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Design of a melting curve analysis (MCA) based on multiplex real-time PCR for detection of Aspergillus terreus and Aspergillus fumigatus in cereals and oilseeds samples

Summary

This study developed a fast molecular test using real-time PCR to detect harmful Aspergillus fungi in grains and seeds. Instead of waiting 3-5 days for traditional culture methods, this new test can identify the fungi in just hours by detecting specific DNA patterns. The test was tested on 140 samples of cereals and oilseeds and proved more accurate than traditional methods, making it useful for food safety in markets and processing plants.

Background

Aspergillus species contamination in cereals and oilseeds is a significant food safety concern that can produce mycotoxins harmful to human health. Traditional phenotypic identification methods require 3-5 days of incubation and have limited accuracy. Molecular techniques like real-time PCR offer faster and more reliable alternatives for detecting fungal contamination in food products.

Objective

To develop and evaluate a multiplex real-time PCR (MRT-PCR) assay combined with melting curve analysis (MCA) for rapid and accurate detection of Aspergillus terreus and Aspergillus fumigatus in cereals and oilseed samples. The study aimed to compare the performance of molecular methods with traditional culture-based phenotypic analysis.

Results

MRT-PCR detected 24 isolates (17.1%) of A. fumigatus and 11 (7.8%) of A. terreus among 140 samples. Melting curve analysis showed lovB gene peaks between 86.2-86.8°C for A. terreus and easL gene peaks from 89.2-89.7°C for A. fumigatus. The molecular method demonstrated superior sensitivity and specificity compared to phenotypic culture methods, which detected 36 A. fumigatus (25.7%) and 21 A. terreus (15%) isolates.

Conclusion

MRT-PCR combined with melting curve analysis is a rapid, sensitive, and specific method for detecting A. terreus and A. fumigatus in cereals and oilseeds, significantly reducing detection time from days to hours. This molecular approach outperforms traditional culture-based methods and provides reliable identification without requiring specialized mycological expertise, making it suitable for routine screening in food processing facilities.
  • Published in:BMC Research Notes,
  • Study Type:Experimental/Methodological Study,
  • Source: 10.1186/s13104-025-07341-w, PMID: 40660338
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