Effect of apo-lactoferrin on leukotoxin and outer membrane vesicles of Mannheimia haemolytica A2
Effect of apo-lactoferrin on leukotoxin and outer membrane vesicles of Mannheimia haemolytica A2
复制标题
DOI：
10.1186/s13567-020-00759-z
复制
发表时间：
2020-03-05
影响因子：
4.4
通讯作者：
de la Garza, Mireya
中科院分区：
文献类型：
--
作者：
Avalos-Gomez, Christian;Reyes-Lopez, Magda;de la Garza, Mireya
溶血性曼氏菌血清型A2是绵羊和山羊家畜中肺炎性曼氏菌病的主要原因;该疾病是由应激和相关病毒引起的免疫抑制的结果，并且是造成全球农场生产中重大经济损失的原因。革兰氏阴性菌如M.溶血性弧菌产生称为外膜囊泡（OMV）的外膜（OM）衍生的球形结构，其含有白细胞毒素和其它生物活性毒力因子。在本研究中，M。溶血素A2与牛乳铁蛋白（BLf）的关系。BLf是一种80 kDa的糖蛋白，具有抑菌和杀菌特性，是哺乳动物先天免疫系统的一部分。Apo-BLf（无铁）对M.溶血性弧菌A2，观察到的最小抑制浓度（MIC）为16 μ M。亚致死剂量（2-8 μ M）的apo-BLf增加了OMV的释放，其通过流式细胞术定量。载脂蛋白BLf修改的OM和OMV的正常结构，通过透射电子显微镜观察。Apo-BLf还诱导细菌释放脂多糖（LPS），破坏OM渗透性和功能性，如通过银染色和SDS和多粘菌素B细胞渗透性测定所测量。westernblot结果显示，apo-BLf可促进M.溶血性A2培养上清液，可能通过其铁螯合活性。相比之下，holo-BLf（含铁）没有这种效果，可能是由于这些蛋白质之间三级结构的差异。总之，apo-BLf影响几种M.溶血性病原体毒力因子，并且可以评估在动物中用作治疗绵羊曼氏病的佐剂。
Mannheimia haemolytica serotype A2 is the principal cause of pneumonic mannheimiosis in ovine and caprine livestock; this disease is a consequence of immune suppression caused by stress and associated viruses and is responsible for significant economic losses in farm production worldwide. Gram-negative bacteria such as M. haemolytica produce outer membrane (OM)-derived spherical structures named outer membrane vesicles (OMVs) that contain leukotoxin and other biologically active virulence factors. In the present study, the relationship between M. haemolytica A2 and bovine lactoferrin (BLf) was studied. BLf is an 80 kDa glycoprotein that possesses bacteriostatic and bactericidal properties and is part of the mammalian innate immune system. Apo-BLf (iron-free) showed a bactericidal effect against M. haemolytica A2, with an observed minimal inhibitory concentration (MIC) of 16 mu M. Sublethal doses (2-8 mu M) of apo-BLf increased the release of OMVs, which were quantified by flow cytometry. Apo-BLf modified the normal structure of the OM and OMVs, as observed through transmission electron microscopy. Apo-BLf also induced lipopolysaccharide (LPS) release from bacteria, disrupting OM permeability and functionality, as measured by silver staining and SDS and polymyxin B cell permeability assays. Western blot results showed that apo-BLf increased the secretion of leukotoxin in M. haemolytica A2 culture supernatants, possibly through its iron-chelating activity. In contrast, holo-BLf (with iron) did not have this effect, possibly due to differences in the tertiary structure between these proteins. In summary, apo-BLf affected the levels of several M. haemolytica virulence factors and could be evaluated for use in animals as an adjuvant in the treatment of ovine mannheimiosis.