Abstract：
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.
摘要：
达克气单胞菌在不同环境下具有双鞭毛系统的运动性。鞭毛介导的运动性是通过细菌最初附着于表面而形成生物膜所必需的，但这在达克氏杆菌中还没有得到阐明。本研究调查了极性(flaH，maf1)和横向(lafB，lafK和lafS）鞭毛基因在从烧伤伤口感染中分离出的临床A.dhakensis菌株WT187的生物膜形成中。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，分别,并使用结晶紫染色和基于实时阻抗的测定来分析运动性和生物膜形成。所有突变体在游泳中显著减少(p<0.0001),蜂群(p<0.0001)和生物膜形成使用结晶紫测定(p<0.05)。基于实时阻抗的分析显示WT187生物膜在6-21小时之间形成，包括早期（6-10小时），中间（11-18小时），和后期（19-21小时）阶段。在22-23小时记录的最高细胞指数为0.0746，生物膜从24小时开始分散。突变体Δmaf1，ΔlafB，与WT187相比，ΔlafK和ΔlafS在6-48小时表现出降低的细胞指数值，这表明生物膜的形成较少。两个互补菌株cmaf1和clafB在游泳中表现出完全恢复到野生型水平，蜂拥而至，用结晶紫分析法形成生物膜，因此表明maf1和lafB基因都通过鞭毛介导的运动和表面附着参与生物膜的形成。我们的研究表明鞭毛在达克曲霉生物膜形成中的作用值得进一步研究。
