1ICMR-National Institute of Pathology (NIOP), Safdarjung Hospital Campus, New Delhi, India.
2Faculty of Health and Biological Sciences, Symbiosis International (Deemed University), Pune, India.
Am J Trop Med Hyg. 2021 Apr 19;104(6):2097-2107. doi: 10.4269/ajtmh.20-1057.
Despite the dwindling number of visceral leishmaniasis (VL) cases in India, there is an urgent need for early and unequivocal diagnostics for controlling and preventing the reemergence of VL. Post-kala-azar dermal leishmaniasis (PKDL), a dermal sequela of VL, serves as a reservoir of the parasite. Diagnosis of PKDL, especially the macular variant, is challenging and poses impediment toward attainment of VL elimination. In this study, a real-time fluorimetry loop-mediated isothermal amplification (RealAmp) assay has been established for the detection of different clinical manifestations of leishmaniasis. The study included 150 leishmaniasis patients (25 VL, 25 cutaneous leishmaniasis [CL], and 100-PKDL) along with 120 controls. The assay demonstrated sensitivity of 100% (95% CI: 86.68-100) for diagnosis of VL and PKDL (95% CI: 79.61-100) and 96% (95% CI: 86.68-100) for CL with 100% specificity. Moreover, considering the cardinal role of PKDL, diagnosis using minimally invasive slit aspirate was explored, which demonstrated remarkable sensitivity of 96% (95% CI: 87.64-98.47). As a test of cure for PKDL, RealAmp successfully detected parasite in two of posttreatment cases who later reported relapse on follow-up. Also, direct sample lysis using slit aspirate was attempted in a small group that yielded sensitivity of 89% (95% CI: 67.20-96.90). RealAmp depicted excellent diagnostic accuracy in the diagnosis of leishmaniasis in concordance with the established SYBR Green I-based (Molecular Probes, Eugene, OR) visual loop-mediated isothermal amplification (LAMP) and the reference comparator real-time PCR. The study endorsed the employment of LAMP either as visual-LAMP or RealAmp for an accurate and expeditious diagnosis of PKDL and as a tool for assessment of cure.
尽管印度内脏利什曼病(VL)病例数量正在减少,但仍迫切需要进行早期、明确的诊断,以控制和预防 VL 的再次出现。Post-kala-azar dermal leishmaniasis(PKDL)是 VL 的皮肤后遗症,是寄生虫的储存库。PKDL 的诊断,特别是黄斑型,具有挑战性,并且是实现 VL 消除的障碍。在这项研究中,建立了一种实时荧光环介导等温扩增(RealAmp)检测方法,用于检测不同临床表现的利什曼病。该研究包括 150 名利什曼病患者(25 例 VL、25 例皮肤利什曼病[CL]和 100 例 PKDL)和 120 名对照。该检测方法对 VL 和 PKDL(95%CI:79.61-100)的诊断灵敏度为 100%(95%CI:86.68-100),对 CL 的灵敏度为 96%(95%CI:86.68-100),特异性为 100%。此外,考虑到 PKDL 的重要作用,探索了使用微创性皮肤划痕抽吸进行诊断,该方法的灵敏度为 96%(95%CI:87.64-98.47)。作为 PKDL 的治疗后检测,RealAmp 成功检测到两名经治疗后复发的病例中的寄生虫。此外,在一小部分病例中尝试了直接用皮肤划痕抽吸进行样本裂解,灵敏度为 89%(95%CI:67.20-96.90)。RealAmp 在利什曼病的诊断中具有出色的诊断准确性,与已建立的基于 SYBR Green I(Molecular Probes,Eugene,OR)的视觉环介导等温扩增(LAMP)和参考比较实时 PCR 相符。该研究支持使用 LAMP 进行视觉-LAMP 或 RealAmp 检测,以实现 PKDL 的准确和快速诊断,并作为评估治疗效果的工具。