Background: Brevinin2 HYba5 (Peptide 29) is a novel cationic peptide identified from an endemic frog, Hydrophylax bahuvistara. Staphylococcus aureus and Enterococcus faecalis are troublesome biofilm-forming pathogens associated with nosocomial and community-acquired infections and contribute to the severity of infections associated with implanted devices and chronic wounds. Co-existence of both pathogens in biofilm mode contributes to an increased antibiotic resistance, treatment failure and hence persistent disease burden. Identifying a novel and stable, less toxic compound targeting multispecies biofilm with a lower probability of acquiring resistance in comparison to antibiotics is highly warranted. Objective: Evaluate the activity of Brevinin2 HYba5 against S. aureus and E. faecalis mixed biofilm. Methods: The anti-biofilm activity of peptide 29 was tested by Crystal violet assay, Confocal laser scanning Microscopy (CLSM) and MTT Assay. Cytotoxicity of the peptide was tested in RBC and L929 fibroblast cell line. Biofilm inhibitory activity of the peptide was evaluated at different temperatures, pH, serum and plasma concentrations. The antibiofilm potential of the peptide was tested against polymicrobial biofilm by Fluorescent in situ hybridisation (FISH) and plate counting on HiCromeTM UTI Agar media. Results: The peptide 29 could inhibit biofilm formation of S. aureus and E. faecalis individually as well as in polymicrobial biofilm at 75 μM concentration. The peptide maintained its antibiofilm potential at different temperatures, serum and plasma concentrations. Activity of the peptide was high at acidic and neutral pH but found to get reduced towards alkaline pH. The peptide is nonhemolytic and does not exhibit significant cytotoxicity against the L929 fibroblast cell line (92.80% cell viability). Conclusion: The biofilm inhibition property makes peptide 29 a promising candidate for the management of S. aureus and E. faecalis biofilm, especially in catheter-associated devices to prevent the initial colonization and thus can ease the burden of pathogenic biofilm-associated infections.

中文翻译：

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抗金黄色葡萄球菌和粪肠球菌多种微生物生物膜的新型 Brevinin2 HYba5 肽

背景：Brevinin2 HYba5（肽 29）是从地方性青蛙 Hydrophylax bahuvistara 中鉴定出的一种新型阳离子肽。金黄色葡萄球菌和粪肠球菌是与医院和社区获得性感染相关的麻烦的生物膜形成病原体，并导致与植入装置和慢性伤口相关的感染的严重性。两种病原体以生物膜模式共存会导致抗生素耐药性增加、治疗失败，从而导致持续的疾病负担。非常有必要鉴定一种新型、稳定、毒性较小的针对多物种生物膜的化合物，并且与抗生素相比，获得耐药性的可能性更低。目的：评价 Brevinin2 HYba5 对金黄色葡萄球菌和粪肠球菌混合生物膜的活性。方法：采用结晶紫法、共聚焦激光扫描显微镜（CLSM）和MTT法检测肽29的抗生物膜活性。在 RBC 和 L929 成纤维细胞系中测试了肽的细胞毒性。在不同温度、pH、血清和血浆浓度下评估肽的生物膜抑制活性。通过荧光原位杂交 (FISH) 和 HiCromeTM UTI 琼脂培养基上的平板计数，测试了肽针对多种微生物生物膜的抗生物膜潜力。结果：肽 29 在 75 μM 浓度下可以单独抑制金黄色葡萄球菌和粪肠球菌的生物膜形成，也可以抑制多种微生物生物膜的形成。该肽在不同温度、血清和血浆浓度下保持其抗生物膜潜力。该肽的活性在酸性和中性 pH 值下较高，但在碱性 pH 值下活性降低。该肽是非溶血性的，并且对 L929 成纤维细胞系（92.80% 细胞活力）没有表现出显着的细胞毒性。结论：生物膜抑制特性使肽 29 成为治疗金黄色葡萄球菌和粪肠球菌生物膜的有希望的候选者，特别是在导管相关装置中防止初始定植，从而减轻致病性生物膜相关感染的负担。