Detection of JC virus DNA by polymerase chain reaction in patients with progressive multifocal leukoencephalopathy

Polymerase chain reaction (PCR) was used in the detection of JC virus (JCV) DNA in formalin-fixed, paraffin-embedded brain tissue sections from 24 patients with progressive multifocal leukoencephalopathy. Brain sections from normal autopsies (n = 17) and other neurologic conditions (n = 4) were used as controls. Specific amplified products were detected in 20 (83%) of 24 patients with progressive multifocal leukoencephalopathy. PCR did not amplify JCV or human beta-globin gene sequences in four patients with characteristic demyelinating lesions of progressive multifocal leukoencephalopathy that were positive for JCV by in situ hybridization or immunocytochemistry. PCR from biopsy sections resulted in more intense amplification signals than PCR from autopsy tissue. Normal brain sections from 17 autopsies were negative by PCR. Low-grade amplification of JCV was observed in one patient with herpes simplex virus encephalitis. PCR served as a more rapid test for confirmation of progressive multifocal leukoencephalopathy than in situ hybridization. However, PCR performance was altered by prolonged formalin fixation of tissue and undetermined inhibitors of the amplification reaction. Laboratories and clinicians should be aware of the difficulties encountered when using paraffin-embedded tissue for PCR.     

Subclinical central nervous system infection with JC virus in patients with AIDS.

Immunocompromised patients, particularly those with AIDS, develop progressive multifocal leukoencephalopathy (PML) due to central nervous system infection with JC virus (JCV). It is unknown whether JCV infection in the central nervous system can occur in the absence of PML symptoms. To address this question, autopsy specimens from patients with AIDS were examined. The brains of a group of patients without AIDS or central nervous system disease were also examined. JCV DNA was detected by the polymerase chain reaction in brain tissue from 4 (31%) of 13 human immunodeficiency virus (HIV)-positive patients. JCV was also detected in 1 elderly HIV-negative patient but not in the 11 other control brains. JCV was not detected in 22 myocardial specimens obtained at autopsy from HIV-negative patients nor 10 peripheral blood specimens from HIV-positive patients. The presence of JCV in brains of patients without clinically evident PML suggests that JCV may be present in the central nervous system without clinical disease.     

Leg weakness in a lung transplant patient

Progressive multifocal leukoencephalopathy (PML) is associated with JC polyomavirus (JCV) infection of central nervous system oligodendrocytes resulting in demyelinization and progressive focal neurologic deficits. Reactivation of dormant JCV occurs in the setting of immunosuppression, most commonly in patients with human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) or hematological malignancies. PML has also been reported in solid organ transplant recipients. We report the case of a 61‐year‐old man after bilateral lung transplantation for chronic hypersensitivity pneumonitis who presented with leg weakness, cognitive decline, and expressive aphasia at 5 months post transplantation. Magnetic resonance imaging and brain biopsy were consistent with PML. Treatment attempt with cytarabine was unsuccessful, and immunomodulation resulted in recurrent grade A3 rejection. The difficulty of managing PML in lung transplant patients is highlighted by the lack of directed therapy and risk of graft rejection or failure with attempts at decreasing immunosuppression.     

Differential expression of mRNAs for JC virus large and small tumor antigens in brain tissues from progressive multifocal leukoencephalopathy patients with and without AIDS.

JC virus (JCV) causes progressive multifocal leukoencephalopathy (PML), the fatal demyelinating infection of oligodendrocytes, in up to 5% of AIDS patients. An intron-differential RNA PCR was developed to study the expression of alternately spliced JCV early mRNAs in brain tissues from PML patients with and without AIDS and in JCV-induced hamster brain tumors. The method utilizes primers that span the large tumor (T) and small tumor (t) antigen introns allowing amplification of specific cDNAs in the presence of contaminating viral genomic DNA. Hybridization with specific junctional probes and DNA sequence analysis confirmed the identity of the PCR products. Sequencing showed that JCV early mRNA is alternatively spliced as previously predicted by analogy to simian virus 40. Large T antigen mRNA was detected in all the brain tissues from PML patients with and without AIDS. The expression of small t antigen mRNA varied depending upon the association of PML with AIDS and upon other unknown factors. Of the 12 PML/AIDS brain tissue samples, 11 (92%) expressed small t antigen mRNA, whereas only 8 of 13 (62%) brain samples from patients with PML alone showed detectable levels of small t antigen mRNA. Human immunodeficiency virus 1 proviral DNA was detected in 10 of 12 PML/AIDS brain samples. The results indicate that alternative splicing of JCV early mRNA is regulated in the human brain and that the production of small t antigen may not be essential for the pathogenesis of PML.     

JC病毒研究进展

JC病毒为小双链DNA病毒,在人群中广泛感染,只有一种血清型,可分为30多个基因型。JC病毒可垂直传播,也可通过呼吸道、消化道传播。严重免疫抑制患者感染JC病毒后可引起进行性多灶性脑白质病(progressive multifocal leukoen-cephalopathy,PML),而CD4+、CD8+T淋巴细胞对感染后是否发病起关键作用。JC病毒对培养细胞和实验动物有很强的致癌潜能,与人类肿瘤存在一定关联性。对感染者的尿液、脑脊液、血液及病变组织进行JC病毒DNA检测和对活组织进行原位杂交及免疫组化检测等为确定JC病毒感染的主要手段,而抗体检测并非确证存在活动性PML的可靠方法。目前没有针对JC病毒有效的抗病毒药物,应用高效抗反转录病毒治疗来获得免疫重建是治疗HIV/AIDS患者感染JC病毒引起PML最好的方法。     

Detection of Toxoplasma gondii, Epstein-Barr virus, and JC virus DNAs in the cerebrospinal fluid in acquired immunodeficiency syndrome patients with focal central nervous system complications.

OBJECT: Toxoplasmic encephalitis (TE), primary central nervous system lymphoma (PCNSL) and progressive multifocal leukoencephalopathy (PML) are major central nervous system (CNS) diseases in patients with acquired immunodeficiency syndrome (AIDS). We assessed the diagnostic value of polymerase chain reaction (PCR) in the detection of DNAs of Toxoplasma gondii (T. gondii), Epstein-Barr virus (EBV) and JC virus (JCV) in the cerebrospinal fluid (CSF). METHODS: We compared the PCR results with those of pathological findings at autopsy. PATIENTS OR MATERIALS: The present study included 23 autopsies representing those in whom CSF samples were obtained before death while the patient was hospitalized or at autopsy. RESULTS: The threshold levels for PCR detection were 4 tachyzoites of T. gondii, 5-15 genomes of EBV and 10 genomes of JCV. We identified T. gondii DNA in 4 out of 5 autopsy-defined cases of TE, EBV DNA in 5 out of 5 cases with PCNSL, and JCV DNA in 2 out of 2 cases with PML. The specificity of PCR was 100% in TE, 78% in PCNSL, and 100% in PML. CONCLUSION: Although the number of cases was relatively small in this study, PCR correctly identified T. gondii DNA in those cases in which PML or PCNSL was the sole clinical diagnosis. Our results indicate that PCR examination of CSF is a clinically useful tool for the diagnosis of focal brain lesions in patients with AIDS.     

JC Virus T-Antigen DNA in Gastrointestinal Mucosa of Immunosuppressed Patients: A Prospective,Controlled Study

Background  

JC virus (JCV), a polyoma virus, is the etiological agent of progressive multifocal leukoencephalopathy in immunosuppressed patients. JCV T-Ag has proven oncogenic potential and is expressed in colonic polyps and carcinomas. We proposed that the prevalence of JCV T-Ag DNA is higher in the normal gastrointestinal (GI) mucosa of immunosuppressed patients compared with their immunocompetent counterparts.     

The effect of potent antiretroviral therapy and JC virus load in cerebrospinal fluid on clinical outcome of patients with AIDS-associated progressive multifocal leukoencephalopathy

A multicenter analysis of 57 consecutive human immunodeficiency virus-positive patients with progressive multifocal leukoencephalopathy (PML) was performed, to identify correlates of longer survival. JC virus (JCV) DNA was quantified in the cerebrospinal fluid (CSF) by polymerase chain reaction. Two months after therapy, 4% of the patients without highly active antiretroviral therapy (HAART) and 26% with HAART showed neurologic improvement or stability (P=.03), and 8% and 57%, respectively, reached undetectable JCV DNA levels in the CSF (P=.04). One-year probability of survival was.04 without HAART and.46 with HAART. HAART and lack of neurologic progression 2 months after diagnosis were independently associated with longer survival. Among HAART-treated patients, a baseline JCV DNA <4.7 log, and reaching undetectable levels after therapy predicted longer survival. Survival of AIDS-related PML is improved by HAART when JCV replication is controlled.     

Therapeutic Experience of Progressive Multifocal Leukoencephalopathy Development during Ofatumumab Therapy for Chronic Lymphocytic Leukemia

A 79-year-old man experienced cognitive impairment and visual field defects during ofatumumab therapy for chronic lymphocytic leukemia refractory to combination chemotherapy. Magnetic resonance imaging revealed T1-weighted low-intensity and T2-weighted high-intensity lesions with patchy gadolinium enhancement in the subcortical white matter. A diagnosis of progressive multifocal leukoencephalopathy was made after the detection of John Cunningham virus (JCV) DNA in his cerebrospinal fluid (CSF). Following plasma exchange and the administration of mirtazapine and mefloquine, the JCV DNA levels in the CSF decreased. However, the patient died 55 days after treatment was initiated. Ofatumumab treatment appears to be associated with the development of progressive multifocal leukoencephalopathy.     

JC virus meningitis in a patient with systemic lupus erythematosus

The human neurotropic JC virus (JCV) is most commonly acquired during childhood, and, because no clinical illness has been associated with primary infection, is presumed to be asymptomatic. In the immunocompromised host, JCV is responsible for progressive multifocal leukoencephalopathy (PML). We describe a patient with longstanding systemic lupus erythematosus who presented with acute meningitis without encephalitis or PML. JCV was the only pathogen found in the cerebrospinal fluid suggesting a primary infection or symptomatic reactivation. Our observation demonstrates the expanding clinical importance of JCV in autoimmune diseases, and diagnostic tests for JCV should be included in the investigative work-up for meningitis or encephalitis in these patients.     

JC virus genotypes in France: molecular epidemiology and potential significance for progressive multifocal leukoencephalopathy

JC virus (JCV) induces progressive multifocal leukoencephalopathy (PML), especially in human immunodeficiency virus (HIV)-infected patients. Although JCV genotypes have primarily been associated with geographic patterns, a distinctive neuropathogenicity was recently attributed to genotype 2. A multicenter study was conducted to describe the distribution of JCV genotypes in France and to investigate correlations between genotypes and PML. Genotypes were determined by sequencing 494 bp in the VP1 capsid gene. Peripheral JCV was studied in 65 urine samples from 43 HIV-infected patients and from 22 control subjects. Genotypes 1, 4, 2, and 3 were detected in 52.3%, 30.8%, 12.3%, and 4.6% of the samples, respectively. In 56 brain or cerebrospinal fluid samples, PML-associated JCV of genotypes 1, 2, 4, and 3 was found in 66%, 19.7%, 8.9%, and 5.4%, respectively. Infection with JCV genotypes 1 or 2 was correlated with PML (odds ratio, 3.29). On the other hand, infection with JCV genotype 4 could represent a lower risk for PML.     

Alpha Interferon in AIDS-Related Progressive Multifocal Leukoencephalopathy

Objective: To determine the efficacy of recombinant interferon alpha in the treatment of progressive multifocal leukoencephalopathy associated with the acquired immunodeficiency syndrome (AIDS). Design: Open label, uncontrolled study. Setting: Neurological unit and clinical AIDS program, Boston City Hospital, Boston, MA. Patients: Four consecutive AIDS patients with pathologically confirmed progressive multifocal leukoencephalopathy. Intervention: Each patient received alpha interferon for 4-12 weeks in a dose of 5-10 million units daily, administered subcutaneously. In addition, two of the four were taking acyclovir 2400 mg/day orally over the same period. Results: None of the patients showed any clinical response to the therapy; the mean survival was 14 weeks. No adverse effects of the treatment were encountered. Conclusions: Despite anecdotal evidence that alpha interferon is effective in the treatment of progressive multifocal leukoencephalo pathy in non-AIDS patients, the experience of these patients suggests that the drug is of no benefit in AIDS-related PML.     

Multifocal progressive leukoencephalopathy occurring after refractory anemia and multiple infectious disorders consecutive to severe lymphopenia

Progressive multifocal leukoencephalopathy (PML) is related to central nervous system infection with JC virus (JCV). This leukoencephalopathy occurs in immunocompromised patients such as those with acquired immunodeficiency syndrome (AIDS) or lymphoid malignancies. We describe here a patient with myelodysplastic syndrome who developed several life-threatening infections including listeriosis, tuberculosis, and PML. Listeriosis and recurrence of tuberculosis preceded the occurrence of PML. Neurologic features associated with major ataxia, speech disorders, and PML were documented by cranial magnetic resonance imaging showing typical features in the cerebellum and proven by polymerase chain reaction (PCR) detection of JCV DNA in the cerebrospinal fluid. No specific treatment was decided because of progression toward acute myeloid leukemia. In this case, PML occurred with no susceptibility and without immunosuppressive treatment. Our case adds further support to the association between the impairment of T-cell immune responses and myelodysplastic disorders.     

Critical role of JC virus-specific CD4 T-cell responses in preventing progressive multifocal leukoencephalopathy

BACKGROUND: JC virus (JCV) is ubiquitous among the general population. However, only individuals with severely impaired immunity, mainly AIDS patients, develop progressive multifocal leukoencephalopathy (PML). Here, we examined the role of specific CD4 T cells in the control of JCV infection. METHODS AND DESIGN: JCV-specific CD4 T-cell responses were investigated by assaying peripheral blood mononuclear cell proliferation in response to the purified virus. Four groups of individuals without PML were examined: 14 HIV-seronegative healthy donors and 25 HIV-infected patients without PML, separated into urinary JCV excretors (active infection) and non-excretors, according to JCV PCR on urine. Two groups of patients with PML were also studied: 14 HIV-infected patients with active PML; and 10 PML survivors on effective and prolonged antiretroviral therapy. All of the patients were PCR-positive for JCV in the cerebrospinal fluid at the time of diagnosis of PML. RESULTS: No significant anti-JCV CD4 T-cell proliferation was found in any of the non-excretors tested. All nine healthy donors and seven of the 13 non-PML HIV-infected patients with urinary JCV excretion had positive JCV-specific CD4 T-cell responses. No significant response was found in the 14 patients with active PML, while nine of the 10 PML survivors had positive responses. Restoration of JCV-specific CD4 T-cell responses was associated with JCV clearance from the cerebrospinal fluid. CONCLUSION: JCV-specific CD4 T-cell responses appear to play a critical role in the control of JCV infection, preventing PML development. Such responses can be restored in PML survivors following effective and prolonged antiretroviral therapy.     

Progressive multifocal leukoencephalopathy in a patient with X-linked agammaglobulinemia

To our knowledge, this is the first case report describing progressive multifocal leukoencephalopathy (PML) associated with X-linked agammaglobulinemia. The JC virus was confirmed at autopsy and PML was diagnosed. Humoral immunodeficiency with normal cellular immunity could be infected with JCV.     

Progressive multifocal leucoencephalopathy in a patient with sarcoidosis--successful treatment with cidofovir and mirtazapine

SIR, Progressive multifocal leukoencephalopathy (PML) representsa demyelinating CNS disease caused by reactivation of JC virus(JCV) in immunocompromised patients including patients post-transplant,with haematological malignancies or AIDS. Recently, the occurrenceof PML was reported in patients with autoimmune diseases onimmunosuppressive treatment [1]. Despite the cessation of immunosuppressionwhen possible, the prognosis of PML is usually poor. A common consensus regarding the therapy of PML has not yetbeen reached. Cidofovir, a nucleotide analogue with in vitroanti-JCV activity represents the most effective single agentin a mouse polyomavirus model [     

Polyomavirus JCV excretion and genotype analysis in HIV-infected patients receiving highly active antiretroviral therapy

OBJECTIVE: To assess the frequency of shedding of polyomavirus JC virus (JCV) genotypes in urine of HIV-infected patients receiving highly active antiretroviral therapy (HAART). METHODS: Single samples of urine and blood were collected prospectively from 70 adult HIV-infected patients and 68 uninfected volunteers. Inclusion criteria for HIV-infected patients included an HIV RNA viral load < 1000 copies, CD4 cell count of 200-700 x 106 cells/l, and stable HAART regimen. PCR assays and sequence analysis were carried out using JCV-specific primers against different regions of the virus genome. RESULTS: JCV excretion in urine was more common in HIV-positive patients but not significantly different from that of the HIV-negative group [22/70 (31%) versus 13/68 (19%); P = 0.09]. HIV-positive patients lost the age-related pattern of JCV shedding (P = 0.13) displayed by uninfected subjects (P = 0.01). Among HIV-infected patients significant differences in JCV shedding were related to CD4 cell counts (P = 0.03). Sequence analysis of the JCV regulatory region from both HIV-infected patients and uninfected volunteers revealed all to be JCV archetypal strains. JCV genotypes 1 (36%) and 4 (36%) were the most common among HIV-infected patients, whereas type 2 (77%) was the most frequently detected among HIV-uninfected volunteers. CONCLUSION: These results suggest that JCV shedding is enhanced by modest depressions in immune function during HIV infection. JCV shedding occurred in younger HIV-positive persons than in the healthy controls. As the common types of JCV excreted varied among ethnic groups, JCV genotypes associated with progressive multifocal leukoencephalopathy may reflect demographics of those infected patient populations.     

Quantification of JC virus DNA in the cerebrospinal fluid of patients with human immunodeficiency virus-associated progressive multifocal leukoencephalopathy--a longitudinal study.

In progressive multifocal leukoencephalopathy (PML) the JC virus (JCV) load in the cerebrospinal fluid (CSF) is discussed as a parameter for disease progression. To investigate the evolution of viral shedding into the CSF, the JCV DNA concentration was quantified by competitive polymerase chain reaction (PCR) in multiple CSF samples from prior to and during an unsuccessful intrathecal salvage therapy in 2 human immunodeficiency virus-infected patients with biopsy-proven PML. With continuous clinical progression the virus load varied considerably intra- and interindividually, ranging from nondetectable to 1.2x108 genome equivalents/10 microliter CSF. Whereas an overall increase during progressive disease was confirmed, the virus burden was either constant or fluctuated irregularly during the intermediate stage of disease. This shows a variability of viral shedding during active disease that must be taken into account when the JCV load is measured by quantitative PCR for both the diagnosis of PML and monitoring under investigational treatment.