Department of Microbiology, School of Medicine, Hamadan University of Medical Sciences, Hamadan, Iran.
Department of Microbiology, School of Medicine, Hamadan University of Medical Sciences, Hamadan, Iran; Brucellosis Research Centers, Hamadan University of Medical Sciences, Hamadan, Iran.
Toxicon. 2020 Aug;183:11-19. doi: 10.1016/j.toxicon.2020.05.004. Epub 2020 May 22.
Staphylococcus aureus is an opportunistic human pathogens, with the ability to produce a series of virulence factors that contribute to the severity of infections. Exfoliative toxins (ETs) are one of the important virulence factors that participating in staphylococcal scalded skin syndrome. Melittin has different biological activities, comprising of antiviral, broad spectrum antibacterial, antiprotozoal, antifungal and anti-inflammatory effects. Twelve clinical isolates of methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) were obtained from wound infection in the burn patients. The MIC plus three sub-inhibitory concentrations (I, II and III) of clindamycin and melittin were tested. Next, the synergistic effects of melittin and clindamycin were evaluated using the broth microdilution checkerboard assay. The detection of exfoliative toxin A and B genes were examined by PCR method. Then the effects of sub-MIC melittin on the expression levels of eta and etb were assessed by quantitative real-time PCR (qRT-PCR) assay. Melittin MIC values against MRSA and MSSA planktonic cells were 0.25-0.5 and 0.25-1 μg/ml, respectively. The clindamycin MIC values against MRSA and MSSA were between 0.5 and 8 μg/ml and 0.5-2 μg/ml, respectively. The results of the time-kill kinetics assay (3.5log and 3log) against MSSA and MRSA planktonic cells were determined within 24 h using melittin. The mean expression of eta in MRSA and MSSA was significantly downregulated to approximately 3.5 and 4 fold, respectively. Moreover, the mean expression of etb in MRSA and MSSA were significantly downregulated to approximately 2.5 and 3 fold, respectively. Hemolytic assay showed that the extracted melittin indicates a strong hemolytic activity (HD = 2 μg/ml). Melittin at 0.5 μg/ml induced cell lysis and stimulated the formation of vesicles in S. aureus strains. Melittin could reduce the expression of eta and etb as encoding exfoliative toxin A and B genes. This component appears to be a good candidate for the treatment of MRSA and MSSA strains. So, melittin in combination with clindamycin can be classified as a complementary treatment of wound infections in burn patients.
金黄色葡萄球菌是一种机会性人类病原体，具有产生一系列毒力因子的能力，这些因子导致感染的严重程度增加。表皮剥脱毒素（ETs）是参与葡萄球菌性烫伤样皮肤综合征的重要毒力因子之一。蜂毒素具有不同的生物学活性，包括抗病毒、广谱抗菌、抗原生动物、抗真菌和抗炎作用。从烧伤患者的伤口感染中获得了 12 株耐甲氧西林金黄色葡萄球菌（MRSA）和甲氧西林敏感金黄色葡萄球菌（MSSA）的临床分离株。测试了克林霉素和蜂毒素的 MIC 加三个亚抑菌浓度（I、II 和 III）。然后，使用肉汤微量稀释棋盘法评估蜂毒素和克林霉素的协同作用。通过 PCR 方法检测表皮剥脱毒素 A 和 B 基因的存在。然后通过实时定量 PCR（qRT-PCR）检测亚 MIC 蜂毒素对 eta 和 etb 表达水平的影响。MRSA 和 MSSA 浮游细胞的蜂毒素 MIC 值分别为 0.25-0.5 和 0.25-1μg/ml。MRSA 和 MSSA 的克林霉素 MIC 值分别在 0.5 至 8μg/ml 和 0.5 至 2μg/ml 之间。在 24 小时内，使用蜂毒素对 MSSA 和 MRSA 浮游细胞进行了时间杀伤动力学（3.5log 和 3log）的测定。MRSA 和 MSSA 的 eta 平均表达水平分别显著下调约 3.5 倍和 4 倍。此外，MRSA 和 MSSA 的 etb 平均表达水平分别显著下调约 2.5 倍和 3 倍。溶血试验表明，提取的蜂毒素显示出很强的溶血活性（HD=2μg/ml）。0.5μg/ml 的蜂毒素诱导了金黄色葡萄球菌菌株的细胞裂解并刺激了小泡的形成。蜂毒素可以降低编码表皮剥脱毒素 A 和 B 基因的 eta 和 etb 的表达。该成分似乎是治疗 MRSA 和 MSSA 菌株的良好候选药物。因此，蜂毒素与克林霉素联合使用可归类为烧伤患者伤口感染的补充治疗。
