School of Biological Sciences, University of the Punjab, Lahore, Pakistan.
Armed Forces Institute of Pathology, Rawalpindi, Pakistan.
PLoS One. 2021 Nov 12;16(11):e0258389. doi: 10.1371/journal.pone.0258389. eCollection 2021.
Serodiagnosis of tuberculosis (TB) can be rapid, reliable and cost-effective if the issue of variable antibody responses of TB patients against different Mycobacterium tuberculosis (Mtb) antigens can be overcome by developing fusion proteins containing epitopes from multiple antigens of Mtb. In this study, Mtb antigens Rv1793, Rv2628, Rv2608 and a truncated variant produced by removing non-epitopic region from N-terminal of Rv2608 (tnRv2608), and the fusion protein Rv1793-Rv2628-tnRv2608 (TriFu64), were expressed in E. coli and purified. Plasma samples from TB patients characterized by sex, age and sputum/culture positivity, were used to compare the sensitivity of the single antigens with the fusion protein. Sensitivity of Rv1793, Rv2628 and Rv2608, was 27.8%, 39% and 36.3%, respectively. Truncation of Rv2608 increased sensitivity by approximately 35% in confirmed TB cases. Sensitivity of the fusion construct, TriFu64 increased to 66% with a specificity of 100%. Importantly, tnRv2608 was better able to detect sputum and culture negative patients, and this carried through to the fusion protein. We demonstrate that fusion of Mtb proteins ensures broad sensitivity across disease types, sex and age groups in a Pakistani population.
如果能够克服结核分枝杆菌（Mycobacterium tuberculosis，Mtb）患者针对不同 Mtb 抗原的抗体反应的变异性问题，通过开发包含 Mtb 多种抗原表位的融合蛋白来克服这一问题，那么结核分枝杆菌（TB）的血清学诊断可以快速、可靠且具有成本效益。在这项研究中，我们在大肠杆菌中表达并纯化了 Mtb 抗原 Rv1793、Rv2628、Rv2608 以及从 Rv2608 的 N 端去除非表位区域后产生的截断变体（tnRv2608），以及融合蛋白 Rv1793-Rv2628-tnRv2608（TriFu64）。我们使用了来自不同性别、年龄和痰/培养阳性的 TB 患者的血浆样本，以比较单一抗原与融合蛋白的敏感性。Rv1793、Rv2628 和 Rv2608 的敏感性分别为 27.8%、39%和 36.3%。Rv2608 的截断增加了确诊 TB 病例的敏感性，约 35%。融合构建体 TriFu64 的敏感性增加到 66%，特异性为 100%。重要的是，tnRv2608 能够更好地检测痰和培养阴性患者，这种情况在融合蛋白中也得到了体现。我们证明了 Mtb 蛋白的融合确保了在巴基斯坦人群中针对不同疾病类型、性别和年龄组具有广泛的敏感性。
