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A new protectant medium preserving bacterial viability after freeze drying.
Microbiological Research ( IF 6.7 ) Pub Date : 2020-03-06 , DOI: 10.1016/j.micres.2020.126454
Sara Bellali 1 , Jacques Bou Khalil 2 , Anthony Fontanini 2 , Didier Raoult 1 , Jean-Christophe Lagier 1
Affiliation  

Freeze-drying technology has been widely considered for decades as a suitable technique to preserve microorganisms. However, protective agents must be added prior to freeze drying to improve the survival and storage stability of the bacteria.

The objective of our study was to evaluate the effect of a new protectant medium containing sucrose (10 %), trehalose (10 %), skimmed milk (10 %) and antioxidants on the viability of gut bacteria under different storage conditions. Two strains were tested, Escherichia coli and Akkermansia muciniphila, as examples of facultative aerobic and anaerobic bacteria, respectively. We studied the cell viability and bacterial morphology in 5 fecal samples in the presence and absence of this protectant medium using plating technique, flow cytometry and scanning electron microscopy.

The results of bacterial viability assessed by plating method showed that the protectant medium yielded higher survival rates for both strains whatever the storage conditions (85–93 %) compared to normal saline solution (0.36–37.50 %). It also showed its effectiveness on fecal samples, where bacterial viability after freeze-drying was 89.47 ± 7.63 % and 84.01 ± 7.44 %, as evidenced by flow cytometry analysis and plating method. However unprotected samples showed the lowest cell viability at 19.01 ± 12.88 % and 13.23 ± 9.56 %, as measured by flow cytometry and plating method. In addition, bacterial size and shape were conserved in the protectant medium. In contrast, storage without protectant medium severely damaged bacterial morphology.

In conclusion, our study is the first to use morphological features as well as culture-dependant and culture-independent tests to evaluate the effectiveness of a new protectant medium.



中文翻译:

一种新的保护性培养基,可在冷冻干燥后保持细菌的生存能力。

数十年来,冷冻干燥技术已被广泛认为是保存微生物的合适技术。但是,必须在冷冻干燥之前添加保护剂,以提高细菌的存活率和储存稳定性。

我们研究的目的是评估一种新的含蔗糖(10%),海藻糖(10%),脱脂牛奶(10%)和抗氧化剂的保护性培养基在不同储存条件下对肠道细菌活力的影响。测试了两种菌株,分别为兼性需氧细菌和厌氧细菌的大肠杆菌粘液阿克曼氏菌。我们使用平板技术,流式细胞仪和扫描电子显微镜研究了5种粪便样品在存在和不存在这种保护性培养基的情况下的细胞活力和细菌形态。

通过平板方法评估的细菌生存力结果表明,与普通盐溶液(0.36-37.50%)相比,无论储存条件如何(85–93%),保护性培养基都能对两种菌株产生更高的存活率。它也对粪便样品显示出有效性,通过流式细胞仪分析和铺板方法证明,粪便样品的冻干后细菌活力分别为89.47±7.63%和84.01±7.44%。然而,如通过流式细胞术和铺板方法所测量的,未保护的样品显示出最低的细胞活力,分别为19.01±12.88%和13.23±9.56%。另外,细菌的大小和形状在保护介质中得以保留。相反,没有保护介质的储存会严重破坏细菌的形态。

总之,我们的研究是第一个使用形态学特征以及依赖于文化和不依赖于文化的测试来评估新型保护剂介质的有效性的研究。

更新日期:2020-03-06
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