基于PacBio SMRT技术的婴幼儿配方奶粉微生物安全性评价
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国家自然科学基金优秀青年科学基金项目(31622043)


Evaluation of Microbial Safety in Infant Formula Based on PacBio SMRT Sequencing
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    摘要:

    目的:婴幼儿配方奶粉中的大量细菌已被灭活,然而灭活微生物仍影响产品品质和货架期。现行纯培养技术只能对食品基质中活菌进行检测,无法全面评估污染微生物组成,而基于宏基因组学策略能够对婴幼儿配方奶粉加工过程中污染微生物进行全面客观的评价。方法:采集6份婴幼儿配方奶粉,使用细菌纯培养、单分子实时测序技术(PacBio SMRT)和微滴式数字PCR(ddPCR)联用技术对其污染微生物组成进行评价。结果:采用纯培养技术在样品中均未检测到大肠菌群、沙门氏菌、金黄色葡萄球菌、阪崎肠杆菌和芽孢。使用PacBio SMRT测序技术,在婴幼儿配方奶粉中共鉴定出14个细菌门、138个细菌属和255个细菌种。其中6份样品均存在过嗜冷菌蜡样芽孢杆菌,其平均相对含量高达23.54%,这一灭活的细菌也会影响产品货架期。此外,在IF3、IF4、IF5和IF6样品中,检测到相对含量较低的致病菌大肠杆菌,其相对含量分别为0.06%,0.01%,0.15%和0.05%。采用ddPCR技术对SMRT测序方法检测到的致病菌蜡样芽孢杆菌和大肠杆菌进行定量分析,发现每克奶粉样品中蜡样芽孢杆菌和大肠杆菌的平均拷贝数取对数后分别为5.45和4.89。结论:在6份婴幼儿配方奶粉中均未检出致病菌,符合食品安全国家标准,然而奶粉中被灭活的蜡样芽孢杆菌和大肠杆菌可能增加产品腐败的风险,影响产品的货架期。本研究结合多个技术全面评价婴幼儿配方奶粉的微生物污染情况,初步建立婴幼儿配方奶粉的微生物检测技术。

    Abstract:

    Objective: A large number of bacteria in infant formula have been inactivated, but they still have adverse effects on the quality and shelf life of products. The traditional pure culture only can detect the viable bacteria, so evaluating the microbial contamination in infant formula by Metagenome strategy is essential and makes up for the disadvantage. Methods: Microbial contamination of six infant formula were evaluated by pure culture, PacBio SMRT (single molecule real-time) and ddPCR (droplet digital PCR). Results: Coliform groups, Salmonella, Staphylococcus aureus, Enterobacter sakazakii and spores were absent in the infant formula samples by traditional pure culture. A total of 14 bacterial phylum, 138 bacterial genus and 255 bacterial species were identified in infant formula, and psychrotrophic bacteria Bacillus cereus that might affect the product shelf life were at some point present in the infant formula samples, the average relative abundance were up to 23.54%. A low relative abundance of Escherichia coli were detected in the IF3, IF4, IF5 and IF6 infant formula samples, and the relative abundance was 0.06%, 0.01%, 0.15% and 0.05%, respectively. The results of ddPCR showed that the copy numbers after taking the logarithm of Bacillus cereus and Escherichia coli were 5.45 and 4.89, respectively. Conclusion: There were no pathogens were detected in six infant formula, in line with national standards. But Bacillus cereus and Escherichia coli that might affect the product shelf life were at some point present in the infant formula samples. This study combined with multiple technologies to fully assess the microbial contamination in infant formula, establish microbial detection method of infant formula.

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边燕飞;席晓霞;郑艺;侯强川;孙天松;孙志宏;.基于PacBio SMRT技术的婴幼儿配方奶粉微生物安全性评价[J].中国食品学报,2019,19(10):212-220

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  • 在线发布日期: 2019-11-21
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