Mayasar I Al-Zaban, Ahlam H Alrokban, Mohamed A Mahmoud
{"title":"建立实时荧光定量PCR和多重荧光定量PCR检测和鉴定玉米储藏中产霉菌毒素真菌的方法。","authors":"Mayasar I Al-Zaban, Ahlam H Alrokban, Mohamed A Mahmoud","doi":"10.1080/21501203.2023.2213704","DOIUrl":null,"url":null,"abstract":"<p><p>This study aimed to identify important mycotoxigenic fungi and accurate detection of mycotoxin in stored maize grains using molecular methods. The current study also optimised the real-time PCR (RT-PCR) assay. The melting curve was established to identify isolated fungal species of <i>Aspergillus</i> (4), <i>Fusarium</i> (3), <i>Penicillium</i> (3), and <i>Alternaria</i> (one). A multiplex polymerase chain reaction (mPCR) technique was developed for the detection and characterisation of mycotoxin producing fungi, mycotoxin metabolic pathway genes, and the determination of eleven mycotoxins in stored maize grains using high-performance liquid chromatography (HPLC). The mPCR results indicated positive signals for potentially mycotoxigenic fungal species tested of <i>Aspergillus, Fusarium, Penicillium</i>, and <i>Alternaria</i>. A protocol for multiplex reverse transcription-polymerase chain reaction (mRT-PCR) was tested to distinguish between free and contaminated, stored maize with aflatoxin B1 (AFB1). The expression pattern of four aflatoxin biosynthetic pathway genes, AFB1 (<i>aflQ, aflP, aflO</i>, and <i>aflD</i>), was a good marker for contaminated, stored maize grains. HPLC analysis showed that maize grain samples were contaminated with mycotoxins, and the concentration was above the detection level. The results indicate that the polyphasic approach might provide a sensitive, rapid, and accurate method for detecting and identifying mycotoxigenic fungal species and mycotoxins in stored maize grains.</p>","PeriodicalId":18833,"journal":{"name":"Mycology","volume":null,"pages":null},"PeriodicalIF":4.6000,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/db/93/TMYC_14_2213704.PMC10424615.pdf","citationCount":"0","resultStr":"{\"title\":\"Development of a real-time PCR and multiplex PCR assay for the detection and identification of mycotoxigenic fungi in stored maize grains.\",\"authors\":\"Mayasar I Al-Zaban, Ahlam H Alrokban, Mohamed A Mahmoud\",\"doi\":\"10.1080/21501203.2023.2213704\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>This study aimed to identify important mycotoxigenic fungi and accurate detection of mycotoxin in stored maize grains using molecular methods. The current study also optimised the real-time PCR (RT-PCR) assay. The melting curve was established to identify isolated fungal species of <i>Aspergillus</i> (4), <i>Fusarium</i> (3), <i>Penicillium</i> (3), and <i>Alternaria</i> (one). A multiplex polymerase chain reaction (mPCR) technique was developed for the detection and characterisation of mycotoxin producing fungi, mycotoxin metabolic pathway genes, and the determination of eleven mycotoxins in stored maize grains using high-performance liquid chromatography (HPLC). The mPCR results indicated positive signals for potentially mycotoxigenic fungal species tested of <i>Aspergillus, Fusarium, Penicillium</i>, and <i>Alternaria</i>. A protocol for multiplex reverse transcription-polymerase chain reaction (mRT-PCR) was tested to distinguish between free and contaminated, stored maize with aflatoxin B1 (AFB1). The expression pattern of four aflatoxin biosynthetic pathway genes, AFB1 (<i>aflQ, aflP, aflO</i>, and <i>aflD</i>), was a good marker for contaminated, stored maize grains. HPLC analysis showed that maize grain samples were contaminated with mycotoxins, and the concentration was above the detection level. The results indicate that the polyphasic approach might provide a sensitive, rapid, and accurate method for detecting and identifying mycotoxigenic fungal species and mycotoxins in stored maize grains.</p>\",\"PeriodicalId\":18833,\"journal\":{\"name\":\"Mycology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.6000,\"publicationDate\":\"2023-01-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/db/93/TMYC_14_2213704.PMC10424615.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Mycology\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1080/21501203.2023.2213704\",\"RegionNum\":2,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"MYCOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Mycology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1080/21501203.2023.2213704","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"MYCOLOGY","Score":null,"Total":0}
Development of a real-time PCR and multiplex PCR assay for the detection and identification of mycotoxigenic fungi in stored maize grains.
This study aimed to identify important mycotoxigenic fungi and accurate detection of mycotoxin in stored maize grains using molecular methods. The current study also optimised the real-time PCR (RT-PCR) assay. The melting curve was established to identify isolated fungal species of Aspergillus (4), Fusarium (3), Penicillium (3), and Alternaria (one). A multiplex polymerase chain reaction (mPCR) technique was developed for the detection and characterisation of mycotoxin producing fungi, mycotoxin metabolic pathway genes, and the determination of eleven mycotoxins in stored maize grains using high-performance liquid chromatography (HPLC). The mPCR results indicated positive signals for potentially mycotoxigenic fungal species tested of Aspergillus, Fusarium, Penicillium, and Alternaria. A protocol for multiplex reverse transcription-polymerase chain reaction (mRT-PCR) was tested to distinguish between free and contaminated, stored maize with aflatoxin B1 (AFB1). The expression pattern of four aflatoxin biosynthetic pathway genes, AFB1 (aflQ, aflP, aflO, and aflD), was a good marker for contaminated, stored maize grains. HPLC analysis showed that maize grain samples were contaminated with mycotoxins, and the concentration was above the detection level. The results indicate that the polyphasic approach might provide a sensitive, rapid, and accurate method for detecting and identifying mycotoxigenic fungal species and mycotoxins in stored maize grains.