Sousa Gabriel S M, De Oliveira Rodrigo S, Souza Alex B, Monteiro Ruan C, Santo Elaine P T E, Franco Filho Luciano C, Moraes Denison L O, De Sá Sarah R, Da Silva Silvia H M
Programa de Pós-Graduação em Biologia de Agentes Infecciosos e Parasitários, Instituto de Ciências Biológicas, Universidade Federal do Pará, Belém 66075-750, Brazil.
Laboratório de Micoses Superficiais e Sistêmicas, Seção de Bacteriologia e Micologia, Instituto Evandro Chagas, Ananindeua 67030-000, Brazil.
J Fungi (Basel). 2024 Aug 4;10(8):548. doi: 10.3390/jof10080548.
The Herpotrichiellaceae family is an important group of dematiaceous filamentous fungi, associated with a variety of pathogenic fungal species causing chromoblastomycosis (CBM) and phaeohyphomycosis (PHM), both with polymorphic clinical manifestations and worldwide incidence. Currently, the identification of this family and determination of the causative agent is challenging due to the subjectivity of morphological identification methods, necessitating the use of molecular techniques to complement diagnosis. In this context, genetic sequencing of the (ITS) has become the norm due to a lack of alternative molecular tools for identifying these agents. Therefore, this study aimed to develop PCR-Multiplex methodologies to address this gap. Sequences from the ITS and (LSU) of ribosomal DNA were used, and after manual curation and in vitro analyses, primers were synthesized for the identification of the targets. The primers were optimized and validated in vitro, resulting in two PCR-Multiplex methodologies: one for identifying the Herpotrichiellaceae family and the bantiana clade, and another for determining the species and . Ultimately, the assays developed in this study aim to complement other identification approaches for these agents, reducing the need for sequencing, improving the management of these infections, and enhancing the accuracy of epidemiological information.
刺盾炱科是一组重要的暗色丝状真菌，与多种致病性真菌物种相关，可引起着色芽生菌病（CBM）和暗色丝孢霉病（PHM），这两种疾病都具有多形性临床表现且在全球范围内均有发病。目前，由于形态学鉴定方法的主观性，对该科的鉴定以及病原体的确定具有挑战性，因此需要使用分子技术来辅助诊断。在此背景下，由于缺乏用于鉴定这些病原体的替代分子工具，核糖体DNA的内转录间隔区（ITS）基因测序已成为常规方法。因此，本研究旨在开发聚合酶链反应-多重（PCR-Multiplex）方法来填补这一空白。使用了核糖体DNA的ITS和大亚基（LSU）序列，经过人工整理和体外分析后，合成了用于鉴定目标的引物。这些引物在体外进行了优化和验证，产生了两种PCR-Multiplex方法：一种用于鉴定刺盾炱科和班替枝孢属分支，另一种用于确定物种[具体物种名称未给出]和[具体物种名称未给出]。最终，本研究开发的检测方法旨在补充针对这些病原体的其他鉴定方法，减少测序需求，改善这些感染的管理，并提高流行病学信息的准确性。
