Generation of a highly attenuated strain of Pseudomonas aeruginosa for commercial production of alginate.,Microbial Biotechnology

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Generation of a highly attenuated strain of Pseudomonas aeruginosa for commercial production of alginate.
Microbial Biotechnology ( IF 4.8 ) Pub Date : 2019-04-21 , DOI: 10.1111/1751-7915.13411
Meagan E Valentine ₁ , Brandon D Kirby ₁ , Thomas R Withers ₁ , Shannon L Johnson ₂ , Timothy E Long ₃ , Youai Hao ₄ , Joseph S Lam ₄ , Richard M Niles ₁ , Hongwei D Yu _{1,

5}

Affiliation

Alginate is an important polysaccharide that is commonly used as a gelling agent in foods, cosmetics and healthcare products. Currently, all alginate used commercially is extracted from brown seaweed. However, with environmental changes such as increasing ocean temperature and the increasing number of biotechnological uses of alginates with specific properties, there is an emerging need for more reliable and customizable sources of alginate. An alternative to seaweed for alginate production is Pseudomonas aeruginosa, a common Gram‐negative bacterium that can form alginate‐containing biofilms. However, P. aeruginosa is an opportunistic pathogen that can cause life‐threatening infections in immunocompromised patients. Therefore, we sought to engineer a non‐pathogenic P. aeruginosa strain that is safe for commercial production of alginate. Using a homologous recombination strategy, we sequentially deleted five key pathogenicity genes from the P. aeruginosa chromosome, resulting in the marker‐free strain PGN5. Intraperitoneal injection of mice with PGN5 resulted in 0% mortality, while injection with wild‐type P. aeruginosa resulted in 95% mortality, providing evidence that the systemic virulence of PGN5 is highly attenuated. Importantly, PGN5 produces large amounts of alginate in response to overexpression of MucE, an activator of alginate biosynthesis. The alginate produced by PGN5 is structurally identical to alginate produced by wild‐type P. aeruginosa, indicating that the alginate biosynthetic pathway remains functional in this modified strain. The genetic versatility of P. aeruginosa will allow us to further engineer PGN5 to produce alginates with specific chemical compositions and physical properties to meet different industrial and biomedical needs.

中文翻译：

产生高度减毒的铜绿假单胞菌菌株，用于海藻酸盐的商业生产。

海藻酸盐是一种重要的多糖，通常用作食品、化妆品和保健产品中的胶凝剂。目前，商业上使用的所有海藻酸盐都是从褐海藻中提取的。然而，随着海洋温度升高等环境变化以及具有特定特性的藻酸盐生物技术用途的增加，人们越来越需要更可靠和可定制的藻酸盐来源。用于生产藻酸盐的海藻替代品是铜绿假单胞菌，这是一种常见的革兰氏阴性细菌，可以形成含藻酸盐的生物膜。然而，铜绿假单胞菌是一种机会性病原体，可在免疫功能低下的患者中引起危及生命的感染。因此，我们试图设计一种可安全用于海藻酸盐商业生产的非致病性铜绿假单胞菌菌株。采用同源重组策略，我们从铜绿假单胞菌染色体上依次删除了五个关键致病基因，产生了无标记菌株PGN5。腹腔注射 PGN5 的小鼠死亡率为 0%，而注射野生型铜绿假单胞菌则死亡率为 95%，这提供了 PGN5 的全身毒力高度减弱的证据。重要的是，PGN5 响应 MucE（藻酸盐生物合成的激活剂）的过度表达而产生大量藻酸盐。 PGN5 产生的藻酸盐在结构上与野生型铜绿假单胞菌产生的藻酸盐相同，表明藻酸盐生物合成途径在该修饰菌株中仍然发挥功能。 P的遗传多样性铜绿假单胞菌将使我们能够进一步设计PGN5，生产具有特定化学成分和物理性质的藻酸盐，以满足不同的工业和生物医学需求。

更新日期：2019-04-21

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