A Pneumocystis jirovecii recombinant synthetic antigen and an ELISA for detection of anti-P. jirovecii antibodies, were developed for the serodiagnosis of severe interstitial pneumonia
Pneumocystis jirovecii is an atypical opportunistic fungus capable of causing severe interstitial pneumonia (PCP) that continues to be the main AIDS-defining disease in Europe and the USA and is an emerging concern in non-HIV-infected patients undergoing immunosuppressive therapies.
The standard laboratory diagnosis of PCP relies on microscopic visualization of stained P. jirovecii organisms and/or DNA detection by PCR in respiratory specimens that are obtained by invasive techniques.
As such, the development of a serological test is mandatory, as it will allow the diagnosis of PCP by means of minimally invasive biological specimens, such as blood.
A recombinant synthetic multiepitope antigen (RSA) of P. jirovecii was produced, based on the major surface glycoprotein (Msg), which contains shared and species-specific epitopes, elicits humoral and cellular protective immune responses and plays a central role in the interaction Pneumocystis-host. The selection of the putative reactive epitopes of the Msg was made targeting the terminal fraction of its middle portion (MsgB) and two reactive antigenic portions of the carboxyl terminal fraction (MsgC), using bioinformatics approaches to analyze electrochemical properties, secondary structure prediction, polarity, relative position to the membrane and hydrophobicity profile of the specific polypeptides. The invention is also aimed to offer an ELISA technique for the detection of Ig, IgG and IgM anti-P. jirovecii antibodies in biological specimens of individuals, namely in serum.
Msg’s RSA is able to detect specific anti-P. jirovecii antibodies. Levels of IgG anti-P. jirovecii can only be used for screening of populations. Levels of IgM anti-P. jirovecii showed a statistically significant increase in PCP patients compared to non-PCP patients; and the ELISA IgM anti-P. jirovecii test showed excellent sensitivity and good specificity, when associated with clinical diagnosis criteria.
With this innovative approach, we succeeded in producing a recombinant synthetic antigen, specific for P. jirovecii, capable of functioning as a PCP biosensor when applied in an ELISA assay for detection of IgM anti-P. jirovecii antibodies. This sensitive and easy to use method will help to decrease the number of PCP cases that still exist in industrialized countries and especially in developing countries where it is difficult to implement laboratory diagnostic techniques and methods to obtain adequate sampling.
- TOMÁS A. L., CARDOSO F., ESTEVES F. & MATOS O. 2016. Serological diagnosis of pneumocystosis: production of a synthetic recombinant antigen for immunodetection of Pneumocystis jirovecii. Scientif Reports, 6, 36287. doi: 10.1038/srep36287
Stage of Development
TRL 4 – Technology validated in laboratory.
This new method may be used as a screening for P. jirovecii infection and diagnostic test for PCP, decreasing the need for biological specimens obtained by invasive techniques, which is a major benefit to the patient’s care and an improvement in the clinical management of the disease.
- Diagnosis of PCP;
- Screening of anti-P. jirovecii antibodies;
- Discriminatory tool between Pneumocystis and other fungal infections.
- Available for licensing;
- Seeking commercial partner.
Pneumocystis pneumonia; major surface glycoprotein; recombinant synthetic antigen; serological diagnosis; ELISA.
Fernando Manuel Cardoso
Olga Guerreiro de Matos
Ana Luísa Chocalheiro