Detection of serum antibodies against Chlamydia pneumoniae by ELISA

Chlamydia pneumoniae causes pneumonia and other respiratory infections in children, adolescents and adults. We tried to evaluate the diagnostic value of detection of serum antibodies by ELISA for C. pneumoniae infections in Japanese children. Serum IgG, IgA and IgM antibodies to C. pneumoniae were determined by the microimmunofluorescence (MIF) test. Serum IgG and IgA antibodies were also determined by ELISA test kits. Results obtained by ELISA were compared with those obtained by MIF test. IgG antibody to C. pneumoniae was detected in 135 (39.5%) by ELISA and in 125 (36.5%) by MIF out of 342 sera from Japanese infants and children without respiratory infections (aged from 2 months old to 15 years old). IgA antibody to C. pneumoniae was detected in 129 (37.7%) by ELISA and in 117 (34.2%) by MIF out of 342 sera tested. Of 342 specimens 113 were IgG-positive by ELISA and MIF (sensitivity: 90.4%, specificity: 89.9%, r = 0.853). Of 342 sera 28 had IgG antibody titers of 1:256 and none had titers 1:512 or higher by MIF. Of 28 infants and children a total of nine were less than 4 years of age. On the other hand, of 342 specimens 99 were IgA-positive by ELISA and MIF (sensitivity: 84.6%, specificity: 86.7%, r = 0.769). Of 342 sera 16 had IgA antibody titers of 1:256 or higher by MIF. Of 16 infants and children, ten were less than 4 years of age. ELISA had excellent sensitivity and specificity relative to MIF test for detection of IgC and IgA antibodies to C. pneumoniae. It was suggested that C. pneumoniae infection in Japanese infants and children under 4 years of age was not infrequent.     

Comparison of serological tests for detection of Mycoplasma gallisepticum antibodies in eggs and chicks hatched from experimentally infected hens

Specific pathogen free hens and males were experimentally infected with Mycoplasma gallisepticum. Eggs were then collected, and a part was incubated and set for hatching. Mycoplasma cultures were performed on infected adults and antibodies to MG were analysed by use of slide agglutination (SA) test and commercial ELISA tests on adults and chicks sera and on yolks from non incubated eggs. Both ELISA tests could detect antibodies in yolks from non incubated eggs laid three weeks after infection. SA and the three ELISA tests revealed positive sera in chicks hatched from eggs laid as soon as one week after infection.     

Detection of Chlamydia trachomatis antibodies by 2 novel tests: rELISA and peptide EIA

The performance of 2 newly developed enzyme immunoassays (EIAs) intended for the routine serological diagnosis of chlamydial infections was evaluated. rELISA is based on a recombinant lipopolysaccharide antigen which detects chlamydia genus-specific antibodies, and Chlamydia trachomatis EIA is based on a peptide derived from major outer membrane protein and is therefore species-specific. Both tests distinguished patients with tubal factor infertility (TFI) or pelvic inflammatory disease (PID) from the controls. The prevalence of IgA antibodies was higher for the PID patients than for the TFI patients; the finding indicates a more active state of infections for the PID patients. Furthermore, C. trachomatis EIA detected more IgG antibodies in the TFI patients than in patients with non-tubal factor infertility. In conclusion, rELISA detected chlamydial antibodies in general, and C. trachomatis EIA detected species-specific antibodies. These EIA tests may be useful in the serodiagnosis of chlamydial infection.     

Evaluation of serological tests for the detection of pseudorabies gE antibodies during early infection

Two enzyme-linked immunosorbent assays (ELISAs) and a particle concentration fluorescence immunoassay (PCFIA) were compared for their ability to detect antibodies against pseudorabies virus (Aujeszky's disease virus) glycoprotein E (gE) in the early stages of infection in pigs previously vaccinated with gE-deleted pseudorabies vaccines. Seventy pigs were included in the study. Five groups of 6 pigs each were vaccinated with one of 5 different pseudorabies virus (PRV) gE-deleted vaccines, and subsequently infected intranasally with 10(5.6) TCID50 of the Iowa 4892 pneumotropic strain of PRV. This entire procedure was repeated using 10(4.6) TCID50 of the Rice strain of PRV. Five unvaccinated control pigs were also challenged with each virus strain. Three control pigs died before seroconverting, leaving 67 pigs for comparison. Blood samples were drawn from experimentally inoculated pigs on the day of vaccination, the day of challenge, and on 4-10, 14, and 21 days postchallenge (DPC). Serology test sensitivity estimates and comparisons among tests were made for each sampling day. Results of this study demonstrated differences among the tests in the time from inoculation to initial antibody detection, and the time to detect 50% and 75% of the infected pigs. The average time until first detection of pigs as seropositive for gE antibodies by PCFIA was 7.5 DPC. The blocking ELISA detected pigs as seropositive an average of 8.8 DPC, and the indirect ELISA first detected gE antibodies by 9.3 DPC. Fifty percent of the pigs were detected as seropositive by days 7, 8, and 9 for the PCFIA, blocking ELISA, and indirect ELISA, respectively. Similarly, 75% of the pigs were detected as seropositive by days 8, 9, and 10 for the PCFIA, blocking ELISA, and indirect ELISA, respectively. All pigs were detected as seropositive by 14 DPC for all 3 tests.     

Chlamydia trachomatis antibodies in serum and ejaculate of male patients without acute urethritis

The effectiveness of the Pediatric Symptom Checklist (PSC) as a psychosocial screening measure to meet Federal Medicaid/Early and Periodic Screening, Diagnosis, and Treatment (EPSDT) requirements was examined in 117 low-income preschool (aged 4-5 years old) Hispanic children during well-child examinations in three clinics over an 8-month period. The PSC identified 7% of the sample as at risk for psychosocial problems. The PSC was significantly associated with parental ratings of the children's problems in functioning, with pediatric clinicians' decisions to make mental health referrals, with degrees of associations similar to those found between PSC scores, and with the same measures with school-aged children in the same clinics. Cronbach's alpha was high (r = .87) and virtually identical in English, Spanish, oral, and written formats. Although it identified a slightly lower rate of psychosocial problems in 4-5-year-olds than it had in school-aged children, the PSC appeared to provide an effective method of screening for psychosocial problems during EPSDT examinations.     

In vitro activity of dirithromycin against Chlamydia trachomatis

Dirithromycin is a new macrolide antibiotic with an active metabolite, erythromycylamine. We evaluated the in vitro activities of both drugs against 16 isolates of Chlamydia trachomatis and compared them with that of doxycycline. In vitro susceptibility testing was performed with McCoy cell monolayers. The MIC was defined as the lowest concentration of antibiotic without inclusions. The MBC was defined as the lowest concentration of antibiotic yielding no inclusions after passage onto 24-h-old antibiotic-free McCoy cell monolayers. Dirithromycin and erythromycylamine appeared to be equally effective against these 16 strains of C. trachomatis (MIC for 90% of strains tested, 1 mg/ml; MBC for 90% of strains tested, 2 micrograms/ml). Both were less active than doxycycline (MIC for 90% of strains tested, 0.06 micrograms/ml; MBC for 90% of strains tested, 0.12 micrograms/ml). The combination of dirithromycin and erythromycylamine appeared to be additive.     

Chlamydia trachomatis antibody detection and diagnosis of reactive arthritis

OBJECTIVE: To investigate whether determining the presence of serum or synovial fluid (SF) IgG and IgA of anti-Chlamydia antibodies with two recent commercially available enzyme-linked immunosorbent assays (ELISA) using synthetic peptides or recombinant antigen could be helpful to detect possible Chlamydia trachomatis (CT)-involved disease in rheumatological patients without evidence of urogenital CT infection. METHODS: The prevalence of such antibodies was determined in samples from patients with well-defined disease, i.e. CT sexually acquired arthritis and from patients with other inflammatory arthropathies unrelated to CT. RESULTS: When considering IgG and/or IgA anti-MOMP or anti-LPS antibodies, a sensitivity of 100% was obtained for serum and SF samples, but with a low specificity. A sensitivity and a specificity equal or close to 80% were observed for the SF IgG anti-MOMP antibodies. CONCLUSION: Clinically, the most appropriate determination was the SF IgG anti-MOMP antibodies. This commercially available ELISA test could be useful for the diagnosis of probable CT reactive arthritis.     

Comparison of two serological methods and a polymerase chain reaction-enzyme immunoassay for the diagnosis of acute respiratory infections with Chlamydia pneumoniae in adults

Chlamydia pneumoniae is a common respiratory tract pathogen. Serological methods currently used for the diagnosis of C. pneumoniae infection lack specificity, give ambiguous results from a single serum sample and often provide only a retrospective diagnosis. A prospective study was undertaken to assess whether PCR could be a useful addition to the serological techniques routinely practised for diagnosis. This study investigated 68 adult patients with a diagnosis of acute respiratory infection. Acute and convalescent serological determination of antibodies to C. pneumoniae were performed by means of an rELISA test and a micro-immunofluorescence (MIF) test. Nasopharyngeal aspirates or bronchoalveolar lavage specimens and bronchial aspirates obtained from the 68 patients were evaluated by PCR-enzyme immunoassay (PCR-EIA) for the presence of C. pneumoniae and by immunofluorescence assay and cell culture for virus identification. Mycoplasma pneumoniae serology was also performed. Eight patients (11.8%) were positive by either rELISA or PCR-EIA, or both, with an infection rate of 5 (18.5%) of 27 in patients with community-acquired pneumonia, 2 (9%) of 22 in asthmatic patients and 1 (5%) of 19 in patients with an exacerbation of chronic obstructive pulmonary disease. Serological evidence of acute infection was found in four of these patients with the rELISA test and in three others with the MIF test. PCR-EIA detected C. pneumoniae DNA in four specimens, but there were concordant results with both rELISA and PCR-EIA in only one patient A positive PCR-EIA was also obtained in a patient who did not show an antibody response in acute serum. The discrepancy between serological and PCR-EIA results reflects the difficulties in routine laboratory diagnosis of C. pneumoniae infection and the necessity for further studies with optimised techniques.     

Detection of serum antibodies to Chlamydia trachomatis in patients with chlamydial and nonchlamydial pelvic inflammatory disease by the IPAzyme Chlamydia and enzyme immunoassay

A novel serological test, IPAzyme Chlamydia (Savyon Diagnostics Ltd., Beer Sheva, Israel), was compared with an enzyme immunoassay (EIA) for the ability to detect serum immunoglobulin G and A antibodies in the diagnosis of acute chlamydial pelvic inflammatory disease. In comparison with cell culture, which is the "gold standard," IPAzyme Chlamydia and EIA exhibited sensitivities of 63 and 68% and specificities of 76 and 87%, respectively. Thus, IPAzyme Chlamydia offers no advantages over the EIA, and neither serological test can be recommended for the diagnosis of acute Chlamydia trachomatis infection. So far, conventional cell culture remains the most reliable diagnostic test for chlamydial pelvic inflammatory disease.     

Comparison of performances of two commercially available tests, a PCR assay and a ligase chain reaction test, in detection of urogenital Chlamydia trachomatis infection

The diagnostic performance of a PCR test (Roche Cobas Amplicor CT/NG Test) and that of a ligase chain reaction (LCR) test (Abbott LCx Chlamydia trachomatis assay) were compared by using endocervical and urethral swab specimen culture as a reference test. First-void urine (FVU) and endocervical and urethral swab specimens were collected from 1,015 unselected patients attending a sexually transmitted disease clinic and a clinic for adolescents in Helsinki, Finland. Chlamydia trachomatis was cultured from samples from the endocervix or urethra. PCR was performed with fresh and frozen urine and the culture transport medium. LCR was performed with fresh and frozen urine and LCx swab transport medium. Diagnostic consistency and diagnostic accuracy were statistically tested. The test results were identical for 984 patients (97%). Discrepant results were observed for 31 patients. Overall, LCR and PCR showed excellent kappa coefficients of consistency for both swab and FVU specimens (0.93 and 0.95, respectively). Sixty-one patients (6%) were culture positive. Testing of FVU by LCR or PCR increased the overall positivity rates to 7.0 and 7.7%, respectively. While PCR of FVU detected the greatest number of C. trachomatis infections (sensitivity, 96.1%), for some PCR-positive FVU specimens the results could not be confirmed (specificity, 99.6%). PCR and LCR were more sensitive than culture (sensitivities, 92 and 93% versus 79% for culture) in the diagnosis of genital C. trachomatis infection. In conclusion, both tests can be recommended for use in the clinical laboratory and for the screening of asymptomatic C. trachomatis infections.     

Rapid detection of Chlamydia pneumoniae by PCR-enzyme immunoassay

Chlamydia pneumoniae is an important human respiratory pathogen. Laboratory diagnosis of infection with this organism is difficult. To facilitate the detection of C. pneumoniae by PCR, an enzyme immunoassay (EIA) for analysis of PCR products was developed. Biotin-labeled PCR products generated from the 16S rRNA gene of C. pneumoniae were hybridized to a digoxigenin-labeled probe and then immobilized to streptavidin-coated microtiter plates. Bound PCR product-probe hybrids were detected with antidigoxigenin peroxidase conjugate and a colorimetric substrate. This EIA was as sensitive as Southern blot hybridization for the detection of PCR products and 100 times more sensitive than visualization of PCR products on agarose gels. The diagnostic value of the PCR-EIA in comparison to cell culture was assessed in throat swab specimens from children with respiratory tract infections. C. pneumoniae was isolated from only 1 of 368 specimens tested. In contrast, 15 patient specimens were repeatedly positive for C. pneumoniae by PCR and Southern analysis. All of these 15 specimens were also identified by PCR-EIA. Of the 15 specimens positive by 16S rRNA-based PCR, 13 specimens could be confirmed by omp1-based PCR or direct fluorescent-antibody assay. Results of this study demonstrate that PCR is more sensitive than cell culture for the detection of C. pneumoniae. The EIA described here is a rapid, sensitive, and simple method for detection of amplified C. pneumoniae DNA.     

A role for interleukin-6 in host defense against murine Chlamydia trachomatis infection

Interleukin-6-deficient (IL-6(-/-)) knockout mice had significantly increased Chlamydia trachomatis levels in lung tissue and increased mortality compared to B6129F2/J controls early after intranasal infection. Gamma interferon production and chlamydia-specific antibody levels were consistent with a decreased but reversible Th1-like response in IL-6(-/-) mice. IL-6 is needed for an optimal early host response to this infection.     

Association of Chlamydia pneumoniae IgA antibodies with recently symptomatic asthma

To determine whether recently diagnosed adult-onset asthma is associated with serologic evidence of chronic Chlamydia pneumoniae infection, we performed a case-control study in a primary care clinic of cases with asthma (25 adults reporting first symptoms of asthma within 2 years of enrollment) and 45 concurrently enrolled sex and age (+/- 10 years) matched non-asthmatic controls with normal pulmonary function. C. pneumoniae-specific IgA, IgG and IgG4 antibodies, and circulating immune complexes (CIC) were measured by microimmunofluorescence testing. Results showed that frequencies of IgG titres > or = 16 (92%), IgG4 titres > or = 16 (20%) and CIC > or = 4 (60%) in asthma patients were not significantly different from those of controls. However, asthmatics had a significantly higher prevalence of C. pneumoniae-specific IgA titres > or = 10 (72% of cases vs 44% of controls, P < 0.05). After adjustment for the effects of age, sex and smoking, the odds ratio for an association of IgA and asthma was 3.7 (95% confidence interval 1.2-11.5). We conclude that recently symptomatic reversible airway obstruction in adults is associated with the presence of C. pneumoniae-specific IgA antibodies, a proposed indicator of chronic respiratory C. pneumoniae infection.     

Improvement of cervical Chlamydia detection in asymptomatic family planning attendees by using Amplicor Chlamydia trachomatis assay

We evaluated the Amplicor C. trachomatis PCR assay (Roche Molecular Systems, N.J.) for the diagnosis of cervical infection in asymptomatic women attending a family planning clinic, aged between 18 and 25 years. Culture onto McCoy cells with fluorescent monoclonal staining was the reference system. Cervical specimens from 485 women were tested. The prevalence of C. trachomatis was 10.5% by culture and 11% by Amplicor. No specimen was positive by culture and negative by PCR. Three PCR-positive, culture-negative specimens were positive by MOMP-PCR and a second plasmid-based PCR. The resolved sensitivity of PCR and culture were 100% and 94.5%, respectively. Specificities for both were 100%, positive and negative predictive values for culture were 100% and 99.3%. Total test efficiency was 99.4%. The Amplicor C. trachomatis assay gave very clear results, quite above or below the cut-off value, and showed high sensitivity and specificity, improved ease of handling and represented a good alternative to culture for large scale diagnosis of asymptomatic C. trachomatis infection.     

Circulating antibodies to Chlamydia trachomatis in women: relationship to antisperm and antichlamydial antibodies in semen of male partners

The relation between antibodies to Chlamydia trachomatis and spermatozoa in sera of 112 asymptomatic female partners of infertile couples with no history of C.trachomatis infections and antichlamydial antibodies in semen or antisperm antibodies on ejaculated spermatozoa of their male partners was examined. Samples were tested for immunoglobulin (Ig)A and IgG antibodies to C.trachomatis by enzyme-linked immunosorbent assay; antisperm antibodies in sera and on motile spermatozoa were assayed by immunobead binding. IgG antibodies to C.trachomatis were detected in 24 (21.4%) of the women; only five (4.5%) women were positive for antichlamydial IgA. Antichlamydial IgG was detected in sera from 10 (40.0%) of 25 women whose partners had antichlamydial IgA in semen as opposed to 14 (16.1%) of 87 women whose partners' semen were negative for this antibody (P = 0.02). Similarly, antichlamydial IgG was detected in sera from five (50%) of 10 women whose partners had antichlamydial IgG in semen as opposed to 19 (18.6%) of 102 women whose partners' semen lacked this antibody (P = 0.03). There was no relation between antichlamydial antibodies in women and circulating antichlamydial antibodies in men. A strong correlation (P = 0.001) was observed between IgG antichlamydial antibodies in a woman's serum and antisperm antibodies on ejaculated spermatozoa of her partner [8 of 14 (57.1%) versus 16 of 98 (16.3%)]. Conversely, antichlamydial antibodies in a woman's serum was unrelated to the presence of antisperm antibodies in either her own serum or her partner's serum. The data demonstrate that chlamydial infections of the male genital tract, which are associated with antisperm antibody formation on ejaculated spermatozoa, are likely to be transmitted to the female partner. In contrast, the presence of antichlamydial antibodies in sera does not necessarily appear to indicate an infection of the genital tract and is not associated with the heterosexual transmission of C.trachomatis.     

Chlamydia psittaci and Chlamydia pneumoniae infection in Caceres

OBJECTIVE: The study of the clinical and epidemiological characteristics of infectious process caused by Chlamydia pneumoniae and Chlamydia psittaci in our medium, Cáceres. METHOD: We are reviewed retrospectively clinical aspects of the patients with infections due to Chlamydia in the las five years. We accepted patients with compatible symptoms and serologic demonstration of recent infection with conventional complement fixation and/or microimmunofluorescence assay, the last used to distinguish Chlamydia pneumoniae. RESULTS: We are studied sixteen patients (9 males and 7 females), sixth median age 46.6 (26-70). Fifteen patients was diagnosed in winter. We found five cases of Chlamydia pneumoniae and three of Chlamydia psittaci. In the other eight cases we didn't distinguish between Chlamydia pneumoniae and Chlamydia psittaci. All patients had fever, accompanied by lung symptoms and pulmonary infiltrates in the 75% of them. The most frequent clinical information was the discord between the pulse and temperature (81%). Splenomegaly was observed in three patients (19%) being the diagnosis of them psitacosis. Nine patients had respiratory insufficiency and eight (50%) disturbance in hepatic enzymes. The clinical presentation in one patients was as unknown origin fever. CONCLUSIONS: The infection produced by Chlamydia in the hospitalary medium isn't much diagnosed in our unit. The months of winter favour the infection. We think that splenomegaly is the only different characteristic in these infections, suggesting psitacosis.