Laboratory AgriFing Department Biosciences and Landscape Architecture, Ghent University College, Voskenslaan 270, Gent B-9000, Belgium.
Food Microbiol. 2010 Feb;27(1):12-8. doi: 10.1016/j.fm.2009.04.006. Epub 2009 May 4.
The predominant lactic acid bacteria (LAB) microbiota associated with three types of modified atmosphere packaged (MAP) sliced cooked meat products (i.e. ham, turkey and chicken) was analyzed at sell-by date using a combination of culturing and molecular population fingerprinting. Likewise routine analyses during industrial MAP production, meat samples were plated on the general heterotrophic Plate Count Agar (PCA) and on the LAB-specific de Man, Rogosa, Sharpe (MRS) agar under different temperature and atmosphere conditions. Subsequently, community DNA extracts were prepared from culturable bacterial fractions harvested from both media and used for PCR targeting the V3 hyper-variable region of the 16S rRNA gene followed by denaturing gradient gel electrophoresis (DGGE) of PCR amplicons (PCR-DGGE). Irrespective of aerobic or anaerobic incubation conditions, V3-16S rDNA DGGE fingerprints of culturable fractions from PCA and MRS medium displayed a high level of similarity indicating that LAB constituted the most dominant group in the culturable bacterial community. Comparison of DGGE profiles of fractions grown at 20, 28 or 37 degrees C indicated that part of the culturable community consisted of psychrotrophs. Four DGGE bands were common among cooked ham, turkey and chicken products, suggesting that these represent the microbiota circulating in the plant where all three MAP product types were sliced and packaged. Based on band sequencing and band position analysis using LAB reference strains, these four bands could be assigned to Lactobacillus sakei and/or the closely related Lactobacillus fuchuensis, Lactobacillus curvatus, Carnobacterium divergens and Leuconostoc carnosum. In conclusion, the PCR-DGGE approach described in this study allows to discriminate, identify and monitor core and occasional LAB microbiota of MAP sliced cooked meat products and provides valuable complementary information to the current plating procedures routinely used in industrial plants.
本研究采用培养法和分子群体指纹图谱法相结合，分析了三种不同气调包装（MAP）切片熟肉（即火腿、火鸡和鸡肉）在销售期时的主要乳酸菌（LAB）菌群。在工业 MAP 生产过程中进行常规分析时，将肉样分别接种于普通异养平板计数琼脂（PCA）和 LAB 专用的 de Man，Rogosa，Sharpe（MRS）琼脂，在不同温度和气氛条件下进行平板培养。随后，从可培养细菌部分提取社区 DNA 提取物，分别从两种培养基中提取，用于针对 16S rRNA 基因 V3 高变区的 PCR 扩增，随后对 PCR 扩增子（PCR-DGGE）进行变性梯度凝胶电泳（DGGE）。无论有氧或厌氧培养条件如何，PCA 和 MRS 培养基中可培养细菌部分的 V3-16S rDNA DGGE 指纹图谱高度相似，表明 LAB 构成了可培养细菌群落中最主要的群体。比较在 20、28 或 37°C 下培养的部分 DGGE 图谱表明，部分可培养菌群由嗜冷菌组成。在火腿、火鸡和鸡肉产品中存在四条共同的 DGGE 条带，表明这些条带代表了在植物中循环的微生物群，所有三种 MAP 产品类型都在该植物中切片和包装。基于测序和使用 LAB 参考菌株的条带位置分析，这四个条带可被分配到清酒乳杆菌和/或密切相关的藤仓乳杆菌、弯曲乳杆菌、异样微球菌和乳脂球明串珠菌。综上所述，本研究中描述的 PCR-DGGE 方法可用于区分、识别和监测 MAP 切片熟肉产品的核心和偶然 LAB 菌群，并为工业工厂中常规使用的平板培养程序提供有价值的补充信息。
