Antibody response to chlamydiae in children with asthma and respiratory illness.

No relation between the occurrence of antibodies to chlamydial agents and asthma in children was found. In asthmatic children, the antibodies to Chlamydia trachomatis occurred in 3.1% and to Chlamydophila pneumoniae in 22.7%, whereas in a control group of children without asthma or other allergic disease in 2.3% and 24.0%, respectively. The occurrence of antibodies of IgA and IgG classes to C. pneumoniae was also very similar; its rise was age-dependent. On the other hand, in the group of children in a pre-school age with respiratory tract infection, anti-chlamydial antibodies were demonstrated significantly more often (18.5% of IgG antibodies to C. trachomatis, 20.0% of IgM antibodies to both C. trachomatis and C. pneumoniae) than in those suffering from other, non-respiratory illness (3.9% of the former and 5.9% of the latter antibodies). However, in these children, we did not succeed in detection of C. trachomatis in conjunctival and nasopharyngeal smears by PCR. Nevertheless, chlamydial agents (C. trachomatis in infants, C. pneumoniae in pre-school children) should be taken into consideration in a differential diagnosis of respiratory tract inflammation.

Detection of antibodies to Chlamydophila pneumoniae by immunoblotting in patients with cardiovascular diseases.

Chlamydophila pneumoniae, one of the most prevalent human pathogens worldwide, is not only a significant cause of pneumonia, but may also be associated with cardiovascular diseases (CVD) as suggested by multiple studies. A total of 228 sera from CVD patients with hypertension, ischemic heart disease or previous reconstructive vascular surgery were screened for the presence of anti-C. pneumoniae IgG and IgA antibodies by ELISA. Out of 150 positive serum samples, 80 with similar IgG and IgA levels were investigated by immunoblot (IB). IgG antibodies were directed predominantly against the 35 kDa and 39 kDa proteins as well as 50-54 and 56-60 kDa proteins of C. pneumoniae. IgA antibodies reacted most frequently with the 50-54 and 56-60 kDa proteins.

Role of structural variations of polysaccharide antigens in the pathogenicity of Gram-negative bacteria.

Pathogenic bacteria employ many strategies to overcome the host immune system for extended survival and propagation in their hosts. Components of the bacterial outer-membrane play an important role in this process. When invading the host, Gram-negative bacteria often use a strategy, known as phase variation, that involves a reversible change in antigenic determinants, frequently polysaccharides. This means that the genes encoding the outer-membrane antigens undergo reversible changes within repeated simple DNA sequence motifs. The antigenic structure of the bacterial outer-membrane is influenced by the character of the host immune system, as well as by the targets for bacterial invasion. When the selection pressure of the immune system is absent or weak, bacteria can fail to synthesise the outer-membrane antigens, which are not needed at that time. Smooth-to-rough (S-R) mutation, an economical and often irreversible process in some Gram-negative bacteria, involves the gradual shortening of the lipopolysaccharide (LPS) O-chain. Under certain conditions, e.g., propagation in embryonated eggs or cell lines, some bacteria will cease synthesis of the complete LPS O-chain because it is an energy-demanding process. A type of gradual shortening of the LPS O-chain by Coxiella burnetii, traditionally called phase variation, is used in serological tests for the diagnosis of Q fever. This review discusses the role and function of polysaccharides, especially LPS produced by some Gram-negative bacteria, in bacterial survival.

Chlamydia pneumoniae antibodies and markers of inflammation in patients with cardiovascular diseases.

BACKGROUND: Chlamydia pneumoniae is suggested to be associated with cardiovascular diseases. OBJECTIVES: To study the presence of IgG, IgA anti-C. pneumoniae antibodies, interleukin-6 (IL-6), and C-reactive protein (CRP) as markers of previous C. pneumoniae infection and inflammation, in sera of patients with acute myocardial infarction (AIM), hypertension (HT), and coronary heart disease (CHD). METHODS: Determination of these markers by ELISA method. RESULTS: Proportion of samples containing both IgG and IgA antibodies as well as IL-6 was significantly higher in all groups of patients than in a control group. The CRP was significantly higher in patients with AIM and HT, however, in other patients, the proportion of positive samples depended on the chosen cut-off value. CONCLUSIONS: The results obtained indicate the feasibility of following chlamydial antibodies on higher number of serum samples extended to direct detection of C. pneumoniae in blood and vascular tissue (Tab. 2, Ref. 24).

Guidelines for the diagnosis of tick-borne bacterial diseases in Europe.

Ticks are obligate haematophagous acarines that parasitise every class of vertebrate (including man) and have a worldwide distribution. An increasing awareness of tick-borne diseases among clinicians and scientific researchers has led to the recent description of a number of emerging tick-borne bacterial diseases. Since the identification of Borrelia burgdorferi as the agent of Lyme disease in 1982, 11 tick-borne human bacterial pathogens have been described in Europe. Aetiological diagnosis of tick-transmitted diseases is often difficult and relies on specialised laboratories using very specific tools. Interpretation of laboratory data is very important in order to establish the diagnosis. These guidelines aim to help clinicians and microbiologists in diagnosing infection transmitted by tick bites and to provide the scientific and medical community with a better understanding of these infectious diseases.

Serum markers of liver fibrogenesis, and liver histology findings in patients with chronic liver diseases.

AIM OF THE STUDY: Investigation of the relationships between the grade and stage of chronic liver diseases irrespective of their etiology using some novel serum markers of liver fibrogenesis, the "classical" serum markers of liver necro-inflammatory injury (such as transaminases), and the histomorphological evaluation of liver biopsies. METHODS: Markers of liver fibrogenesis: serum metalloproteinase 1 (MMP-1), tissue inhibitor of MMP-1 (TIMP-1), and N-terminal propeptide of the procollagen III (PIIINP); "liver function tests" (LFTs): bilirubin, transaminases ALT, AST; ALP, GMT; and liver morphology findings: necro-inflammatory activity, fibrosis; were studied in the series of 32, 'naive', i.e. yet untreated patients (women/men--11/21) with various CLDs: chronic viral hepatitis B or C 13 (CHB 3, CHC 10), non-alcoholic steatohepatitis 9, liver steatosis 4, primary biliary cirrhosis 5, drug-induced hepatitis. The diagnoses were based on the clinical, laboratory and liver imaging (ultrasonography) findings and confirmed by the liver biopsy. CONCLUSIONS: Investigation of liver fibrogenesis serum markers (PIIINP, MMP-1, TIMP-1) in patients with various CLDs has shown statistically significant correlations of these parameters with "classical" serum markers of liver necro-inflammation (ALT, AST) and the results of histomorphological evaluation of the necro-inflammatory activity (parameters NAI, MEF) and fibrosis (parameter FI) in liver biopsies. (Tab. 4, Ref. 31.)

Detergent extract from chlamydial elementary bodies used as antigen for enzyme-linked immunosorbent assay.

Soluble antigen (SA) from chlamydial elementary bodies (EBs) was extracted with N-lauroylsarcosine. The extracted SA composed of lipopolysaccharide (LPS) and proteins was compared with EBs using an enzyme-linked immunosorbent assay (ELISA). Patient sera from natural chlamydial infections exhibited ELISA mean absorbance (A(492) and A(405/650)) values 2-5 times higher with SA than with EBs, resulting in a better discrimination between positive and negative human sera.

Q fever--still a query and underestimated infectious disease.

Coxiella burnetii (C.b.) is a strictly intracellular, Gram-negative bacterium. It causes Q fever in humans and animals worldwide. The animal Q fever is sometimes designated "coxiellosis". This infection has many different reservoirs including arthropods, birds and mammals. Domestic animals and pets, are the most frequent source of human infections. Q fever may appear basically in two forms, acute and chronic (persistent). The latter form of Q fever in animals is characteristic by shedding C.b. into the environment during parturition or abortion. Human Q fever results usually from inhalation of contaminated aerosols originating mostly from tissue and body fluids of infected animals. Q fever may appear in humans either in an acute form accompanied mainly by fever (pneumonia, flu-like disease, hepatitis) or in a chronic form (mainly endocarditis). Diagnosis of Q fever is based on isolation of the agent in cell culture, its direct detection, namely by PCR, and serology. Detection of high phase II antibodies titers 1-3 weeks after the onset of symptoms and identification of IgM antibodies are indicative to acute infection. High phase I IgG antibody titers >800 as revealed by microimmunofluorescence offer evidence of chronic C.b. infection. For acute Q fever, a two-weeks-treatment with doxycycline is recommended as the first-line therapy. In the case of Q fever endocarditis a long-term combined antibiotic therapy is necessary to prevent relapses. Application of Q fever vaccines containing or prepared from phase I C.b. corpuscles should be considered at least for professionally exposed groups of the population. Infections caused by C.b. are spread worldwide and may pose serious and often underestimated health problems in human but also in veterinary medicine. Though during the last decades substantial progress in investigation of C.b. has been achieved and many data concerning this pathogen has been accumulated, some questions, namely those related to the pathogenesis of the disease, remain open.

[Comparison of various diagnostic kits and antigens for the detection of chlamydial antibodies in the serum of patients with respiratory diseases] . / Porovnanie rôznych diagnostických súprav a antigénov pre dokaz chlamýdiových protilátok v sérach pacientov s respiracnými ochoreniami.

Examination of sera of patients with respiratory diseases did not reveal any substantial difference in detection of chlamydial antibodies by ELISA using two commercial kits (Chlamydia STAT and rRLISA) demonstrating IgG antibodies and two corpuscular antigens prepared from Chlamydia psittaci and Chlamydia pneumoniae in the Institute of Virology, SAS, in Bratislava detecting whole serum antibodies. However, higher sensitivity of rELISA diagnostic kit was found when analyzing immunoglobulin classes, namely in case of IgG antibodies. Results of ELISA were in a great majority of cases, confirmed by examination of sera by microimmunofluorescence test, corpuscular antigens from the Institute of Virology being somewhat more sensitive than the commercial Micro-IF test kit. (Tab. 4, Ref. 23.)

Participation of Chlamydia pneumoniae in an epidemic of respiratory infections in Slovakia.

In human sera collected in three regions of Slovakia during an epidemic of respiratory infections, both genus-specific chlamydial and species-specific anti-Chlamydia pneumoniae antibodies, as detected by enzyme-linked immunosorbent assay and microimmunofluorescence test, respectively, were found. Based on seroconversion or significant rise of antibody titers and detection of antibodies of IgM class, an acute C. pneumoniae infection was indicated in 21 of 298 (7.0%) patients tested. The results obtained bring the first evidence on the role of C. pneumoniae as respiratory pathogen in Slovakia.

Rickettsial diseases and their serological diagnosis.

Rickettsial diseases (typhus and spotted fever group rickettsioses, scrub typhus and Q fever) may pose a serious public health problem, namely when they are non-diagnosed or misdiagnosed. Although rickettsiae can be isolated from or detected in clinical specimens, serological tests still remain an indispensable tool in the diagnosis of rickettsial diseases. The complement fixation test widely used in the past is being replaced by other tests which make differentiation of immunoglobulin classes possible. Of these tests microimmunofluorescence is considered the test of choice followed by the latex agglutination, indirect hemagglutination, immunoperoxidase assay, and enzyme-linked immunosorbent assay. The last one is also suitable for seroepidemiological studies. Immunoblot analysis can be used to confirm the results of other tests. The use of the low-specific and low-sensitive Weil-Felix test should be reserved only for situations in which other serologic tests are not available.

Comparison of manganese superoxide dismutase precursor induction ability in human hepatoma cells with or without hepatitis B virus DNA insertion.

In a previous study (Hajnická et al., Acta virol. 38, 55-57 (1994)), we described synthesis of a 23 K protein in high amounts in the PLC/PRF/5 human hepatoma cell line after stimulation with sera of patients suffering from liver cirrhosis. In this study we identified this protein as manganense superoxide dismutase (Mn-SOD). When PLC/PRF/5 cells stimulated by various cytokines (interleukin-1 alpha (IL-1 alpha), IL-1 beta, tumor necrosis factor-alpha (TNF-alpha), TNF-beta, IL-6, tumor growth factor-alpha (TGF-alpha), TGF-beta, and interferon-gamma (IFN-gamma) were compared, the most effective was IL-1, followed by TNF-alpha and IL-6. Other cytokines had no effect on the stimulation of Mn-SOD. IL-1 alpha was selected for stimulation of Mn-SOD production in four human hepatoma cell lines (PLC/PRF/5, Hep-3B, Hep-G2 and Sk-Hep 1). Maximum Mn-SOD production occurred in PLC/PRF/5 cells. In other cell lines, Mn-SOD production was lower, reaching 35.7% and 31.5% in Hep-3B and Sk-Hep-1 cells, respectively, while it was only 4.3% in Hep-G2 cells.

Q fever in Bulgaria and Slovakia.

As a result of dramatic political and economic changes in the beginning of the 1990s, Q-fever epidemiology in Bulgaria has changed. The number of goats almost tripled; contact between goat owners (and their families) and goats, as well as goats and other animals, increased; consumption of raw goat milk and its products increased; and goats replaced cattle and sheep as the main source of human Coxiella burnetii infections. Hundreds of overt, serologically confirmed human cases of acute Q fever have occurred. Chronic forms of Q fever manifesting as endocarditis were also observed. In contrast, in Slovakia, Q fever does not pose a serious public health problem, and the chronic form of infection has not been found either in follow-ups of a Q-fever epidemic connected with goats imported from Bulgaria and other previous Q-fever outbreaks or in a serologic survey. Serologic diagnosis as well as control and prevention of Q fever are discussed.

Cross-reactivity between Coxiella burnetii and chlamydiae.

A cross-reactivity among some strains of Coxiella burnetii and chlamydiae with immune rabbit and mouse sera in ELISA and immunoblot analysis was observed. In the latter, the cross-reactivity disappeared after a treatment of C. burnetii or C. psittaci with proteinase K, which indicates that only proteins were involved. The observed cross-reactivity was not influenced by host chick embryo yolk sac proteins. After adsorption of immune rabbit sera with homologous corpuscular antigens the cross-reactivity disappeared. The possibility of influence of such cross-reactivity on serological diagnosis of C. burnetii or chlamydiae infections is discussed.

Clinical and serological analysis of a Q fever outbreak in western Slovakia with four-year follow-up.

In the spring of 1993, an outbreak of respiratory infection affected 113 persons (103 males) in Jedl'ové Kostol'any, a village located in a hilly area of western Slovakia. Q fever, manifested as a flu-like illness with atypical pneumonia and hepatic involvement, was diagnosed using four serological tests (microimmunofluorescence, microagglutination, complement fixation, and enzyme immunoassay). Aborting goats were proven as a source of infection. During a 4-year follow-up study, no chronic form of Q fever could be demonstrated, either clinically or by tests to detect phase I Coxiella burnetii antibodies.

Suitability of various Coxiella burnetii antigen preparations for detection of serum antibodies by various tests.

Comparison of four serological tests (complement fixation (CF) test, microagglutination (MA) test, microimmunofluorescence (MIF) test, and enzyme-linked immunosorbent assay (ELISA)) for detection of post-infection antibody response in human and animal sera revealed a low sensitivity of the CF test with acute Q fever human, goat and sheep sera but not with chronic Q fever human sera and sera of aborting cows. The remaining three tests gave similar results with human (both acute and chronic) and cow sera, but the ELISA was more sensitive than the MA and MIF tests with goat and sheep sera. A treatment of phase I Coxiella burnetii (C.b.) cells with chloroform-methanol, potassium periodate and trichloroacetic acid (TCA), and mild acidic hydrolysis did not result in increase of the sensitivity of the tests when compared with the natural phase I and phase II C.b. cells, respectively. The suitability of various C.b. antigen preparations for the abovementioned serological tests with various sera is discussed.

Monoclonal antibody based differentiation of Coxiella burnetii isolates.

Isolates of Coxiella burnetii from different geographic regions in Europe, USA, Japan and Africa were compared in their binding properties to the monoclonal antibody (MoAb) 1/4/H directed against the lipopolysaccharide (LPS) of C. burnetii strain Priscilla. Immunoblot analysis and enzyme-linked immunosorbent assay (ELISA) revealed different binding patterns of C. burnetii isolates under study. Most of the isolates tested did react with MoAb 1/4/H. Only four of 20 groups of isolates and one isolate of an otherwise positively reacting group did not react with MoAb 1/4/H. The results indicate a significant variation of LPS structure of the C. burnetii isolates studied.

Serological examination of human and animal sera from six countries of three continents for the presence of rickettsial antibodies.

Forty samples each of human sera collected in Guinea Bissau, Cape Verde, El Salvador and Iran, and animal sera (goat and cattle from Sri Lanka and sheep from Tanzania) were examined for the presence of antibodies to typhus group (TG) rickettsiae, spotted fever group (SFG) rickettsiae and Coxiella burnetii by enzyme-linked immunosorbent assay (ELISA) and indirect fluorescent antibody (IFA) test. Of human sera tested, a higher proportion of positive sera were found with ELISA and IFA test for TG, SFG rickettsiae and C. burnetii in El Salvador (42.5 vs 20.0%, 40.0 vs 32.5%, and 27.5 vs 27.5%, respectively) and in Iran (25.0 vs 15.0%, 45.0 vs 27.5%, and 27.5 vs 25.0%, respectively), than in Guinea Bissau and Cape Verde, where they were less than 20.0% except for antibodies to SFG rickettsiae in Guinea Bissau (25.0% with ELISA and 20.0% with IFA test). While all animal sera were negative for the presence of antibodies to TG rickettsiae, a high proportion of sera from Sri Lanka reacted in ELISA and IFA test with SFG rickettsiae and C. burnetii (37.5 vs 20.0% and 27.5 vs 25.0% for goat sera, and 40.0 vs 30.0%, and 17.5 vs 15.0% for cattle sera, respectively). The results obtained indicate that the studied rickettsial diseases can be spread in given territories and may pose a public health problem requiring greater attention than has been paid so far. The suitability of ELISA and IFA test for serological survey of rickettsial antibodies is discussed.

Improved method of isolation of Coxiella burnetii proteins.

An improved method of isolation of Coxiella burnetti proteins was developed. It consists of a combination of detergent (sodium dodecyl sulphate (SDS) or sodium deoxycholate (DOC) and hot phenol treatments. The resulting phenol phase (PP) contained either lipopolysaccharide-(LPS) free proteins (DOC extraction) or proteins contaminated with LPS (SDS extraction), while the water phase (WP) contained LPS. Isolated C. burnetii proteins induced in mice and rabbits antibodies reacting in immunoblot analysis with both phase I and II C. burnetii corpuscles. A rabbit serum against C. burnetii prepared by DOC-phenol extraction did not react with purified I C. burnetii LPS in immunoblot analysis.