Comparison of PCR-southern hybridization and quantitative real-time PCR for the detection of JC and BK viral nucleotide sequences in urine and cerebrospinal fluid

Ryschkewitsch, Caroline F.; Jensen, Philip J.; Hou, Jean; Fahle, Gary A.; Fischer, Steven H.; Major, Eugene O.

doi:10.1016/j.jviromet.2004.06.021

Cited by 115 publications

(100 citation statements)

References 12 publications

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“…We used quantitative JCV PCR (qPCR) to detect and quantify JCV DNA in all samples as previously described (22). The limit of detection of the assay was 10 copies of JCV/g of cellular DNA.…”

Section: Methodsmentioning

confidence: 99%

JC Virus Latency in the Brain and Extraneural Organs of Patients with and without Progressive Multifocal Leukoencephalopathy

Tan

Ellis

Wüthrich

et al. 2010

J Virol

106

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“…We used quantitative JCV PCR (qPCR) to detect and quantify JCV DNA in all samples as previously described (22). The limit of detection of the assay was 10 copies of JCV/g of cellular DNA.…”

Section: Methodsmentioning

confidence: 99%

JC Virus Latency in the Brain and Extraneural Organs of Patients with and without Progressive Multifocal Leukoencephalopathy

Tan

Ellis

Wüthrich

et al. 2010

J Virol

106

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“…All CSF samples were sent to the Mayo Clinic (Rochester, MN, USA) for JCV PCR-Southern hybridization testing. The sensitivity of this JCV PCR assay was determined to be 1-10 genome equivalents per reaction [25-250 copies (ml sample) 21 ] (in-house Mayo Clinic procedure manual; Ryschkewitsch et al, 2004). All other tests were performed at our clinical laboratory.…”

Section: Methodsmentioning

confidence: 99%

Effective use of JC virus PCR for diagnosis of progressive multifocal leukoencephalopathy

Wang

Qian

2009

Journal of Medical Microbiology

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In a retrospective review of data from 168 patients with suspected progressive multifocal leukoencephalopathy (PML) between 1996 and 2006, JC virus (JCV) PCR on cerebrospinal fluid (CSF) samples was positive only in human immunodeficiency virus (HIV)-infected patients with low CD4 cell counts and in severely immunocompromised patients with radiographic lesions consistent with PML or infectious processes generally. Of note, one HIV patient with a very low CD4 cell count had a positive JCV PCR despite a normal magnetic resonance imaging exam. We concluded that JCV PCR testing on CSF specimens should therefore be targeted to these high-risk patients.

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“…2A; Table 1) (20). Viral DNA was isolated as previously described (21,22). The NCCR was amplified by PCR directly from the DNA isolate and sequenced by the Division of Intramural Research Sequencing Facility, NINDS.…”

Section: -Fold In Cd34mentioning

confidence: 99%

Lymphocyte Gene Expression and JC Virus Noncoding Control Region Sequences Are Linked with the Risk of Progressive Multifocal Leukoencephalopathy

Marshall

Ferenczy

Daley

et al. 2014

J Virol

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Progressive multifocal leukoencephalopathy (PML)-derived noncoding control region (NCCR) sequences permitted greater early viral gene expression than kidney-associated NCCR sequences. This was driven in part by binding of the transcription factor Spi-B to unique PML-associated Spi-B binding sites. Spi-B is upregulated in developing B cells in response to natalizumab therapy, a known risk factor for PML. Naturally occurring JCV sequence variation, together with drug treatment-induced cellular changes, may synergize to create an environment leading to an increased risk of PML. The incidence of progressive multifocal leukoencephalopathy (PML) has risen dramatically in recent years because of the AIDS pandemic and the increased use of immunomodulatory therapies (1). In particular, natalizumab (Tysabri; Biogen Idec), a very effective multiple sclerosis (MS) treatment, has been highly associated with PML (2).Natalizumab treatment alters the expression profiles of peripheral blood mononuclear cells (PBMCs), including the expression of the transcription factor Spi-B (3). Spi-B is required for normal B-cell receptor signaling and maturation (4-7). Spi-B levels are higher in cells permissive to JC virus (JCV) transcription and replication, and overexpression of Spi-B increases viral gene expression (8). Spi-B binds to target sites in the JCV noncoding control region (NCCR) isolated from PML brain tissue but not in the archetype NCCR commonly detected in the urine of asymptomatic healthy individuals (8-11). Importantly, mutation of these sites in PML-associated NCCRs decreases Spi-B protein binding and viral gene expression (8,12). Taken together, these results suggest that Spi-B binding to sequences in the JCV NCCR have functional effects on viral gene expression and may play a role in the activation of JCV in peripheral blood cells (13).JCV DNA has been detected in CD19 ϩ B cells and CD34 ϩ hematopoietic progenitor cells (14, 15) isolated from peripheral blood and bone marrow. These CD34 ϩ cells are strikingly increased in the peripheral blood of natalizumab-treated patients (16, 17). Immunomagnetic separation was used to isolate CD3 ϩ , CD19 ϩ , and CD34 ϩ cells from PBMCs of normal donors and MS patients treated with natalizumab. Total RNA was isolated from each cell subset, and Spi-B gene expression was measured by quantitative reverse transcription (qRT)-PCR using the endogenous control PUM1 for normalization. Spi-B gene expression was more variable in natalizumab patients. It was upregulated up to 2-fold in CD19 ϩ B cells in some patients treated with natalizumab (Fig. 1C). Spi-B expression was increased ϩ , CD19 ϩ , and CD34 ϩ cells were isolated by immunomagnetic separation from PBMC samples from patients treated with natalizumab or normal donors. The remaining cells from the separation are labeled the negative fraction. Total RNA was isolated from each fraction of cells, and the Spi-B gene expression level was measured by qRT-PCR. Spi-B mRNA levels were normalized between samples by using the endogenous control P...

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Comparison of PCR-southern hybridization and quantitative real-time PCR for the detection of JC and BK viral nucleotide sequences in urine and cerebrospinal fluid

Cited by 115 publications

References 12 publications

JC Virus Latency in the Brain and Extraneural Organs of Patients with and without Progressive Multifocal Leukoencephalopathy

JC Virus Latency in the Brain and Extraneural Organs of Patients with and without Progressive Multifocal Leukoencephalopathy

Effective use of JC virus PCR for diagnosis of progressive multifocal leukoencephalopathy

Lymphocyte Gene Expression and JC Virus Noncoding Control Region Sequences Are Linked with the Risk of Progressive Multifocal Leukoencephalopathy

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