Detection of human IgG antibodies against Mycoplasma genitalium using a recombinant MG075 antigen

Mycoplasma genitalium is a sexually transmitted pathogen that can cause a range of reproductive tract diseases in both men and women. To disentangle the relationship between M. genitalium infection(s) and subsequent reproductive health complications at the population level, accurate serological tools are needed. The major challenge in developing specific M. genitalium serological tests is the extensive cross-reactivity with the closely related ubiquitous respiratory tract pathogen, Mycoplasma pneumoniae. In this report, we describe the development of an immunoblot assay based on a recombinant fragment of the M. genitalium MG075 protein present in lipid-associated membrane extracts. A sensitivity of 87.1% was achieved based on the testing antibody responses in sera from 101 adults with PCR-confirmed M. genitalium infection. A specificity of 95.2% was obtained through the evaluation of sera from 166 children under 15 years of age with and without M. pneumoniae infection, who were unlikely to have been exposed to sexually transmitted M. genitalium. The development of a serological assay capable of accurately distinguishing M. genitalium and M. pneumoniae will enable a better understanding of associations between M. genitalium and adverse reproductive sequelae.

Mycoplasma genitalium is the second most common sexually transmitted bacterial infection after chlamydia. The long-term consequences of the infection are still under investigation, but reliable tools for monitoring exposure by detection of antibodies have been lacking specificity due to the presence of cross-reacting antibodies to the closely related Mycoplasma pneumoniae. Here, we describe a novel diagnostic antigen with promising sensitivity (87%) and specificity (95%) based on testing of sera from patients with PCR-confirmed M. genitalium infection and children with and without M. pneumoniae infection, respectively. The MG075F1 antigen was expressed in Escherichia coli, and the recombinant antigen was used in a western line-blot. Due to the insolubility of the antigen, harsh denaturing conditions were needed, making an enzyme-linked immunosorbent assay (ELISA) format impossible. Future work should explore shorter fragments or protein engineering to allow for assay designs better suited for high-throughput screening.