Department of Bacteriology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
Biosun Pharmed Factory, Barkat Pharmaceutical group, Tehran, Iran.
Microb Pathog. 2020 Oct;147:104246. doi: 10.1016/j.micpath.2020.104246. Epub 2020 Jun 17.
Anti-adhesion therapy and anti-adhesin immunity are meant to diminish the interaction between pathogens and host tissues, either by prevention or by exclusion of bacterial adhesion and entrance to cells. Azurin is a scaffold protein possessing antiviral, antiparasitic, and anticancer activities. The purpose of the present study was to determine the effect of recombinant Azurin (rAzurin) on the adhesion and invasion capacity of invasive (Shigella sonnei, Shigella flexneri, Campylobacter jejuni) and non-invasive (Vibrio cholerae) enteric bacteria to cells. The non-toxic dose of rAzurin and the best MOI (Multiplicity of Infection) of bacterial species was assessed by MTT assay. Bacterial species were used at MOIs of 20:1 and Azurin was applied at the concentrations of 5 and 25 μg/mL and added to Caco-2 cells in competition and replacement assay to assess the anti-adhesion and anti-invasion properties of rAzurin. The protein caused significant decrease in the adhesion rate of S. sonnei, S. flexneri, C. jejuni, and V. cholerae strains to Caco-2 cells by 43, 39, 72, and 38% in competition and 45, 46, 75, and 48% in replacement assays, respectively. Also, S. sonnei, S. flexneri, and C. jejuni strains invasion rate was reduced to 50, 50, and 70% in anti-invasion assay, respectively. The inhibitory effect of Azurin against C. jejuni and V. cholerae strains adhesion was more significant (p < .001) compared to Shigella spp. (p < .05) which may be due to smaller size of the former bacteria. On the contrary, in invasion assay, rAzurin showed a greater inhibitory effect against Shigella spp. (p < .001) compared to C. jejuni (p < .05), which may probably be due to the interaction of rAzurin with several effectors or ligands, involved in Shigella invasion and internalization. The findings of the present study opens new insights of rAzurin as a new and potent candidate for reducing or probably preventing enteric bacterial attachment, invasion, and pathogenesis.
抗黏附治疗和抗黏附素免疫旨在通过预防或排除细菌黏附和进入细胞来减少病原体与宿主组织之间的相互作用。铜绿假单胞菌蓝蛋白是一种支架蛋白，具有抗病毒、抗寄生虫和抗癌活性。本研究的目的是确定重组铜绿假单胞菌蓝蛋白（rAzurin）对侵袭性（福氏志贺菌、宋内志贺菌、空肠弯曲菌）和非侵袭性（霍乱弧菌）肠杆菌黏附细胞和入侵细胞的黏附及入侵能力的影响。通过 MTT 测定法评估 rAzurin 的无毒剂量和最佳细菌物种 MOI（感染复数）。将细菌物种以 MOI 20:1 的浓度使用，并将 rAzurin 以 5 和 25 μg/mL 的浓度添加到 Caco-2 细胞中，以竞争和替代测定法评估 rAzurin 的抗黏附和抗入侵特性。该蛋白显著降低了 S. sonnei、S. flexneri、C. jejuni 和 V. cholerae 菌株与 Caco-2 细胞的黏附率，竞争测定法分别为 43%、39%、72%和 38%，替代测定法分别为 45%、46%、75%和 48%。此外，S. sonnei、S. flexneri 和 C. jejuni 菌株的侵袭率分别降低到 50%、50%和 70%。与志贺菌属（p<0.05）相比，rAzurin 对空肠弯曲菌和霍乱弧菌的黏附抑制作用更为显著（p<0.001），这可能是由于前者细菌的体积较小。相反，在侵袭测定中，rAzurin 对志贺菌属的抑制作用大于空肠弯曲菌（p<0.001），这可能是由于 rAzurin 与参与志贺菌侵袭和内化的几种效应物或配体相互作用。本研究的结果为 rAzurin 作为减少或可能预防肠杆菌黏附、侵袭和发病机制的新型有效候选物提供了新的见解。
