Polyomavirus Nephropathy Research Articles

Urine cytology screening for polyoma virus infection following renal transplantation: the Oxford experience

Objective: To review the first year of a monthly urine cytology screening service, introduced to identify renal transplant patients at risk of polyoma virus nephropathy (PVN), at an early, potentially...

Polyomavirus nephropathy in native kidneys and renal allografts: an update on an escalating threat

Polyomavirus nephropathy, also termed BK-virus nephropathy (BKN) after the main causative agent, the polyoma-BK-virus strain, is a significant complication after kidney transplantation. BKN is the most common viral infection that affects renal allografts with a prevalence of 1-9% on average 8-13 months post surgery. It can also occur sporadically in native kidneys. Viral nephropathy is caused by the (re)activation of latent BK viruses that enter into a replicative cycle under sustained and intensive immunosuppression. Pure productive kidney infections with JC- and SV-40 polyomaviruses are exceptionally rare. BKN is morphologically defined by the presence of intranuclear viral inclusion bodies in epithelial cells and tubular injury, which is the morphological correlate for renal dysfunction. Renal disease can progress through different histologic stages (from early BKN stage A to late fibrotic stage C) that carry prognostic significance; disease stages B and C often result in chronic kidney (allograft) dysfunction and end-stage renal disease. The clinical goal is to diagnose viral nephropathy in disease stage A and to limit chronic renal injury. Strategies to recognize, classify, and manage BKN are critically discussed including ancillary techniques for risk assessment and patient monitoring: (i) urine cytology and the search for so-called 'decoy cells'; (ii) PCR analyses for viral load measurements in the plasma and urine; and (iii) negative staining urine electron microscopy to identify viral particles.

Urine Cytology as a Useful Screening Method for Polyoma Virus Nephropathy in Renal Transplant Patients: A Single-Center Experience

Polyoma virus nephropathy occurs in 3% to 4% of renal transplant recipients, causing graft loss in 50% of cases. In this study we sought to identify the incidence of polyoma virus infection among our transplanted patients on the basis of age, sex, creatinine level, and postoperative period. During this study the 1086 urine samples collected from 362 patients were centrifuged and stained with the Papaniclaou method. All slides were classified as negative or positive (>1 decoy cell/sample). Among 1086 urine cytologies from 241 men and 121 women, decoy cells were identified in 26.6% (96) of patients, including 29.9% ( n = 72) men and 20% ( n = 24) women. The incidence of decoy cells (26.6%) was increased among men and associated with a longer transplantation period ( P < .05). A significant relation was detected between older age and positive urine cytology. The patients with positive urine cytology for decoy cells showed a greater incidence of abnormal plasma creatinine values (26%) compared with patients showing a negative urine cytology (13.5%). In conclusion, identification of cells with viral inclusions (decoy cells) may help with the diagnosis of viral replication or active infection, therefore, routine urine cytology may be used as screening method for the detection of polyoma virus infection.

Inhibitory Effect of Gamma Interferon on BK Virus Gene Expression and Replication

BK virus (BKV) is widely accepted to be the causative agent of polyomavirus nephropathy. In immunocompromised individuals, especially kidney transplant recipients, BKV can replicate in kidney epithelial cells, causing loss of renal function and eventual destruction of the graft. Advances in immunosuppressive therapies may be partially responsible for the increasing incidence of polyomavirus nephropathy among transplant recipients by more effectively eliminating components of the immune system, such as gamma interferon (IFN-gamma)-producing lymphocytes, that keep BKV infections at a subclinical level. In this study, we investigated the role of IFN-gamma in regulating lytic infection by BKV. Treatment with IFN-gamma inhibited the expression of the viral early protein large tumor antigen (TAg) and the late protein VP1 in a dose-dependent manner. We detected 1.6- and 12-fold reductions in TAg transcripts at 48 and 96 h postinfection, respectively, with 250 U/ml IFN-gamma, suggesting that IFN-gamma-mediated inhibition occurs at the level of transcription. Furthermore, IFN-gamma inhibited the level of viral progeny production as much as 50-fold at a multiplicity of infection (MOI) of 0.5 and 80-fold at an MOI of 0.1. The inhibitory effects of IFN-gamma were similar for three different strains of BKV examined. These results indicate an important role for IFN-gamma in regulating BKV lytic infection.

Nephron segment localization of polyoma virus large T antigen in renal allografts

This study uses CD10 and epithelial membrane antigen (EMA) as respective proximal and distal tubular segment markers to localize polyoma virus replicative activity, as determined by large T antigen (TAg) expression, in allograft polyoma virus nephropathy (PVN). Sixteen biopsies and 2 nephrectomy specimens with PVN had serial 2-mum paraffin sections stained using monoclonal antibodies to polyoma virus TAg by immunoperoxidase with diaminobenzidine as chromogen. A second immunolabeling step used CD10 as a proximal nephron marker or EMA as a distal tubular marker, and alkaline phosphatase with fast red as chromogen. BK viral DNA was detected in blood in 16 of 18 tested. Fourteen of 16 had cortex and medulla, and 2 had cortex only in the biopsy sample. Fourteen (100%) of 14 had double-positive EMA and TAg expression (EMA+TAg+) in medullary collecting ducts. T antigen was evident in loops of Henle in nephrectomies. Thirteen (81.3%) of 16 had EMA+TAg+, and 10 (62.5%) of 16 had CD10+TAg+ cortical tubular segments. CD10+TAg+ cells were observed in the parietal epithelium of Bowman's capsule in 3 biopsies (18.75%). T antigen was observed in 5.1% of CD10+ tubular profiles compared with 21% of EMA+ profiles in renal cortex (P < .0001). Polyoma virus TAg was observed in medullary collecting ducts, in distal and proximal cortical tubules, and in Bowman's capsule, in decreasing order of frequency. Loops of Henle also had TAg. This distribution suggests potential for an ascending route of infection of allograft tubules in PVN.

BKV Replication and Cellular Immune Responses in Renal Transplant Recipients

BKV Replication and Cellular Immune Responses in Renal Transplant Recipients

Polyomavirus Nephropathy in the Native Kidneys of an Unrelated Cord Blood Transplant Recipient Followed by A Disseminated Polyomavirus Infection

Polyomavirus Nephropathy in the Native Kidneys of an Unrelated Cord Blood Transplant Recipient Followed by A Disseminated Polyomavirus Infection

Urinary HLA-DR and CD54 expression—indicators for inflammatory activity in decoy cell shedding patients

Polyomavirus (PV) nephropathy may coexist with or follow acute renal transplant rejection. The aim of this study was to evaluate whether HLA-DR and CD54 are useful cellular markers for surveillance of acute rejection in PV-infected patients. A prospective study was conducted using 205 renal transplant patients. Urine samples were collected at a regular interval post-transplantation for routine cytology and immunocytochemistry. Urinary levels of tumour necrosis factor alpha, soluble interleukin-2 receptor and interleukin-6 were used as adjunctive markers for acute rejection. Of the 699 total samples, decoy cells were identified in 100 samples of 50 patients. Patients with decoy cell-positive (DCP) samples had higher serum creatinine levels than decoy cell-negative (DCN) samples (1.55 vs 1.41 mg/dl, respectively; P = 0.006). DCP samples were also more likely to be HLA-DR positive (50.0 vs 32.4%; P = 0.029), as well as CD54 positive (17.4 vs 6.9%; P = 0.038). However, serum creatinine levels did not correlate with HLA-DR or CD54 positivity among DCP samples. Instead, CD54 positivity correlated with decoy cell grades. Immunosuppression decreased in 11 DCP patients, and HLA-DR was negatively converted in three of them. None of the patients developed acute clinical rejection. Urinary cytokine levels did not correlate with serum creatinine levels, nor did they correlate with HLA-DR or CD54 status among DCP patients. Urinary tubular HLA-DR and CD54 expression increased in decoy cell shedding patients but did not indicate a concomitant acute rejection. These markers may instead indicate renal inflammatory activity associated with viral reactivation, which has the potential to progress to PV interstitial nephritis.

Expression of p53 in virally infected tubular cells in renal transplant patients with polyomavirus nephropathy

Polyomavirus (PV) infection is associated with ureteral stenosis, hemorrhagic cystitis, and interstitial nephritis in renal transplant patients. The 3 PVs detected in human beings-BK virus, JC virus, and simian virus 40-each encode highly homologous forms of a large T antigen, a transcriptional and replicational regulatory protein. We describe immunohistochemical findings in 5 renal transplant patients who developed PV nephropathy (PVN) and a sixth patient with both PVN and PV infection of the bladder mucosa. Polyomavirus infection was confirmed by immunohistochemical detection of T antigen in kidney and bladder biopsies. We report on the expression of p53 specific to virally infected cells in all biopsies positive for T antigen. Examination of posttransplant biopsies obtained from these 6 patients before they were diagnosed with PVN revealed no expression of T antigen or p53. Accumulation of p53 in PV-infected cells may occur in response to binding of p53 by T antigen, resulting in stabilization of p53. These results provide the first evidence for intracellular actions of PV T antigen in the context of nonneoplastic diseases.

The prevalence of BK viremia and urinary viral shedding in a pediatric renal transplant population: A single‐center retrospective analysis

Polyomavirus-induced nephropathy has emerged as an important cause of renal graft dysfunction. Limited pediatric data are available for this disease. We therefore reviewed the results of the first year of polyomavirus screening in our pediatric renal transplant recipients to determine the prevalence of polyomavirus viremia and urinary shedding. Screening included detection of polyomavirus in plasma by polymerase chain reaction (PCR) and in urine by electron microscopy (EM). In patients with a positive screening test, an assessment of graft dysfunction was made. Fifty-two patients met the inclusion criteria. Urinary EM was performed in 205 samples and polyomavirus was detected in 10 patients, representing 19% of the study population. PCR was performed on 222 samples and was positive for the BK virus in plasma from seven patients or 13.4% of the study population. Eight patients had a positive screening test and increased creatinine. All these patients underwent renal transplant biopsy. This revealed evidence of polyomavirus nephropathy in four patients. Our findings reveal a high prevalence of polyomavirus in both urine and plasma that is frequently associated with graft dysfunction. These findings support the routine screening of pediatric post-renal transplant patients for polyomavirus replication.

No indication for activation of exogenous retroviruses in patients with renal allograft rejection

Reactivation of latent BK virus in kidney-transplanted patients results in severe graft dysfunction. The role of retroviruses infecting also latently target cells is not investigated so far in this setting. We determined the presence or induction of retroviruses in sera of immunosuppressed patients with renal allografts at the timepoint of organ rejection or ongoing polyomavirus nephropathy. Sera of patients with acute kidney rejection or polyomavirus nephropathy (n=25) and controls (n=8) were tested for reverse transcriptase activity by the ultrasensitive product enhanced reverse transcriptase (PERT) assay. In parallel, kidney biopsies were investigated for histological signs of kidney rejection or polyomavirus nephropathy confirmed by either immunofluorescence or immunohistochemistry. None of the investigated sera, specifically those of patients with ongoing BK virus nephropathy, indicated reverse transcriptase activity. Our results do not support the idea of the induction of known or unknown retroviruses in patients with kidney transplantation, even under highly immunosuppressive therapies.

Renal transplant patient with polyoma virus bladder infection and subsequent polyoma virus nephropathy

Polyoma virus nephropathy (PVN) is a significant cause of renal allograft dysfunction in transplant patients. A 58-year-old male received a cadaveric renal transplant and 12 weeks later presented with fever, diarrhea, and dysuria. He was diagnosed with a polyoma virus infection of the bladder by a transurethral bladder biopsy. One year post-transplant, he presented with renal allograft dysfunction and was diagnosed by biopsy with PVN of the non-native kidney. The diagnosis of a polyoma virus infection was confirmed by immunoreactivity to the polyoma T-antigen. We suggest that polyoma virus infection of the bladder be included in the differential diagnosis of urinary dysfunction in post-transplant patients, as such infections might be an under-recognized comorbidity in individuals with PVN.

Medical Care of Kidney Transplant Recipients after the First Posttransplant Year

Kidney transplantation is the treatment of choice for patients with ESRD. Despite improvements in short-term patient and graft outcomes, there has been no major improvement in long-term outcomes. The use of kidney allografts from expanded-criteria donors, polyoma virus nephropathy, underimmunosuppression, and incomplete functional recovery after rejection episodes may play a role in the lack of improvement in long-term outcomes. Other factors, including cardiovascular disease, infections, and malignancies, also shorten patient survival and therefore reduce the functional life of an allograft. There is a need for interventions that improve long-term outcomes in kidney transplant recipients. These patients are a unique subset of patients with chronic kidney disease. Therefore, interventions need to address disease progression, comorbid conditions, and patient mortality through a multifaceted approach. The Kidney Disease Outcomes Quality Initiative from the National Kidney Foundation, the European Best Practice Guidelines, and the forthcoming Kidney Disease: Improving Global Outcomes clinical practice guidelines can serve as a cornerstone of this approach. The unique aspects of chronic kidney disease in the transplant recipient require the integration of specific transplant-oriented problems into this care schema and a concrete partnership among transplant centers, community nephrologists, and primary care physicians. This article reviews the contemporary aspects of care for these patients.

Interplay of Cellular and Humoral Immune Responses against BK Virus in Kidney Transplant Recipients with Polyomavirus Nephropathy

Reactivation of the polyomavirus BK (BKV) causes polyomavirus nephropathy (PVN) in kidney transplant (KTx) recipients and may lead to loss of the renal allograft. We have identified two HLA-A*0201-restricted nine-amino-acid cytotoxic T lymphocyte (CTL) epitopes of the BKV major capsid protein VP1, VP1(p44), and VP1(p108). Using tetramer staining assays, we showed that these epitopes were recognized by CTLs in 8 of 10 (VP1(p44)) and 5 of 10 (VP1(p108)) HLA-A*0201+ healthy individuals, while both epitopes elicited a CTL response in 10 of 10 KTx recipients with biopsy-proven PVN, although at variable levels. After in vitro stimulation with the respective peptides, CTLs directed against VP1(p44) were more abundant than against VP1(p108) in most healthy individuals, while the converse was true in KTx recipients with PVN, suggesting a shift in epitope immunodominance in the setting of active BKV infection. A strong CTL response in KTx recipients with PVN appeared to be associated with decreased BK viral load in blood and urine and low anti-BKV antibody titers, while a low or undetectable CTL response correlated with viral persistence and high anti-BKV antibody titers. These results suggest that this cellular immune response is present in most BKV-seropositive healthy individuals and plays an important role in the containment of BKV in KTx recipients with PVN. Interestingly, the BKV CTL epitopes bear striking homology with the recently described CTL epitopes of the other human polyomavirus JC (JCV), JCV VP1(p36) and VP1(p100). A high degree of epitope cross-recognition was present between BKV and corresponding JCV-specific CTLs, which indicates that the same population of cells is functionally effective against these two closely related viruses.

Polyoma virus-induced hemorrhagic cystitis in renal transplantation patient with polyoma virus nephropathy

We report a biopsy proven case of hemorrhagic cystitis in a cadaveric renal transplant patient with hematuria. Because more and more polyoma virus infection is being diagnosed in kidney transplant recipients, clinicians should be aware that gross hematuria in a recent transplant recipient may represent polyoma virus-induced hemorrhagic cystitis.

Immunocytological Urinalysis and Monocyte Chemotactic Peptide-1 in Renal Transplant Recipients With Polyomavirus Replication

In some patients polyomavirus replication induces chronic tubulointerstitial inflammation in the transplanted kidney. The aim of this study was to investigate whether immunocytological urinalysis and monocyte chemoattractant protein-1 (MCP-1) assays could be used for an early diagnosis of nephropathy for patients with polyomavirus replication. We analyzed 1189 urine sediments from 174 renal allograft recipients who were transplanted between 2000 and 2005. Decoy cells were identified by an immunofluorescence method using specific antibodies (JC/BK monoclonal antibody). A similar method was used to detect CD3 +, CD14 +, and HLA-DR + cells with appropriate antibodies. The urinary excretion of MCP-1 was assayed by enzyme-linked immunosorbent assay. The results of urine sediment analysis and MCP-1 concentrations were compared with those of patients with stable graft function (control group n = 65). In 17 patients (10%) decoy cells were identified in urine. In 12 patients polyomavirus DNA was detected in plasma or urine by a polymerase chain reaction method. Polyomavirus nephropathy was diagnosed in eight patients by the presence of intranuclear viral inclusions or immunohistochemical staining with SV40 large T-antigen specimens from a renal biopsy, as well as by clinical and histopathological evidence (group I). Polyomavirus replication was diagnosed in four patients by urinary excretion of decoy cells and polyomavirus DNA detection (group III). In five patients only decoy cells were found. The patients of groups I and II showed an increased number of CD3, CD14, HLA-DR surface antigen-positive cells and greater excretion of MCP-1 compared with the control group ( P < .02). The number of excreted cells was higher among patients with more severe infiltration. The results of patients from group III were similar to the control group. In conclusion, increased excretion of cells with CD3, CD14, and HLA-DR surface antigens and of MCP-1 were associated with intragraft tubulointerstitial inflammation in patients with polyomavirus nephropathy. Asymptomatic polyomavirus replication was associated with hidden tubulointerstitial inflammation. Monitoring cell excretion and chemokine content may be utilized for early detection of polyomavirus-induced nephropathy.

Long-Term Trends in Allograft Survival

Kidney transplantation has changed dramatically over the past decade. Many advances have occurred that result in shorter hospital stays, decreased acute rejection, less infectious morbidity, and improved long-term allograft survival. Many factors are responsible for the improvement in kidney transplantation, and these factors are discussed in this review. These successes have led to a paradox in the care of the kidney-transplant recipient, however. As the short-term complications are prevented or successfully managed, more patients will develop chronic problems with the function of the allograft.

Transplantation: Polyomavirus Nephropathy and the Risk of Specific Immunosuppression Regimens

BK virus is ubiquitously present in the latent state in humans, and awareness of the importance of BK polyomavirus is emerging among the kidney transplant community. First discovered in 1971 in the urine of a renal transplant recipient, BK virus nephropathy (BKVN) has come to be recognized as a significant cause of genitourinary disease and potential graft loss in the kidney transplant patient. In this review, we discuss the risk factors, available methods of diagnosis and therapeutic monitoring, and current approaches to therapy of BKVN.

Negative-staining Electron Microscopy of the Urine for the Detection of Polyomavirus Infections

Negative-staining electron microscopy (EM) has played a pivotal role in diagnostic virology. It is a rapid technique for viral detection in the urine and can provide an easy means for monitoring viral activity and productive infections. EM of urine for the detection of polyomaviruses has hitherto not been systematically evaluated as a screening tool for renal transplant patients at risk for BK polyomavirus nephropathy (BKN). Here, the authors discuss technical aspects of negative-staining EM of urine (n = 76 samples) and present a simple and rapid protocol for the semiquantitative evaluation of patient samples. In two patient populations (either with (n = 15 samples) or without (n = 15 samples) an established diagnosis of BKN), EM results were compared with two previously established techniques for monitoring polyomavirus activation: (1) cytology for the quantitation of decoy cells, and (2) quantitative PCR assays for the detection of BK virus DNA load levels. In both patient groups, the dynamics of decoy cell shedding by urine cytology closely paralleled free viral particle shedding by EM, and viral load levels as measured by PCR. A trend toward higher readings was observed in patients with BKN (median values, control versus BKN groups: decoy cells 21 versus 50/slide; free virions by EM: 32 versus 66 viral particles/10 high-power fields; PCR: 3.5 × 108 versus 5.4 × 108 BK virus copies/ml; all differences not statistically significant). The authors conclude that negative-staining EM and the semiquantitative assessment of free viral particles in the urine can be a useful clinical method to identify patients at increased risk for BKN. EM can be used alone or in combination with urine cytology or PCR assays.

Re: Detection of JC virus sequences in colorectal cancers in Japan

To the Editors: We read “Detection of JC virus DNA sequences in colorectal cancers in Japan” by Hori et al. with great excitement. These authors offer convincing evidence that polyoma T antigen may be integrated into the cellular genome of colon carcinoma tissues, although the T-antigen protein is not expressed [4]. We enthusiastically agree with their conclusions and would like to offer additional data to advance this discussion. First, we have investigated the presence of polyomavirus (PV) T antigen in tumors by immunohistochemistry (IHC), using a primary antibody that reacts with the large T antigen of both the JC and BK viruses [1, 5]. T antigen was not detected in any of 50 cases of colon carcinoma, 18 cases of gastric cancer, or 15 cases of bladder carcinoma. In addition, we examined by IHC the presence of T antigen in six cases of polyoma virus nephropathy (PVN) in renal transplant patients. In all of these six patients, robust nuclear staining specific for T antigen was observed in scattered tubular cells. The infected tubular cells were characterized by distinct cytopathological features, including anisonucleosis, hyperchromasia with clumped chromatin, and intranuclear inclusions. Such cellular changes represent a “permissive” infection, i.e., a state of viral replication. These histopathological features are absent in our 83 cases of carcinoma. We agree with these authors that if PV is present in tumors, it is likely to be integrated into the cellular genome, representing a “nonpermissive” infection in which viral replication does not occur. We conclude that future IHC studies of tumors are likely to be uninformative because we fail to detect viral antigens in nonpermissive infections, which are precisely the category of infections hypothesized to drive oncogenesis. Although recent studies have detected T antigen by IHC in esophageal [2] and colon [3] carcinoma, it remains to be seen whether these data truly represent evidence for a viral etiology in these tumors, or, alternatively, a sequelae of the patient’s immunocompromised state. We look forward to hearing the results of future studies from Dr. Hori’s team.