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Article(id=1153986714663117290, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1153986709126635984, articleNumber=null, orderNo=null, doi=10.19812/j.cnki.jfsq11-5956/ts.20240624001, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1719158400000, receivedDateStr=2024-06-24, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1753061472812, onlineDateStr=2025-07-21, pubDate=1737734400000, pubDateStr=2025-01-25, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1753061472812, onlineIssueDateStr=2025-07-21, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1753061472812, creator=13701087609, updateTime=1753061472812, updator=13701087609, issue=Issue{id=1153986709126635984, tenantId=1146029695717560320, journalId=1149652044408987649, year='2025', volume='16', issue='2', pageStart='1', pageEnd='324', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=0, createTime=1753061471492, creator=13701087609, updateTime=1760345674980, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1184538872999457117, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1153986709126635984, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1184538872999457118, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1153986709126635984, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=302, endPage=308, ext={EN=ArticleExt(id=1153986715321623023, articleId=1153986714663117290, tenantId=1146029695717560320, journalId=1149652044408987649, language=EN, title=Research on fungal culturomics methods in food based on 3 kinds of different culture media, columnId=1151895321388347923, journalTitle=Journal of Food Safety & Quality, columnName=Food Analysis and Detection, runingTitle=null, highlight=null, articleAbstract=

Objective To establish a solution for fungal culturomics in food by comparing the performance of 3 fungal culture media and optimising fungal culturomics techniques. Methods The 24 food samples and 2 proficiency test samples were tested using commonly used fungal culture media for food testing, including potato dextrose agar (PDA), rose Bengal agar, and a novel Pan Fungi medium (PF). The cultured fungi were isolated and identified using matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results Yeast phase fungi could grow well on all 3 kinds of media, and the colony growth rate, colony morphology size and colony numbers had no significant difference. Filamentous fungi were able to grow well on the 3 kinds of medias, but growth rate and colony morphology were significantly different. There were 12 kinds of fungal species identified in the 24 samples, including 2 kinds of yeast phase fungi and 10 kinds of filamentous fungi. Conclusion With excellent accuracy and stability and self-limited growth of cultured colonies, PF fungal medium is favourable for isolation and identification of single colonies, and can be used as a supplement to the commonly used fungal medium for fungal culturomics studies in food.

, correspAuthors=Yuan YUE, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, authorCompany=null, fund=null, authors=null, authorsList=Yuan YUE, Meng-Shi ZHOU, Xiu-Juan FENG, Shu-Zhen HE, Yang SU, Wei HE, Rui LI), CN=ArticleExt(id=1153986730035241919, articleId=1153986714663117290, tenantId=1146029695717560320, journalId=1149652044408987649, language=CN, title=基于3种不同培养基的食品中真菌培养组学方法研究, columnId=1151895321958773274, journalTitle=食品安全质量检测学报, columnName=食品分析与检测, runingTitle=null, highlight=null, articleAbstract=

目的 通过对比3种真菌培养基的性能, 优化真菌培养组学技术, 建立食品中真菌培养组学解决方案。方法 利用食品检测常用的真菌培养基马铃薯葡萄糖琼脂(potato dextrose agar, PDA)、孟加拉红琼脂(rose Bengal agar)和1种新型Pan-Fungal真菌培养基(Pan-Fungal medium, PF)检测24份食品样本和2份能力验证样品。采用基质辅助激光解吸电离-飞行时间质谱法(matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, MALDI-TOF MS)对培养出的真菌进行分离鉴定。结果 酵母相真菌在3种培养基上均能良好生长, 菌落生长速度、菌落形态、大小及菌落数基本一致。丝状真菌在3种培养基上能正常生长, 但生长的速度和菌落形态均有明显的差异。24个样品共鉴定出12种真菌, 其中酵母相真菌2种, 丝状真菌10种。结论 PF真菌培养基具有良好的准确性和稳定性, 且培养出的菌落呈自限性生长, 有利于单菌落的分离鉴定, 可作为食品中常用真菌培养基的补充, 用于食品中真菌的培养组学研究。

, correspAuthors=岳苑, authorNote=null, correspAuthorsNote=
* 岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:
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#岳苑和周梦诗为共同第一作者

岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:

周梦诗(1995—), 女, 主要研究方向为病原微生物的诊断技术。E-mail:

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岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:

"}, bioImg=null, bioContent=

岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:

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周梦诗(1995—), 女, 主要研究方向为病原微生物的诊断技术。E-mail:

"}, bioImg=null, bioContent=

周梦诗(1995—), 女, 主要研究方向为病原微生物的诊断技术。E-mail:

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Ningxia Hui Autonomous Region Food Testing Institute, Key Laboratory of Quality Safety of Chinese Wolfberry and Wine, State Administration for Market Regulation, National Wine Product Quality Inspection and Testing Centre (Ningxia), Yinchuan 750002, China, bio=null, bioImg=null, bioContent=null, aboutCorrespAuthor=null), CN=AuthorExt(id=1184566796850971379, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, authorId=1184566796632867567, language=CN, stringName=冯秀娟, firstName=null, middleName=null, lastName=null, prefix=null, suffix=null, authorComment=null, nameInitials=null, affiliation=null, department=null, xref=1, address=1.宁夏回族自治区食品检测研究院, 国家市场监管重点实验室(枸杞与葡萄酒质量安全), 国家葡萄酒产品质量检验检测中心(宁夏), 银川 750002, bio=null, bioImg=null, bioContent=null, aboutCorrespAuthor=null)}, companyList=[AuthorCompany(id=1184566795693343447, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, xref=null, ext=[AuthorCompanyExt(id=1184566795701732056, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, companyId=1184566795693343447, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1. 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International Journal of Environmental Research and Public Health, 2020, 17(9): 3104., articleTitle=The gut microbiota of the Egyptian mongoose as an early warning indicator of ecosystem health in Portugal, refAbstract=null)], funds=[Fund(id=1184566800218997568, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, awardId=2023SJKY0002, language=CN, fundingSource=宁夏回族自治区市场监管厅科技计划项目(2023SJKY0002), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1184566795693343447, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, xref=null, ext=[AuthorCompanyExt(id=1184566795701732056, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, companyId=1184566795693343447, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1. 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Xi'an Biorealcoming Biomedical Center Co., Ltd., Xi'an 710000, China)), AuthorCompanyExt(id=1184566795819172573, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, companyId=1184566795798201051, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2.西安博睿康宁生物医学中心有限公司, 西安 710000 )])], figs=[ArticleFig(id=1184566798675493667, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Fig.1, caption=Growth of grape samples on different media (5 d), figureFileSmall=YxNCX8SQssngurwQTzBUZg==, figureFileBig=qCqgi/nLM96l0MxpSB2sSQ==, tableContent=null), ArticleFig(id=1184566798771962661, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=图1, caption=葡萄样本在不同培养基上的生长情况(5 d), figureFileSmall=YxNCX8SQssngurwQTzBUZg==, figureFileBig=qCqgi/nLM96l0MxpSB2sSQ==, tableContent=null), ArticleFig(id=1184566798843265831, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Fig.2, caption=Growth of isolates on different media (day 5), figureFileSmall=AV1AlugTyzlkGwm799eIOw==, figureFileBig=IfK1wyonX23iwluA7uj5Xg==, tableContent=null), ArticleFig(id=1184566798910374697, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=图2, caption=分离株在不同培养基上的生长情况(第5 d), figureFileSmall=AV1AlugTyzlkGwm799eIOw==, figureFileBig=IfK1wyonX23iwluA7uj5Xg==, tableContent=null), ArticleFig(id=1184566799048786731, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Table 1, caption=

Results of mould and yeast counts in different media in food

, figureFileSmall=null, figureFileBig=null, tableContent=
样品 PDA 孟加拉红琼脂 PF培养基
菌落数(×10-1) 结果 菌落数(×10-1) 结果 菌落数(×10-1) 结果
老式面包/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
大列巴/(CFU/g) 1/2 20 1/1 10 1/2 20
无水蛋糕/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
手撕软麻花/(CFU/g) 1/1 10 1/0 10 1/0 10
槽子糕/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
炉馍/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
江米条/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
麻辣锅巴/(CFU/g) 2/1 20 1/1 10 1/1 10
苹果脯/(CFU/g) 1/1 10 0/0 <10 0/0 10
枸杞果糕/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
炒瓜子/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
五香瓜子/(CFU/g) 1/0 10 0/0 <10 0/0 <10
麻辣条/(CFU/g) 0/0 <10 0/0 <10 1/0 10
麻辣片/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
原味发酵乳/(CFU/mL) 0/0 <10 0/0 <10 0/0 <10
发酵乳/(CFU/mL) 0/0 <10 0/0 <10 0/0 <10
玉米淀粉/(CFU/g) 4/3 40 4/4 40 5/6 60
生粉/(CFU/g) 2/2 20 2/2 20 2/1 20
枸杞1(干果)/(CFU/g) 7/6 70 8/8 80 9/10 100
枸杞2(干果)/(CFU/g) 14/16 150 15/14 150 19/17 180
黑果枸杞(干果)/(CFU/g) 5/5 50 5/4 50 5/5 50
葡萄1(鲜果)/(CFU/g) 11/10 110 9/9 90 14/13 140
葡萄2(鲜果)/(CFU/g) 18/17 180 23/17 200 31/25 280
红枣(鲜果)/(CFU/g) 10/11 110 12/12 120 12/12 120
), ArticleFig(id=1184566799149450029, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=表1, caption=

食品中不同培养基霉菌和酵母计数结果

, figureFileSmall=null, figureFileBig=null, tableContent=
样品 PDA 孟加拉红琼脂 PF培养基
菌落数(×10-1) 结果 菌落数(×10-1) 结果 菌落数(×10-1) 结果
老式面包/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
大列巴/(CFU/g) 1/2 20 1/1 10 1/2 20
无水蛋糕/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
手撕软麻花/(CFU/g) 1/1 10 1/0 10 1/0 10
槽子糕/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
炉馍/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
江米条/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
麻辣锅巴/(CFU/g) 2/1 20 1/1 10 1/1 10
苹果脯/(CFU/g) 1/1 10 0/0 <10 0/0 10
枸杞果糕/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
炒瓜子/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
五香瓜子/(CFU/g) 1/0 10 0/0 <10 0/0 <10
麻辣条/(CFU/g) 0/0 <10 0/0 <10 1/0 10
麻辣片/(CFU/g) 0/0 <10 0/0 <10 0/0 <10
原味发酵乳/(CFU/mL) 0/0 <10 0/0 <10 0/0 <10
发酵乳/(CFU/mL) 0/0 <10 0/0 <10 0/0 <10
玉米淀粉/(CFU/g) 4/3 40 4/4 40 5/6 60
生粉/(CFU/g) 2/2 20 2/2 20 2/1 20
枸杞1(干果)/(CFU/g) 7/6 70 8/8 80 9/10 100
枸杞2(干果)/(CFU/g) 14/16 150 15/14 150 19/17 180
黑果枸杞(干果)/(CFU/g) 5/5 50 5/4 50 5/5 50
葡萄1(鲜果)/(CFU/g) 11/10 110 9/9 90 14/13 140
葡萄2(鲜果)/(CFU/g) 18/17 180 23/17 200 31/25 280
红枣(鲜果)/(CFU/g) 10/11 110 12/12 120 12/12 120
), ArticleFig(id=1184566799245919023, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Table 2, caption=

Number of isolated bacterial colonies and identification results

, figureFileSmall=null, figureFileBig=null, tableContent=
名称 PDA 孟加拉红琼脂
 PF
菌落数/CFU 鉴定结果/% 菌落数/CFU 鉴定结果/% 菌落数/CFU 鉴定结果/%
近平滑假丝酵母 15 100.0 12 100.0 17 100.0
酿酒酵母 8 87.5 9 88.9 9 100.0
普通青霉 5 60.0 5 60.0 7 85.7
草酸青霉 6 100.0 6 83.3 6 100.0
产黄青霉 5 100.0 5 100.0 5 100.0
黄曲霉 4 100.0 4 100.0 7 100.0
卷枝毛霉 6 83.3 5 100.0 6 100.0
塔宾曲霉 29 96.6 32 97.0 40 97.6
总状毛霉 8 75.0 6 66.7 11 84.6
互格交链链格孢 14 100.0 17 100.0 17 100.0
黑根霉 7 100.0 5 100.0 8 100.0
枝孢样枝孢霉 9 90.0 8 88.9 11 84.6
), ArticleFig(id=1184566799354970928, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=表2, caption=

分离的菌落数与鉴定结果

, figureFileSmall=null, figureFileBig=null, tableContent=
名称 PDA 孟加拉红琼脂
 PF
菌落数/CFU 鉴定结果/% 菌落数/CFU 鉴定结果/% 菌落数/CFU 鉴定结果/%
近平滑假丝酵母 15 100.0 12 100.0 17 100.0
酿酒酵母 8 87.5 9 88.9 9 100.0
普通青霉 5 60.0 5 60.0 7 85.7
草酸青霉 6 100.0 6 83.3 6 100.0
产黄青霉 5 100.0 5 100.0 5 100.0
黄曲霉 4 100.0 4 100.0 7 100.0
卷枝毛霉 6 83.3 5 100.0 6 100.0
塔宾曲霉 29 96.6 32 97.0 40 97.6
总状毛霉 8 75.0 6 66.7 11 84.6
互格交链链格孢 14 100.0 17 100.0 17 100.0
黑根霉 7 100.0 5 100.0 8 100.0
枝孢样枝孢霉 9 90.0 8 88.9 11 84.6
), ArticleFig(id=1184566799426274098, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Table 3, caption=

Measurement results of moulds in proficiency testing infant formulae (CFU/g)

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样品编号 稀释度 霉菌
PDA 孟加拉红琼脂 PF
SP25 10-1 多不可计 多不可计 多不可计 多不可计 多不可计 多不可计
10-2 138 145 多不可计 多不可计 多不可计 多不可计
10-3 13 14 14 16 15 15
10-4 1 1 1 2 1 2
10-5 0 0 0 0 0 0
结果 14000 15000 15000
), ArticleFig(id=1184566799556297523, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=表3, caption=

能力验证婴幼儿配方奶粉中霉菌的测定结果(CFU/g)

, figureFileSmall=null, figureFileBig=null, tableContent=
样品编号 稀释度 霉菌
PDA 孟加拉红琼脂 PF
SP25 10-1 多不可计 多不可计 多不可计 多不可计 多不可计 多不可计
10-2 138 145 多不可计 多不可计 多不可计 多不可计
10-3 13 14 14 16 15 15
10-4 1 1 1 2 1 2
10-5 0 0 0 0 0 0
结果 14000 15000 15000
), ArticleFig(id=1184566799707292470, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Table 4, caption=

Determination of yeast in proficiency testing infant formulae (CFU/g)

, figureFileSmall=null, figureFileBig=null, tableContent=
样品编号 稀释度 酵母
PDA 孟加拉红琼脂 PF
22NP3 A 10-1 0 0 0 0 0 0
10-2 0 0 0 0 0 0
10-3 0 0 0 0 0 0
10-4 0 0 0 0 0 0
10-5 0 0 0 0 0 0
结果 <10 <10 <10
B 10-1 0 0 0 0 0 0
10-2 0 0 0 0 0 0
10-3 0 0 0 0 0 0
10-4 0 0 0 0 0 0
10-5 0 0 0 0 0 0
结果 <10 <10 <10
C 10-1 65 69 68 70 71 67
10-2 6 7 7 8 9 8
10-3 0 0 0 0 1 0
10-4 0 0 0 0 0 0
10-5 0 0 0 0 0 0
结果 670 690 690
), ArticleFig(id=1184566799866676024, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=表4, caption=

能力验证婴幼儿配方奶粉中酵母的测定结果(CFU/g)

, figureFileSmall=null, figureFileBig=null, tableContent=
样品编号 稀释度 酵母
PDA 孟加拉红琼脂 PF
22NP3 A 10-1 0 0 0 0 0 0
10-2 0 0 0 0 0 0
10-3 0 0 0 0 0 0
10-4 0 0 0 0 0 0
10-5 0 0 0 0 0 0
结果 <10 <10 <10
B 10-1 0 0 0 0 0 0
10-2 0 0 0 0 0 0
10-3 0 0 0 0 0 0
10-4 0 0 0 0 0 0
10-5 0 0 0 0 0 0
结果 <10 <10 <10
C 10-1 65 69 68 70 71 67
10-2 6 7 7 8 9 8
10-3 0 0 0 0 1 0
10-4 0 0 0 0 0 0
10-5 0 0 0 0 0 0
结果 670 690 690
), ArticleFig(id=1184566800026059579, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=EN, label=Table 5, caption=

Evaluation of the results of mould and yeast determination in infant formulae

, figureFileSmall=null, figureFileBig=null, tableContent=
检测项目 样品编号 测定结果
/(CFU/g)		 log10
(CFU/g)		 Z值
霉菌 SP25 15000 4.18 1.31
酵母 22NP3/样品A <10 - -
22NP3/样品B <10 - -
22NP3/样品C 690 2.84 -0.02
), ArticleFig(id=1184566800097362749, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1153986714663117290, language=CN, label=表5, caption=

婴幼儿配方奶粉中霉菌和酵母测定的结果评价

, figureFileSmall=null, figureFileBig=null, tableContent=
检测项目 样品编号 测定结果
/(CFU/g)		 log10
(CFU/g)		 Z值
霉菌 SP25 15000 4.18 1.31
酵母 22NP3/样品A <10 - -
22NP3/样品B <10 - -
22NP3/样品C 690 2.84 -0.02
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基于3种不同培养基的食品中真菌培养组学方法研究
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岳苑 1, #, * , 周梦诗 2, # , 冯秀娟 1 , 何淑桢 1 , 苏洋 1 , 何微 1 , 李睿 2
食品安全质量检测学报 | 食品分析与检测 2025,16(2): 302-308
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食品安全质量检测学报 | 食品分析与检测 2025, 16(2): 302-308
基于3种不同培养基的食品中真菌培养组学方法研究
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岳苑1, #, * , 周梦诗2, # , 冯秀娟1, 何淑桢1, 苏洋1, 何微1, 李睿2
作者信息
  • 1.宁夏回族自治区食品检测研究院, 国家市场监管重点实验室(枸杞与葡萄酒质量安全), 国家葡萄酒产品质量检验检测中心(宁夏), 银川 750002
  • 2.西安博睿康宁生物医学中心有限公司, 西安 710000

    • 岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:

    周梦诗(1995—), 女, 主要研究方向为病原微生物的诊断技术。E-mail:

通讯作者:

* 岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:
Research on fungal culturomics methods in food based on 3 kinds of different culture media
Yuan YUE1, * , Meng-Shi ZHOU2 , Xiu-Juan FENG1, Shu-Zhen HE1, Yang SU1, Wei HE1, Rui LI2
Affiliations
  • 1. Ningxia Hui Autonomous Region Food Testing Institute, Key Laboratory of Quality Safety of Chinese Wolfberry and Wine, State Administration for Market Regulation, National Wine Product Quality Inspection and Testing Centre (Ningxia), Yinchuan 750002, China
  • 2. Xi'an Biorealcoming Biomedical Center Co., Ltd., Xi'an 710000, China)
出版时间: 2025-01-25 doi: 10.19812/j.cnki.jfsq11-5956/ts.20240624001
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摘要
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目的 通过对比3种真菌培养基的性能, 优化真菌培养组学技术, 建立食品中真菌培养组学解决方案。方法 利用食品检测常用的真菌培养基马铃薯葡萄糖琼脂(potato dextrose agar, PDA)、孟加拉红琼脂(rose Bengal agar)和1种新型Pan-Fungal真菌培养基(Pan-Fungal medium, PF)检测24份食品样本和2份能力验证样品。采用基质辅助激光解吸电离-飞行时间质谱法(matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, MALDI-TOF MS)对培养出的真菌进行分离鉴定。结果 酵母相真菌在3种培养基上均能良好生长, 菌落生长速度、菌落形态、大小及菌落数基本一致。丝状真菌在3种培养基上能正常生长, 但生长的速度和菌落形态均有明显的差异。24个样品共鉴定出12种真菌, 其中酵母相真菌2种, 丝状真菌10种。结论 PF真菌培养基具有良好的准确性和稳定性, 且培养出的菌落呈自限性生长, 有利于单菌落的分离鉴定, 可作为食品中常用真菌培养基的补充, 用于食品中真菌的培养组学研究。

真菌  /  培养组学  /  分离培养

Objective To establish a solution for fungal culturomics in food by comparing the performance of 3 fungal culture media and optimising fungal culturomics techniques. Methods The 24 food samples and 2 proficiency test samples were tested using commonly used fungal culture media for food testing, including potato dextrose agar (PDA), rose Bengal agar, and a novel Pan Fungi medium (PF). The cultured fungi were isolated and identified using matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results Yeast phase fungi could grow well on all 3 kinds of media, and the colony growth rate, colony morphology size and colony numbers had no significant difference. Filamentous fungi were able to grow well on the 3 kinds of medias, but growth rate and colony morphology were significantly different. There were 12 kinds of fungal species identified in the 24 samples, including 2 kinds of yeast phase fungi and 10 kinds of filamentous fungi. Conclusion With excellent accuracy and stability and self-limited growth of cultured colonies, PF fungal medium is favourable for isolation and identification of single colonies, and can be used as a supplement to the commonly used fungal medium for fungal culturomics studies in food.

fungi  /  culturomics  /  isolation and culture
岳苑, 周梦诗, 冯秀娟, 何淑桢, 苏洋, 何微, 李睿. 基于3种不同培养基的食品中真菌培养组学方法研究. 食品安全质量检测学报, 2025 , 16 (2) : 302 -308 . DOI: 10.19812/j.cnki.jfsq11-5956/ts.20240624001
Yuan YUE, Meng-Shi ZHOU, Xiu-Juan FENG, Shu-Zhen HE, Yang SU, Wei HE, Rui LI. Research on fungal culturomics methods in food based on 3 kinds of different culture media[J]. Journal of Food Safety & Quality, 2025 , 16 (2) : 302 -308 . DOI: 10.19812/j.cnki.jfsq11-5956/ts.20240624001
正文
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0 引言
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微生物安全在食品工业产品质量控制中至关重要[1]。导致食品变质的微生物会产生代谢物, 从而改变产品的感官特性, 如外观、质地、味道和气味等, 降低其营养价值[2-3], 给食品行业造成巨大经济损失的同时对人类的健康和生命构成了严重的威胁[4-6]。
培养组学(culturomics)是一种基于微生物培养与高通量鉴定技术相结合的新方法, 其基础是利用非选择性或选择性富集和接种培养基, 优化不同的培养条件[7]。在分离可培养菌落的同时, 基于16S核糖体RNA (16S ribosomal RNA, 16S rRNA)基因或核糖体RNA内转录间隔区(ribosomal DNA internal transcribed spacer, ITS rRNA)变异区的测序结果和/或基质辅助激光解吸/电离-飞行时间质谱法(matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, MALDI-TOF MS)进行分子鉴定[8]。这种培养方法旨在模拟不同的自然条件, 检测边缘种群、评估其活力, 并利用可培养的分离物进行下游研究[9]。然而, 一些菌株需要非常特殊的体外生长条件, 在这种情况下, 需要定制培养组学方法。在很多真菌生物资源的应用场景中, 获得纯培养菌株有助于真正了解该真菌的生物学特性以及对组学研究提出的各种功能假说进行验证[10-11]。由此可见, 从样品中直接挑选菌落的精确度、适合菌落生长的培养基组分、培养条件的优化等是影响培养组学准确性的重要因素[12]。
霉菌和酵母菌同属于真菌, 广泛分布于自然界中, 具有较强的新陈代谢能力, 属于食品正常菌群的一部分。因此, 检测机构常选择霉菌和酵母菌作为指示菌, 通过其数量判断食品的污染程度, 评价食品的卫生质量[13]。微生物纯度的分析包括检测、鉴定和确定微生物数量[14]。用于分离食品中微生物的传统方法是以表型技术为基础的, 如观察微生物的形态和生长情况、镜检及生化分析等[15-16]。虽然这些方法可以有效地识别菌落, 但是操作复杂、耗时且需要检测人员丰富的工作经验, 这些缺点给微生物的常规分析带来困难[17]。本研究通过检测24种食品的霉菌和酵母计数, 比较2种食品检测常用的真菌培养基马铃薯葡萄糖琼脂(potato dextrose agar, PDA)、孟加拉红琼脂(rose Bengal agar)和1种新型Pan-Fungal真菌培养基(Pan-Fungal medium, PF), 结合MALDI-TOF MS鉴定结果, 建立常见食品中真菌培养组学解决方案, 以期用于食品中病原真菌的培养组学研究。
1 材料与方法
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1.1 样品
本研究部分样本来自宁夏回族自治区食品检测研究院2024年监督检验的样品, 包括面包、无水蛋糕、麻花、锅巴、麻辣条、枸杞果糕、发酵乳、淀粉、瓜子、枸杞等, 另一部分样本为购自农贸市场的葡萄、红枣等, 详见表1。
1.2 主要试剂
孟加拉红琼脂、PDA培养基(北京路桥技术股份有限公司); PF(西安博睿康宁生物医学中心有限公司); 质谱样本预处理试剂(郑州安图生物工程股份有限公司); 无水乙醇(分析纯, 国药集团化学试剂有限公司); 甲酸[质谱纯, 赛默飞世尔科技公司(中国)]。
1.3 主要仪器
MALDI-TOF MS基质辅助激光解吸电离-飞行时间质谱仪(郑州安图生物工程股份有限公司); KBF 240(E5.2)恒温恒湿培养箱[宾德环境试验设备(上海)有限公司]; Esco Labculture A2型生物安全柜(新加坡益世科企业发展有限公司)。
1.4 方法
1.4.1 样品检测
食品中霉菌和酵母的检测按照GB 4789.15—2016《食品安全国家标准 食品微生物学检验 霉菌和酵母计数》方法进行。以无菌操作称取样品25 g, 加入225 mL无菌稀释液(生理盐水)于无菌均质袋中, 用拍击式均质器拍打1~2 min, 制成1:10 (m:V)的样品匀液。取1 mL 1:10 (m:V)的样品匀液注入含有9 mL无菌稀释液的试管中, 在涡旋混合器上混匀, 此液为1:100 (m:V)样品匀液。按以上操作制备10倍递增系列稀释样品匀液。
将10倍递增系列稀释样品匀液每个稀释度分别吸取1 mL样品于两个无菌平皿内, 将20~25 mL冷却至46 ℃的PDA和孟加拉红琼脂倾注平皿, 并转动平皿使其混合均匀。PF为即用型培养基, 将10倍递增系列稀释样品匀液每个稀释度分别吸取1 mL样品于PF, 涂布。琼脂凝固后, 正置平板, 于28 ℃±1 ℃培养5 d后计数。PF于36 ℃±1 ℃培养48 h后计数。
1.4.2 分离纯化菌株
将从PDA、孟加拉红和PF培养基上所有单个菌落分别接种到新的PDA、孟加拉红和PF培养基上, 分别于28 ℃±1 ℃和36 ℃±1 ℃温度下培养直至生长出单个菌落。
1.4.3 分离菌株的鉴定
在1.5 mL离心管中加入1 mL 75%乙醇, 在生物安全柜中挑取所有纯化后的单个菌株的少量菌丝体或孢子于离心管中, 避免挑入培养基, 振荡混匀。13000 r/min离心2 min, 弃上清。加入30 µL甲酸, 振荡1 min。吸取溶液于靶板上, 待自然晾干后用MALDI-TOF MS进行检测。
1.4.4 能力验证样本的检测
依据GB 4789.15—2016, 对能力验证计划“婴幼儿配方奶粉中霉菌和酵母菌的测定”样品SP25和22NP3(包括A、B、C)进行检测。其中霉菌测定样品来自英国Laboratory of the Government Chemist (LGC), 酵母测定样品来自澳大利亚IFM Quality Services Pty Ltd (IFM)。
1.5 数据处理
霉菌和酵母菌落计数的方法按照GB 4789.15—2016进行。能力验证Z比分数(Z-score)是由能力验证的指定值和能力评定标准差计算的实验室偏倚的标准化度量, 依据CNAS—GL002《能力验证结果的统计处理和能力评价指南》(4.4能力统计量的计算)的规定计算。
2 结果与分析
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2.1 样品的检测结果
依据GB 4789.15—2016, 对24个样品中的霉菌和酵母进行计数, 结果见表1。
在食品样本的检测中, 酵母相真菌在3种培养基上均能良好生长, 菌落生长速度、菌落形态、大小及菌落数基本一致。丝状真菌在3种培养基上能正常生长, 但生长的速度和菌落形态均有明显的差异。丝状真菌在PDA和孟加拉红培养基上培养3 d后能观察到明显的菌落, 5 d能够看到菌落呈典型的弥漫性生长; 丝状真菌在PF 36 ℃±1 ℃培养2 d后能够观察到明显的菌落, 菌落形态紧致, 边缘清晰, 呈限制性生长。详见图1。
对于预包装食品而言, 糕点、炒货、发酵乳以及调味面制品等的卫生状况较好, 检测的结果均符合产品标准的要求。由于培养基上生长的菌落数较少, 无法对3种培养的生长情况进行比较。而对于枸杞、葡萄和红枣这些干果、鲜果而言, 受采摘晾晒等环节的影响, 检测出的菌落数较预包装食品更多。对于PF, 其说明书标注的培养时间为48 h, 根据生长情况可适当延长时间: (1)培养基上的菌落呈自限性生长, 所以延长培养时间不会影响菌落的计数和分离鉴定; (2)延长培养时间能够考虑到不同菌种具有不同的生长周期。
2.2 分离菌株的鉴定结果
将从PDA、孟加拉红和PF培养基上分离的单个菌落经预处理后用MALDI-TOF MS检测。MALDI-TOF MS的得分说明: 分值9.5~10.0为种水平置信, 可能的亚种; 分值9.0~9.5为种水平置信; 分值6.0~9.0为属水平置信; 分值0.0~6.0为不可置信。挑取的单个菌落数与鉴定结果见表2, 其中选择鉴定结果分值大于9.0的菌落(鉴定结果/%=鉴定结果分值大于9.0的菌落数/挑取菌落数)。
24个样品共鉴定出12种真菌, 其中酵母相真菌2种: 近平滑假丝酵母(Candida parapsilosis)和酿酒酵母(Saccharomyces cerevisiae); 丝状真菌10种: 普通青霉(Penicillium commune)、草酸青霉(Pcnicillium oxalicum)、产黄青霉(Penicillium chrysogenum)、黄曲霉(Aspergillus flavus)、卷枝毛霉(Mucor circinelloides)、塔宾曲霉(Aspergillus tubingensis)、总状毛霉(Mucor racemosus)、互格交链链格孢(Alternaria alternata)、黑根霉(Rhizopus nigricans)、枝孢样枝孢霉(Cladosporium cladosporioides), 详见图2。
通过表2、图2可知: 近平滑假丝酵母、草酸青霉、产黄青霉、黄曲霉、互格交链链格孢和黑根霉的分离的菌落全部达到鉴定分值大于9.0, 而鉴定结果百分比较低的是普通青霉和总状毛霉。3种培养基菌落计数和质谱鉴定结果存在差异的原因在于: (1)国标规定使用的孟加拉红琼脂中添加的孟加拉红成分可以作为着色剂, 霉菌菌落吸收后呈现出黄、绿、黑、白、青等多种颜色[18], 颜色可能会影响后续质谱鉴定的准确性。而PF上的菌落多为白色、乳白色或灰白色, 由于挑取单菌落的原则并不是根据菌落形态来区分菌种的, 所有的单菌落都应该被挑取鉴定。(2)丝状真菌在PDA培养基和孟加拉红琼脂培养基上菌落大且疏松, 呈现网状或毛絮状, 菌丝体在培养基表面呈蔓延生长, 尤其是青霉属、曲霉属和毛霉属, 给菌落计数工作带来了很大困难。而PF上的菌落呈自限性生长, 便于计数。(3)使用MALDI-TOF MS时, 分析前的样品制备方案可能会影响方法的结果[19]。首先为了减少蛋白质的降解, 保证核糖体蛋白的质量和良好的鉴定评分值, 用于MALDI-TOF MS鉴定的微生物菌落要尽量新鲜; 其次, 对于细胞壁更坚固的微生物(如真菌、粘性细菌等), 在样品制备过程中难以破坏, 需要在与基质混合之前进行蛋白质提取程序以获得良好的生物分子电离并提高其识别性[20-21]。
MALDI-TOF MS是一种快速、灵敏的微生物鉴定和表征分析方法, 与传统的生化自动化系统相比, MALDI-TOF MS的使用改善了微生物学实验室的工作流程, 并将获得微生物诊断的时间大幅度缩短[22]。对于威胁生命的感染或生长缓慢的菌株, 缩短病原细菌或真菌的鉴定时间显得尤为重要[23]。据估计, 真菌腐败造成的食品损失占全球损失总量的5%~10%[24]。QUÉRO等[25]使用MALDI-TOF MS技术快速识别食品变质霉菌。他们根据MALDI-TOF MS获得的136个物种的618个真菌菌株质谱图创建了一个数据库。该数据库在种水平上鉴定真菌的正确率大约为95%, 并且不受培养时间和培养基的影响(基于交叉验证的评估)。MOOTHOO-PADAYACHIE等[26]证明, MALDI-TOF MS技术在鉴定工业酵母菌方面是一种比分子方法更好的生物分型工具。他们在属和种水平上对酿酒酵母鉴定的正确率达到100%。
2.3 能力验证样本的检测结果
收到样品后, 确认样品编号、状态、是否存在破损或被污染的情况后, 根据样品的作业指导书分别对婴幼儿配方奶粉中的霉菌和酵母进行检测。PDA和孟加拉红琼脂28 ℃±1 ℃培养5 d、PF 36 ℃±1 ℃培养2 d后, 记录样品在3种培养基上的菌落数, 其中霉菌的测定结果见表3, 酵母的测定结果见表4。根据GB 4789.15—2016的要求, 选择菌落数在10~150 CFU的平板分别计数, 菌落数均大于150 CFU记录为多不可计, 菌落数在10~100之间时采用两位有效数字报告。最终霉菌的上报结果为15000 CFU/g, 酵母3个样品的上报结果分别为: <10 CFU/g、<10 CFU/g、690 CFU/g。
2.4 能力验证结果评价
能力验证评价结果见表5。能力验证评价准则为: |Z|值≤2.0时, 实验结果为满意; 2.0<|Z|<3.0时, 实验结果为可疑; |Z|≥3.0时, 实验结果为不满意。样品SP25和22NP3的评价结果均为满意。
在能力验证样本的检测过程中, PDA和孟加拉红琼脂在培养24 h后, 需要每天观察, 直至第5 d。培养时间越长, 菌丝蔓延的范围越大, 给菌落计数带来困难, 易造成计数结果偏低的情况。研究者们认为, 对于微生物而言, 自然环境是一个寡营养环境, 而人工培养基营养过高, 快速生长的微生物, 特别是真菌, 会导致绝大部分生长较慢的微生物被遮盖或失去生长空间而难以分离[27]。而霉菌在PF真菌培养基上, 菌落呈限制性生长, 即便超过了培养时间(48 h), 菌落也不会呈现蔓延趋势, 便于实验人员计数及后续的分离培养。能力验证评价结果Z值说明3种培养基准确度的一致性较好。
3 结论
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霉菌和酵母由于其较强的环境适应能力, 可以造成许多食物的腐败, 因此食品中霉菌和酵母的检测尤为重要。本研究通过比较3种真菌培养基(PDA、孟加拉红琼脂和PF真菌培养基), 结合MALDI-TOF MS鉴定结果, 建立食品中真菌培养组学解决方案。结果显示: 3种真菌培养基都具有良好的准确性和稳定性, 但在菌落数较多或卫生状况较差的条件下, PF真菌培养基的菌落边缘清晰, 呈自限性生长, 培养时间短, 有利于单个菌株的分离鉴定以及真菌菌落的计数。PF真菌培养基可作为食品中常用真菌培养基的补充, 用于食品中真菌的培养组学研究。培养组学是一种依赖培养的方法, 它使用大量不同培养条件的组合和高通量手段(即质谱法或DNA条形码)鉴定分离出菌株。广泛依赖培养的方法不仅能对以前无法培养的菌种进行培养和特征描述, 还能检测微生物群落中从未报道过的物种[28-30]。优化实验设计、探索新兴真菌分离培养技术对于培养组学具有十分重要的意义。
基金
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  • 宁夏回族自治区市场监管厅科技计划项目(2023SJKY0002)
文献
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doi: 10.19812/j.cnki.jfsq11-5956/ts.20240624001
  • 接收时间：2024-06-24
  • 首发时间：2025-07-21
  • 出版时间：2025-01-25
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  • 收稿日期：2024-06-24
基金
宁夏回族自治区市场监管厅科技计划项目(2023SJKY0002)
作者信息
    1.宁夏回族自治区食品检测研究院, 国家市场监管重点实验室(枸杞与葡萄酒质量安全), 国家葡萄酒产品质量检验检测中心(宁夏), 银川 750002
    2.西安博睿康宁生物医学中心有限公司, 西安 710000

通讯作者:

* 岳苑(1982—), 女, 博士, 高级工程师, 主要研究方向为食品中微生物及分子生物学检测。E-mail:
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2种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
Genus		 种数
Number of
species		 占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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