Chemical analysis of cellular and extracellular carbohydrates of a biofilm-forming strain Pseudomonas aeruginosa PA14.
Chemical analysis of cellular and extracellular carbohydrates of a biofilm-forming strain Pseudomonas aeruginosa PA14.
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DOI：
10.1371/journal.pone.0014220
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发表时间：
2010-12-03
期刊：
影响因子：
3.7
通讯作者：
Sadovskaya I
中科院分区：
文献类型：
--
作者：
Coulon C;Vinogradov E;Filloux A;Sadovskaya I
铜绿假单胞菌是一种革兰氏阴性细菌和机会性病原体，可引起囊性纤维化（CF）患者持续危及生命的感染。生物膜的生长方式有利于其在各种环境中生存。大多数铜绿假单胞菌分离株，包括非粘液样实验室菌株PA14，能够在静液培养中形成厚膜，从而在气液（a -l）界面形成表面相关的生物膜。胞外多糖（EPS）被认为是形成这种生物膜的关键成分。在非黏液性P. aeruginosa菌株PA14中，生物膜基质的“脚手架”多糖和负责刚性A-L生物膜结构完整性的分子尚未确定。此外，LPS在这一过程中的作用尚不清楚，PA14的LPS o抗原的化学结构尚未阐明。本文对铜绿假单胞菌PA14的胞外碳水化合物进行了系统的分析。我们还通过化学方法和二维核磁共振光谱分析了LPS o抗原的化学结构。我们的结果表明，它由线性三糖重复单元组成，与P. aeruginosa Lanýi型O:2a，c （Lanýi-Bergman O-血清组10a， 10c； IATS血清型19）相同，具有以下结构：-4)-α- l - galnaca -(1- 3)-α- d - quinac -(1- 3)- α-L-Rha-(1-)。此外，在最小培养基中静态培养的PA14上清液中鉴定出EC o抗原多糖（EC O-PS）和甘油磷酸化的环β-(1,3)-葡聚糖。最后，在A-L界面形成的厚生物膜的细胞外基质除了含有eDNA外，还含有大量的lps样物质（至少占干重的20%）。我们对LPS o -抗原的化学结构进行了表征，表明o -抗原多糖是PA14丰富的胞外碳水化合物。我们提出的证据表明，脂多糖样物质被发现为铜绿假单胞菌的生物膜基质的组成部分。
Pseudomonas aeruginosa is a Gram-negative bacterium and an opportunistic pathogen, which causes persisting life-threatening infections in cystic fibrosis (CF) patients. Biofilm mode of growth facilitates its survival in a variety of environments. Most P. aeruginosa isolates, including the non-mucoid laboratory strain PA14, are able to form a thick pellicle, which results in a surface-associated biofilm at the air-liquid (A–L) interface in standing liquid cultures. Exopolysaccharides (EPS) are considered as key components in the formation of this biofilm pellicle. In the non-mucoid P. aeruginosa strain PA14, the “scaffolding” polysaccharides of the biofilm matrix, and the molecules responsible for the structural integrity of rigid A–L biofilm have not been identified. Moreover, the role of LPS in this process is unclear, and the chemical structure of the LPS O-antigen of PA14 has not yet been elucidated. In the present work we carried out a systematic analysis of cellular and extracellular (EC) carbohydrates of P. aeruginosa PA14. We also elucidated the chemical structure of the LPS O-antigen by chemical methods and 2-D NMR spectroscopy. Our results showed that it is composed of linear trisaccharide repeating units, identical to those described for P. aeruginosa Lanýi type O:2a,c (Lanýi-Bergman O-serogroup 10a, 10c; IATS serotype 19) and having the following structure: -4)-α-L-GalNAcA-(1–3)-α-D-QuiNAc-(1–3)- α-L-Rha-(1-. Furthermore, an EC O-antigen polysaccharide (EC O-PS) and the glycerol-phosphorylated cyclic β-(1,3)-glucans were identified in the culture supernatant of PA14, grown statically in minimal medium. Finally, the extracellular matrix of the thick biofilm formed at the A-L interface contained, in addition to eDNA, important quantities (at least ∼20% of dry weight) of LPS-like material. We characterized the chemical structure of the LPS O-antigen and showed that the O-antigen polysaccharide is an abundant extracellular carbohydrate of PA14. We present evidence that LPS-like material is found as a component of a biofilm matrix of P. aeruginosa.
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