School of Agricultural Sciences/College of Agricultural, Life & Physical Sciences, Southern Illinois University, 1205 Lincoln Dr., Carbondale, IL 62901, USA.
Botany Department, Zagazig University, Zagazig 44519, Egypt.
Lett Appl Microbiol. 2023 Jan 23;76(1). doi: 10.1093/lambio/ovac023.
Fusarium oxysporum,F. graminearum,F. acuminatum,F. equiseti,F. proliferatum,F. solani, and Rhizoctonia solani are soil-borne fungal pathogens that cause substantial yield loss in a widespread list of crops worldwide. The objective of this study was to develop a panel of TaqMan assays for the detection and quantification of these six widespread soil-borne fungal species using real-time polymerase chain reaction (qPCR). The primers and probes were designed based on the intergenic spacer ribosomal RNA and translation elongation factor 1-alpha gene (tef1). These assays, although not multiplexed, can be performed simultaneously as they have similar reaction conditions, allowing more efficiency when targeting multiple pathogens in a sample. The assays presented high efficiency (94.3%-108.9%) and sensitivity, with a limit of detection of 0.05 picograms (50 femtograms) of target DNA. Results from an assay targeting 19 non-target and closely related species confirmed the specificity of the developed assays. The assays were also evaluated to detect the target species in different matrices, such as soil and plant material. This panel of qPCR assays is an additional tool that can be used by plant pathologists, microbiologists, plant breeders, diagnostic clinics, and other researchers interested in these fungal species.
尖孢镰刀菌、禾谷镰刀菌、层出镰刀菌、燕麦镰刀菌、串珠镰刀菌、茄病镰刀菌和立枯丝核菌是土壤传播的真菌病原体，它们在全球范围内广泛种植的作物中造成了大量的产量损失。本研究的目的是开发一组 TaqMan 检测和定量这些六种广泛存在的土壤传播真菌物种的实时聚合酶链反应 (qPCR) 方法。引物和探针是基于基因间核糖体 RNA 和翻译延伸因子 1-α（tef1）设计的。这些检测方法虽然没有多重化，但可以同时进行，因为它们具有相似的反应条件，在一个样本中针对多个病原体时更有效率。这些检测方法具有高效性（94.3%-108.9%）和灵敏度，检测限为 0.05 皮克（50 飞克）的目标 DNA。针对 19 种非目标和密切相关物种的检测证实了所开发检测方法的特异性。还评估了这些检测方法在不同基质（如土壤和植物材料）中检测目标物种的能力。该 qPCR 检测方法组是植物病理学家、微生物学家、植物育种家、诊断诊所和其他对这些真菌物种感兴趣的研究人员的另一种工具。
