Department of Food Hygiene and Environmental Health, PO Box 66, FI-00014, University of Helsinki, Finland.
Int J Food Microbiol. 2011 Jan 5;144(3):360-6. doi: 10.1016/j.ijfoodmicro.2010.10.018. Epub 2010 Nov 17.
Characterization of psychrotrophic lactic acid bacteria (LAB) and Brochothrix thermosphacta communities is needed to understand the microbial ecology of spoilage of modified atmosphere-packed (MAP) meats. To overcome the limitations of the currently used methods for the characterization of psychrotrophic bacterial communities in meat, we developed a culture-independent, 16S rRNA gene-targeted terminal restriction fragment length polymorphism (T-RFLP) method. An identification library consisting of 100 Gram-positive and 30 Gram-negative meat-associated bacterial strains was set up to identify the terminal restriction fragments derived from the communities. The taxonomic resolution level of the T-RFLP method was in between genus and species within the investigated LAB strains and within family and genus within the investigated Gram-negative strains. The established library was applied to identify the members of bacterial communities in MAP minced meat at the end of the shelf life. The T-RFLP results and plate counts on Man-Rogosa-Sharpe, Violet Red Bile Glucose, and Streptomycin sulfate thallium acetate actidione agars indicated that LAB and B. thermosphacta predominated in meat. The bacterial taxa associated with the T-RFLP results were compared to those identified among plate-grown LAB isolates by numerical ribopattern analysis. Both methods agreed that Leuconostoc spp. and Carnobacterium spp. prevailed in the LAB community in minced meat followed by Lactobacillus algidus, Lactococcus spp. and Weissella spp. Colony identification revealed that Leuconostoc gasicomitatum, L. gelidum, Carnobacterium divergens and C. maltaromaticum were the predominant LAB species. The T-RFLP results were shown to correlate with viable counts of Leuconostoc spp. and B. thermosphacta. The T-RFLP method was found to be a useful tool enabling rapid and high-throughput characterization of psychrotrophic bacteria prevailing in MAP meat.
需要对低温乳酸菌（LAB）和嗜冷性芽孢杆菌（Brochothrix thermosphacta）群落进行特征描述，以了解改性气氛包装（MAP）肉类腐败的微生物生态学。为了克服目前用于描述肉类低温细菌群落的方法的局限性，我们开发了一种不依赖培养的 16S rRNA 基因靶向末端限制性片段长度多态性（T-RFLP）方法。建立了一个由 100 株革兰氏阳性和 30 株革兰氏阴性与肉相关的细菌菌株组成的鉴定文库，以鉴定从群落中获得的末端限制性片段。T-RFLP 方法的分类分辨率水平在研究的 LAB 菌株的属和种之间以及研究的革兰氏阴性菌株的科和属之间。建立的文库用于鉴定货架期结束时 MAP 肉末中细菌群落的成员。T-RFLP 结果和在 Man-Rogosa-Sharpe、Violet Red Bile Glucose、Streptomycin sulfate thallium acetate actidione 琼脂上的平板计数表明，LAB 和 B. thermosphacta 在肉中占优势。与平板培养的 LAB 分离物的 T-RFLP 结果相关的细菌分类群通过数值核糖体图谱分析进行了比较。这两种方法都认为，在肉末中的 LAB 群落中，Leuconostoc spp. 和 Carnobacterium spp. 占优势，其次是 Lactobacillus algidus、Lactococcus spp. 和 Weissella spp.。菌落鉴定表明，Leuconostoc gasicomitatum、L. gelidum、Carnobacterium divergens 和 C. maltaromaticum 是主要的 LAB 物种。T-RFLP 结果与 Leuconostoc spp. 和 B. thermosphacta 的活菌计数相关。T-RFLP 方法被证明是一种有用的工具，可快速高通量地描述 MAP 肉中占优势的低温细菌。
