Department of Medicine & Epidemiology, School of Veterinary Medicine, University of California, Davis, California 95616-5270, USA.
Dis Aquat Organ. 2021 May 6;144:175-185. doi: 10.3354/dao03582.
Systemic phaeohyphomycosis, aka 'fluid belly', is one of the most important emergent diseases in sturgeon Acipenser spp. aquaculture. The etiologic agent is the saprobic, dematiaceous fungus Veronaea botryosa. Effective vaccines and chemotherapeutic treatments are currently unavailable. Additionally, the fungus is a slow-growing organism, taking from 10-15 d for colonies to be observed in agar media. To this end, a specific quantitative PCR (qPCR) targeting the V. botryosa β-tubulin gene was developed and validated. The specificity of the assay to V. botryosa was initially confirmed in silico and in vivo against common fungal fish pathogens, including closely related members of the order Chaetothyriales (Exophiala spp.) and other black pigmented fungi (Alternaria spp. and Cladosporium spp.), as well as tissues from uninfected sturgeon. The assay possessed high clinical specificity (100%) and clinical sensitivity (74%) in detecting V. botryosa DNA in splenic tissues from laboratory-infected sturgeon. Using V. botryosa genomic DNA as a template, the limit of detection was equivalent to 10 conidia, and the method was found suitable for the detection of fungal DNA in fresh and formalin-fixed tissues. In addition, the presence of non-target DNA from white sturgeon did not influence assay sensitivity. The developed qPCR assay is a sensitive, specific, and rapid diagnostic method for the detection and quantification of V. botryosa DNA from white sturgeon tissues.
系统性暗色丝孢霉病，又名“腹水病”，是鲟鱼水产养殖中最重要的新兴疾病之一。病原体是腐生性、暗色真菌 Veronaea botryosa。目前尚无有效的疫苗和化学治疗方法。此外，真菌的生长速度较慢，在琼脂培养基中观察到菌落需要 10-15 天。为此，开发并验证了针对 V. botryosa β-微管蛋白基因的特定定量 PCR（qPCR）。该方法在针对常见真菌鱼类病原体的计算机模拟和体内实验中，对 V. botryosa 具有特异性，包括 Chaetothyriales 目（Exophiala 属）和其他黑色色素真菌（Alternaria 属和 Cladosporium 属）的密切相关成员，以及未感染的鲟鱼组织。该检测方法在检测实验室感染鲟鱼脾脏组织中的 V. botryosa DNA 时具有高临床特异性（100%）和临床敏感性（74%）。使用 V. botryosa 基因组 DNA 作为模板，检测限相当于 10 个分生孢子，该方法适用于检测新鲜和福尔马林固定组织中的真菌 DNA。此外，白鲟的非靶 DNA 不会影响检测的敏感性。所开发的 qPCR 检测方法是一种敏感、特异、快速的检测和定量白鲟组织中 V. botryosa DNA 的方法。
