Direct 16S/ITS rRNA Gene PCR Followed by Sanger Sequencing for Detection of Mycetoma Causative Agents in Dakar, Senegal: A Pilot Study Among Patients with Mycetoma Attending Aristide Le Dantec University Hospital.

IF 3.6 3区 生物学 Q2 MYCOLOGY
Khadim Diongue, Jean-Noël Dione, Abdoulaye Diop, Jihane Kabtani, Mamadou Alpha Diallo, Coralie L'Ollivier, Mame Cheikh Seck, Mouhamadou Ndiaye, Aida Sadikh Badiane, Daouda Ndiaye, Stéphane Ranque
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引用次数: 0

Abstract

Mycetoma can be caused either by fungi or aerobic Actinomycetes. A precise identification of the causal agents is critical for the therapeutic outcome. Thus, this study aimed to identify the pathogens of mycetoma using 16S/ITS rRNA gene polymerase chain reaction (PCR) followed by Sanger sequencing directly on grains. In sum, 32 samples including 15 black grains, 12 red grains, and five white/yellow grains collected from patients with mycetoma at the Aristide Le Dantec University Hospital in Dakar, Senegal, between October 2014 and September 2020 were submitted to PCR/sequencing. For black grain eumycetoma, the ITS rRNA region was targeted. Similarly, the 16S rRNA gene was targeted for red grain actinomycetoma. These two regions were targeted in parallel for white/yellow grains, which could be of either bacterial or fungal origin. The age of the patients ranged from 14 to 72 years with a mean age of 36 ± 14 years. Thirteen (86%) of the 15 samples with black grains, were successfully sequenced with only one established eumycetoma pathogen, Madurella mycetomatis identified in 11 (73%). Cladosporium sphaerospermum was identified in one sample. For the 16S rRNA sequencing of red grains, a 58.3% (7/12) success rate was obtained with Actinomadura pelletieri identified in six samples. Among the five samples sequenced twice, the 16S rRNA allowed us to identify the causative agent in 2 cases, A. madurae in one, and A. geliboluensis in the other. The ITS rRNA identified 3 fungi, of which none was a mycetoma agent. Overall, direct 16S/ITS rRNA sequencing of the grains for detecting and identifying mycetoma pathogens was successful in 59.4% of cases. Fungi, led by M. mycetomatis, were the predominant pathogens identified. Two probable new mycetoma agents, C. sphaerospermum, and A. geliboluensis were identified and both deserve to be confirmed in further studies.

直接 16S/ITS rRNA 基因 PCR 和 Sanger 测序用于检测塞内加尔达喀尔的霉菌瘤致病因子:在阿里斯蒂德-勒丹特克大学医院就诊的霉菌性瘤患者中开展的试点研究。
霉菌瘤可由真菌或需氧放线菌引起。准确识别病原体对治疗效果至关重要。因此,本研究旨在利用 16S/ITS rRNA 基因聚合酶链式反应(PCR),然后直接对谷物进行桑格测序,以确定霉菌瘤的病原体。2014年10月至2020年9月期间,塞内加尔达喀尔阿里斯蒂德-勒丹泰克大学医院从霉菌瘤患者身上采集了32份样本,其中包括15颗黑色谷粒、12颗红色谷粒和5颗白色/黄色谷粒,并对这些样本进行了PCR/测序。对于黑粒真菌瘤,目标是 ITS rRNA 区域。同样,红粒放线菌也以 16S rRNA 基因为目标。这两个区域同时也是白粒/黄粒的目标区域,白粒/黄粒可能来自细菌或真菌。患者年龄从 14 岁到 72 岁不等,平均年龄为 36 ± 14 岁。在 15 份带有黑色颗粒的样本中,有 13 份(86%)成功测序,仅在 11 份(73%)样本中发现了一种已确定的真菌病原体--马杜雷拉霉菌(Madurella mycetomatis)。在一个样本中发现了 Cladosporium sphaerospermum。在对红色谷物进行 16S rRNA 测序时,成功率为 58.3%(7/12),在 6 个样本中鉴定出 Actinomadura pelletieri。在测序两次的 5 个样本中,16S rRNA 使我们确定了 2 个病原体,一个是 A. madurae,另一个是 A. geliboluensis。ITS rRNA 鉴定出 3 种真菌,其中没有一种是霉菌瘤病原体。总体而言,对谷物进行 16S/ITS rRNA 直接测序以检测和鉴定霉菌瘤病原体的成功率为 59.4%。以霉菌为首的真菌是主要的病原体。发现了两种可能的新霉菌瘤病原体:C. sphaerospermum 和 A. geliboluensis,这两种病原体都有待进一步研究证实。
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来源期刊
Mycopathologia
Mycopathologia 生物-真菌学
CiteScore
6.80
自引率
3.60%
发文量
76
审稿时长
3 months
期刊介绍: Mycopathologia is an official journal of the International Union of Microbiological Societies (IUMS). Mycopathologia was founded in 1938 with the mission to ‘diffuse the understanding of fungal diseases in man and animals among mycologists’. Many of the milestones discoveries in the field of medical mycology have been communicated through the pages of this journal. Mycopathologia covers a diverse, interdisciplinary range of topics that is unique in breadth and depth. The journal publishes peer-reviewed, original articles highlighting important developments concerning medically important fungi and fungal diseases. The journal highlights important developments in fungal systematics and taxonomy, laboratory diagnosis of fungal infections, antifungal drugs, clinical presentation and treatment, and epidemiology of fungal diseases globally. Timely opinion articles, mini-reviews, and other communications are usually invited at the discretion of the editorial board. Unique case reports highlighting unprecedented progress in the diagnosis and treatment of fungal infections, are published in every issue of the journal. MycopathologiaIMAGE is another regular feature for a brief clinical report of potential interest to a mixed audience of physicians and laboratory scientists. MycopathologiaGENOME is designed for the rapid publication of new genomes of human and animal pathogenic fungi using a checklist-based, standardized format.
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