaA, identified a novel protein not previously reported in S. epidermidis. SsaA showed strong homology with two other staphylococcal proteins: SceB from Staphylococcus carnosus and a staphyloxanthin biosynthesis protein from Staphylococcus aureus. Further investigation revealed SsaA to be a highly antigenic protein that was expressed in vivo and could be recovered from whole cells and from the culture supernatant. A combination of Western blot analysis and PCR screening identified SsaA or a homologue in 103/103 staphylococcal strains. SsaA-like genes were not detected in other Gram-positive bacteria of medical importance or a number of Gram-negative organisms. Elevated anti-SsaA IgG antibody levels were detected in sera of five patients with S. epidermidis endocarditis but not in patients with other S. epidermidis infections, endocarditis of other aetiologies or patients with no evidence of infection. The expression of SsaA during episodes of S. epidermidis endocarditis suggests a virulence role specific to the pathogenesis of this infectious disease.     

Antibody response to Staphylococcus aureus whole cell, lipase and staphylolysin in patients with S. aureus infections

Abstract Three assays to measure antibodies against Staphylococcus aureus whole cells, lipase and staphylolysin were used to try to discriminate between complicated and uncomplicated S. aureus septicaemia. Sera were examined from 8 patients with S. aureus endocarditis, 23 patients with complicated S. aureus septicaemia, 12 patients with uncomplicated S. aureus septicaemia and 93 febrile non-septicaemic controls. No single assay could distinguish between complicated and uncomplicated S. aureus septicaemia. If the criterion for a positive result is defined as positive antibody level in the anti-lipase ELISA as well as in at least 1 of the other 2 assays, 10/31 patients with S. aureus endocarditis or complicated septicaemia were positive compared to 0/93 non-septicaemic patients and 0/12 patients with uncomplicated S. aureus septicaemia. Therefore, the combined use of serological assays in the diagnosis of complicated S. aureus septicaemia, one of which is the anti-lipase ELISA, is recommended.     

Staphylococcus aureus cholecystitis: a report of three cases with review of the literature

Infection of the hepatobiliary system is most commonly due to enteric bacteria. We report three unusual cases of acute cholecystitis in which Staphylococcus aureus was the primary pathogen. Infection of the gallbladder with this organism has been rarely described and may be associated with gallstones and obstructive disease as well as acalculous cholecystitis in the setting of staphylococcal bacteremia and endocarditis. Two of our patients had multiple chronic medical conditions and were infected with oxacillin-resistant S. aureus (ORSA) suggesting nosocomial acquisition. Including our cases with a review of the literature, three of nine reports of S. aureus cholecystitis were associated with infectious endocarditis. Thus, the finding of S. aureus cholecystitis with bacteremia is rare and should prompt an investigation for a possible endovascular focus of infection.     

IgM antibodies to Epstein-Barr virus-associated membrane antigen in sera of infectious mononucleosis patients

By the indirect immunofluorescence technique, IgM antibodies to the cell surface of an Epstein-Barr virus (EBV) producer cell line, P3HR-1, were detected in sera from infectious mononucleosis (IM) patients but not in sera from patients with Burkitt lymphoma or nasopharyngeal carcinoma nor in sera from healthy adult donors having antibodies to EBV-specific viral capsid antigen (VCA). Titers of the IgM antibodies were higher in the earlier stages of IM, a pattern similar to that for IgM antibodies to VCA. The IgM antibodies to the cell surface were identified as being those against the EBV-specific membrane antigen (MA) by the following criteria: (1) The antibodies were reactive to MA-positive cell preparations but to MA-negative cell preparations. (2) Titers of the IgM antibodies were not significantly affected after absorption of sera with sheep red blood cells which could completely eliminate heterophil antibodies in the same sera. Detection of the IgM antibodies to MA may have a particular diagnostic value for providing evidence of a recent EBV infection.     

Identification of the secreted macromolecular immunogens of Staphylococcus aureus by analysis of serum

The ability of sera to recognise secreted macromolecules of Staphylococcus aureus was examined by ELISA and Western immunoblotting. Individual secreted proteins were also studied using both human sera and sera from rabbits immunised with secreted macromolecules. Patients sera showed a wide range of IgG antibody titres to secreted macromolecules and whole bacteria. Controls showed a significantly lower IgG response. Western immunoblotting revealed that a significant number of secreted proteins were recognised by circulating IgG antibodies. Surprisingly, both the sera from controls and from patients recognised similar macromolecules including a number of potential virulence factors. The major difference was in the IgG binding to a 16-kDa component, which was recognised by the majority of the sera from infected individuals, but only by a small number of sera from healthy controls. The higher incidence of antibodies recognising the 16 kDa component may be related to our earlier finding that the major bone resorbing component of S. aureus is a heterodimeric protein containing a 16-kDa subunit, the activity of which could be blocked by sera.     

Late-stage polyribitol phosphate wall teichoic acid biosynthesis in Staphylococcus aureus

Wall teichoic acids are cell wall polymers that maintain the integrity of the cellular envelope and contribute to the virulence of Staphylococcus aureus. Despite the central role of wall teichoic acid in S. aureus virulence, details concerning the biosynthetic pathway of the predominant wall teichoic acid polymer are lacking, and workers have relied on a presumed similarity to the putative polyribitol phosphate wall teichoic acid pathway in Bacillus subtilis. Using high-resolution polyacrylamide gel electrophoresis for analysis of wall teichoic acid extracted from gene deletion mutants, a revised assembly pathway for the late-stage ribitol phosphate-utilizing enzymes is proposed. Complementation studies show that a putative ribitol phosphate polymerase, TarL, catalyzes both the addition of the priming ribitol phosphate onto the linkage unit and the subsequent polymerization of the polyribitol chain. It is known that the putative ribitol primase, TarK, is also a bifunctional enzyme that catalyzes both ribitol phosphate priming and polymerization. TarK directs the synthesis of a second, electrophoretically distinct polyribitol-containing teichoic acid that we designate K-WTA. The biosynthesis of K-WTA in S. aureus strain NCTC8325 is repressed by the accessory gene regulator (agr) system. The demonstration of regulated wall teichoic acid biosynthesis has implications for cell envelope remodeling in relation to S. aureus adhesion and pathogenesis.     

The effect of immunotherapy with cell-free vaccines made from the antigens of opportunistic microorganisms on the dynamics of DNA antibody formation]

The sera of patients subjected to immunotherapy with staphylococcal vaccine and with multicomponent vaccine (i.e. the mixture of the antigenic preparations of Staphylococcus aureus, Klebsiella pneumoniae, Proteus vulgaris, Escherichia coli) were studied by the method of the enzyme immunoassay on the basis of cattle spleen DNA. Immunotherapy with staphylococcal vaccine was given to patients with dermal diseases, chronic obstructive bronchitis and pulmonary abscess. Multicomponent vaccine was introduced to patients with the infectious allergic form of bronchial asthma, moderate or severe. Immunotherapy with both preparations under study was shown to produce no accumulation of antibodies to native and denatured DNA.