Development and evaluation of a novel visual and rapid detection assay for toxigenic Fusarium graminearum in maize based on recombinase polymerase amplification and lateral flow analysis

Liang, Xiaofeng; Zhang, Xiu; Haseeb, Hafiz Abdul; Tang, Tingting; Shan, Jianguo; Yin, Biaolin; Guo, Wei

doi:10.1016/j.ijfoodmicro.2022.109682

Cited by 4 publications

(1 citation statement)

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“…Recent literature has demonstrated the successful application of RPA-based techniques in the identification of various fungal species, such as Candida albicans (Wang et al, 2021), Candida tropicalis , Candida glabrata , Cryptococcus neoformans/C. gattii (Ma et al, 2019), Aspergillus fumigatus (Li et al, 2022), and Fusarium graminearum (Lei et al, 2022;Liang et al, 2022;, thereby fulfilling the demand for rapid, specific, and highly sensitive detection within a timeframe of 20-30 min (Supplementary Table S2). However, to date, there have been no published studies investigating the use of RPA for identifying fungi belonging to the order Mucorales.…”

Section: Introductionmentioning

confidence: 99%

Rapid detection of Mucorales based on recombinase polymerase amplification and real-time PCR

Xu,

Li,

Zhao

et al. 2023

Front. Microbiol.

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Mucormycosis, an invasive fungal disease with severe consequences, poses a significant threat to immunocompromised individuals. However, the timely and accurate identification of Mucorales infection continues to present difficulties. In this study, novel detection techniques utilizing recombinase polymerase amplification (RPA) and quantitative real-time polymerase chain reaction (qPCR) were developed, specifically targeting the mitochondrial rnl gene, in order to address this challenge. The specificity of the RPA and qPCR assay was assessed by adding genomic DNAs extracted from 14 non-targeted strains, as well as human and mouse blood. No false-positive results were observed. Additionally, genomic DNAs from 13 species in five genera of order Mucorales were tested and yielded positive results in both methods. To further evaluate the sensitivity of the assays, DNAs from Rhizopus oryzae, Mucor racemosus, Absidia glauca, Rhizomucor miehei, and Cunninghamella bertholletiae were utilized, with concentrations ranging from 1 ng/μL to 1 fg/μL. The limit of detection (LoD) for the RPA assay was determined to be 1 pg., with the exception of Rhizomucor miehei which had a LoD of 1 ng. The LoD for the qPCR assay varied between 10 fg and 1 pg., depending on the specific species being tested. Sensitivity analysis conducted on simulated clinical samples revealed that the LoD for RPA and qPCR assays were capable of detecting DNA extracted from 103 and 101 colony forming units (CFU) conidia in 200 μL of blood and serum, respectively. Consequently, the real-time RPA and qPCR assays developed in this study exhibited favorable sensitivity and specificity for the diagnosis of mucormycosis.

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