New Molecular Markers Distinguishing Fonsecaea Agents of Chromoblastomycosis

Gabriela X Schneider,Israella R Colombo,Amanda Bombassaro,Renata R Gomes,Bruna J F S Lima,Giovanna Z Cândido,Vânia A Vicente,Mohammad J Najafzadeh,Bruna S Soley,Aniele C R Leão,Flávia F Costa,Conceição M P S De Azevedo,Sirlei G Marques,Kassiely Zamarchi,Morgana F Voidaleski,Jiufeng Sun,G Sybren De Hoog

doi:10.1007/s11046-019-00359-2

Abstract

The species belonging to the genus Fonsecaea are the main causative agents of chromoblastomycosis. The invasive potential of Fonsecaea differs significantly among its various sibling species. Moreover, the lack of clarity on the virulence and availability of precise markers to distinguish and detect Fonsecaea species is attributed to the different ways of dissemination and pathogenicity. Therefore, the present study aimed to propose new molecular tools to differentiate between sibling species causing chromoblastomycosis. We used an infection model of chromoblastomycosis in BALB/c to study species-specific molecular markers for the in vivo detection of Fonsecaea species in biological samples. Specific primers based on the CBF5 gene were developed for Fonsecaea pedrosoi, Fonsecaea monophora, Fonsecaea nubica, and Fonsecaea pugnacius. In addition, a padlock probe was designed for F. pugnacius based on ITS sequences. We also assessed the specificity of Fonsecaea species using in silico, in vitro, and in vivo assays. The results showed that markers and probes could effectively discriminate the species in both clinical and environmental samples, enabling bioprospecting of agents of chromoblastomycosis, thereby elucidating the infection route of the disease.

Highlights

Human chromoblastomycosis (CBM) is a skin disease that is exclusively caused by the members of order Chaetothyriales
The oligonucleotide primers used for F. monophora (FOMO), F. nubica (FONU), F. pedrosoi (FOPE), and F. pugnacius (FOPU) were specific to the target species studied
The in silico assays demonstrated that the rolling circle amplification (RCA) padlock probe designed was specific to F. pugnacius (Fig. 2)

Summary

Introduction

Human chromoblastomycosis (CBM) is a skin disease that is exclusively caused by the members of order Chaetothyriales (black yeasts and relatives). The agents of the disease may be morphologically indistinguishable from their environmental counterparts. The disease is one of the most prevalent implantation mycoses in the world and occurs in the (sub)tropical regions of South America, Africa, and China [1]. The genus Fonsecaea comprises cryptic species related to the disease, known as F. pedrosoi, F. monophora, and F. nubica [2]. All these species are found in the environment and infect humans, and F. pugnacius is a recently described agent of CBM [3]

Methods

Results

Conclusion