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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 to detect harmful fungi (Aspergillus species) in grains and seeds. Traditional fungal identification takes 3-5 days, but the new PCR-based method can produce results in hours. Testing 140 grain and seed samples, the new method successfully identified two dangerous Aspergillus species that produce toxic compounds harmful to human health. This advancement helps ensure food safety by enabling quicker detection of contamination in food production facilities.

Background

Aspergillus species contamination in cereals and oilseeds poses significant health risks through mycotoxin production. Early detection of contamination is crucial to prevent harm to human health and economic losses. Traditional phenotypic methods for fungal identification are time-consuming, requiring 3-5 days of incubation and specialized expertise.

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 oilseeds samples.

Results

Among 140 samples, phenotypic analysis identified 36 A. fumigatus isolates (25.7%) and 21 A. terreus isolates (15%). MRT-PCR detected 24 A. fumigatus isolates (17.1%) and 11 A. terreus isolates (7.8%). The lovB gene showed melt peaks at 86.2-86.8°C for A. terreus and easL gene at 89.2-89.7°C for A. fumigatus with sensitivity down to 0.01 ng DNA.

Conclusion

MRT-PCR combined with melting curve analysis provides superior speed, sensitivity, and accuracy compared to traditional phenotypic methods for detecting A. terreus and A. fumigatus in food samples. The technique can reduce identification time from days to hours and is suitable for implementation in laboratories and food processing facilities.
  • Published in:BMC Research Notes,
  • Study Type:Experimental Laboratory Study,
  • Source: 10.1186/s13104-025-07341-w; PMID: 40660338
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