A prospective longitudinal study of polyomavirus shedding in lung-transplant recipients

BACKGROUND: Polyomavirus infection causes renal dysfunction after kidney transplantation, but it has not been thoroughly investigated in nonrenal solid-organ transplantation. METHODS: Fifty lung-transplant recipients provided prospective urine and blood samples over the course of 17 months. Samples were analyzed for BK virus (BKV), JC virus (JCV), and simian virus 40 (SV40) using conventional polymerase chain reaction (PCR), sequence analysis, and quantitative real-time PCR. RESULTS: Thirty-one (62%) of 50 patients had polyomavirus detected in at least 1 urine specimen, including 16 (32%) for BKV, 12 (24%) for JCV, and 6 (12%) for SV40. Mean BKV loads (5.0 log(10) copies/mL) did not differ from those of JCV (5.7 log(10) copies/mL; P=.38), but SV40 loads (2.5 log(10) copies/mL) were lower than those of BKV (P=.006) and JCV (P=.002). Blood samples were negative. Infection with individual polyomaviruses or polyomavirus infection in aggregate was not associated with reduced creatinine clearance. Patients not shedding polyomavirus had better survival than patients shedding polyomavirus (P=.049). CONCLUSIONS: Polyomaviruses BKV and JCV were commonly detected in urine from lung-transplant recipients. SV40 was found in 12% of patients but was shed at a lower frequency and with lower viral loads than the other viruses. Polyomavirus infection was not associated with renal dysfunction.     

JC polyomavirus nephropathy confirmed by using an in‐house polymerase chain reaction method

We report the case of an isolated JC virus (JCV) infection, without co‐infection by polyoma BK virus (BKV), associated with nephropathy 4 years after kidney transplantation. Clinical suspicion followed the observation of a decrease in estimated glomerular filtration rate (eGFR) and a renal allograft biopsy revealing polyomavirus‐associated tubulointerstitial nephritis and positivity for SV40. An in‐house real‐time polymerase chain reaction assay, targeting the presence of JCV and the absence of BKV in biopsy tissue, confirmed diagnosis. Thirteen months after diagnosis, and following therapeutic measures, eGFR remains stable.     

Polyomavirus JC Urinary Shedding in Kidney and Liver Transplant Recipients Associated With Reduced Creatinine Clearance

Background.?Polyomavirus reactivation can cause significant morbidity in solid organ transplant recipients, particularly BK virus (BKV) in kidney transplant patients. Less is known about dynamics of John Cunningham virus (JCV) in nonkidney organ transplant patients. Methods.?We examined the frequency of urinary shedding of polyomaviruses BKV and JCV and their relationship to creatinine clearance (CrCl) in a longitudinal study of 41 kidney and 33 liver transplant recipients. Results.?Any polyomavirus urinary shedding was more frequent in liver than kidney recipients (64% vs 39%; P?=?.03). JCV was excreted more frequently by liver than kidney recipients (71% vs 38%), whereas BKV was shed more often by kidney than liver patients (69% vs 52%). Mean JCV loads were significantly higher than those of BKV in both patient groups (P?相似文献   

The dynamics of herpesvirus and polyomavirus reactivation and shedding in healthy adults: a 14-month longitudinal study

Humans are infected with viruses that establish long-term persistent infections. To address whether immunocompetent individuals control virus reactivation globally or independently and to identify patterns of sporadic reactivation, we monitored herpesviruses and polyomaviruses in 30 adults, over 14 months. Epstein-Barr virus (EBV) DNA was quantitated in saliva and peripheral blood mononuclear cells (PBMCs), cytomegalovirus (CMV) was assayed in urine, and JC virus (JCV) and BK virus (BKV) DNAs were assayed in urine and PBMCs. All individuals shed EBV in saliva, whereas 67% had >or=1 blood sample positive for EBV. Levels of EBV varied widely. CMV shedding occurred infrequently but occurred more commonly in younger individuals (P<.03). JCV and BKV virurias were 46.7% and 0%, respectively. JCV shedding was age dependent and occurred commonly in individuals >or=40 years old (P<.03). Seasonal variation was observed in shedding of EBV and JCV, but there was no correlation among shedding of EBV, CMV, and JCV (P>.50). Thus, adults independently control persistent viruses, which display discordant, sporadic reactivations.     

A kidney transplant recipient with renal medullary viral cytopathic changes

We present a case of JC polyomavirus (JCV)‐associated nephropathy (PyVAN) in an asymptomatic deceased‐donor kidney transplant recipient. Despite the presence of viral cytopathic effect in the kidney biopsy and positive BK polyomavirus (BKV) in situ hybridization (ISH), BKV real‐time polymerase chain reaction (PCR) results of plasma and urine were negative. JCV ISH was performed and was found to be positive. JCV real‐time PCR on urine, plasma, and the kidney biopsy tissue was positive. Reduction in immunosuppression resulted in resolution of JCV viremia. This case highlights that JC‐PyVAN is a distinct clinical entity and is likely to have a better clinical outcome than BK‐PyVAN. Concurrent infection with BKV and JCV may occur, but may be difficult to confirm due to the potential for cross‐reactivity between BKV and JCV ISH stains.     

BKV-DNA and JCV-DNA in CSF of Patients with Suspected Meningitis or Encephalitis

Abstract. Background: Few studies have looked for the polyoma viruses JC or BK virus in the central nervous system (CNS) of patients without neurological symptoms or with neurological symptoms other than progressive multifocal leukoencephalopathy (PML). PCR-microplate hybridization method was employed for the detection of BKV-DNA or JCV-DNA in cerebrospinal fluid (CSF) specimens from patients with suspected meningitis or encephalitis. Materials and Methods: A total of 181 CSF specimens from 151 patients with suspected meningitis or encephalitis was examined for BKV or JCV using PCR-microplate hybridization method. None of the patients had (clinically diagnosed) PML. A control group consisting of 20 CSF specimens from normal subject was also included. Results: BKV DNA was found in five out of 131 (3.8%) and JCV DNA in two out of 131 (1.5%) of the patients with suspected meningitis or encephalitis by PCR ELISA. BKV or JCV DNA was not detected in CSF samples of any of 19 HIVpositive patients. BKV and JCV DNAs were detected respectively in two CSF samples in which Mycobacterium tuberculosis (TB) PCR was also positive. Another patient who was positive for JCV PCR died with a diagnosis of cerebral lymphoma. Among the BK virus infected patients there was a patient with a previous history of hemolytic uremia and acute renal failure. Neither BKV nor JCV DNA was found in any of the 20 CSF samples from normal patients undergoing lumbar puncture for myelography as a part of an investigation of lower back pain. Conclusion: These results suggest that BK virus may be associated with neurological diseases either in immunocompetent or immunocompromised patients. Detection of BKV and JCV DNA in the CSF of the patients suspected to have either meningitis or encephalitis suggests that these viruses may have an etiological role. Thus, diagnostic tests for BK and JC viruses should be included in the investigative program for meningitis or encephalitis patients.     

SV40 and HIV sequences in the cerebrospinal fluid of a patient with AIDS dementia complex

Central Nervous System (CNS) diseases that occur in HIV-positive patients are mainly due to HIV itself or to opportunistic microorganisms. Polyomavirus JCV, BKV and SV40 have been associated with encephalopathies in both HIV-positive and HIV-negative patients. To investigate the presence of Polyomavirus DNA sequences in patients affected by CNS disorders, 82 CSF samples from 70 HIV-positive and 12 HIV-negative patients were analyzed by PCR. A double HIV and SV40 infection was found in one patient suffering with AIDS dementia complex. SV40 DNA sequence analysis showed the homology with wild type SV40 strain. SV40 should be considered as a potential causal agent of CNS disorders in AIDS patients.     

JC virus DNA sequences are frequently present in the human upper and lower gastrointestinal tract

BACKGROUND & AIMS: JC virus (JCV), a human polyomavirus, has been found in a limited number of normal human tissues and cancers. The oncogenic potential of this virus is mediated by a transforming protein, the T antigen (TAg). We have previously demonstrated the presence of JCV-TAg in colorectal cancers, in adjacent normal colonic mucosa from these patients, and in the human colon cancer cell line SW480. The mode of transmission of this virus is unclear, and we hypothesized that the gastrointestinal (GI) tract may be a reservoir for the virus. METHODS: DNA was extracted from 129 normal GI tissue samples collected from 33 patients. Topoisomerase I-assisted polymerase chain reaction (PCR) was used to detect the virus using exact and degenerate primers. Nested PCR and Southern blot analysis confirmed the identity of the PCR products. Single-stranded conformation polymorphism (SSCP) analysis and sequencing were used to evaluate the presence of viral quasispecies. RESULTS: JCV sequences were found in 75.8% of patients (70.6% of upper GI and 81.2% of colonic samples); no significant differences in rates of infection were found by site. The use of degenerate primers combined with topoisomerase I treatment led to viral detection in 58.9% of samples, compared with 27.9% of samples using exact primers and topoisomerase I (P < 0.01). SSCP and sequencing analysis confirmed the amplification of viral quasispecies and the authenticity of TAg sequences. CONCLUSIONS: The results show that JCV DNA sequences are highly prevalent in the human upper and lower gastrointestinal tract of immunocompetent individuals.     

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.     

Evaluation of a quantitative real-time PCR assay for the detection of JC polyomavirus DNA in cerebrospinal fluid without nucleic acid extraction

The JC polyomavirus (JCV) is the causative agent of progressive multifocal leukoencephalopathy (PML), a fatal demyelinating disease. The current diagnostic standard for PML is real-time PCR testing of extracted DNA for assessing the presence of JCV DNA in cerebrospinal fluid (CSF). This study was aimed at evaluating the feasibility of a real-time PCR assay without nucleic acid extraction for the rapid quantification of JCV DNA in CSF. CSF samples were heat-treated or treated with DNAzol Direct, a commercially available reagent for direct PCR, and the performances of the real-time PCR assays using templates obtained by either treatment were compared with that using DNA extracts. JCV DNA was detected in the heat- or DNAzol Direct-treated samples containing only a few copies of the viral genome per reaction, and a linear relationship was noted between the copy number detected and the amount of input virus ascertained by the DNA extraction method. The sensitivities of the assays using the heat and DNAzol Direct treatments were 85.7 and 90.5%, respectively, with the results of the DNA extraction method being used as reference. These data demonstrate that the real-time PCR assay introduced in this study can serve as a rapid and cost-effective method of testing for JCV without DNA extraction and thereby facilitate the assessment of PML.     

Adenovirus is a key pathogen in hemorrhagic cystitis associated with bone marrow transplantation.

Late-onset hemorrhagic cystitis (HC) is a well-known complication of bone marrow transplantation (BMT) that is mainly attributed to infection with BK virus (BKV) and adenovirus (AdV). From 1986 through 1998, 282 patients underwent BMT, and 45 of them developed HC. Urine samples tested positive for AdV in 26 patients, of which 22 showed virus type 11. Among patients who underwent allogeneic BMT, logistic regression analysis revealed acute graft-versus-host disease (grade, > or = 2) to be the most significant predictive factor for HC (P < .0001). In addition, a total of 193 urine samples regularly obtained from 26 consecutive patients who underwent allogeneic BMT were examined for BKV, JC virus (JCV), and AdV by means of polymerase chain reaction. Of patients without HC, approximately 30% of the specimens tested positive for BKV (58 samples) and JCV (55 samples), whereas 5 (3%) tested positive for AdV. Of the 3 samples obtained from patients with HC, the numbers of positive results for BKV, JCV, and AdV were 3, 1, and 1, respectively; the numbers of positive results increased to 14 of 17, 9 of 17, and 10 of 17, respectively, when we added another 14 samples obtained from 14 patients with HC (P < .0001, P = .026, and P < .0001, respectively). In conclusion, there was significant correlation between AdV and HC in the patients we studied.     

Prospective monitoring of BK virus reactivation in renal transplant recipients in North India

R. Thakur, S. Arora, R. Nada, M. Minz, K. Joshi. Prospective monitoring of BK virus reactivation in renal transplant recipients in North India.
Transpl Infect Dis 2011: 13: 575–583. All rights reserved Abstract: Background. BK nephropathy (BKN) is an important complication of renal transplantation with a reported incidence between 1% and 10% in different parts of the world. Early diagnosis is important to plan early therapeutic strategies. The epidemiology and evolution of BKN is relatively unknown in India and hence, the present study has been designed to prospectively monitor the activation of BK virus (BKV) in renal transplant recipients in India. Patients and methods. In this study, 32 renal allograft recipients were prospectively monitored with protocol biopsies of allografts, BKV DNA load in plasma, and viral particles in urine by electron microscopy (EM) on day 1, and at 1, 3, and 6 months. Additionally, the baseline BKV DNA load in plasma was quantitated in 21 corresponding donors. Results. On follow‐up in 32 recipients, 9.7%, 23.8%, 19.2%, and 13.3% of patients showed viral profiles by EM at day 1, 1 month, 3 months, and 6 months, respectively. BKV DNA positivity in plasma was 25.8%, 42.9%, 15.4%, and 20% at day 1, 1 month, 3 months, and 6 months, respectively, with mean BKV copy number/mL plasma of 1796, 1029, 2611, and 3318, respectively. A total of 15.7% (17/108) urine samples of 32 renal recipients were positive by urine EM. Out of 100 protocol biopsies, none developed histologically demonstrable cytopathic effects of BKN, although 8% biopsies were SV‐40 large T antigen (SV‐40 T Ag) positive. By quantitative real‐time polymerase chain reaction assay, 27/108 (25%) of recipients' plasma samples were positive for BKV. Peak viremia and viruria occurred at 1–3 months post transplantation. The baseline viremia in donors was predictive of viremia positivity in the post‐transplantation period at 1 month. Twenty‐four episodes of graft dysfunction were attributed mainly to rejection. Conclusion. The study shows a total of 15.7% and 25% urine and plasma samples were positive for BKV at any time during a 6‐month follow‐up. The highest incidence of BK viruria and viremia occurred at 1 month. In protocol biopsies, focal positivity of SV‐40 T Ag was seen in 8% biopsies.     

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.     

Detection of human neurotropic JC virus DNA sequence and expression of the viral oncogenic protein in pediatric medulloblastomas

Medulloblastoma represents greater than 25% of childhood intracranial neoplasms and is considered a highly malignant tumor. This tumor, which arises predominantly in the cerebellar vermis, preferentially affects children between the ages of 5 and 15. Although the etiology of medulloblastomas in humans remains unknown, results from several experiments have indicated that the human neurotropic JC virus (JCV) is able to induce cerebellar neoplasms in rodents that exhibit a phenotype similar to that of human medulloblastomas. JCV is a polyomavirus that is widespread in the human population, with infection occurring most frequently in early childhood. In this study, we have examined the possible association of JCV with human medulloblastomas. By using PCR techniques we demonstrate that 11 of 23 samples of tumor tissue contain DNA sequences corresponding to three different regions of the JCV genome. More importantly, we demonstrate the presence of DNA sequences encoding the N- and C-terminal regions of the JCV oncogenic protein, T antigen, in 11 of 23 samples and the production of T antigen in the nuclei of 4 samples of tumor tissue. These observations provide evidence for a possible association of JCV with human medulloblastomas.     

A longitudinal molecular surveillance study of human polyomavirus viremia in heart, kidney, liver, and pancreas transplant patients

In this study of 263 heart, kidney, liver, and pancreas transplant patients, BK virus (BKV) and JC virus (JCV) DNAemia were observed most commonly in kidney and/or pancreas transplant patients (26%), although they were also observed, to a lesser extent, in heart (7%) and liver (4%) transplant patients. The majority of episodes of polyomavirus DNAemia were subclinical, although, in some cases, BKV DNAemia was associated with kidney rejection, and JCV DNAemia was accompanied by nonspecific symptoms. Hence, BKV and JCV DNAemia are not uncommon during the first year after kidney, heart, liver, and pancreas transplantation, and they could be associated with certain clinical syndromes in transplant patients.     

Quantitation of DNA of polyomaviruses BK and JC in human kidneys

BACKGROUND: Renal allograft recipients can be monitored for polyomavirus-associated nephropathy (PVAN) using urine samples. Because virus in urine can be derived from the kidneys, ureter, or urinary bladder, we evaluated whether measurement of intrarenal concentrations of viral DNA might serve as a more reliable monitoring tool. METHODS: Real-time quantitative polymerase chain reaction was used to quantitate DNA of polyomaviruses BK (BKV) and JC (JCV) in renal tissue obtained from various clinical settings. RESULTS: Renal biopsy samples from 28 nonimmunosuppressed patients contained very low viral copy numbers. Minimally higher BKV loads (mean +/- SE, 3.4 +/- 1.7 copies/cell) were observed in 74 renal biopsy samples from renal allograft recipients with BKV viruria. The BKV DNA concentration was approximately 10-fold higher in renal allograft recipients with BKV viruria, but 58 (50.4%) of 115 renal biopsy samples tested negative for BKV DNA, reflecting the focal nature of infection. JCV DNA was found in only 2 renal biopsy samples. CONCLUSIONS: The BKV load is better measured in urine than in tissue, because a urine sample represents material from the entire kidney. An increase in the BKV load is usually not accompanied by a proportional increase in the JCV load, which indicates that these 2 related polyomaviruses are subject to different mechanisms regulating viral replication.     

BK and JC viruses in patients with systemic lupus erythematosus: prevalent and persistent BK viruria, sequence stability of the viral regulatory regions, and nondetectable viremia.

A role for polyomaviruses in the pathogenesis of systemic lupus erythematosus (SLE) has been suggested. BK virus (BKV) and JC virus (JCV) were demonstrated in single urine specimens from 7 (16%) of 44 and 5 (11%) of 44 patients with SLE and 0/88 and 18 (21%) of 88 matched healthy controls, respectively. During a 1-year follow-up study, episodes of polyomaviruria were detected in 16 (80%) of 20 patients, BKV in 13, and JCV in 3 patients. A group of 12 (60%) of 20 patients demonstrated persistent or recurrent polyomaviruria, BKV viruria (n=9), or JCV viruria (n=3) in 180 (70%) of 256 specimens. Polyomaviruria was not significantly associated with immunosuppressive therapy. The BKV and JCV isolates revealed predominantly stable archetypal regulatory regions over 3 years, indicating viral persistence rather than reinfection as a cause for urinary shedding. The demonstration of nondetectable viremia and stable archetypal BKV and JCV noncoding control regions during persistent viruria argue against the urinary tract as a focus for the creation of rearranged regulatory region variants.     

Persistence of DNA sequences of BK virus and JC virus in normal human tissues and in diseased tissues

Available evidence suggests that BK virus (BKV) and JC virus (JCV) persist in the kidneys of healthy individuals after primary infection and may reactivate when the host's immune response is impaired. Data supporting this hypothesis are presented. A previous study had shown BKV to be present in the kidneys of eight (57%) of 14 subjects. In the present study, which extended the investigation to a total of 30 subjects, BKV DNA was found in the renal tissues of 10 (33%) subjects, and JCV DNA was found in the renal tissues of three (10%) subjects. The viral DNA detected appeared not to be integrated with host DNA and to be isolated in foci. Investigation of normal and diseased brain tissue, including tissue from six subjects with multiple sclerosis, failed to reveal the presence of either JCV DNA or BKV DNA.     

Prevalence of infection by JC and BK polyomaviruses in kidney transplant recipients and patients with chronic renal disease

E.P. Pires, C.V. Bernardino‐Vallinoto, D.M. Alves, S.R.C. Migone, L.F.A. Machado, M.O.G. Ishak, R. Ishak, I.M.V. Cayres‐Vallinoto, A.C.R. Vallinoto. Prevalence of infection by JC and BK polyomaviruses in kidney transplant recipients and patients with chronic renal disease.
Transpl Infect Dis 2011: 13: 633–637. All rights reserved Abstract: The present study investigated the prevalence of infection by JC and BK polyomaviruses (JCV and BKV) in patients with chronic renal disease (CRD), kidney transplant recipients, and a control group of asymptomatic subjects. We tested a total of 295 urine samples. After DNA extraction, polymerase chain reaction assay was used to amplify a fragment of 173 bp of the polyomavirus T antigen, followed by analysis using the BamHI restriction endonuclease. Infection by polyomavirus was detected in 17.6% (52/295 subjects) of the subjects. Whereas 30.5% (18/59) of transplant recipients were infected, the frequency was only 22.4% (30/134) in the control subjects, and 3.9% (4/102) in the CRD group (all JCV). The vast majority of infections (88.9%; 16/18) in transplant recipients were of the BKV type, whereas this type was absent in CRD patients, and made up only 10.0% (3/30) of infections in the control group. The risk of BKV infection was 72 times greater in renal transplant patients than in asymptomatic subjects. The low frequency of infection found in CRD patients may have been related to elevated levels of urea excreted in the urine, together with reduced urine volume and cell content. These factors may combine to reduce viral load or inhibit amplification. The results of the study indicate a need for the routine screening for polyomavirus in pre‐ and post‐transplant patients, as well as organ donors, considering that BKV infection has been associated with graft rejection in kidney transplants.