Lima Mayara Ingrid Sousa, Corrêa Meydson Benjamim Carvalho, Moraes Emilly Caroline Dos Santos, Oliveira Jaqueline das Dores Dias, de Souza Santos Paula, de Souza Aline Gomes, Goulart Isabela Maria Bernardes, Goulart Luiz Ricardo
Laboratory of Genetics and Molecular Biology, Department of Biology, Federal University of Maranhão, São Luís, MA, Brazil.
Postgraduate Program on Health and Environment and Postgraduate Program on Health Sciences, Federal University of Maranhão, São Luís, MA, Brazil.
AMB Express. 2023 Oct 27;13(1):120. doi: 10.1186/s13568-023-01625-9.
The early diagnosis of leprosy serves as an important tool to reduce the incidence of this disease in the world. Phage display (PD) technology can be used for mapping new antigens to the development of immunodiagnostic platforms. Our objective was to identify peptides that mimic Mycobacterium leprae proteins as serological markers using phage display technology. The phages were obtained in the biopanning using negative and positive serum from household contacts and leprosy patients, respectively. Then, the peptides were synthesized and validated in silico and in vitro for detection of IgG from patients and contacts. To characterize the native protein of M. leprae, scFv antibodies were selected against the synthetic peptides by PD. The scFv binding protein was obtained by immunocapture and confirmed using mass spectrometry. We selected two phase-fused peptides, MPML12 and MPML14, which mimic the HSP60 protein from M. leprae. The peptides MPML12 and MPML14 obtained 100% and 92.85% positivity in lepromatous patients. MPML12 and MPM14 detect IgG, especially in the multibacillary forms. The MPML12 and MPML14 peptides had positivity of 11.1% and 16.6% in household contacts, respectively. There was no cross-reaction in patient's samples with visceral leishmaniasis, tuberculosis and other mycobacteriosis for both peptides. Given these results and the easy obtainment of mimetic antigens, our peptides are promising markers for application in the diagnosis of leprosy, especially in endemic and hyperendemic regions.
麻风病的早期诊断是降低全球该疾病发病率的重要手段。噬菌体展示（PD）技术可用于绘制新抗原，以开发免疫诊断平台。我们的目标是利用噬菌体展示技术鉴定模拟麻风分枝杆菌蛋白的肽段作为血清学标志物。分别使用来自家庭接触者和麻风病患者的阴性和阳性血清进行生物淘选，从而获得噬菌体。然后，合成肽段并在计算机模拟和体外实验中进行验证，以检测患者和接触者的IgG。为了表征麻风分枝杆菌的天然蛋白，通过噬菌体展示针对合成肽段选择单链抗体（scFv）。通过免疫捕获获得scFv结合蛋白，并使用质谱法进行确认。我们选择了两个模拟麻风分枝杆菌HSP60蛋白的融合肽段MPML12和MPML14。MPML12和MPML14在瘤型麻风患者中的阳性率分别为100%和92.85%。MPML12和MPM14可检测IgG，尤其是在多菌型麻风病中。MPML12和MPML14肽段在家庭接触者中的阳性率分别为11.1%和16.6%。这两种肽段在患者样本中与内脏利什曼病、结核病和其他分枝杆菌病均无交叉反应。鉴于这些结果以及模拟抗原易于获得，我们的肽段有望成为麻风病诊断的标志物，尤其是在地方病和高度流行地区。
