BKV-associated Nephropathy Research Articles

Total nephroureterocystectomy and urethrectomy due to urothelial carcinoma associated with the BK polyomavirus infection after kidney transplantation: a case report with literature review

BackgroundBK polyomavirus (BKPyV) infection after kidney transplantation is an important cause of graft failure among kidney transplant recipient and may cause malignant tumor, although the association between BKPyV infection and malignant tumor has been controversial yet.Case presentationWe report a case of a 39-year-old-male kidney transplantation (KTx) recipient with urine BKPyV replication who developed a graft pelvic tumor with the positive Simian virus 40 large T antigen (SV40 TAg). The patients received a living-related KTx from his 65-year-old mother. A protocol biopsy at 14 months after KTx showed BKPyV-associated nephropathy. Therefore, the dose of immunosuppressants was reduced, resulting in improved BKPyV viremia, but viruria persisted. About 117 months after KTx, urine cytology showed atypical cells suspicious for malignancy. Cystoscopy revealed a tumor on the neck of the bladder. Transurethral resection of the bladder tumor (TUR-BT) was performed; however, the diagnosis of malignancy was not confirmed at that time. Six months after the TUR-BT, urine cytology showed atypical cells definite for malignancy. Computed tomography and retrograde pyelography showed no evidence of urinary tract tumor and metastasis. Subsequently, total nephroureterocystectomy and urethrectomy were performed. Histological examination of the graft ureter revealed a high-grade urothelial carcinoma, with glandular differentiation, pT1. Immunohistochemically, the tumor showed positivities for SV40 TAg and p53, along with increased Ki67 labeling cells were increased. By contrast, nonneoplastic cells were negative for SV40 TAg. At the time of writing the present manuscript, the patient is free from recurrence or residual tumor and being closely monitored without additional therapy, 32 months after the surgery.ConclusionThe relationship between BKPyV infection after KTx and bladder carcinogenesis remains to be elucidated. However, when the KTx recipients who continue to have BKPyV infection for a long time are treated, the possibility of risk factors for renourinary carcinoma should always be carefully considered.

Donor-derived Cell-free DNA and the Prediction of BK Virus-associated Nephropathy.

Background.Approximately 15% of kidney transplant recipients (KTRs) develop BK viremia (BKV), with 1%–10% developing BK virus-associated nephropathy (BKVAN), which histologically resembles rejection. The Diagnosing Acute Rejection in Kidney Transplant Recipients (DART) study showed that donor-derived cell-free DNA (dd-cfDNA) levels <1% have a negative predictive value of 85% for active allograft rejection. Using data from this study, we evaluated the association of dd-cfDNA with plasma BK viral loads and biopsy findings to determine if dd-cfDNA can distinguish asymptomatic BKV from BKVAN.Methods.Data on dd-cfDNA, plasma BK viral loads, and biopsy findings from patients from the DART study were retrospectively examined. BKV was defined as 500–10 000 copies/mL. Presumptive BKVAN was defined as BK >10 000 copies/mL.Results.Of 102 participants with biopsies, 10 patients with BKV and BKVAN had paired dd-cfDNA, and viral loads available for analysis. Patients diagnosed with BKV and BKVAN had a median dd-cfDNA of 0.58% (IQR 0.43–1.15) and 3.38% (IQR 2.3–4.56, P = 0.001), respectively. dd-cfDNA titers correlated with BK PCR viral loads (R = 0.874, P = 0.01) and the presence of histologic evidence of BKVAN (100% sensitivity, 50% specificity). Five of 7 patients with BKVAN, but only 2 of 7 with BKV, had biopsies meeting Banff criteria for T-cell–mediated rejection. Median dd-cfDNA in nonrejection patients was 0.43% versus 2.84% in rejection patients (P = 0.001).Conclusion.Higher dd-cfDNA titers were associated with higher BK viral loads, biopsy-diagnosed BVAN, as well histologic changes meeting Banff criteria for as T-cell–mediated rejection. dd-cfDNA may be a useful noninvasive test to assess for progression of BKV to BKVAN.

Nephrotoxicity of calcineurin inhibitors as a risk factor for BK polyomavirus replication after kidney transplantation.

BK polyomavirus-associatednephropathy (PyVAN) is responsible for a significant percentage of transplanted kidneys prematurely terminating their function. Its occurrence is closely related to the intensity of immunosuppressive therapy. In a group of 161 newly transplanted patients, we prospectively evaluated 457 protocol renal biopsies performed within the first year after transplantation. Using the calcineurin inhibitors (CI) nephrotoxicity score, the incidence of nephrotoxicity was monitored as a manifestation of excessive immunosuppression. Findings were correlated with clinical evidence of active BK polyomavirus (BKPyV) replication and PyVAN. Compared to the normal histology, nephrotoxicity was associated with more frequent BKPyV viremia and viruria (p = .01 and p < .01, respectively) and more common occurrence of PyVAN. The persistence of toxicity in the subsequent biopsy proved to be a negative risk factor of viremia and viruria (p = .03 and p < .01, respectively), independently of the initial BKPyV status. Toxicity could also be used as a predictor of viremia and viruria (p = .04 and p < .01, respectively) even in the absence of viral replication at the time of initial biopsy. The early histological manifestation of CI nephrotoxicity was associated with significant BKPyV reactivation in the risky first posttransplant year.

Clinical Efficacy of Intravenous Immunoglobulin for BK Polyomavirus-Associated Nephropathy After Living Kidney Transplantation.

PurposeBK polyomavirus (BKPyV)-associated nephropathy (BKPyVAN) is one of the most difficult infections to be treated after kidney transplantation. Although patients with BKPyVAN usually received a reduction of immunosuppressive agents, the majority of these patients undergo the loss of the graft kidney without any effective treatment afterward. Therefore, development of more effective therapy for BKPyVAN is eagerly expected.Patients and MethodsAmong patients who underwent a kidney transplantation between January 2016 and April 2019 at our hospital, there were five cases of BKPyVAN. After the initial diagnosis, all patients discontinued administration of mycophenolate mofetil (MMF), which was not enough to diminish decoy cells in urine cytology test. Therefore, all patients received additional intravenous immunoglobulin (IVIG) (100 mg/kg/day) therapy for five days and were evaluated for the therapeutic effect of IVIG with immunohistochemical examination using re-biopsy samples of the graft kidney.ResultsAfter IVIG therapy, 2 cases showed negative decoy cells in urine and 3 cases showed a drastic decrease of plasma BK virus load. Importantly, simian virus (SV) 40 large T antigens diminished after IVIG administration in all cases, which degraded polyomavirus nephropathy classification.ConclusionAlthough it is difficult to treat BKPyVAN after kidney transplant, IVIG therapy was considered to a promising treatment to improve severity of BKPyVAN especially in cases that dose reduction of immunosuppressive agents was ineffective.

A Case of BK Polyomavirus-Associated Nephropathy of Native Kidneys in a 19-Year-Old Woman with a History of Orthotopic Cardiac Allotransplantation

A Case of BK Polyomavirus-Associated Nephropathy of Native Kidneys in a 19-Year-Old Woman with a History of Orthotopic Cardiac Allotransplantation

Detection of Proximal Tubule Involvement by BK Polyomavirus in Kidney Transplant Recipients With Urinary Sediment Double-Immunostaining

BackgroundThe extent and depth of BK polyomavirus (BKPyV) infection in renal allograft correlate with prognosis. This study was designed to evaluate the value of urinary sediment double-immunostaining for predicting BKPyV infection in proximal tubular epithelium.Materials and methodsA total of 76 urine sediment cell blocks, as well as the corresponding transplanted kidney tissues with BK polyomavirus associated-nephropathy (BKPyVAN), were evaluated by automatic double-immunostaining with anti-58-kDa Golgi protein (58K, a proximal renal tubular marker) + anti-SV40-T and anti-homogentisate 1, 2-dioxygenase (HGD, a renal tubular marker) + anti-SV40-T.ResultsImmunohistochemical staining demonstrated that 58K was expressed in proximal tubular epithelium but not in distal tubular epithelium or transitional epithelium. Of the 76 patients, 28 (36.8%) had urinary 58K(+)/SV40-T(+) cells and HGD(+)/SV40-T(+) cells, 41 (53.9%) had only HGD(+)/SV40-T(+) cells, one (1.3%) had only 58K(+)/SV40-T(+) cells, and six (7.9%) had only 58K(−)/HGD(−)/SV40-T(+) cells. The presence of urinary 58K(+)/SV40-T(+) cells was correlated with BKPyV infection in proximal tubular epithelium (P < 0.001, r = 0.806). The mean extent of SV40-T staining was significantly more extensive in patients with urinary 58K(+)/SV40-T(+) cells than those without urinary 58K(+)/SV40-T(+) cells (21.4 vs. 12.0%, P < 0.001). The positive predictive value, negative predictive value, sensitivity, and specificity of urinary 58K(+)/SV40-T(+) cells for predicting BKPyV infection in proximal tubular epithelium were 89.7% (95% CI: 71.5–97.3%), 91.5% (95% CI: 78.7–97.2%), 86.7% (95% CI: 68.4–95.6%), and 93.5% (95% CI: 81.1–98.3%), respectively.ConclusionUrinary sediment double-immunostaining with anti-58K and anti-SV40-T is valuable for predicting the extent and depth of BKPyV infection in renal allograft.

DIFFERENTIAL IMPACT OF ALLOGRAFT REJECTION ON KIDNEY TRANSPLANT PATIENTS WITH BKV INFECTION

Background: BK virus-associated nephropathy (BKVAN) is a known risk factor for allograft dysfunction and graft failure in kidney transplant recipients. The mainstay of treatment for BKVAN is reduction of immunosuppression and this may increase risk for acute rejection. We proposed to observe the effects of acute rejection on patients with BKVAN. Methods: Using data from the Korean Organ Transplantation Registry, a nationwide organ transplantation database, we com pared graft function, and allograft survival in BKVAN patients with or without biopsy proven acute rejection (BPAR). Results: Among the 5,403 patients who received kidney transplantation between 2014 and June 2019, a total of 97 patients were diagnosed with BKVAN. Twenty-six patients (27%) developed BPAR within 6 months of BKVAN diagnosis, 71 patients did not. There were no differences in baseline characteristics, immunosuppression or BKVAN treatment methods between the BPAR and no BPAR groups. There was a significant decrease in allograft function in the BPAR group compared to the no BPAR group in both the 1-year (BPAR creatinine [Cr], 2.5±1.9 mg/dL vs. no BPAR Cr, 1.9±0.9 mg/dL; P=0.044) and 2-year follow-up period (BPAR Cr, 3.8±3.6 mg/dL vs. no BPAR Cr, 2.2±1.0 mg/dL; P=0.015). The BPAR group had lower allograft survival rates compared to the no BPAR group, although not statistically significant (P=0.474). On multivariate Cox regression analysis, MMF discontinuation was observed as a significant risk factor for rejection in BKVAN patients (hazard ratio, 4.000; 95% confidence interval [CI], 1.014- 15.775; P=0.048). Conclusions: Acute rejection with BKVAN is associated with poorer allograft function and survival. Also, discontinuation of MMF as treatment for BKVAN increases risk for acute rejection.

GENOTYPIC CHARACTERISTICS, GENOMIC MUTATIONS OF BK POLYOMAVIRUS IN PATIENTS AFTER KIDNEY TRANSPLANTATION: A CROSS-SECTIONAL STUDY IN VIETNAM

Purpose: BK polyomavirus (BKPyV) frequently reactivates in kidney transplant recipients during immunosuppressive therapy and triggers BKPyV-associated nephropathy (BKVN) and graft rejection. The virulence and the ability of viral replication to be related to the mutation status of BKPyV. The study aims to determine genotypic characteristics and genomic mutations of BKPyV in patients after kidney transplantation. Materials and Methods: A total of 201 patients underwent kidney transplantation were enrolled in the study. Serum, urine, and kidney biopsy samples were collected to detect BKPyV infection and BKVN. Genotypic and genomic mutations identified based on the VP1 (Virus Protein 1) gene region by sequencing and phylogenetic analysis. The presence of any single-nucleotide polymorphisms (SNPs) and changes in amino acid gene sequences VP1 (1630-1956) was recorded. Results: BKPyV infection was determined in 84 patients (41.8%). Among them, 8 patients (3.9%) were diagnosed with BKPyV-associated nephropathy. Genotypic analysis was performed in 40 patients with an average age was 39.7 years old, 82.1% male, the average duration of post-transplantation 1.56 ± 2.82 years, average serum creatinine 115.82 ± 29.85 µmol/L. Genotypic characteristics showed that BKPyV-I accounted for 25/40 (62.5%), BKPyV-IV observed in 15/40 (37.5%). Of note, BKPyV-I/b-1 was (100%) with BKV genotype I, whereas BKPyV-IV/a-1 (26.67%), BKPyV-IV/a-2 (6.67%), BKPyV-IV/c-1 (66.66%). Sequence analysis of VP1 region of BKPyV-I genotypes identified 5 mutant positions, of BKPyV-IV genotype presented 17 mutant positions, in which at position 1745 of BKPyV-IV appeared to change from A to G with the frequency of 13/15 samples (86.67%) resulted in amino acid change from N to S. There was no difference between the BKPyV genotypes with age, sex, viral load, and BKPyV-associated nephropathy. Conclusion: This study indicated that the BKPyV-I genotype was more common than BKPyV-IV, however genomic mutations that occur in the VP1 region of BKPyV-IV strains were more common than those of BKPyV-I.

Intravenous immunoglobulin as a preventive strategy against BK virus viremia and BKV-associated nephropathy in kidney transplant recipients-Results from a proof-of-concept study.

BK virus (BKV) replication occurs frequently in kidney transplant recipients (KTR), potentially leading to BKV-associated nephropathy (BKVAN) and graft loss. Patients with high titers of BKV-neutralizing antibodies (NAbs) are protected against BKV replication, and intravenous immunoglobulin (IVIg) infusion can increase NAb titers. We investigated whether early IVIg administration prevents BKV replication in patients with low NAb titers (<4 log10 against the BKV-specific genotype). Based on NAb titers on the day of transplantation, KTR followed in the Strasbourg University Hospital (n=174) were retrospectively divided into the following 3 risk categories for BKV replication: (1) patients with low NAb titers ("high-risk") who received IVIg for the first 3 posttransplant months (n=44), (2) patients with low NAb titers ("high-risk") who did not undergo IVIg treatment (n=41), and (3) patients with high NAb titers ("low-risk") who did not receive IVIg (n=89). At 12 posttransplant months, the incidence of BKV viremia in the high-risk group treated with IVIg (6.8%) was similar to that observed in the low-risk group (10.1%) and markedly lower than that of the untreated high-risk group (36.6%; P<.001). Similar results were observed with regard to BKVAN. We conclude that IVIg may be a valuable strategy for preventing BKV replication.

Urine Donor-Derived Cell-Free DNA Helps Discriminate BK Polyomavirus-Associated Nephropathy in Kidney Transplant Recipients With BK Polyomavirus Infection.

Background: Studies have shown that plasma donor–derived cell-free DNA (dd-cfDNA) can predict renal allograft antibody-mediated rejection. This study was performed to evaluate the value of urine dd-cfDNA concentration and dd-cfDNA fraction (%) for discriminating BK polyomavirus-associated nephropathy (BKPyVAN) in kidney transplant recipients with urinary BK polyomavirus (BKPyV) infection.Methods: In this retrospective single-center observational study, we enrolled kidney transplant recipients who were diagnosed with urine BKPyV infection between August 2018 and May 2019 at the First Affiliated Hospital of Sun Yat-sen University. Urine dd-cfDNA was measured by using a novel target region capture sequencing methodology. The pathological diagnosis of BKPyVAN was confirmed by anti-SV40-T immunohistochemical staining and classified using the American Society for Transplantation schema. Receiver operating characteristic curve analysis was used to investigate the relations of urine dd-cfDNA and dd-cfDNA% to intrarenal allograft BKPyV infection states.Results: In total, 93 patients were enrolled, including 40 cases of proven BKPyVAN, seven cases of probable BKPyVAN, 23 cases of possible BKPyVAN, and 23 cases of resolving BKPyVAN. Urine dd-cfDNA level in proven BKPyVAN (22.09 ± 21.27 ng/ml) was comparable to that in probable BKPyVAN (15.64 ± 6.73 ng/ml, P = 0.434) but was significantly higher than that in possible BKPyVAN (5.60 ± 3.53 ng/ml) and resolving BKPyVAN (5.30 ± 3.34 ng/ml) (both Ps < 0.05). Urine dd-cfDNA% of proven BKPyVAN (0.71 ± 0.21) was lower than that of probable BKPyVAN (0.91 ± 0.04, P < 0.001), but was significantly higher than that of possible BKPyVAN (0.56 ± 0.30) and resolving BKPyVAN (0.46 ± 0.28) (both Ps < 0.05). For distinguishing biopsy-proven BKPyVAN from biopsy-excluded BKPyVAN, the discrimination capacity of urine dd-cfDNA (AUC: 0.842, 95% CI: 0.735, 0.918) was superior to that of plasma BKPyV DNA load (AUC: 0.660, 95% CI: 0.537, 0.769) with 0.181 (95% CI: 0.043, 0.319) difference between areas under ROC curves (P = 0.010).Conclusion: The elevated urine dd-cfDNA level may help discriminate BKPyVAN in kidney transplant recipients with BKPyV viruria.

BK virus-associated nephropathy in a lung transplant patient: case report and literature review

BackgroundBK virus-associated nephropathy (BKVAN) is a relatively common cause of renal dysfunction in the first six months after renal transplantation. It arises from reactivation of the latent and usually harmless BK virus (BK virus) due to immunosuppression and other factors including some that are unique to renal transplantation such as allograft injury. BKVAN is much rarer in non-renal solid organ transplantation, where data regarding diagnosis and management are extremely limited.Case presentationWe report a case of a 58-year-old man found to have worsening renal dysfunction nine months after bilateral sequential lung transplantation for chronic obstructive pulmonary disease (COPD). He had required methylprednisolone for acute allograft rejection but achieved good graft function. Urine microscopy and culture and renal ultrasound were normal. BK virus PCR was positive at high levels in urine and blood. Renal biopsy subsequently confirmed BKVAN. The patient progressed to end-stage renal failure requiring haemodialysis despite reduction in immunosuppression, including switching mycophenolate for everolimus, and the administration of intravenous immunoglobulin (IVIG).ConclusionsThis very rare case highlights the challenges presented by BK virus in the non-renal solid organ transplant population. Diagnosis can be difficult, especially given the heterogeneity with which BKV disease has been reported to present in such patients, and the optimal approach to management is unknown. Balancing reduction in immunosuppression against prevention of allograft rejection is delicate. Improved therapeutic options are clearly required.

Fluoroquinolones for BK viral complication after transplantation: Meta‐analysis

BK polyomavirus (BKV) causes two distinct complications after transplantation, hemorrhagic cystitis (BKV-HC) after hematopoietic stem cell transplantation (HSCT), and BKV-associated nephropathy (BKVAN) after kidney transplantation (KT). Although fluoroquinolones show efficacy against BKV proliferation in vitro, the clinical effect remains uncertain; thus, we performed meta-analysis to assess its efficacy in the prophylaxis. Articles published before March 2020 were searched from PubMed, the Cochrane Library, ISRCTN registry, and ClinicalTrials.gov. Primary outcomes were BKV-HC after HSCT and BKVAN after KT. Secondary outcomes were BK viremia, viruria after KT, and fluoroquinolone-related adverse events. Three trials with 281 patients post-HSCT and 11 trials with 1882 patients post KT were included as for prophylaxis. Fluoroquinolone prophylaxis did not show effects on BKV-HC (OR 0.54, 95% CI 0.13-2.25), BKVAN (OR 0.74, 95% CI 0.35-1.55), and BK viremia (OR 0.79, 95% CI 0.49-1.28), but significantly decreased BK viruria (OR 0.64, 95% CI 0.45-0.91). Fluoroquinolone prophylaxis was associated with the higher percentage of fluoroquinolone-resistant infection among identified bacteria (OR 2.38, 95% CI 1.16-4.88), but the incidence of fluoroquinolone-resistant infection was similar (OR 1.15, 95% CI 0.71-1.86), due to the decrease of infection itself (OR 0.52, 95% CI 0.34-0.81). This meta-analysis showed that fluoroquinolones did not prevent BKV-HC after HSCT or BKVAN after KT, although the effect against BKV-HC should be further investigated by randomized controlled trials. Fluoroquinolones could reduce the rate of BK viruria to some extent but may not have clinically sufficient effects.

Pathological characteristics of BK polyomavirus-associated nephropathy with glomerular involvement.

BackgroundThis study aimed to investigate the pathological characteristics of BK polyomavirus (BKPyV)-associated nephropathy (BKPyVAN) with glomerular involvement in kidney transplant recipients.MethodsForty-four patients with glomerular BKPyV infection were retrospectively included for analysis. Immunohistochemical (IHC) staining was performed on paraffin sections using monoclonal mouse anti-SV40 large T antigen antibody.ResultsIn BKPyV-infected glomeruli, the glomerular parietal epithelial cells (GPECs) were swollen, hyperchromatic, and enlarged, with an increased nuclear to cytoplasm (N/C) ratio and smudgy basophilic intra-nuclear viral inclusions. IHC staining revealed the distribution of BKPyV involvement in GPECs, podocytes, and shedding cells within Bowman’s space. Notably, BKPyV affected GPEC proliferation and caused crescent formation (7 biopsies, 15.9%). Three biopsies exhibited fibrous crescents and the absence of viral inclusions. The other 4 biopsies exhibited cellular and fibro-cellular crescents, with viral cytopathic changes and positive IHC staining in the proliferative GPECs. Electron microscopy showed viral particles in both GPECs and podocytes. BKPyV-infected GPECs were degenerative, with mitochondrial swelling, endoplasmic reticulum expansion, and multi-layered membranous structure formation. Twelve (27.3%) patients received repeat biopsies within 1.6 to 39.5 months (median: 13.5 months), but none revealed persistent glomerular BKPyV infection.ConclusionsDistinct glomerular changes in BKPyVAN biopsies should raise the possibility of glomerular involvement.

Significance of revised criteria for chronic active T cell-mediated rejection in the 2017 Banff classification: Surveillance by 1-year protocol biopsies for kidney transplantation.

Diagnostic criteria for chronic active T cell-mediated rejection (CA-TCMR) were revised in the Banff 2017 consensus, but it is unknown whether the new criteria predict graft prognosis of kidney transplantation. We enrolled 406 kidney allograft recipients who underwent a 1-year protocol biopsy (PB) and investigated the diagnostic significance of Banff 2017. Interobserver reproducibility of the 3 diagnosticians showed a substantial agreement rate of 0.68 in Fleiss's kappa coefficient. Thirty-three patients (8%) were classified as CA-TCMR according to Banff 2017, and 6 were previously diagnosed as normal, 12 as acute TCMR, 10 with borderline changes, and 5 as CA-TCMR according to Banff 2015 criteria. Determinant factors of CA-TCMR were cyclosporine use (vs tacrolimus), previous acute rejection, and BK polyomavirus-associated nephropathy. In survival analysis, the new diagnosis of CA-TCMR predicted a composite graft endpoint defined as doubling serum creatinine or death-censored graft loss (log-rank test, P<.001). In multivariate analysis, CA-TCMR was associated with the second highest risk of the composite endpoint (hazard ratio: 5.42; 95% confidence interval, 2.02-14.61; P<.001 vs normal) behind antibody-mediated rejection. In conclusion, diagnosis of CA-TCMR in Banff 2017 may facilitate detecting an unfavorable prognosis of kidney allograft recipients who undergo a 1-year PB.

BK polyomavirus genotypes and progression of BKV disease in renal transplant recipients

Background. BK virus (BKV) reactivation is a significant cause of BK Polyomavirus associated nephropathy (BKPyVN) resulting in acute graft rejection in 1-10% of post-renal transplant recipients. However, the association of BKV genotypes with development of BKPyVN is poorly understood. Here we aimed to determine the prevalence of BKV genotypes in post-renal transplant recipients, and its association with BKPyVN disease progression. Method. Two methods were utilised to genotype BKV. A 800bp fragment of virus VP1 antigen region was amplified using nested (PCR) followed by sequencing. The genotypes were determined according to a previously developed algorithm based on analysing 100bp region of the VP1 gene. Furthermore, the logarithm results were validated with the constructed phylogenetic tree. The results were correlated with patient viral loads and development of BKPyVN. Results. BK virus DNA was detected in 32 (69%) of 46 post-renal transplant recipients with BK viremia, while BKPyVN was only reported in two (4.3%)patients. 30 out of 32 samples were successfully genotyped (93.7%) with 23 (76.6%) belonging to the BKV Ib-2 subtype and seven (23.3%) belonging to the BKV Ib-1 subtype and with no cases representing genotype II, III and IV. All cases with confirmed BKPyVN matched to the BKV Ib-2 genotype. Additionally, no significance differences were observed between BKV genotypes in regards to viral loads, development of viremia, HLA mismatch, age or sex. Conclusion. The results indicate no correlations between BKV genotypes and the development of BKPyVN. Furthermore,a high distribution of BKV genotype Ib-2 was found among BKV infected patients within this cohort.

Investigating the mechanisms of BK polyomavirus egress and virus-host interactions

BK polyomavirus (BKPyV) is a small, non-enveloped dsDNA virus that infects 70-90% of the world's population and causes a lifelong, silently persistent infection. In immunocompromised individuals, BKPyV replication can result in serious pathology. Bone marrow transplant patients can develop a haemorrhagic cystitis, and in kidney transplant patients BKPyV replication can provoke a nephropathy that leads to deterioration of allograft function and eventual loss of the transplanted organ. There are currently no antiviral treatments with clinical efficacy against BKPyV associated nephropathy. While the life cycles of non-enveloped viruses are often assumed to require cell lysis to release progeny virions, we have evidence to suggest that BKPyV exits the cell via non-lytic means using an unconventional secretory pathway. We have investigated the effects of knocking out cellular genes thought to be involved in unconventional secretory pathways that bypass the Golgi apparatus on the release of BKPyV. We observe decreased BKPyV release from cells that have undergone CRISPR-mediated knockout of Golgi Reassembly Stacking Protein (GORASP) 1 or 2. Investigation of BKPyV-induced changes to the plasma membrane of infected cells demonstrated increased cell surface expression of transmembrane proteins normally resident in the endoplasmic reticulum. This appears to be inhibited by the knockout of GORASP1 or 2, suggesting that virions and ER markers are secreted via a common pathway in infected cells. These experiments are uncovering novel virus-host interactions that, when targeted, could help prevent BKPyV-associated nephropathy and allograft loss.

P1639DEVELOPMENT OF SELF-LIMITED LOW-LEVEL BK VIREMIA/VIRURIA SUGGESTS IDEAL IMMUNOSUPPRESSION TO SUPPRESS T-CELL AND ANTIBODY-MEDIATED REJECTION AND PRESERVE KIDNEY ALLOGRAFT FUNCTION

Abstract Background and Aims Polyomavirus BK (BKV) viremia has been suggested as a surrogate marker of overimmunosuppression. Due to recent advances in monitoring strategies and pre-emptive therapy, progression to BKV-associated nephropathy (BKVN) has been reduced. Reduction of maintenance immunosuppression during BKVN, however, has been associated with both T-cell mediated rejection (TCMR), antibody-mediated rejection (ABMR), and inferior kidney allograft outcomes. Although the administration of intravenous immunoglobulins (IVIG) failed to treat BKVN, IVIG may have properties to reduce allosensitization. Method We studied 860 kidney transplant recipients (KTRs) predominantly under tacrolimus-based triple-drug maintenance immunosuppression from 2009 to 2018. We identified 130 KTRs (15.1%) with high-level BK viremia &amp;gt;10,000 copies/mL (presumptive BKVN), 180 KTRs (20.9%) with low-level BK viremia &amp;lt;10,000 copies/mL, 85 KTRs (9.9%) with isolated BK viruria, and 465 KTRs (54.1%) with no BK viruria/viremia. Kidney allograft outcomes with respect to TCMR, ABMR, development of de novo donor-specific antibodies (DSA), kidney allograft survival, and estimated glomerular filtration rate (eGFR) slope were analysed. Due to the increased incidence of TCMR and ABMR among KTRs with presumptive BKVN, 48 of 130 KTRs (36.9%) with high-level BK viremia starting from 2015 received IVIG (0.5 g/kg of body weight) on a biweekly basis during BKV replication. Results Very interestingly, KTRs with low-level BK viremia/viruria showed a significantly lower incidence of TCMR compared to KTRs with no BK viremia/viruria and KTRs with high-level BK viremia (p&amp;lt;0.05). In addition, KTRs with low-level BK viremia/viruria showed a significantly lower incidence of ABMR compared to KTRs with high-level BK viremia (p&amp;lt;0.05). No differences were observed with respect to the development of de novo DSA (MFI&amp;gt;5000) between KTRs with low-level BK viremia/viruria, KTRs with no BK viremia/viruria, and KTRs with high-level BK viremia (p&amp;gt;0.05). KTRs with low-level BK viremia/viruria showed superior kidney allograft survival and lower eGFR slope compared to KTRs with no BK viremia/viruria and KTRs with high-level BK viremia (p&amp;lt;0.05). The administration of IVIG during high-level BK viremia didn’t impact the development of TCMR, ABMR, development of de novo DSA, eGFR slope, and kidney allograft survival (p&amp;lt;0.05). Conclusion Our results show that low-level BK viremia/viruria is associated with suppression of TCMR and ABMR in the early period posttransplantation, and preservation of kidney allograft function in the long-term. The administration of IVIG didn’t prove beneficial to reduce allosensitization among KTRs with high-level BK viremia.

BK Virus RNA in Renal Allograft Biopsies.

BK polyomavirus-associated nephropathy (BKpyVAN) remains a cause of graft loss in kidney transplant recipients on immunosuppressive therapy. Its diagnosis relies on the identification of BK virus (BKV) in the renal allograft biopsy by positive immunohistochemical (IHC) stain for the viral SV40 large T antigen, although in situ hybridization (ISH) for viral DNA is used in some centers. We examined tissue detection of BKV RNA by RNAscope, a novel, automated ISH test, in 61 allograft biopsies from 56 patients with BKpyVAN. We found good correlation between the estimate of BKV tissue load by RNAscope ISH and SV40 IHC (R2 = 0.65, p<0.0001). RNAscope ISH showed 88% sensitivity and 79% specificity and, as an alternative test, could confirm the presence of BKV tissue in presumed BKpyVAN and rule out BKV as the causative agent in JC virus nephropathy. We also used tissue BK viral load estimates by both RNAscope ISH and SV40 IHC to examine the relation between tissue and plasma BK levels and found significant correlation only between BK viremia and tissue BK measured by RNAscope ISH. Our findings suggest that the RNAscope ISH assay could be a reliable test for BKV detection in allograft biopsies.

High expression of JC polyomavirus-encoded microRNAs in progressive multifocal leukoencephalopathy tissues and its repressive role in virus replication.

JC polyomavirus (JCPyV, JCV) causes progressive multifocal leukoencephalopathy (PML) in immunocompromised hosts. JCPyV replicates in oligodendrocytes within the brain tissue of patients with PML. The JCPyV genome encodes a microRNA (miRNA) in the region encoding the large T antigen. JCPyV-encoded miRNA (miR-J1) has been detected in the tissue and cerebrospinal fluid samples of patients with PML; however, there are no reports describing the localization of polyomavirus-encoded miRNA in histological samples of patients with virus-associated diseases. In the present study, we detected high miR-J1 expression in the nuclei of JCPyV-infected cells in PML tissue samples via in situ hybridization. Additionally, in situ hybridization also revealed the expression of BK polyomavirus (BKPyV, BKV)-encoded miRNA in lesions of BKPyV-associated nephropathy. In situ hybridization for miR-J1-5p and -3p showed positive signals in 24/25 (96%) of PML tissues that were positive for JCPyV by immunohistochemistry. Higher copy numbers of miR-J1 were detected in PML tissues than in non-PML tissues by real-time reverse transcription PCR. Next generation sequencing showed that miR-J1-5p, a mature miRNA of primary miRNA, was predominant in the lesions compared with miR-J1-3p, another mature miRNA. Deletion or mutation of miR-J1 in recombinant JCPyV promoted the production of JCPyV-encoded proteins in cells transfected with JCPyV DNA, suggesting that polyomavirus-encoded miRNA may have a repressive role in viral replication in PML tissues. In situ hybridization for viral miRNA may be a useful diagnostic tool for PML.

Usefulness of BK virus-specific interferon-γ enzyme-linked immunospot assay for predicting the outcome of BK virus infection in kidney transplant recipients.

Background/AimsTo investigate if BK virus (BKV)-specific T cell immunity measured by an interferon-γ enzyme-linked immunospot (ELISPOT) assay can predict the outcome of BK virus infection in kidney transplant recipients (KTRs). MethodsWe included 68 KTRs with different viremia status (no viremia [n = 17], BK viremia [n = 27], and cleared viremia [n = 24]) and 44 healthy controls (HCs). The BK viremia group was divided into controller (< 3 months) and noncontroller (> 3 months) according to sustained duration of BKV infection. We compared BKV-ELISPOT results against five BKV peptides (large tumor antigen [LT], St, VP1-3). ResultsBKV-ELISPOT results were higher in three KTRs groups with different BKV infection status than the HCs group (p < 0.05). In KTR groups, they were higher in cleared viremia group than no viremia or BK viremia group. Within the BK viremia group, controller group had higher LT-ELISPOT results compared to noncontroller group (p = 0.032). Also, KTRs without BK virus-associated nephropathy (BKVN) had higher LT, St, VP1, and VP2-ELISPOT results than those with BKVN (p < 0.05). ConclusionsBKV-ELISPOT assay may be effective in predicting clinical outcomes of BKV infection in terms of clearance of BK virus and development of BKVN.