Background: Paracoccidioidomycosis (PCM) is a systemic mycosis caused by dimorphic
fungi Paracoccidioides brasiliensis and Paracoccidioides lutzii. It is prevalent in Latin American,
mainly in Brazil. Therefore, PCM has fundamental impact on the Brazilian global economy, especially
in public health system, since it is affecting economical active population in different country regions.
Objective: The present study aimed to standardize the Real Time-Polymerase Chain Reaction (rt-PCR)
for an efficient and safe PCM diagnosis amplifying the recombinant protein PB27 gene, only expressed
by specimens of Paracoccidioides genus.
Methods: To standardize a methodology of rt-PCR using species-specific primers and probe designed for
annealing in this specific region of the fungi´s genome, amplifying the recombinant protein PB27 gene,
only expressed by specimens of Paracoccidioides genus. Followed by design in silico, experiments were
performed in vitro to determine rt-PCR specificity, efficiency and genome detection limit.
Results: The primers and probe sequences were deposited in Brazilian Coordination of Technological
Innovation and Transfer (CTIT), under patent reference number BR1020160078830. The present study
demonstrated the rt-PCR applicability for support on diagnosis of paracoccidioidomycosis, presenting
low cost, which makes it affordable for public health services in developing countries as Brazil. It is
noteworthy that it is necessary to validate this methodology using clinical samples before to use as a
safe method of diagnosis. A review of all patents related to this topic was performed and it was shown
that, to date, there are no records of patent on kits for paracoccidioidomycosis´s diagnostic. Indeed,
there is still a lot to go to reach this goal.
Conclusion: The reaction developed was standardized and patented, opening perspectives to molecular
diagnosis development for paracoccidioidomycosis, since rt-PCR can be applied to a broad spectrum of
infectious diseases. It would need to be tested in biological samples in order to validate this method and
then generate a diagnostic kit for Paracoccidioidomycosis.