Pertti Terho

Analysis and detection of chlamydial DNA

Elementary bodies of lymphogranuloma venereum (LGV) strains of Chlamydia trachomatis contain, in addition to the genomic DNA, a 6.7 kb plasmid. The plasmid from serovar L2 (434-B) was cloned at the BamHI site of pBR327 into Escherichia coli and a restriction cleavage map of this pLGV125 recombinant plasmid determined. All 15 C. trachomatis serovars contained DNA sequences that hybridized with pLGV125. When total DNA from L2 elementary bodies was used as a probe in Southern blotting and spot hybridization, serovars L1, L2 and L3 exhibited significant homology. The detection level of homologous DNA was 100 pg and LGV DNA was detectable in infected cells when total L2 probe was used in the nucleic acid hybridization test. These DNA probes may be useful as investigative and diagnostic reagents for C. trachomatis.

Detection of Chlamydia trachomatis in clinical specimens by nucleic acid spot hybridization.

A nucleic acid spot hybridization assay was used to detect Chlamydia trachomatis DNA. The hybridization probes included DNA isolated from elementary bodies of lymphogranuloma venereum (LGV) strains and cloned fragments of both chromosomal and plasmid DNA. The sensitivity of the test was in the range 10 to 100 pg homologous DNA and 10 in vitro infected cells. Cross-reactivity with bacterial DNA was avoided when purified chlamydia-specific DNA fragments were used as probes. C. trachomatis was detectable in most of the clinical specimens with large amounts of infectious particles. Also some isolation-negative specimens gave a positive signal in the test.

Cervical and serum IgA and serum IgG antibodies to Chlamydia trachomatis and herpes simplex virus in threatened abortion: a prospective study

Summary. The aetiological role of Chlamydia trachomatis (CT) and herpes simplex virus (HSV) was investigated in 189 patients with threatened abortion. Assessment of infection was based on isolation, and on determination of serum immunoglobulin (Ig)G and IgA antibodies as well as cervical IgA antibody levels with new sensitive radioimmunoassay (RIA) techniques. One third of the women were delivered of a healthy infant and two thirds aborted, but the two groups were otherwise clinically similar. By isolation, only 2.7% of the patients were CT‐positive, but increased cervical IgA antibody level to CT was detected in 41.3%. The mean level of these local antibodies was similar in both study groups, but the mean levels of serum IgA and IgG antibodies were somewhat higher in the patients who aborted although the difference was not significant. None of the cervical specimens was positive for HSV by isolation but the cervical IgA antibody level to HSV was raised in 47.1% of the patients. Both cervical and serum IgA antibody levels to HSV were significantly raised among the patients who aborted, but there were no differences between the patients with spontaneous abortion and those with a blighted ovum. There was no clear association between CT and abortion, but an association between HSV and abortion is possible. The incidence of raised levels of both CT and HSV IgA antibodies in the cervix was surprisingly high in both groups and the significance of this finding remains to be investigated.

Detection of Chlamydia Trachomatis Antigen by Radio Immunoassay

A sensitive indirect radioimmunoassay (RIA) was developed for detection of chlamydial antigens in infected, irradiated McCoy cell culture. Polystyrene beads were used as the solid phase, guinea pig antichlamydial immunoglobulins were used as the captive antibodies, rabbit antichlamydial immunoglobulins were used as the secondary antibodies, and 125I- labelled sheep antirabbit immunoglobulin was used as the indicator antibodies. The immunizations were done intracutaneously with purified genital and lymphogranuloma venereum Chlamydia trachomatis strains grown in the yolk sacs of embryonated eggs. The bound radioactivity was a function of the amount of chlamydial antigen and the method demonstrated the antigen approximately 20 hours post infection. Also noninfectious chlamydial antigen was detectable by RIA. The sensitivity of the assay was about 10 ng/ml for purified antigen and less than 10 inclusions in the cell culture. Each chlamydial serotype could be detected. The RIA was found to be more sensitive than iodine staining and as rapid and sensitive as immunofluorescence method to demonstrate chlamydial infection in cell culture.