Lau Tien-Tien Vicky, Puah Suat-Moi, Tan Jin-Ai Mary Anne, Merino Susana, Puthucheary S D, Chua Kek-Heng
Department of Biomedical Science, Faculty of Medicine, Universiti Malaya, 50603, Kuala Lumpur, Malaysia.
Department of Biomedical Science, Faculty of Medicine, Universiti Malaya, 50603, Kuala Lumpur, Malaysia.
Microb Pathog. 2023 Apr;177:106059. doi: 10.1016/j.micpath.2023.106059. Epub 2023 Mar 5.
Aeromonas dhakensis possesses dual flagellar systems for motility under different environments. Flagella-mediated motility is necessary for biofilm formation through an initial attachment of bacteria to the surface, but this has not been elucidated in A. dhakensis. This study investigates the role of polar (flaH, maf1) and lateral (lafB, lafK and lafS) flagellar genes in the biofilm formation of a clinical A. dhakensis strain WT187 isolated from burn wound infection. Five deletion mutants and corresponding complemented strains were constructed using pDM4 and pBAD33 vectors, respectively, and analyzed for motility and biofilm formation using crystal violet staining and real-time impedance-based assays. All mutants were significantly reduced in swimming (p < 0.0001), swarming (p < 0.0001) and biofilm formation using crystal violet assay (p < 0.05). Real-time impedance-based analysis revealed WT187 biofilm was formed between 6 to 21 h, consisting of early (6-10 h), middle (11-18 h), and late (19-21 h) stages. The highest cell index of 0.0746 was recorded at 22-23 h and biofilms began to disperse starting from 24 h. Mutants Δmaf1, ΔlafB, ΔlafK and ΔlafS exhibited reduced cell index values at 6-48 h when compared to WT187 which indicates less biofilm formation. Two complemented strains cmaf1 and clafB exhibited full restoration to wild-type level in swimming, swarming, and biofilm formation using crystal violet assay, hence suggesting that both maf1 and lafB genes are involved in biofilm formation through flagella-mediated motility and surface attachment. Our study shows the role of flagella in A. dhakensis biofilm formation warrants further investigations.
达卡气单胞菌拥有双鞭毛系统，以便在不同环境中运动。鞭毛介导的运动对于细菌通过初始附着于表面形成生物膜是必要的，但这在达卡气单胞菌中尚未得到阐明。本研究调查了极性（flaH、maf1）和侧向（lafB、lafK和lafS）鞭毛基因在从烧伤创面感染中分离出的临床达卡气单胞菌菌株WT187生物膜形成中的作用。分别使用pDM4和pBAD33载体构建了五个缺失突变体和相应的互补菌株，并使用结晶紫染色和基于实时阻抗的测定法分析了运动性和生物膜形成。所有突变体在游泳（p < 0.0001）、群游（p < 0.0001）以及使用结晶紫测定法的生物膜形成方面均显著降低（p < 0.05）。基于实时阻抗的分析表明，WT187生物膜在6至21小时之间形成，包括早期（6 - 10小时）、中期（11 - 18小时）和晚期（19 - 21小时）阶段。在22 - 23小时记录到最高细胞指数为0.0746，并且生物膜从24小时开始分散。与WT187相比，突变体Δmaf1、ΔlafB、ΔlafK和ΔlafS在6 - 48小时表现出降低的细胞指数值，这表明生物膜形成较少。两个互补菌株cmaf1和clafB在游泳、群游以及使用结晶紫测定法的生物膜形成方面表现出完全恢复到野生型水平，因此表明maf1和lafB基因均通过鞭毛介导的运动性和表面附着参与生物膜形成。我们的研究表明鞭毛在达卡气单胞菌生物膜形成中的作用值得进一步研究。
