Department of Orthopedics, West China Hospital, Sichuan University, No.37 Guoxue Alley, Chengdu City, 610041, Sichuan, China.
West China School of Public Health, Sichuan University, Chengdu, China.
J Orthop Surg Res. 2019 Dec 12;14(1):433. doi: 10.1186/s13018-019-1508-z.
Methicillin-resistant Staphylococcus aureus (MRSA) strains present an urgent medical problem in osteomyelitis cases. Our previous study indicated that the YycFG two-component regulatory pathway is associated with the bacterial biofilm organization of MRSA strains. The aim of this study was to investigate the regulatory roles of ASyycG in the bacterial biofilm formation and the pathogenicity of MRSA strains using an antisense RNA strategy.
An ASyycG-overexpressing MRSA clinical isolate was constructed. The bacterial growth was monitored, and the biofilm biomass on bone specimens was examined using scanning electron microscopy and confocal laser scanning microscopy. Furthermore, quantitative RT-PCR (QRT-PCR) analysis was used to measure the expression of yycF/G/H and icaA/D in the MRSA and ASyycG strains. The expression of the YycG protein was quantified by Western blot assays. We validated the role of ASyycG in the invasive ability and pathogenicity of the strains in vivo using histology and peptide nucleic acid fluorescent in situ hybridization.
The results showed that overexpression of ASyycG lead to a reduction in biofilm formation and exopolysaccharide (EPS) synthesis compared to the control MRSA strains. The ASyycG strains exhibited decreased expression of the yycF/G/H and icaA/D genes. Furthermore, Western blot data showed that the production of the YycG protein was inhibited in the ASyycG strains. In addition, we demonstrated that ASyycG suppressed the invasive ability and pathogenicity of the strain in vivo using an SPF (specific pathogen free) rat model.
In summary, the overexpression of ASyycG leads to a reduction in biofilm formation and bacterial pathogenicity in vivo, which provides a potential target for the management of MRSA-induced osteomyelitis.
耐甲氧西林金黄色葡萄球菌（MRSA）菌株在骨髓炎病例中是一个紧迫的医学问题。我们之前的研究表明，YycFG 双组分调控途径与 MRSA 菌株的细菌生物膜组织有关。本研究旨在通过反义 RNA 策略研究 ASyycG 在 MRSA 菌株细菌生物膜形成和致病性中的调控作用。
构建了一个 ASyycG 过表达的 MRSA 临床分离株。通过扫描电子显微镜和共聚焦激光扫描显微镜观察骨标本上的细菌生物膜生物量，监测细菌生长。此外，采用定量 RT-PCR（QRT-PCR）分析测量 MRSA 和 ASyycG 菌株中 yycF/G/H 和 icaA/D 的表达。通过 Western blot 测定定量 YycG 蛋白的表达。我们通过组织学和肽核酸荧光原位杂交验证了 ASyycG 在体内菌株侵袭能力和致病性中的作用。
结果表明，与对照 MRSA 菌株相比，ASyycG 的过表达导致生物膜形成和胞外多糖（EPS）合成减少。ASyycG 株的 yycF/G/H 和 icaA/D 基因表达降低。此外，Western blot 数据表明 ASyycG 菌株中 YycG 蛋白的产生受到抑制。此外，我们使用 SPF（无特定病原体）大鼠模型证明 ASyycG 抑制了菌株在体内的侵袭能力和致病性。
总之，ASyycG 的过表达导致体内生物膜形成和细菌致病性降低，为管理 MRSA 引起的骨髓炎提供了一个潜在的靶点。
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