Differential avidity determination of IgG directed towards the receptor-binding domain (RBD) of SARS-CoV-2 wild-type and its variants in one assay: Rational tool for the assessment of protective immunity.

The avidity (binding strength) of IgG directed towards the receptor-binding domain (RBD) of spike protein has been recognized as a central marker in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serology. It seems to be linked to increased infection-neutralization potential and therefore might indicate protective immunity. Using a prototype line assay based on the established recomLine SARS-CoV-2 assay, supplemented with RBD of the delta and the omicron variant, differential avidity determination of IgG directed towards RBD of wild-type (WT) SARS-CoV-2 and distinct variants was possible within one assay. Our data confirm that natural SARS-CoV-2 infection or one vaccination step lead to low avidity IgG, whereas further vaccination steps gradually increase avidity to high values. High avidity is not reached by infection alone. After infection with WT SARS-CoV-2 or vaccination based on mRNA WT, the avidity of cross-reacting IgG directed towards RBD of the delta variant only showed marginal differences compared to IgG directed towards RBD WT. In contrast, the avidity of IgG cross-reacting with RBD of the omicron variant was always much lower than for IgG RBD WT, except after the third vaccination step. Therefore, parallel avidity testing of RBD WT and omicron seems to be mandatory for a significant assessment of protective immunity towards SARS-CoV-2.

Analysis of humoral immune responses to recombinant Chlamydia pneumoniae antigens.

OBJECTIVES: Chlamydia pneumoniae is a difficult to diagnose respiratory pathogen. This study was performed to systematically characterize humoral immune responses to selected C. pneumoniae antigens in order to provide novel serodiagnostic perspectives for clinical and epidemiological issues. METHODS: Based on a literature search, gene library screening, and serological proteome analysis, 15 immunogenic surface-associated, virulence-associated, and hypothetical C. pneumoniae antigens were selected, recombinantly expressed, and lined on a nitrocellulose strip. Specific IgM and IgG reactivity was measured in a total of 172 PCR- and micro-immunofluorescence testing (MIF)-characterized serum samples from patients with respiratory infections. A theoretical model was conceived to approximate a putative course of C. pneumoniae antigen expression and assess the potential of early and late antigens. RESULTS: While surface antigens performed poorly, the virulence-associated TARP was a reliable antigen for IgM detection, with a sensitivity of 80.0% and a diagnostic specificity of 90.2%. The hypothetical protein YwbM proved powerful for IgG detection with MIF-correlative sensitivities of up to 94.4% and a diagnostic specificity of 95.1%. CONCLUSIONS: This study provides new insights into antibody profiles to immunogenic proteins in C. pneumoniae infection. The study findings offer antigen candidates for more reliable and standardized serological investigations of C. pneumoniae infections, including studies on seroprevalence and epidemiology.

Analysis of Humoral Immune Responses to Surface and Virulence-Associated Chlamydia abortus Proteins in Ovine and Human Abortions by Use of a Newly Developed Line Immunoassay.

The obligate intracellular bacterium Chlamydia abortus is the causative agent of enzootic abortion of ewes and poses a significant zoonotic risk for pregnant women. Using proteomic analysis and gene expression library screening in a previous project, we identified potential virulence factors and candidates for serodiagnosis, of which nine were scrutinized here with a strip immunoassay. We have shown that aborting sheep exhibited a strong antibody response to surface (MOMP, MIP, Pmp13G) and virulence-associated (CPAF, TARP, SINC) antigens. While the latter disappeared within 18 weeks following abortion in a majority of the animals, antibodies to surface proteins persisted beyond the duration of the study. In contrast, nonaborting experimentally infected sheep developed mainly antibodies to surface antigens (MOMP, MIP, Pmp13G), all of which did not persist. We were also able to detect antibodies to these surface antigens in C abortus-infected women who had undergone septic abortion, whereas a group of shepherds and veterinarians with occupational exposure to C abortus-infected sheep revealed only sporadic immune responses to the antigens selected. The most specific antigen for the serodiagnosis of human C abortus infections was Pmp13G, which showed no cross-reactivity with other chlamydiae infecting humans. We suggest that Pmp13G-based serodiagnosis accomplished by the detection of antibodies to virulence-associated antigens such as CPAF, TARP, and SINC may improve the laboratory diagnosis of human and animal C abortus infections.

Serological profiling with Chlamycheck, a commercial multiplex recombinant antigen Western blot assay of chlamydial infections.

A new chlamydial test system, the Chlamycheck assay, which uses 4 purified recombinant antigens of Chlamydia trachomatis and Chlamydophila pneumoniae and one antigen of Chlamydophila psittaci, has been developed and commercialized. We investigated the reactivities of the recombinant antigens with sera from a group of 30 patients with acute Chlamydia trachomatis infection, 88 patients consulting for sexually transmitted infections, and 46 patients with serological evidence of Chlamydophila pneumoniae infection. The results obtained from human and infected mouse sera suggest that Chlamycheck serology against multiple proteins may provide additional useful information that is not available by conventional whole elementary body microimmunofluorescence or single-antigen enzyme-linked immunosorbent assay serology. Specific serological profiles were associated with acute versus past Chlamydia trachomatis infection or with Chlamydia trachomatis primo-infection versus infection in a Chlamydophila pneumoniae history context.