背景：群体感应（QS）分子的酶促降解（称为群体猝灭，QQ）被认为是治疗生物膜相关感染和对抗抗生素耐药性的有前途的抗毒力疗法。据报道，最近发现的QQ酶MomL可有效降解各种革兰氏阴性病原体的不同N-酰基高丝氨酸内酯（AHLs）。在这里，我们调查了MomL对两种重要的医院病原体形成的生物膜的影响，铜绿假单胞菌和鲍曼不动杆菌。
方法：MomL在大肠杆菌BL21中表达并纯化。测试了MomL对具有羟基取代基的AHL的活性。铜绿假单胞菌PAO1和不动杆菌菌株的生物膜在96孔微量滴定板中形成。通过结晶紫染色评估生物膜的形成，电镀和荧光显微镜。还测试了MomL对生物膜对抗生素的敏感性的影响。我们进一步评估了由铜绿假单胞菌和鲍曼不动杆菌形成的双物种生物膜中的MomL，以及在伤口模型中形成的生物膜中。还在秀丽隐杆线虫模型中测试了MomL对鲍曼不动杆菌毒力的影响。
结果：MomL减少了体外在微量滴定板中形成的铜绿假单胞菌PAO1和鲍曼不动杆菌LMG10531的生物膜中的生物膜形成，并增加了对不同抗生素的生物膜敏感性。然而,在双物种生物膜和伤口模型生物膜中未检测到显着差异。此外,在秀丽隐杆线虫模型中，MomL不影响鲍曼不动杆菌的毒力。最后,MomL对不动杆菌菌株生物膜的影响似乎是菌株依赖性的。
结论：我们的结果表明，尽管MomL对微量滴定板中形成的铜绿假单胞菌和鲍曼不动杆菌生物膜显示出有希望的抗生物膜作用，在更可能模拟现实生活情况的条件下，对生物膜形成的影响不那么明显，甚至不存在。我们的数据表明，为了更好地了解QQ酶治疗生物膜相关感染的潜在适用性，需要使用更复杂的模型系统。
BACKGROUND: The enzymatic degradation of quorums sensing (QS) molecules (called quorum quenching, QQ) has been considered as a promising anti-virulence therapy to treat biofilm-related infections and combat antibiotic resistance. The recently-discovered QQ enzyme MomL has been reported to efficiently degrade different N-acyl homoserine lactones (AHLs) of various Gram-negative pathogens. Here we investigated the effect of MomL on biofilms formed by two important nosocomial pathogens, Pseudomonas aeruginosa and Acinetobacter baumannii.
METHODS: MomL was expressed in E.coli BL21 and purified. The activity of MomL on AHLs with hydroxyl substituent was tested. Biofilms of P. aeruginosa PAO1 and Acinetobacter strains were formed in 96-well microtiter plates. Biofilm formation was evaluated by crystal violet staining, plating and fluorescence microscopy. The effect of MomL on biofilm susceptibility to antibiotics was also tested. We further evaluated MomL in dual-species biofilms formed by P. aeruginosa and A. baumannii, and in biofilms formed in a wound model. The effect of MomL on virulence of A. baumannii was also tested in the Caenorhabditis elegans model.
RESULTS: MomL reduced biofilm formation and increased biofilm susceptibility to different antibiotics in biofilms of P. aeruginosa PAO1 and A. baumannii LMG 10531 formed in microtiter plates in vitro. However, no significant differences were detected in the dual-species biofilm and in wound model biofilms. In addition, MomL did not affect virulence of A. baumannii in the C. elegans model. Finally, the effect of MomL on biofilm of Acinetobacter strains seems to be strain-dependent.
CONCLUSIONS: Our results indicate that although MomL showed a promising anti-biofilm effect against P. aeruginosa and A. baumanii biofilms formed in microtiter plates, the effect on biofilm formation under conditions more likely to mimic the real-life situation was much less pronounced or even absent. Our data indicate that in order to obtain a better picture of potential applicability of QQ enzymes for the treatment of biofilm-related infections, more elaborate model systems need to be used.
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