Abstract
Talaromyces flavus is a soilborne fungus that can contaminate fruits. It constitutes serious influence on heat-processed food spoilage, as T. flavus belongs to the heat-resistant fungi group, which are able to survive the pasteurization process. Moreover T. flavus has been reported to be capable of mycotoxigenicity, therefore they have a serious threat to human health. To maintain the safety of food production, sensitive method for T. flavus detection was developed. The loop mediated amplification, abbreviated LAMP, reactions were designed as specific for detection of DNA replication licensing factor gene of T. flavus. The specificity of assay was confirmed by use of 5 T. flavus strains and 35 other fungal isolates. The achieved limit of detection was 1fg of T. flavus genomic DNA and 64 ascospores in 1 g of strawberry fruits or soil samples.
Highlights
Fungal spoilage of heat-processed food emerges as a major problem in food industry
Positive amplification results were observed with all five T. flavus isolates: T. flavus DSM 63536, T. flavus NBRC 31879, T. flavus NBRC 30574, T. flavus NBRC 7233 and T. flavus NBRC 102293, and with T. flavus DSM 63536 DNA mixed with Byssochlamys nivea LMEM G49/11 DNA in ratio of 100fg:[10] ng of genomic DNA, while no amplification was observed with DNA templates isolated from other tested 35 fungal isolates (Fig. 2a)
Positive reaction observed in samples containing mixed DNA of T. flavus and B. nivea in ratio of 1:100000 suggest, that presence of external DNA does not interfere with amplification
Summary
Fungal spoilage of heat-processed food emerges as a major problem in food industry. Estimated losses caused to fruit juice production by heat-resistant fungi (HRF) cost food industry millions of USD solely in United States of America[1,2]. Successful methods based on PCR and qPCR techniques to detect HRF of genus Neosartorya, Byssochlamys and species Talaromyces flavus were developed[18,19,20]. Emergence of isothermal amplification techniques may provide the possibility to develop HRF detection methods that can be regarded as easier and cheaper assay to deploy within food industry and on the field. The aim of this study was to develop LAMP method for detection of Talaromyces flavus that provides specificity and rapidity of achieving results. The DNA replication licensing factor has already been reported as a possible marker and target gene for detection of T. flavus by qPCR reaction[20]
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