Abstract
BackgroundJC polyomavirus (JCV) is the causative agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the central nervous system in immunosuppressed patients. PML usually has a poor prognosis. Detection and quantification of the JCV genome in cerebrospinal fluid (CSF) is an efficacious tool for the diagnosis and management of PML, for which proper therapeutic interventions are required.MethodsA loop-mediated isothermal amplification (LAMP) assay was applied for the quantitative detection of JCV. The LAMP assay was evaluated for the efficacy in diagnosis of PML in comparison with the TaqMan-based quantitative real-time PCR (qPCR) assay using 153 CSF specimens collected from patients with suspected PML.ResultsThe LAMP assay showed no cross-reactivity against other polyomavirus plasmids, viral DNA, and viral RNA, which causes encephalitis, and detected 1 copy of the standard DNA per reaction. Among 50 qPCR-positives, 42 specimens (containing JCV genome ranged from 3.2 × 100 to 3.2 × 106 copies/reaction) showed positive reactions and 8 specimens (containing 0.9 to 19.9 copies/reaction) showed negative in the LAMP assay. Furthermore, 3 of 103 qPCR-negative specimens showed positive reactions in the LAMP assay. The sensitivity, specificity, positive predictive value, and negative predictive values of the LAMP assay were 84% (42/50), 97% (100/103), 93% (42/45), and 93% (100/108), respectively. The kappa statistic was 0.83. The JCV loads determined by the LAMP assay showed a strong positive correlation with those determined by the qPCR assay for 33 specimens with copy numbers of ≥1 copies/reaction (r = 0.89). Additionally, the LAMP assay could monitor the JCV genome copy number in CSF for sequential samples equivalently to qPCR assay.ConclusionsThe newly developed LAMP assay is highly specific against JCV and detect the JCV genome in the sample DNA containing 20 or more copies of JCV genome per reaction with 100% sensitivity (n = 29), which corresponds to ≥3 × 103 copies/mL of CSF. The LAMP assay is useful for the diagnosis and offers valuable information for the evaluation and management of PML in the clinical setting.
Highlights
JC polyomavirus (JCV) is the causative agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the central nervous system in immunosuppressed patients
It is assumed that the development of PML is a result of the reactivation of JCV from latency in immunosuppressed patients, including hematopoietic stem cell transplant recipients, those with human immunodeficiency virus (HIV) infection, those with hematologic malignancies, and those treated with immunosuppressive therapy [2, 4]
Plasmids containing the complete genome of other polyomaviruses, including the BK virus (BKV), simian virus 40 (SV40), and murine polyomavirus (MPyV) A2 strain, were purchased from the American Type Culture Collection (Manassas, VA, USA)
Summary
JC polyomavirus (JCV) is the causative agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the central nervous system in immunosuppressed patients. Detection and quantification of the JCV genome in cerebrospinal fluid (CSF) is an efficacious tool for the diagnosis and management of PML, for which proper therapeutic interventions are required. JC polyomavirus (JCV), a non-enveloped DNA virus, belonging to the Polyomaviridae family, is the causative agent of progressive multifocal leukoencephalopathy (PML), a fatal demyelinating disease of the central nervous system [1]. It is assumed that the development of PML is a result of the reactivation of JCV from latency in immunosuppressed patients, including hematopoietic stem cell transplant recipients, those with human immunodeficiency virus (HIV) infection, those with hematologic malignancies, and those treated with immunosuppressive therapy [2, 4]. PML was found in approximately 4% of all HIV-infected patients before the introduction of highly active antiretroviral therapy (HAART) [7]
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