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The human JC polyomavirus (JCV) is a widespread, neurotropic, opportunistic pathogen responsible for progressive multifocal leukoencephalopathy (PML) as well as other diseases in immunosuppressed individuals, including granule cell neuronopathy, JCV-associated nephropathy, encephalitis, and meningitis in rare cases. JCV classification is still unclear, where the ICTV (International Committee on Taxonomy of Viruses) has grouped all the strains into human polyomavirus 2, with no classification on clade and subclade levels. Therefore, JCV strains were previously classified using different genomic regions, e.g., full-length, VP1, and the V-T intergenic region etc., and the strains were grouped into several types related to various geographic locations and human ethnicities. However, neither of these classifications and nomenclature contemplates all the groups described so far. Herein, we evaluated all the available full-length coding genomes, VP1, and large T antigen nucleotide sequences of JCV reported during 1993-2023 and classified them into four major phylogenetic clades, i.e., GI-GIV, where GI is further grouped into two types GI.1 and GI.2 with five sub-clades each (GI.1/GI.2 a-e), GII into three (GII a-c), GIII as a separate clade, and GIV into seven sub-clades (GIV a-g). Similarly, the phylogeographic network analysis indicated four major clusters corresponding to GI-GIV clades, each with multiple subclusters and mutational sub-branches corresponding to the subclades. GI and GIV clusters are connected via GI.1-e reported from Europe and America, GII, GIII and GIV clusters are connected by GII-b and GII-c strains reported from Africa, while GIV cluster strains are connected to the Russia-Italy JCV haplotype. Furthermore, we identified JCV-variant-GS/B-Germany-1997 (GenBank ID: AF004350.1) as an inter-genotype recombinant having major and minor parents in the GI.1-e and GII-a clades, respectively. Additionally, the amino acid variability analysis revealed high entropy across all proteins. The large T antigen exhibited the highest variability, while the small t antigen showed the lowest variability. Our phylogenetic and phylogeographic analyses provide a new approach to genotyping and sub-genotyping and present a comprehensive classification system of JCV strains based on their genetic characteristics and geographic distribution, while the genetic recombination and amino acid variability can help identify pathogenicity and develop effective preventive and control measures against JCV infections., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Progressive multifocal leukoencephalopathy (PML) caused by the JC virus is the main limitation to the use of disease modifying therapies for treatment of multiple sclerosis (MS)., Methods: To assess the PML risk in course of ocrelizumab, urine and blood samples were collected from 42 MS patients at baseline (T0), at 6 (T2) and 12 months (T4) from the beginning of therapy. After JCPyV-DNA extraction, a quantitative-PCR (Q-PCR) was performed. Moreover, assessment of JCV-serostatus was obtained and arrangements' analysis of non-coding control region (NCCR) and of viral capsid protein 1 (VP1) was carried out., Results: Q-PCR revealed JCPyV-DNA in urine at all selected time points, while JCPyV-DNA was detected in plasma at T4. From T0 to T4, JC viral load in urine was detected, increased in two logarithms and, significantly higher, compared to viremia. NCCR from urine was archetypal. Plasmatic NCCR displayed deletion, duplication, and point mutations. VP1 showed the S269F substitution involving the receptor-binding region. Anti-JCV index and IgM titer were found to statistically decrease during ocrelizumab treatment., Conclusions: Ocrelizumab in JCPyV-DNA positive patients is safe and did not determine PML cases. Combined monitoring of ocrelizumab's effects on JCPyV pathogenicity and on host immunity might offer a complete insight towards predicting PML risk.

JC virus (JCV) , and BK virus (BKV) can remain latency in kidney and excrete via urine asymptomatically. JCV has been associated with colorectal and bladder cancers. BKV has been linked with lung, pancreas, liver, urogenital tract, head and neck cancers. Therefore, the frequency of JCV DNA and BKV DNA are essential to evaluate in urine samples of healthy individuals., Materials and Methods: Hundred sixty four urine samples were collected from healthy subjects [96 females and 68 males]. DNA was extracted and detection of JCV DNA and BKV DNA was carried out by PCR . The analysis of sequencing and construction of phylogenetic tree were performed for the samples positive for JCV DNA and BKV DNA., Results: Ten (6.09%) urine samples [5/96(5.2%) females and 5/68( 8.82) males] were tested positive for JCV DNA (P= 0.814). The results of sequencing and phylogenetic tree showed the isolated JCV DNA were cluster with 3A genotype. 21/164 (12.8%) samples were tested positive for BKV DNA [11/96(11.45%) females and males 10/68(14.7%)] ( P= 0.63). The results of sequencing and phylogenetic tree showed that the isolated BKV was cluster with genotype III., Conclusion: In the present study 6.09% and 12.8% of the healthy individuals showed positive for JCV DNA (genotype 3A) and BKV DNA(genotype III) respectively. With regard to life threating diseases by BKV and JCV in immunocomprsied patients , the screening BKV DNA and JCV DNA should be implemented for patients with cancer /autoimmune diseases /organ recipient/ multiple sclerosis (MS), prior to immunosuppression therapy or immunomodulatory agents treatment., .

Background: JC polyomavirus (JCPyV) establishes a stable and successful interaction with the host, causing progressive multifocal leukoencephalopathy (PML) in immunocompromised subjects. Recently, it has been reported that JCPyV, like other viruses, may exploit extracellular vesicles (EV) in cell cultures., Objective: To investigate the presence of JCPyV-DNA in EV circulating in human plasma obtained from patients at risk for PML., Study Design: JCPyV-DNA status was studied in EV obtained from 170 plasma samples collected from 120 HIV positive patients and 50 healthy donors. EV were extracted from plasma and characterized by Nanoparticle tracking analysis, by western blot for presence of tetraspanin CD63, CD81, annexin II, cythocrome C protein and, finally, by immunoelectron microscopy (IEM). Presence and quantitation of JCPyV-DNA were assessed with Multiplex real-time TaqMan PCR assay., Results: The JCPyV-DNA plasma prevalence in 120 HIV positive patients and 50 healthy donors was 28% and 4%, respectively. The investigation performed on well-characterized plasma EV reported JCPyV-DNA detection in 15 out of 36 (42%) of the viremic samples (14 were from HIV patients and 1 from healthy people) at a mean level of 23.5 copies/mL. The examination of EV selected samples reported the percentage of JCPyV-DNA in EV of 5.4% of the total viral load. Moreover, IEM reported the presence of JCPyV Vp1 antigen in plasma-derived EV., Conclusion: The potential role of EV-associated JCPyV-DNA open new avenues and mechanistic insights into the molecular strategies adopted by this polyomavirus to persist in the host and spread to the central nervous system., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

JC polyomavirus (JCPyV) is one of the most prevalent human viruses. Findings based on the geographic distribution of viral subtypes suggested that JCPyV codiverged with human populations. This view was however challenged by data reporting a much more recent origin and expansion of JCPyV. We collected information on ∼1,100 worldwide strains and we show that their geographic distribution roughly corresponds to major human migratory routes. Bayesian phylogeographic analysis inferred a Subsaharan origin for JCPyV, although with low posterior probability. High confidence inference at internal nodes provided strong support for a long-standing association between the virus and human populations. In line with these data, pairwise FST values for JCPyV and human mtDNA sampled from the same areas showed a positive and significant correlation. Likewise, very strong relationships were found when node ages in the JCPyV phylogeny were correlated with human population genetic distances (nuclear-marker based FST). Reconciliation analysis detected a significant cophylogenetic signal for the human population and JCPyV trees. Notably, JCPyV also traced some relatively recent migration events such as the expansion of people from the Philippines/Taiwan area into Remote Oceania, the gene flow between North-Eastern Siberian and Ainus, and the Koryak contribution to Circum-Arctic Americans. Finally, different molecular dating approaches dated the origin of JCPyV in a time frame that precedes human out-of-Africa migration. Thus, JCPyV infected early human populations and accompanied our species during worldwide dispersal. JCPyV typing can provide reliable geographic information and the virus most likely adapted to the genetic background of human populations., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: The use of human and viral genetic markers offers a novel way to study human migration in multiethnic populations of Latin America., Objectives: Our goal was to characterize the genetic diversity and geographical origins of JC Polyomavirus (JCPyV) and the genetic ancestry of mitochondrial DNA (mtDNA) in inhabitants from 25 de Mayo, Misiones-Argentina, a small village of largely German ancestry located close to the border with Brazil. We also evaluated the extent of agreement between viral and mtDNA markers for the different ancestry components of this population., Study Design: 68 individuals were analyzed for JCPyV and mtDNA diversity. JCPyV detection and typing was conducted in urine samples by PCR amplification, sequencing and phylogenetic analysis of the VP1 gene. mtDNA ancestry was assessed through HVS1 sequencing, with the resulting haplotypes being classified into haplogroups of Amerindian, European and African origin. The distribution of JCPyV diversity and mtDNA ancestry in the population was statistically evaluated by Fisher exact test and the level of agreement of both markers at the individual level was evaluated by Cohen's kappa coefficient., Results: Our analysis showed that 57.4% of the samples were positive for JCPyV. Of these, the 47.6% were Asian-American Type 2, 33.3% European Type 1 and 19.1% African Type 3 in origin. The mtDNA ancestry of the study participants was 33.3% Amerindian and 66.7% European. There was a significant difference among the distribution of JCPyV diversity and mtDNA ancestry (p = 0.009) and at the individual level there was no correlation between the distribution of the both markers (κ = 0.154, p = 0.297)., Conclusion: The apparent incongruence between JCPyV diversity and mtDNA ancestry may reflect the original settlement process and more recent migration to 25 de Mayo, the latter involving viral spread through migrants from Brazil. Some potential limitations to our interpretations are also discussed., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Before kidney transplantation, donors and recipients are routinely screened for viral pathogens using specific tests. Little is known about unrecognized viruses of the urinary tract that potentially result in transmission. Using an open metagenomic approach, we aimed to comprehensively assess virus transmission in living-donor kidney transplantation., Methods: Living kidney donors and their corresponding recipients were enrolled at the time of transplantation. Follow-up study visits for recipients were scheduled 4-6 weeks and 1 year thereafter. At each visit, plasma and urine samples were collected and transplant recipients were evaluated for signs of infection or other transplant-related complications. For metagenomic analysis, samples were enriched for viruses, amplified by anchored random polymerase chain reaction (PCR), and sequenced using high-throughput metagenomic sequencing. Viruses detected by sequencing were confirmed using real-time PCR., Results: We analyzed a total of 30 living kidney donor and recipient pairs, with a follow-up of at least 1 year. In addition to viruses commonly detected during routine post-transplant virus monitoring, metagenomic sequencing detected JC polyomavirus (JCPyV) in the urine of 7 donors and their corresponding recipients. Phylogenetic analysis confirmed infection with the donor strain in 6 cases, suggesting transmission from the transplant donor to the recipient, despite recipient seropositivity for JCPyV at the time of transplantation., Conclusions: Metagenomic sequencing identified frequent transmission of JCPyV from kidney transplant donors to recipients. Considering the high incidence rate, future studies within larger cohorts are needed to define the relevance of JCPyV infection and the donor's virome for transplant outcomes., (© The Author(s) 2018. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Colorectal cancer is the most repetitious malignancies with high mortality worldwide. JC virus (JCV) is ubiquitous Polyomavirus, with seroprevalence rates ranging from 70% to 90% in adult population. Recently the role of JCV have been reported in many malignant tumors worldwide. The association of JCV was reported in patients with colon and rectum cancers. Thus this study was conducted to evaluate the association of JCV DNA in patients with colon cancer type Adenocarcinoma. Material and Methods: A total of 120 formalin-fixed paraffin-embedded tissue blocks samples were collected including 20/40(50%) males, 20/40(50%) females patients with Colorectal Cancer(CRC), and 80 (50% males, 50% females) patients with benign tumor as a control. DNA was extracted for all the samples. Nested PCR was carried out for detection of Vp1/T-Ag junction genome in JCV genome by Nested-PCR assay. Randomly, PCR products of 6 samples were sequenced to analysis the partial JCV DNA. The phylogeny tree was constructed to determine homology identity with other JCV. Results: 4/40(10%) samples of test group and 10/80 (12.5%) of control samples were positive for JCV DNA (P= 0.69). Out of 4 samples positive for JC DNA, 3(7.5%) were males and 1(2.4%) female (P=0.29). The frequency of JCV DNA in age group> 50 years was 4/32(10%), while in age group <50 years was 0/8 (0%) (p= 0.29). Conclusion: prevalence of JCV DNA was among 10% patients with CRC and 12.5% benign tumors (p=0.69). The distribution of JCV DNA was among 7.5% male and 2.5% female (p= 0.29). The frequency of JCV DNA was among 10% cases of age group >50 years and 0% of age group <50 years (P= 0.29). The subsequent T-Ag protein expression might explain the increased risk of colorectal cancer and requires further investigation., (Creative Commons Attribution License)

Objectives Human JC polyomavirus (JCPyV) infection has an increased risk of developing progressive multifocal leukoencephalopathy (PML). Different JCPyV subtypes differ in the virulence with which they cause PML. Currently, the JCPyV infection status and subtype distribution in patients with human immunodeficiency virus-1 (HIV-1) in China are still unclear. This study aimed to investigate the epidemiology and subtype distribution of JCPyV in HIV-1-infected patients in China. Methods Urine samples from 137 HIV-1-infected patients in Zhejiang Province in China were tested for the presence of JCPyV DNA. The detected VP1 sequences were aligned and analysed using BioEdit and MEGA software. Results Among urine samples from HIV-1-infected patients, 67.2% were positive for JCPyV DNA (92/137). Primarily, the type 7 strains of JCPyV were detected, among which 45.5% (15/33) were subtype 7A, 30.3% (10/33) were 7B, and 24.2% (8/33) were 7C. Six nucleotide mutations, as well as one amino acid substitution, were isolated from the patients. Conclusions Urine samples from HIV-1-infected patients from Zhejiang Province show a high JCPyV infection rate. The most common JCPyV strains are subtypes 7A, 7B, and 7C.

BKV and JCV belong to the Polyomaviridae family and are opportunistic agents associated with complications in immunocompromised individuals. Although a single screening assay for both viruses would be convenient, the diversity of BKV and JCV serotypes and genotypes is a methodological challenge. In this paper, we developed a PCR method able to detect and segregate BKV and JCV, despite these genetic discrepancies. A duplex semi-nested PCR (duplex snPCR) was designed to target a conserved region (639nt-1516nt) within the VP2 gene. In the first PCR, a primer set common to all BKV and JCV serotypes/ genotypes was used, followed by a semi-nested PCR with internal primers for BKV and JCV segregation. The limit of detection of the duplex snPCR was as low as 10 copies of BKV or JCV plasmids/μL. Specific products were observed when JCV and BKV plasmids were mixed in the same reaction. In field sample testing, the duplex snPCR detected and distinguished both viruses in different biological samples. Results were confirmed by Sanger's sequencing. The geographical complexity of BKV and JCV serotypes and genotypes imposes limits to a simple and universal method that could detect each virus. However, we describe here a sensitive and reliable PCR technique for BKV and JCV diagnosis that overcomes these limitations and could be universally applied.

In healthy individuals John Cunningham virus is latent without any clinical signs, but in the cases of the use of immunosuppressive drugs in graft recipients, autoimmune diseases and also increasing of age, that the immune system is suppressed it may cause disease in reactivation. Progressive multifocal leukoencephalopathy (PML) is the well-known disease caused by the virus. It has also been associated with nephropathy and tumorogensis. At present, based on vp1 capsid gene 7 genotypes have been detected. Genetic variations of JC virus in different geographical areas and the presence of different subtypes is a useful tool for reconstructing of the genetic information of JC virus and understanding of its evolution. The aim of this study was to investigate different genotypes of the JC virus in the urine of 100 kidney transplant recipients, 43 rheumatoid arthritis patients, and 100 healthy individuals as control group in Isfahan. DNA was extracted by phenol-chloroform method and subjected to a nested PCR using specific primer for vp1 capsid gene designed by Oligo 7 software. Fisher's exact test was used for statistical analyses. Using MEGA 6 software the sequences were aligned using Clustal W tool and phylogenetic trees were constructed by neighbor joining method. Thirty-one positive samples were sequenced. Genotypes 1, 3, and 4 of the virus were detected for the first time in Iran. For the first time genotype 3 was reported as the dominant genotype in Iran. For the first time in the world, genotype 4 was detected in rheumatoid arthritis patients. J. Med. Virol. 89:337-344, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Recent studies have established that the human urine contains a complex microbiome, including a virome about which little is known. Following immunosuppression in kidney transplant patients, BK polyomavirus (BKV) has been shown to induce nephropathy (BKVN), decreasing graft survival. In this study we investigated the urine virome profile of BKV+ and BKV- kidney transplant recipients. Virus-like particles were stained to confirm the presence of VLP in the urine samples. Metagenomic DNA was purified, and the virome profile was analyzed using metagenomic shotgun sequencing. While the BK virus was predominant in the BKV+ group, it was also found in the BKV- group patients. Additional viruses were also detected in all patients, notably including JC virus (JCV) and Torque teno virus (TTV) and interestingly, we detected multiple subtypes of the BKV, JCV and TTV. Analysis of the BKV subtypes showed that nucleotide polymorphisms were detected in the VP1, VP2 and Large T Antigen proteins, suggesting potential functional effects for enhanced pathogenicity. Our results demonstrate a complex urinary virome in kidney transplant patients with multiple viruses with several distinct subtypes warranting further analysis of virus subtypes in immunosuppressed hosts.

Objective: Symptomatic infections of the central nervous system (CNS) with JC polyomavirus (JCV) usually occur as a result of immunocompromise and manifest as progressive multifocal leukoencephalopathy (PML) or granule cell neuronopathy (GCN). After immune reconstitution, some of these cases may show long-term persistence of JCV and delayed clinical improvement despite inflammation., Methods: We followed 4 patients with multiple sclerosis, who developed natalizumab-associated PML or GCN with regard to JC viral load and JCV-specific T-cell responses in the CNS. All of them experienced immune reconstitution inflammatory syndrome (IRIS), but in 2 cases JCV persisted > 21 months after IRIS accompanied by delayed clinical improvement., Results: Persistence of JCV was associated with a lack of JCV VP1-specific T-cell responses during immune reconstitution in 1 of the patients. Detailed analysis of the brain infiltrate in another patient with neuronal persistence of JCV revealed strong infiltration of CD8(+) T cells and clonal expansion of activated CD8(+) effector T cells with a CD4(dim) CD8(+) phenotype, both exhibiting exquisite specificity for conserved epitopes of JCV large T antigen. However, clearance of JCV was not efficient, because mutations in the major capsid protein VP1 caused reduced CD4(+) T-cell responses against the identified JCV variant and subsequently resulted in a decline of CD8(+) T-cell responses after IRIS., Interpretation: Our findings suggest that efficient CD4(+) T-cell recognition of neurotropic JCV variants is crucial to support CD8(+) T cells in combating JCV infection of the CNS., (© 2016 American Neurological Association.)

The objective of this study was to evaluate the prevalence, genotypic characterization, and determination of the patterns of shedding of human polyomavirus JC (JCPyV) and BK (BKPyV) in consecutive urine samples collected from healthy adults. Urine samples collected monthly over a 6 month period were screened by polymerase chain reaction (PCR) with two sets of primers complementary to the VP1 protein region specific for the JCPyV or BKPyV genome. The viral load of JCPyV and BKPyV in positive samples was determined by quantitative real time PCR. Seventy-one healthy individuals (ages between 18 and 65) were included in the study. Polyomavirus DNA urinary shedding was identified in 44 (62%) of the 71 individuals evaluated: BKPyV only in 16 (22.5%); JCPyV only in 19 (26.7%); and both in 9 (12.7%). Among the 28 individuals shedding JCPyV, the shedding was nearly continuous in 13 (46.4%) and sporadic in 15 (53.6%), whereas all BKPyV shedding was sporadic. A total of 45 (19 BKPyV and 26 JCPyV) strains were identified. Of the BKPyV strains, individuals were observed that excreted all genotypes except genotype 3 and the JCPyV strains, excretion of 5 different genotypes. Evaluating the age of individuals who excrete JCPyV and BKPyV, mostly are young adults, with a slight increase with increasing age and observing the viral load can not draw any parallel between the increase or decrease of age or excreted genotype as there was a wide variation both in the excretion of BKPyV and JCPyV. The high occurrence of isolated or simultaneous urinary shedding of JCPyV and BKPyV in healthy individuals merits further study., (© 2015 Wiley Periodicals, Inc.)

Progressive multifocal leukoencephalopathy (PML) is caused by JC polyomavirus (JCPyV). Because a reciprocal relationship has been described between antibody levels to JCPyV and BK polyomavirus (BKPyV), we performed a nested case control study with pre-diagnostic serum samples from HIV infected subjects to examine the relationship between BKPyV capsid antibodies and the risk of PML. Serum samples collected 0.5-2 years before PML diagnosis from 25 cases (66 samples) and 80 matched controls (204 samples) were tested in ELISA for JCPyV, BKPyV type 1 and type 4 capsid antibodies. High levels of BKPyV 1 and 4 antibodies were associated with a lower risk of PML (BKPyV 1 OR, 0.56, 95% CI, 0.35-0.89; BKPyV 4 OR, 0.40, 95% CI, 0.24-0.0.67). Our study demonstrates that antibodies to BKPyV capsids are an immunological marker of protection against development of PML. Further studies are needed to define the mechanism., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Progressive multifocal leukoencephalopathy (PML) represents a rare but potentially fatal reactivation of JC-polyomavirus (JCPyV) recently also reported in patients with autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis (RA) treated with rituximab. The aim of the present study was to analyse the pattern of JCPyV infections in patients with RA undergoing treatment with biologic agents. Urine and blood samples were analysed from 80 patients for antibody levels and/or the presence of JCPyV DNA. Genotyping of the control region and VP1 was performed for all JCPyV DNA-positive specimens. Viremia of JCPyV was only temporarily detected in two patients, and these viruses did not carry any mutations associated with the occurrence of PML. JCPyV DNA was prevalent in initial urine samples of 33% of all patients. RA patients who have consecutively been treated with two or more biologic agents revealed significantly higher prevalence of JCPyV DNA in the urine compared to RA patients treated with their first biologic agent. The presence of JCPyV DNA in the urine closely correlated to JCPyV antibody positivity, and therefore, antibody titres were higher in RA patients who had consecutively received two or more biologic agents over time. Therefore, the overall number of biologic agents had an impact on the pattern of JCPyV detection in this study. Hence, JCPyV antibody screening might be useful as part of the PML risk stratification for RA patients in the future.

Unlabelled: Opportunistic infection of oligodendrocytes by human JC polyomavirus may result in the development of progressive multifocal encephalopathy in immunocompromised individuals. Neurotropic JC virus generally harbors reorganized noncoding control region (NCCR) DNA interspersed on the viral genome between early and late coding genes. By applying 454 sequencing on NCCR DNA amplified from body fluid samples (urine, plasma, and cerebrospinal fluid [CSF]) from 19 progressive multifocal leukoencephalopathy (PML) patients, we attempted to reveal the composition of the JC polyomavirus population (the quasispecies, i.e., the whole of the consensus population and minor viral variants) contained in different body compartments and to better understand intrapatient viral dissemination. Our data demonstrate that in the CSF of PML patients, the JC viral population is often a complex mixture composed of multiple viral variants that contribute to the quasispecies. In contrast, urinary JC virus highly resembled the archetype virus, and urine most often did not contain minor viral variants. It also appeared that archetype JC virus could sporadically be identified in PML patient brain, although selection of rearranged JC virus DNA was favored. Comparison of the quasispecies from different body compartments within a given patient suggested a strong correlation between the viral population in plasma and CSF, whereas the viral population shed in urine appeared to be unrelated. In conclusion, it is shown that the representation of viral DNA in the CSF following the high-level DNA replication in the brain underlying PML has hitherto been much underestimated. Our data also underscore that the hematogenous route might play a pivotal role in viral dissemination from or toward the brain., Importance: For the first time, the JC polyomavirus population contained in different body compartments of patients diagnosed with progressive multifocal encephalopathy has been studied by deep sequencing. Two main findings came out of this work. First, it became apparent that the complexity of the viral population associated with PML has been highly underestimated so far, suggestive of a highly dynamic process of reorganization of the noncoding control region of JC polyomavirus in vivo, mainly in CSF and blood. Second, evidence showing viral dissemination from and/or toward the brain via the hematogenous route was provided, confirming a hypothesis that was recently put forward in the field., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Previous studies have demonstrated that infection with human polyomavirus, such as JCPyV and BKPyV, might be associated with various human tumors. However, an association between human JCPyV and BKPyV infection and diffuse large B-cell lymphoma (DLBCL) has not been reported. The purpose of this study was to examine DLBCLs of the gastrointestinal tract for evidence of human polyomavirus infection. Nested PCR and DNA sequencing were employed for viral DNA detection and viral genotype identification. In addition, two viral proteins, the large tumor antigen (LT) and the major structural protein (VP1), were detected by immunohistochemistry (IHC). Human JCPyV and BKPyV DNA was detected in 14 out of 16 tissue samples (87.5%), whereby nine cases were infected with JCPyV and five cases were infected with BKPyV. Both archetypal and rearranged genotypes of JCPyV and BKPyV were detected in the tissues. LT was detected in 11 tissue samples (68.75%). However, VP1 was not detected in any of the tissue samples. The presence of human JCPyV and BKPyV DNA and protein in DLBCL tissues of gastrointestinal tract were first reported in this study. The current results provide evidence of a possible association between human JCPyV and BKPyV infection and DLBCL.

Background: JC polyomavirus (JCPyV) is a widespread human polyomavirus that usually resides latently in its host. It can be reactivated under immunomodulating conditions and cause Progressive Multifocal Leukoencephalopathy (PML). Circulating microRNAs (miRNAs) are emerging as promising biomarkers for several pathologies. In this study, we have investigated whether circulating miRNAs exist that are differentially expressed between JCPyV seropositive and JCPyV seronegative on the one hand or between JCPyV shedders and JCPyV non-shedders on the other hand., Methods: Human miRNA expression profiling was performed in a small set of plasma samples obtained from seronegative subjects, seropositive shedders and seropositive non-shedders. A set of 10 miRNAs was selected for further analysis in a larger group of samples., Results: Based on the plasma profiling experiment of 30 samples, 6 miRNAs were selected that were possibly differentially expressed between seropositive and seronegative subjects and 4 miRNAs were selected that were possibly differentially expressed between shedders and non-shedders. Subsequently, expression of these 10 selected miRNAs was assessed in an independent set of 100 plasma samples. Results indicated that none of them were differentially expressed., Conclusion: This study could not identify circulating human miRNAs that were differentially expressed between plasma from JCPyV seropositive and JCPyV seronegative subjects or between JCPyV shedders and JCPyV non-shedders.

The JC virus is a widely infected human polyomavirus. Recent foreign researches showed that the JC virus infection is correlated with tumors of nervous system and digestive system, while, and study on the relationship between JC virus infection and gynecological tumor is seldom reported. In this study, we first establish the nucleic acid detection methods and procedures for JC virus and its highly homologous BK virus. The JC and BK viruses infection was evaluated by detect the viral DNA in samples including biopsy tissues, serum as well as urine of myoma of uterus (98 cases), cervical cancer (84 cases), endometrial cancer (40 cases) and ovarian tumor (72 cases) patients. The BK viral DNA positive rate was significantly higher in urine samples than that of blood and biopsy samples, and there is no significant difference of the BK viral DNA positive rate among all patient groups. The JC viral DNA positive rate is almost 0 in serum samples and biopsy. tissues, however, viral DNA positive rate is more than 50% in urine samples. In fibroids group, the JC viral DNA positive rate is up to 65. 3% which is significantly higher than that in other patients groups and healthy control. Further gynecological tumor associated viruses detection showed that only human papilloma virus infection is associated with cervical cancer, the herpes simplex virus, EB virus and cytomegalovirus infection is extremely low in our patient groups. No synergistic effect on gynecological tumor caused by viruses co-infection was observed. Our study showed that JC virus infection is highly related to the pathogenesis of uterine fibroids.

Certain factors lead to increased reactivation of JC virus (JCV) and immunodeficiency seems to be the most important. JCV isolates can be classified into eight different genotypes and several subtypes based on nucleotide difference in the VP1 gene. JCV genotypes are strongly associated with particular ethnic groups and frequently used as genetic markers for human evolution and migration. The aim of this study was to determine the frequency of JCV urinary shedding and genotype distribution in Serbia among patients infected with HIV and healthy donors. Urine samples from 107 healthy donors and 93 patients infected with HIV were collected. PCR followed by sequence analysis was carried out using primers specific for VP1 and NCRR of the virus genome. Excretion rate of JCV-DNA in urine was higher in patients infected with HIV than in healthy donors (44.1% vs. 31.7%) although statistical significance was not found. Within the group infected with HIV, the degree of immunosuppression (measured by CD4(+) cell count) did not influence JCV excretion rate. Sequence analysis of JCV NCRR from both patients infected with HIV and healthy donors showed a pattern identical to archetype structure. In healthy Serbian donors the predominant genotype was 1 (41.2%), followed by 4 (32.4%) and 2 (26.4%). On the other hand, genotype distribution pattern was different in patients infected with HIV: 2 (43.9%), 1 (31.7%), and 4 (24.4%). This study showed that European, Eurasian, and Indian types are circulating in Serbia and that distribution corresponds to the origin of the inhabitants of Serbia., (© 2013 Wiley Periodicals, Inc.)

There is a resurgence of interest in the study of occurrence, genotype and pathogenic associations of human Polyomaviruses in recent years. In the present study, we have ascertained the presence of human Polyomavirus JC (JCV) in the urine and peripheral blood leukocytes of tribal populations, for the first time in the North-Eastern part of West Bengal State of India. We have also characterized the prevalent genotypes of the non-coding controlregions (NCCRs) of these natural isolates. The result suggests a high incidence of JCV reactivation in the populations assayed. Approximately 25% of the non-immunocompromized tribal men and women, tested positive based on polymerase chain reaction (PCR) analysis, and these results were further confirmed by sequencing of PCR products. Pairwise sequence comparison and alignment of the NCCR sequence of these Indian strains appeared to be comparable and related to the archetypal JCV (CY) and the Tibetan LH3 strains, with some alterations in few key positions. The sequence analyses were done with regard to transcription factor binding to DNA sequence elements of endemic JCV NCCRs.

BK virus (BKV) and JC virus (JCV) are human polyomaviruses that cause asymptomatic latent infections. Under immunosuppression, BKV-associated nephropathy has been documented in Kuwait and elsewhere. Even though different BKV and JCV genotypes with distinct geographical distribution have been described, the genotype of polyomavirus detected in Kuwait is still unknown. The aim of this study was to determine the genotypes of BKV and JCV detected in renal transplant recipients. The detection of polyomavirus DNA was carried out in serum and urine samples of 200 post-transplant recipients during a 1-year follow-up period. Fifty-one (25.5%) post-transplant recipients were tested positive for polyomavirus DNA by semi-nested PCR. JCV DNA could be detected in 29 (57%) patients, and BKV DNA in 22 (43%) patients. In two renal transplant recipients, both BKV and JCV were detected. According to the Bayesian phylogenetic analysis of polyomavirus VP1 sequences, the majority of detected BKV sequences were most closely related to genotypes I and IV, whereas the majority of JCV sequences were most closely related to genotype 3. Polyomavirus VP1 sequences showed strong stability for up to 12 months in most patients; however, in one patient, an amino acid substitution in the BKV VP1 protein was identified over time. The results suggest a close relationship of BKV sequences with the Asian and European strains, and of JCV sequences with the African strains. Long follow-up studies are needed to investigate the association of polyomavirus polymorphism or genotypic shift with the development of nephropathy., (Copyright © 2013 Wiley Periodicals, Inc.)

JC virus is a human polyomavirus that infects the majority of people without apparent symptoms in healthy subjects and it is the causative agent of progressive multifocal leucoencephalopathy (PML), a disorder following lytic infection of oligodendrocytes that mainly manifests itself under immunosuppressive conditions. A hallmark for JC virus isolated from PML-brain is the presence of rearrangements in the non-coding control region (NCCR) interspersed between the early and late genes on the viral genome. Such rearrangements are believed to originate from the archetype JC virus which is shed in urine by healthy subjects and PML patients. We applied next generation sequencing to explore the non-coding control region variability in urine of healthy subjects in search for JC virus quasispecies and rearrangements reminiscent of PML. For 61 viral shedders (out of a total of 254 healthy subjects) non-coding control region DNA and VP1 (major capsid protein) coding sequences were initially obtained by Sanger sequencing. Deletions between 1 and 28 nucleotides long appeared in ∼24.5% of the NCCR sequences while insertions were only detected in ∼3.3% of the samples. 454 pyrosequencing was applied on a subset of 54 urine samples demonstrating the existence of JC virus quasispecies in four subjects (∼7.4%). Hence, our results indicate that JC virus DNA in urine is not always restricted to one unique virus variant, but can be a mixture of naturally occurring variants (quasispecies) reflecting the susceptibility of the non-coding control region for genomic rearrangements in healthy individuals. Our findings pave the way to explore the presence of viral quasispecies and the altered viral tropism that might go along with it as a potential risk factor for opportunistic secondary infections such as PML.

JC polyomavirus (JCPyV) was the first of now 12 PyVs detected in humans, when in 1964, PyV particles were revealed by electron microscopy in progressive multifocal leukoencephalopathy (PML) tissues. JCPyV infection is common in 35-70% of the general population, and the virus thereafter persists in the renourinary tract. One third of healthy adults asymptomatically shed JCPyV at approximately 50,000 copies/mL urine. PML is rare having an incidence of <0.3 per 100,000 person years in the general population. This increased to 2.4 per 1000 person years in HIV-AIDS patients without combination antiretroviral therapy (cART). Recently, PML emerged in multiple sclerosis patients treated with natalizumab to 2.13 cases per 1000 patients. Natalizumab blocks α4-integrin-dependent lymphocyte homing to the brain suggesting that not the overall cellular immunodeficiency but local failure of brain immune surveillance is a pivotal factor for PML. Recovering JCPyV-specific immune control, e.g., by starting cART or discontinuing natalizumab, significantly improves PML survival, but is challenged by the immune reconstitution inflammatory syndrome. Important steps of PML pathogenesis are undefined, and antiviral therapies are lacking. New clues might come from molecular and functional profiling of JCPyV and PML pathology and comparison with other replicative pathologies such as granule cell neuronopathy and (meningo-)encephalitis, and non-replicative JCPyV pathology possibly contributing to some malignancies. Given the increasing number of immunologically vulnerable patients, a critical reappraisal of JCPyV infection, replication and disease seems warranted., (© 2013 APMIS. Published by John Wiley & Sons Ltd.)

Very little is known about JCV infection and genotype distribution with respect to the different demographic and clinical characteristics of the Italian population. A cross-sectional study was carried out on the prevalence of JCV genotypes in 323 Caucasian subjects (mean age: 37.5 years, range: 2-70 years). Urine samples from 200 immunocompromised patients, including patients affected by Multiple Sclerosis (MS), Human Immunodeficiency Virus (HIV), colon cancer, inflammatory diseases and Progressive Multifocal Leukoencephalopathy (PML), and 123 immunocompetent individuals were tested by quantitative real time PCR. Sequencing of the JCV viral protein 1 (VP1) and transcriptional control region (TCR) was performed. In this series, the overall prevalence of JCV excretion was 32.8% without significant differences between males and females. JCV was detected in 39.5% of patients and in 22% of healthy individuals (P = 0.004). The most prevalent JCV genotype excreted was genotype 1 (69.8%), followed by genotype 2 (22.6%) and genotype 4 (7.5%). Distribution of genotypes between patients and healthy subjects showed a statistically significant difference for the type 1 compared to the other variants (P < 0.01). Of note, JCV genotype 2 was found to be associated to young patients (P = 0.0001) and to patients treated with immunomodulator drugs (P = 0.0001), but not to PML subjects. The non-pathogenic archetype IIS (singular, insert) form was present in all JCV strains detected. This result allows to hypothesize a possible JCV genotype selection in response to pressure by immunomodulatory drugs., (Copyright © 2013 Wiley Periodicals, Inc.)

Background: JC virus (JCV) is the etiologic agent for progressive multifocal leukoencephalopathy (PML), a demyelinating disease occurring in the brain of patients with underlying immune compromised states. All viable JCV genomes contain a conserved region in the T protein coding nucleotide sequence that when detected by PCR in CSF is a confirmatory diagnostic marker for PML along with clinical and neuroradiological evidence. The non-coding regulatory region (NCRR) is hypervariable, as evidenced by nucleotide sequence of the non-virulent variant, which is predominantly excreted in urine, versus that of virulent variants found in brain and CSF of PML patients. All variants can be found in blood., Objective: A single assay that quantifies and identifies JCV DNA in clinical samples and discriminates between variants has significant value to physicians and patients at risk for PML., Study Design: Separate primer pairs were tested together to quantitatively detect conserved viral DNA nucleotide sequence in patient samples, while simultaneously detecting the NCRR specific for the non-virulent variant., Results: In testing using control plasmids and patients' CSF, blood, and urine, PML patients predictably demonstrated the non-virulent, archetype NCRR in urine, but virulent NCRR variants in CSF and blood., Conclusion: The JCV qPCR multiplex assay targets two regions in JCV genomes to simultaneously identify and measure viral DNA, as well as distinguish between variants associated with PML and those that are not. The multiplex results could signal risk for PML if patients are viremic with JCV variants closely associated with PML pathogenesis., (Published by Elsevier B.V.)

Treatment with biological drugs is associated with increased susceptibility to viral infections. Reactivation of JC virus (JCV) and human cytomegalovirus (HCMV) in adults after therapy has been documented. The long-term effects of biological and conventional therapy on human herpesviruses and polyomaviruses infections in young patients were assessed. One hundred eighty-six samples [urine, serum, and blood cells (PBMCs)] from 62 patients (15.8 ± 6.2 years old) with Crohn's disease, ulcerative rectocolitis or juvenile rheumatoid arthritis treated with immunotherapy or conventional therapy for over 12 months were tested by real time PCR. One hundred twenty-four samples (urine and blood) from 62 matched healthy volunteers (13.8 ± 8.6 years old) were included as controls. Sequencing of the JCV viral protein 1 (VP1) and transcriptional control region (TCR) was performed. Herpes simplex virus 1/2 and varicella zoster virus genomes were not detected in any patients, whereas Epstein-Barr virus, HCMV, and human herpesvirus-6 genomes were detected in 4.8%, 3.2%, and 1.6% of the patients, respectively. JCV was detected in 22.6% (14/62) of urine samples from patients and in 8% (5/62) from controls, in 50% (7/14) of sera from patients shedding JCV, and in 71.4% (5/7) of matched PBMCs. There was a significant association between infliximab treatment and excretion of JCV genotype 2. Subclinical infection/reactivation of JCV genotype 2 in young patients during infliximab therapy was demonstrated. Conversely, increased susceptibility to herpesviruses infection was not shown. Future studies are warranted to investigate the effects of JCV reactivation on the health of young patients treated with infliximab., (Copyright © 2013 Wiley Periodicals, Inc.)

The polyomavirus JC (JCPyV) is a ubiquitous virus in humans, causing progressive multifocal leukoencephalopathy, a fatal demyelinating disease. JCPyV propagates in the adult kidney and excretes its progeny in urine, from which its DNA can be recovered readily. JCPyV isolates worldwide can be classified into 14 subtypes or genotypes, each associated with a specific geographical region. The European genotypes EU-a-b-c are spread throughout Europe and Mediterranean areas. The major African genotype Af2 is spread not only throughout Africa but also in West and South Asia. A minor African genotype (Af1) occurs in Central and West Africa. Partially overlapping domains in Asia were occupied by various genotypes (e.g., B1-a, -b, -d, B2, CY, MY, and SC). To characterize the subtypes of JCPyV prevalent in Tunisia, the presence of the virus was investigated by real-time PCR in urine samples from 98 renal transplant recipients. For subtype identification, a 610 bp typing region of the JCPyV genome was amplified from each urine sample, and its DNA sequence was determined. In the patients studied, the major African subtype Af2 was the predominant (62.5%), followed by the European subtype EU (33.5%). Only one case clustering with the Asian genotype SC (4%) was identified. The presence of the European subtype with high prevalence in this population suggests that the epidemiological distribution of JCPyV virus sequences in North Africa is related partially to the epidemiological data in Europe., (Copyright © 2012 Wiley Periodicals, Inc.)

JC polyomavirus (JCV) is a member of the Polyomaviridae family. It presents a tropism to kidney cells, and the infection occurs in a variety of human population groups of different ethnic background. The present study investigated the prevalence of JCV infection among human populations from the Brazilian Amazon region, and describes the molecular and phylogenetic features of the virus. Urine samples from two urban groups of Belém (healthy subjects), one Brazilian Afro-descendant "quilombo" from the Rio Trombetas region, and native Indians from the Wai-Wai, Urubu-Kaapor, Tembé, Assurini, Arara do Laranjal, Aukre, Parakanã, Surui and Munduruku villages were investigated for the presence of the virus by amplifying VP1 (230 bp) and IG (610 bp) regions using a polymerase chain reaction. Nucleotide sequences (440 nucleotides, nt) from 48 samples were submitted to phylogenetic analysis. The results confirmed the occurrence of types A (subtype EU), B (subtypes Af-2, African and MY, Asiatic) and C (subtype Af-1) among healthy subjects; type B, subtypes Af-2 and MY, among the Afro-Brazilians; and type B, subtype MY, within the Surui Indians. An unexpected result was the detection of another polyomavirus, the BKV, among Afro-descendants. The present study shows, for the first time, the occurrence of JC and BK polyomaviruses infecting humans from the Brazilian Amazon region. The results show a large genetic variability of strains circulating in the region, infecting a large group of individuals. The presence of European, Asiatic and African subtypes associated to the ethnic origin of the population samples investigated herein, highlights the idea that JCV is a fairly good marker for studying the early migration of human populations, reflecting their early and late history. Furthermore, the identification of the specific mutations associated to the virus subtypes, suggests that these mutations have occurred after the entrance of the virus in the Amazon region of Brazil.

JC polyomavirus (JCV) infects 80% of the population. In immunosuppressed patients viral replication may be enhanced, resulting in progressive multifocal leukoencephalopathy. There are at least eight distinct strains of JCV of which distribution varies between ethnic groups however there remain issues with identifying viral strains due to the insufficient yield and sensitivity of Sanger sequencing (SS) protocols. In this study matrix assisted laser desorption/ionisation time of flight mass spectrometry (MALDI TOF MS) is presented as an alternative to SS. DNA from 19 urine samples was assessed. Successful amplification and strain identification was possible in all samples with >10 copies of starting material indicating a high sensitivity assay, in contrast to SS (>20 copies). The genotypes defined for each sample via MALDI TOF MS were identical where SS resulted in a readable trace. MALDI TOF MS identified eight novel SNPs in the VP1 gene. This study represents the first application of MALDI TOF MS to viral comparative sequence analysis., (Crown Copyright (c) 2010. Published by Elsevier B.V. All rights reserved.)

JC virus (JCV) is ubiquitous in the human population, infecting children asymptomatically. After primary infection, JCV persists in the host throughout life and is often excreted in the urine. Two hundred thirty-four urine samples and 78 serum samples, collected from 171 healthy individuals and 63 patients infected with HIV, were used to characterize JCV infection in a Portuguese population. Using PCR, JCV DNA was detected in 38% of the urine samples. A significant difference in the excretion rate was observed between patients infected with HIV (51%) and healthy individuals (33%). The frequency of JCV viruria increased with age in healthy individuals, but not in patients infected with HIV. JCV urinary load was determined by real-time quantitative PCR and was independent of gender, age, HIV infection, and CD4+ cell count. Overall, the JCV genotype detected most commonly was 1B, followed by genotypes 2B and 4. The detection and quantitation of JCV-specific antibodies were performed in serum samples by an established enzyme immunoassay (EIA). Antibodies to JCV were observed in 91% of the patients tested, irrespective of HIV infection. A positive correlation between JCV urinary load and antibody titers was demonstrated. The present study provides the first characterization of seroprevalence and urinary excretion of JCV in a Portuguese population and revealed similar results to those observed in other European countries. A comparison between healthy individuals and patients infected with HIV, despite identical values of seroprevalence, showed some differences in the pattern of urinary excretion. J. Med. Virol. 82:494-504, 2010. (c) 2010 Wiley-Liss, Inc.

Hemorrhagic cystitis is characterized by hematuria due to inflammation of the bladder. In bone marrow transplants, this disease is linked to the infection by human polyomavirus BK, whereas the role of the human polyomavirus JC is unclear. The transcriptional control regions of both viruses contain important cellular transcription factor binding sites that undergo rearrangement process generating suitable variants that could be more active for viral replication and for the onset of hemorrhagic cystitis. In this study urine obtained from seven patients with bone marrow transplant were examined. Polyomavirus genomes were quantified by PCR and viral loads were compared. The transcriptional regions of both viruses were amplified and sequenced to determine the presence of variants. Subtypes of polyomaviruses were determined by amplification and sequencing of the viral protein 1 region. The results showed that four of seven patients were positive for BK DNA, two of seven patients had BK and JC DNA and one of seven had JC DNA. Positive samples were amplified and sequenced successively for transcriptional regions. The viral archetype was always found in both viruses. Finally, typing showed that BK virus subtype I infected patients with BK, whereas JC virus genotype IA and genotype 1B were found in patients infected with JC. The data suggest that new and different approaches are required to improve the morbidity and mortality caused by polyoma-associated hemorrhagic cystitis, since it known that BK virus is involved in the onset of hemorrhagic cystitis, whereas the role of JC virus should be investigated further., ((c) 2009 Wiley-Liss, Inc.)

JC virus (JCV) causes progressive multifocal leukoencephalopathy under conditions of immunosuppression, especially associated with HIV. Despite the high prevalence of HIV-1 infection, few cases of progressive multifocal leukoencephalopathy have been reported and only a small number of JCV strains have been characterized in AIDS patients in Southern Africa. Diagnosis of progressive multifocal leukoencephalopathy through PCR detection of JCV DNA in CSF may result in false negative results if variable regions of the genome are targeted. Characterization of circulating JCV genotypes in Southern Africa may assist with designing appropriate diagnostic assays and characterizing the molecular epidemiology of JCV in African AIDS patients. In this study, a new real-time PCR using fluorescence resonance energy transfer hybridization probes targeting the conserved large T-antigen was developed and compared to a conventional nested PCR targeting the variable VP1 region. Validation against known JCV positive specimens confirmed its specificity while amplification of a serial dilution of JCV control DNA suggests that the new real-time PCR can detect lower concentrations of JCV than the VP1 nested PCR. Investigation of CSF specimens from 44 suspected progressive multifocal leukoencephalopathy patients suggest that the new assay is more sensitive being able to detect JCV in 12 specimens versus 5 specimens with the VP1 nested PCR. Sequence analysis confirmed that the T-antigen region is completely conserved while phylogenetic analysis of the five VP1 products identified two genotype 3, one genotype 1 and two genotype 4 strains, the latter two identified for the first time in South African AIDS patients.

JC polyomavirus (JCV) isolates worldwide are classified into three super-lineages (A, B and C), with A and B further split into several lineages and sub-lineages. The transcriptional control region (TCR) of the JCV genome generally has the archetypal configuration, but rearranged TCRs have occasionally been detected in isolates from immunocompetent individuals. To investigate the phylogenetic significance of these rearrangements, we analyzed 298 TCR sequences all derived from complete JCV genomes directly cloned from the urine of non-immunocompromised individuals. While sporadic rearrangements were found in many lineages and sub-lineages, common rearrangements were identified in all, or essentially all, isolates belonging to particular lineages or sub-lineages. Interestingly, several common rearrangements were also detected as sporadic rearrangements in other lineages or sub-lineages. This observation suggests that during the course of JCV evolution, JCV strains with sporadic rearrangements became predominant over archetypal TCRs in some JCV lineages or sub-lineages.

Although JC virus (JCV), a human polyomavirus, has been detected in colon cancers, the association between JCV and colon cancer remains controversial. In Taiwan, the prevalence of JCV infection in colon cancer patients has not been reported. Thus, the purpose of this study was to investigate JCV infection in colon cancers in Taiwan. Formalin-fixed, paraffin-embedded tissues from 22 colon cancer patients were examined in this study. Nested PCR was performed to detect viral genomic DNA. The product of the nested PCR flanking the JCV regulatory region was sequenced further. Viral large tumor protein, LT, and late capsid protein, VP1, were examined by immunohistochemistry (IHC). Nested PCR revealed JCV genomic DNA in 86.4% (19/22) of the colon cancer tissue samples. Both rearranged and archetypal genotypes of JCV were identified. Expression of JCV LT was positive in 63.6% (14/22) of the examined colon cancer tissue samples but not in any adjacent normal region. Expression of viral capsid protein VP1 was not detected in any of the tissues examined. The current study demonstrates that JCV genomic DNA was present in the examined colon cancer tissues. The genotypes of JCV in colon cancer tissues were also identified. Expression of viral early protein but not structural capsid protein was detected in the examined colon cancer tissues. Furthermore, a high prevalence of JCV infection in colon cancer tissues in Taiwan was also demonstrated.

Background/purpose: Among the genotypes of human polyomavirus JC (JCV) reported in Taiwan, CY, TW1, TW2 and TW3 are the most commonly correlated with human diseases. JCV is usually detected using nucleotide sequencing and restriction fragment length polymorphism (RFLP) analysis. The aim of this study was to detect the rate of positivity and genotype of the JCV genome in urine by RFLP or capillary electrophoresis (CE) in renal transplant patients and healthy volunteers., Methods: We compared CE analysis to the methods of nucleotide sequencing and RFLP analysis for detection of JCV viruria among 60 renal transplant patients and 50 unrelated healthy controls. Genotyping of the positive PCR products was performed using CE and RFLP analysis simultaneously., Results: The urine JCV-positive rate was significantly higher in renal transplant patients than in healthy volunteers (40% [24/60] vs. 20% [10/50]; p=0.0238). In addition, multiple genotypes of JCV could be detected by CE, but only one genotype could be detected by RFLP. In our study, 20% (2/10) of urine JCV-positive samples from healthy volunteers had two different genotypes. In renal transplant patients 66% (16/24) of JCV-positive samples had two different genotypes and 12% (3/24) had three different genotypes., Conclusion: In comparison with RFLP, CE can detect multiple genotypes in urine JCV-positive samples and requires only 1/200 of the volume of specimen required for RFLP analysis. The CE method has sensitivity and specificity suitable for use in the clinical laboratory, and identifies more genotypes than RFLP analysis.

Microbial pathogens, and viruses in particular, can serve as important complements to traditional genetic markers when investigating the population histories of their human host. The range of mutation rates for DNA viruses suggests that DNA viruses can be useful markers of both recent and ancient events in their host histories. Here, we assess the utility of a well known DNA virus, JC virus (JCV), for investigating human history and demography. Using complete coding viral genomes, we confirm the phylogeographic structure of JCV in populations worldwide and provide coalescent estimates of its evolutionary rate under two alternative models of its history. Using these rate estimates, we compare Bayesian skyline plots of population size changes for JCV to those of its human host as estimated with coding mitochondrial genomes of the latter. These comparisons, when combined with other evidence including a log Bayes Factor model test, show that JCV is evolving rapidly and is therefore tracking the recent history of its human host. These results support the hypothesis that post-World War II societal changes are most likely responsible for the recent demographic patterns observed among different regional JCV populations. In sum, fast evolving DNA viruses, such as JCV, can complement RNA viruses to provide novel insights about the recent history and demography of their human host.

JC virus (JCV)-specific CD8+ cytotoxic T lymphocytes (CTL) are associated with a favorable outcome in patients with progressive multifocal leukoencephalopathy (PML) and cross-recognize the polyomavirus BK virus (BKV). We sought to determine the frequency and phenotype in fresh blood of CD8+ T cells specific for two A*0201-restricted JCV epitopes, VP1(p36) and VP1(p100), and assess their impact on JC and BK viremia and viruria in 15 healthy subjects, eight human immunodeficiency virus-positive (HIV+) individuals, and nine HIV+ patients with PML (HIV+ PML patients) classified as survivors. After magnetic pre-enrichment of CD8+ T cells, epitope-specific cells ranged from 0.001% to 0.022% [corrected] by tetramer staining, with no significant difference among the three study groups. By use of seven-color flow cytometry, there was no predominant differentiation phenotype subset among JCV-specific CD8+ T cells in healthy individuals, HIV+ subjects, or HIV+ PML patients. However, in one HIV+ PML patient studied in the acute phase, there was a majority of activated effector memory cells. BKV DNA was undetectable in all blood samples by quantitative PCR, while a low JC viral load was found in the blood of only one HIV+ and two HIV+ PML patients. JCV and BKV DNA were detected in 33.3% and 13.3% of all urine samples, respectively, independent of the presence of JCV-specific CTL. The detection of JCV DNA in the urine was associated with the presence of a JCV VP1(p100) CTL response. Immunotherapies aiming at increasing the cellular immune response against JCV may be valuable in the treatment of HIV+ individuals with PML.

Human polyomavirus JC (JCV) infects the worldwide population, remains latent in the kidney, and is excreted in the urine. A longitudinal study was performed in order to evaluate JCV excretion, to characterize molecularly the virus and to determine if its presence in urine is a consequence of viral reactivation or merely of epithelial squamous cell shedding. The presence of cellular sediment and the JCV genome were examined in 333 urine samples collected periodically for 3 months from 17 healthy subjects; molecular characterization, and quantitation of the virus were also undertaken. JCV DNA was detected in 40.2% of the samples, with a significant difference (P<0.001) observed between males and females. JCV shedding was independent of the presence of cellular sediment in every individual. JCV genotype 1 was the genome detected most frequently, while all of the amplified strains showed archetypal organization of the transcriptional control region (TCR). No clinical symptoms have been associated with JCV excretion and no microbial load was detected in the urine samples. The lack of correlation between JCV DNA detection and the presence of squamous cells in urine sediment indicates that viruria is regulated by the life cycle of JCV. Thus, the virus is eliminated as consequence of its reactivation., ((c) 2007 Wiley-Liss, Inc.)

The ubiquitous human polyomavirus JC (JCV) is a small double-stranded DNA virus that establishes a persistent infection, and it is often transmitted from parents to children. There are at least 14 subtypes of the virus associated with different human populations. Because of its presumed codivergence with humans, JCV has been used as a genetic marker for human evolution and migration. Codivergence has also been used as a basis for estimating the rate of nucleotide substitution in JCV. We tested the hypothesis of host-virus codivergence by (i) performing a reconciliation analysis of phylogenetic trees of human and JCV populations and (ii) providing the first estimate of the evolutionary rate of JCV that is independent from the assumption of codivergence. Strikingly, our comparisons of JCV and human phylogenies provided no evidence for codivergence, suggesting that this virus should not be used as a marker for human population history. Further, while the estimated nucleotide substitution rate of JCV has large confidence intervals due to limited sampling, our analysis suggests that this virus may evolve nearly two orders of magnitude faster than predicted under the codivergence hypothesis.

Mixtures of polyomaviruses can be present in the central nervous system, the gastrointestinal tract, the genitourinary tract, blood, and urban sewage. We have developed 12 primer/probe sets (four per virus) for real-time, quantitative PCR assays (TaqMan) that can specifically detect BKV, JCV, and SV40 genomes present in mixtures of these viruses. The specificities of these primer/probe sets were determined by evaluating their level of interaction with the DNA from other polyomaviruses and their ability to estimate the number of copies of homologous viral DNA in blinded samples of defined mixtures of three polyomaviral DNAs. Three early region and three late region primer/probe sets determined, within a two-fold range, the number of copies of their respective DNAs. Four sets of SV40 primer/probes also detected 1.1-2.4 copies of SV40 DNA per COS-1 cell, cells estimated to contain a single copy of SV40 DNA. Three JCV primer/probe sets detected 3.7-4.2 copies per cell of JCV DNA in the JCV-transformed cell line M1-HR, cells estimated to contain between 0.5 and 1 copy of the JCV genome. We suggest that the virus-specific primer/probe sets in this study be considered sufficiently characterized to initiate the quantification of polyomavirus DNA in biological samples.

Many human neurological diseases involve demyelination of the central and/or peripheral nervous systems. These include the hereditary leukodystrophies--which have a genetic basis; multiple sclerosis (MS)--where the underlying cause of demyelination remains unknown; and progressive multifocal leukoencephalopathy (PML)--where the etiology is well-established as being viral. The human neurotropic polyomavirus--JC virus (JCV)--is the etiologic agent of PML, a fatal demyelinating disease of the central nervous system that occurs mainly in immunosuppressed patients, especially those with HIV/AIDS. JCV belongs to the polyomavirus family of tumor viruses that are characterized by non-enveloped icosahedral capsids containing small, circular, double-stranded DNA genomes. Serological studies have shown that JCV is widespread throughout the human population, but infections are usually restricted by the immune system, particularly cell-mediated immunity, causing the virus to enter a latent phase. An important corollary of this is that situations of severe immunosuppression may permit JCV to replicate and are thus a risk factor for PML.

Herpes zoster (HZ) is a neurocutaneous disease caused by Varicella-zoster virus (VZV) as a consequence of declined cell-mediated immunity, immune suppression and immunodeficiency. As reactivation of JC polyomavirus (JCPyV) might be linked with immunodeficiency or immunosuppressive therapy, the relationship between HZ and JCPyV reactivation was investigated. The incidence of JCPyV in urine samples from 102 patients with HZ and 100 healthy individuals from South Korea was determined by PCR. The incidence values for HZ patients and control individuals did not differ significantly (24.5% vs. 20.0%, respectively, P = 0.5391). When different age groups were monitored, the positivity values of 21.1%, 20.0%, and 30% were found for 20-39, 40-59 and over 60 year-old patients, respectively. In order to determine the genotype of JCPyV isolates, their VP1-large T antigen (VT)-intergenic region was PCR amplified, sequenced and analyzed. Three distinct types, namely 1, 2A and 7B were found in 8%, 24%, and 68% of were found among 25 isolates from HZ patients. Using phylogenetic analysis, the type 1 isolates were assigned to the 1C subtype. These results indicate that HZ does not play an important role in JCPyV reactivation and is not associated with JCPyV.

The etiology of brain tumors and meningiomas is still unknown. Several factors have been considered, such as genetic predisposition and environmental risk factors, but the hypothesis that one or more infectious agents may play a role in tumor pathogenesis has also been investigated. Therefore, emphasis was placed on the neurooncogenic family Polyomaviridae and the presence of human polyomavirus DNA sequences and JCV mRNA were examined in malignant human brain biopsies. Italian patients affected with different types of neoplasias of the brain and its covering were enrolled. The patients underwent surgical tumor excision and the presence of the polyomavirus genome in biopsy and other body fluids was evaluated by PCR. In addition, the genomic organization of JCV was examined in depth, with the aim of providing information on genotype distribution and TCR rearrangements in the population affected with intracranial neoplasms. On the whole, polyomavirus DNA was found in 50% of the biopsy specimens studied, JC virus DNA and BK virus DNA were amplified in 40.6% mainly glioblastomas and 9.4% of the tissue specimens, respectively, while none of the biopsy specimens tested contained Simian virus 40 DNA. Genotype 1 and Mad 4 TCR organization were the most frequent in the population enrolled. Although a cause and effect was not demonstrated and the specific role of the viruses remains unknown, the findings appear to confirm the hypothesis that JCV and BKV could be important co-factors in tumor pathogenesis., ((c) 2005 Wiley-Liss, inc.)

JC polyomavirus (JCPyV) causes progressive multifocal leukoencephalopathy (PML) in patients with decreased immune competence. To elucidate genetic changes in JCPyV associated with the pathogenesis of PML, multiple complete JCPyV DNA clones originating from the brains of three PML cases were established and sequenced. Although unique rearranged control regions occurred in all clones, a low level of nucleotide variation was also found in the coding region. In each case, a parental coding sequence was identified, from which variant coding sequences with nucleotide substitutions would have been generated. A comparison between the parental and variant coding sequences demonstrated that all 12 detected nucleotide substitutions gave rise to amino acid changes. Interestingly, seven of these changes were located in the surface loops of the major capsid protein (VP1). Finally, 16 reported VP1 sequences of PML-type JCPyV (i.e. derived from the brain or cerebrospinal fluid of PML patients) were compared with their genotypic prototypes, generated as consensus sequences of representative archetypal isolates belonging to the same genotypes; 13 VP1 proteins had amino acid changes in the surface loops. In contrast, VP1 proteins from isolates from the urine of immunocompetent and immunosuppressed patients rarely underwent mutations in the VP1 loops. The present findings suggest that PML-type JCPyV frequently undergoes amino acid substitutions in the VP1 loops. These polymorphisms should serve as a new marker for the identification of JCPyV isolates associated with PML. The biological significance of these mutations, however, remains unclear.

Recently genotyping of JC virus (JCV) DNA in renal tissue was reported to be useful to identify the geographic origin of unidentified cadavers. In the above study, autopsied tissue samples without storage or stored in a frozen state were used. This study examined JCV DNA sequence modifications caused by formalin-fixation, in an attempt to elucidate whether formalin-fixed, paraffin-embedded tissue samples can also be used to determine the genotypes of JCV DNA in the kidney. In four cases, a 610 bp typing region of the JCV genome was PCR-amplified from renal tissues stored for 1 year in three different states: frozen at -80 degrees C [Amaker, B.H., Chandler, F.W., Huey, L.O., Colwell, R.M., 1997. Molecular detection of JC virus in embalmed, formalin-fixed, paraffin-embedded brain tissue. J. Forensic Sci., 1157-1159], formalin-fixed, paraffin-embedded [Ault, G.S., Stoner, G.L., 1992. Two major types of JC virus defined in progressive multifocal leukoencephalopathy brain by early and late coding region DNA sequences. J. Gen. Virol. 73, 2669-2678], and soaked in 5% formalin [Baksh, F.K., Finkelstein, S.D., Swalskey, P.A., Stoner, G.L., Ryschkewitsch, C.F., Randhawa, P.R., 2001. Molecular genotyping of BK and JC virus in human polyomavirus-associated interstitial nephritis after renal transplantation. Am. J. Kidney Dis. 38 (2), 354-365]. The amplified fragments were cloned, and the resultant clones were sequenced. In frozen samples, single sequences ('original' sequences) were detected in all cases. In formalin-fixed, paraffin-embedded samples, not only the original sequences but also those with 1-6 base substitutions were detected. From formalin-soaked samples, the original sequences and those with 1-5 and 10-13 substitutions were detected. The genotyping of JCV DNA was not hampered by the presence of 1-6 substitutions, but a shift in JCV genotypes was observed in sequences with 10-13 substitutions. Thus, it was concluded that the genotypes of JCV DNA in the kidney can be determined only with specimens stored in a frozen state or formalin-fixed for a short time.

JC virus (JCV) is the cause of progressive multifocal leukoencephalophathy (PML) in immunocompromised patients. The paucity of reports from Africa has led to the hypothesis that PML is rare because of an absence of virus genotypes associated with the condition. Genotypes 3 and 6 have been identified in East and West Africa but the distribution of types across the rest of Africa is unknown. Full-length sequences of five JCV cerebrospinal fluid samples from PML patients in South Africa are reported here. Three isolates from African AIDS patients grouped with type 3A or 3B, and one with type 7, while one from a Caucasian leukaemia patient grouped with type 2D. Widespread distribution of type 3 on the continent may reflect migration patterns in antiquity, but this is the first report of type 7 in an African individual. Type 2D has only been isolated previously in South Asia, although transmission of this genotype to Europeans who later settled in South Africa is not unlikely.

We have further characterized the Asian genotypes (Types 2 and 7) and subtypes of JC virus (JCV). Urine samples from 224 individuals with Han and Mongolian populations were collected in five regions in eastern China: Kunming, Chengdu, Shenyang, Chifeng, and Manzhouli. Also, 99 urine samples were collected from coastal and hill groups in Kerala, southern India, and 23 urine samples from Seoul, Korea. PCR products of four typing fragments were sequenced, including two in the VP1 gene, as well as one each in the VT intergenic region and regulatory region. It was possible to clone and sequence a total of 42 JCV whole genomes (approximately 5120 bp). Five genotypes of JCV (Types 7A, 7B, 7C, 2D, and 4) were found in China, four genotypes (Types 2D, 7C, 4, and 1B) in southern India, and three genotypes (Types 7B, 2A, and 1A) in Korea. Type 7A was most prevalent in South China (59-64%) and Type 7B was predominant in northeast China and Inner Mongolia (67-77%). Type 7C strains were spread throughout North and South China (3-14%), while Type 2D strains were found only in the two Mongolian groups (9-10%). In southern India, Type 2D was predominant in the coastal group (95%), and two major types, Type 7C (50%) and Type 2D (35%), were prevalent in the tribal hill groups. In Korea two major genotypes were found: Type 7B (50%) and Type 2A (43%). Phylogenetic reconstruction places the Chinese genotypes in the Afro-Asiatic supercluster, but distinct from the Mongolian and Indian strains (Type 2D), as well as the Korean and Japanese genotype (Type 2A) that predominates in the Americas.