A refined technique for extraction of extracellular matrices from bacterial biofilms and its applicability.
A refined technique for extraction of extracellular matrices from bacterial biofilms and its applicability.
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DOI：
10.1111/1751-7915.12155
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发表时间：
2015-05
影响因子：
5.7
通讯作者：
Mizunoe Y
中科院分区：
文献类型：
--
作者：
Chiba A;Sugimoto S;Sato F;Hori S;Mizunoe Y
生物被膜细菌嵌入由多糖、蛋白质和/或胞外DNA组成的聚合物细胞外基质(ECM)中，对抗菌剂和宿主免疫系统具有高度的抵抗力。为了了解生物膜形成和维持的分子机制，并制定针对慢性生物膜相关感染的治疗对策，可靠的分离ECM的方法是不可避免的。本研究对最近报道的细胞外基质提取方法进行了改进，并对其适用性进行了评价。使用三种金黄色葡萄球菌生物膜，其中蛋白质、多糖或EDNA是其完整性的主要贡献者，使用盐和洗涤剂提取ECM。我们发现，用1.5 M氯化钠(NaC L)提取细胞外基质蛋白以及多糖和EDNAS是最优的。此外，细胞外基质的有效分离不需要长时间的培养。氯化锂(LiCl)与氯化钠相当，但价格更高。与十二烷基硫酸钠相比，氯化钠几乎不会引起细胞内蛋白质的泄漏，也不会影响生物膜内细菌细胞的活力。此外，该方法还适用于革兰氏阳性表皮葡萄球菌、革兰氏阴性大肠埃希菌和铜绿假单胞菌等细菌。因此，这种精制的方法非常简单、快速、低成本和非侵入性，可以广泛应用。
Biofilm-forming bacteria embedded in polymeric extracellular matrices (ECMs) that consist of polysaccharides, proteins and/or extracellular DNAs (eDNAs) acquire high resistance to antimicrobial agents and host immune systems. To understand molecular mechanisms of biofilm formation and maintenance and to develop therapeutic countermeasures against chronic biofilm-associated infections, reliable methods to isolate ECMs are inevitable. In this study, we refined the ECM extraction method recently reported and evaluated its applicability. Using three Staphylococcus aureus biofilms in which proteins, polysaccharides or eDNAs are major contributors to their integrity, ECMs were extracted using salts and detergents. We found that extraction with 1.5 M sodium chloride (NaCl) could be optimum for not only ECM proteins but also polysaccharides and eDNAs. In addition, long-time incubation was not necessary for efficient ECM isolation. Lithium chloride (LiCl) was comparative to NaCl but is more expensive. In contrast to SDS, NaCl hardly caused leakage of intracellular proteins and did not affect viability of bacterial cells within biofilms. Furthermore, this method is applicable to other bacteria such as Gram-positive Staphylococcus epidermidis and Gram-negative Escherichia coli and Pseudomonas aeruginosa. Thus, this refined method is very simple, rapid, low cost and non-invasive and could be used for a broad range of applications.
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影响因子：
3.2
作者：
Jucker, BA;Harms, H;Zehnder, AJB
通讯作者：
Zehnder, AJB
DOI：
10.1073/pnas.0910560107
发表时间：
2010-02-02
影响因子：
11.1
作者：
Romero, Diego;Aguilar, Claudio;Kolter, Roberto
通讯作者：
Kolter, Roberto
影响因子：
2.6
作者：
Bae, T;Schneewind, O
通讯作者：
Schneewind, O
EDNA释放和降解的调节会影响金黄色葡萄球菌生物膜成熟。
影响因子：
3.7
作者：
Mann EE;Rice KC;Boles BR;Endres JL;Ranjit D;Chandramohan L;Tsang LH;Smeltzer MS;Horswill AR;Bayles KW
通讯作者：
Bayles KW
影响因子：
2.7
作者：
Allison, DG
通讯作者：
Allison, DG