Search Results (91)

Polyomaviruses have the potential to cause significant morbidity not only in transplant medicine, but also in other forms of disease or variants of immunosuppression. In kidney transplant recipients or recipients of human stem cell transplants, the BK-Virus is the major proponent of manifestations such as BKPyV-associated nephropathy or hemorrhagic cystitis. As no polyomavirus-specific drug with proven in vivo effects has been developed so far, methods to screen for such drugs are important. This work describes the establishment of a virus-secreting cell line. By infecting a pre-established monkey kidney cell line (COS-1) with a non-rearranged human BK polyomavirus isolated from a kidney transplant patient suffering from BKPyV-associated nephropathy, a continuously replicating cell type with consistent virus secretion could be established and was termed COSSA. Measurements of BKPyV replication, virion production, and secretion were performed both intracellularly and in the cell supernatant. Viral proteins such as VP1 and LTAg were accurately tracked by confocal microscopy, as well as by immunoblot and qPCR. An intracellular flow cytometry (FACS) assay detecting VP1 protein was established and revealed an expanded range of positive intracellular signals. The viruses produced proved to be infectious in human tubular epithelial cell lines. Long-range sequencing of the COSSA genome using Oxford Nanopore Technology revealed a total of five distinct BKPyV integration events. One integration of a partial BKPyV genome was located upstream of the epidermal growth factor receptor gene. The second and third, both truncated forms of integration, were close to histocompatibility gene locuses, while the fourth was characterized by a ninefold and the fifth by a fourfold tandem repeat of the BKPyV genome. From both of the repeat forms, virus replicates were derived showing deletions/duplications on early and late genes and inversions within the non-coding control region (NCCR). This pattern of repetitive viral genome integration is a potential key driver of enhanced viral replication and increased virion assembly, ultimately supporting efficient virus egress. Quantitative PCR analysis confirmed the release of approximately 10⁸/mL viral units per 48 h from 2 × 10⁵ COSSA cells into the culture supernatant. Notably, the NCCR region of the most frequent copies of circular virus and the integrated tetrameric tandem repeat exhibited a rearranged configuration, which may contribute to the observed high replication dynamics. The establishment of a consistent methodology to generate and secrete BKPyV from a cell line is expected to significantly facilitate antiviral drug development. Full article

BK virus (BKV) reactivation is a significant complication in renal transplant recipients, often leading to BK viremia and BK virus-associated nephropathy (BKVAN), which can compromise graft survival. While the routine monitoring of BKV DNA in blood aids in early detection, identifying pre-transplant risk factors remains a challenge. This study investigates the role of pre- and post-transplant anti-BKV IgG levels and human leukocyte antigen (HLA) alleles in predicting BKV reactivation. The hospital-based cross-sectional study was conducted on 38 renal transplant recipients, stratified into viremic, non-viremic, and BKVAN groups. Anti-BKV IgG levels were measured pre-transplant, at viremia onset, and post-viremia using ELISA. BKV DNA was detected via qPCR, and HLA typing was performed using sequence-specific oligonucleotide probe (SSOP) hybridization. Statistical analyses included Kaplan–Meier survival curves and Cox regression models. Pre-transplant anti-BKV IgG seropositivity was higher in viremic (94%) and BKVAN (100%) patients than in non-viremic recipients (66.6%). Post-transplant IgG levels increased significantly in viremic recipients (p < 0.05). HLA-B44 and HLA-DR15 were significantly associated with increased BKV viremia risk (p = 0.02 and p = 0.01, respectively). Pre-transplant anti-BKV IgG levels and specific HLA alleles influence BKV reactivation risk. These findings highlight the potential for integrating serological and genetic screening into pre-transplant assessments to improve risk stratification and post-transplant monitoring strategies. Full article

Background/Objectives: This study evaluated infectious complications and immune reconstitution in 253 adults undergoing peripheral blood allogeneic hematopoietic cell transplantation (allo-HCT) with post-transplant cyclophosphamide (PTCY)-based GVHD prophylaxis. Methods: Patients received grafts from HLA-matched donors (47.4%), mismatched unrelated donors (MMUD, 33.2%), or haploidentical donors (19.4%). Results: The estimated 2-year non-relapse mortality (NRM) was 11.8%, 26.4%, and 22.4%, respectively (p = 0.0528). The cumulative incidence (Cum.Inc) of acute and chronic GVHD, immunosuppression duration, and post-transplant outcomes were similar across donor types. The day +30 Cum.Inc of bacterial bloodstream infections (BSI) tended to be higher in HLA-matched transplants (49.2%, p = 0.073), while HHV-6 reactivation showed a trend toward higher frequency in haploidentical transplants (22.4%, p = 0.068). Cytomegalovirus (CMV) reactivation occurred between days +30 and +100, with the highest Cum.Inc in MMUD (59.5%, p = 0.033). BK virus-associated hemorrhagic cystitis showed a trend toward higher incidence in MMUD (22.3%, p = 0.056). Respiratory and fungal infections were most frequent in the first 100 days, with comparable rates across donor types. By day +180, most patients achieved immune reconstitution, with normalization of CD4+ T cells, CD8+ T cells, and IgG levels, independent of donor type. Conclusions: Patients undergoing allo-HCT with PTCY-based prophylaxis experience a high infectious density rate early post-transplant, which decreases after 6 months as immune reconstitution progresses, regardless of donor type. Full article

Polyomaviruses (PyVs) are non-enveloped double-stranded DNA viruses that can cause significant morbidity in allogeneic hematopoietic stem cell transplant (allo-HSCT) recipients, particularly BK polyomavirus (BKPyV) and JC polyomavirus (JCPyV). BKPyV is primarily associated with hemorrhagic cystitis (HC), while JCPyV causes progressive multifocal leukoencephalopathy (PML). The pathogenesis of these diseases involves viral reactivation under immunosuppressive conditions, leading to replication in tissues such as the kidney, bladder, and central nervous system. BKPyV-HC presents as hematuria and urinary symptoms, graded by severity. PML, though rare after allo-HSCT, manifests as neurological deficits due to JCPyV replication in glial cells. Diagnosis relies on nucleic acid amplification testing for DNAuria or DNAemia as well as clinical criteria. Management primarily involves supportive care, as no antiviral treatments have proven consistently effective for either virus and need further research. This review highlights the virology, clinical presentations, and management challenges of PyV-associated diseases post-allo-HSCT, emphasizing the need for improved diagnostic tools and therapeutic approaches to mitigate morbidity and mortality in this vulnerable population. Full article

BK polyomavirus (BKPyV) is a pathogen responsible for infectious complications in hematopoietic stem cell transplant (HSCT) recipients. This review aims to give an insight into recent data about the structure and genomic organization, epidemiology, clinical manifestations, diagnosis, and current treatment options of BKPyV infections in children after HSCT. News regarding viral replication and pathogenesis include the generation of miRNA, new mechanisms of viral shedding by releasing infectious particles via extracellular vesicles, and human bladder microvascular endothelial cells probably acting as viral reservoirs enabling low-level viral replication and persistence. In studies conducted over the past five years, BKPyV hemorrhagic cystitis (BKPyV-HC) has a prevalence rate of 4 to 27% in children undergoing HSCT. Diagnostics still has unsolved dilemmas like whole blood or plasma samples as well as the standardization of molecular methods to allow for reporting in international units. In terms of treatment, new approaches have been used in the past five years, including the use of mesenchymal stem cells (MSCs), virus-specific T cells (VSTs), and recombinant human keratinocyte growth factor (rH-KGF), although the efficacy of some of these treatments has only been documented in isolated studies. This complication continues to pose a substantial clinical challenge, characterized by an absence of effective preventive and therapeutic measures. Full article

Kidney transplant recipients require a lifelong protocol of immunosuppressive therapy to prevent graft rejection. However, these same medications leave them susceptible to opportunistic infections. One pathogen of particular concern is human polyomavirus 1, also known as BK virus (BKPyV). This virus attacks kidney tubule epithelial cells and is a direct threat to the health of the graft. Current standard of care in BK virus-infected transplant recipients is reduction in immunosuppressant therapy, to allow the patient’s immune system to control the virus. This requires a delicate balance; immune suppression must be strong enough to prevent rejection, yet weak enough to allow viral clearance. We seek to model viral and immune dynamics with the ultimate goal of applying optimal control methods to this problem. In this paper, we begin with a previously published model and make simplifying assumptions that reduce the number of parameters from 20 to 14. We calibrate our model using newly available patient data and a detailed sensitivity analysis. Numerical results for multiple patients are given to show that the newer model reflects observed dynamics well. Full article

Polyomavirus BK (BKPyV)-associated nephropathy (BKPyV-nephropathy) remains a significant cause of premature kidney allograft failure. In the absence of effective antiviral treatments, current therapeutic approaches rely on immunosuppression (IS) reduction, possibly at the risk of inducing alloimmunity. Therefore, we sought to explore the long-term effects of a tailored viro-immunologic surveillance and treatment program for BKPyV on the development of alloimmunity and kidney graft outcome. Forty-five pediatric kidney transplant recipients were longitudinally monitored for BKPyV replication, virus-specific immunity, and donor-specific HLA antibodies (DSAs). DNAemia developed in 15 patients who were treated with stepwise IS reduction. Among the other 30 patients, 17 developed DNAuria without DNAemia and 13 always resulted as BKPyV-negative. All patients with DNAemia cleared BKPyV after having mounted a virus-specific cellular immune response, and no biopsy-proven BKPyV-nephropathy was observed. The presence of cytotoxic populations directed to the BKPyV Large-T (LT) antigen early after transplantation protected kidney recipients from developing BKPyV replication, and the appearance of LT-specific T cells in viruric patients prevented the development of BKPyV-DNAemia. In our cohort, no significant correlation was observed between BKPyV-DNAemia and the development of DSA and antibody-mediated rejection. However, patients who experienced and cleared BKPyV-DNAemia had a worse allograft survival at a median follow-up of 18.9 years (p = 0.048). These data need to be confirmed in larger cohorts. Full article

Cell culture underpins virus isolation and virus neutralisation tests, which are both gold-standard diagnostic methods for foot-and-mouth disease (FMD). Cell culture is also crucial for the propagation of inactivated foot-and-mouth disease virus (FMDV) vaccines. Both primary cells and cell lines are utilised in FMDV isolation and propagation. Widely used cell lines for FMDV and isolation and propagation include baby hamster kidney cells (BHK-21), swine kidney cells (IB-RS-2), foetal goat tongue (ZZ-R 127), foetal porcine kidney cells (LFBKvB6), bovine kidney cells (BK), human telomerase reverse transcriptase bovine thyroid (hTERT-BTY) and porcine kidney-originating PK-15 or SK 6 cell lines. This review highlights how different receptors and molecules—integrins, heparan sulphate (HS), and the Jumonji C-domain containing Protein 6 (JMJD6)—found on the surface of different cell types contribute to differences experienced with susceptibility and sensitivity of the cells to infection with different serotypes of FMDV. This review specifically focuses on Southern African territory (SAT) serotypes, which are unique to the Southern African context and are often under-investigated in cell line development for FMDV isolation and propagation. Full article

BK viremia (BKPyV-DNAemia) and nephropathy (BKPyVAN) are significant causes of morbidity and mortality in kidney transplant recipients (KTRs). Vitamin D supports immune function, yet low 25-hydroxyvitamin D [25(OH)D] is common among KTRs. The association between serum 25(OH)D, measured 61 days to 2 years post-transplant, and subsequent incident BKPyV-DNAemia and BKPyVAN was examined in KTRs without previous BKPyV-DNAemia or BKPyVAN, respectively. Out of 3308 KTRs, 399 (12%) were vitamin D deficient [25(OH)D ≤ 20 ng/mL], and 916 (27.7%) were insufficient [25(OH)D 21–29 ng/mL]. A total of 184 KTRs developed BKPyV-DNAemia and 44 developed BKPyVAN. The incidence rate (/100 person-years) for BKPyV-DNAemia was 2.88 in the 25(OH)D sufficient group, 2.22 in the insufficient group, and 2.37 in the deficient group. The incidence rate (/100 person-years) for BKPyVAN was 0.30 in the 25(OH)D sufficient group, 0.75 in the insufficient group, and 1.28 in the deficient group. Vitamin D deficiency (adjusted hazard ratio [aHR] compared to 25(OH)D sufficiency: 3.92; 95% CI: 1.66–9.23) and insufficiency (aHR: 2.22; 95% CI: 1.11–4.45) remained significantly associated with the incidence of BKPyVAN after adjustment for baseline characteristics. Low serum 25(OH)D was associated with an increased risk of BKPyVAN but not BKPyV-DNAemia. Full article

Though antimicrobial stewardship programs (ASPs) are required for hospitals, the involvement of transplant recipients in programmatic interventions, protocols, and metrics has historically been limited. Though there is a growing interest in studying stewardship practices in transplant patients, optimal practices have not been clearly established. A component of ASPs, antiviral stewardship (AVS), specifically targeting cytomegalovirus (CMV), has been more recently described. Understanding AVS opportunities and interventions is particularly important for transplant recipients, given the morbidity and mortality associated with viral infections, challenging clinical syndromes, ultrasensitive molecular diagnostic assays, antiviral resistance, and costs of viral disease and medications, as well as antiviral drug toxicities. This review highlights opportunities for AVS for CMV, EBV, HSV, VZV, SARS-CoV-2, respiratory syncytial virus, and BK polyomavirus in transplant patients. Full article

Chronic kidney disease is a really important heath issue, and transplantation is an intervention that can greatly increase patient quality of life and survival. The aim of this study was to perform a comprehensive evaluation of the BK virus, CMV, and EBV in kidney transplant recipients (KTRs); to assess the prevalence of infections; and to test if our detection method would be feasible for use in follow-ups with KTRs. A total of 157 KTRs registered at the Clinical Hospital “Dr. C. I. Parhon”, Iași, Romania, were selected using specific inclusion/exclusion criteria. We tested the blood samples from each patient for BK, EBV, and CMV using a multiplex real-time PCR (qPCR) assay and the TaqMan PCR principle. The highest prevalence was detected for BKV (11/157, 7%), followed by CMV (9/157, 5.7%) and EBV (5/157, 3.2%). By simultaneously detecting three possible nephropathic viruses and oncogenes in KTRs using multiplex real-time PCR, we aimed to optimize their monitoring and follow-up. The prevalence of the tested nephropathogenic viruses—BKV, CMV, and EBV—was comparable to that analyzed in other studies. We demonstrate that the use of qPCR for viral detection in KTRs is a robust, cost-effective method for case monitoring. Full article

Background/Objectives: With kidney transplant immunosuppression, physicians must balance preventing rejection with minimizing infection and malignancy risks. Steroids have been a mainstay of these immunosuppression regimens since the early days of kidney transplantation, yet their risks remain debated. Our study looks at the clinical outcomes of patients undergoing early steroid withdrawal (ESW) vs. steroid continuous (SCI) maintenance immunosuppression in adult kidney transplant recipients. Methods: A retrospective case-control study, utilizing propensity score-matching, was performed using the US Collaborative Network Database within TriNetX to evaluate renal transplant outcomes at one year in first-time kidney transplant adult patients (>18 years old) who were prescribed an ESW regimen (no steroids after post-transplant day 7 with maintenance tacrolimus [tac] + mycophenolic acid [MMP]/mycophenolate mofetil [MMF]) vs. SCI (tac + MMF/MMP + prednisone). Cohorts were matched on demographics, comorbidities, previously described risk factors for rejection, and induction immunosuppression. Primary outcomes included viral infections, pyelonephritis, and sepsis. Secondary outcomes included renal transplant rejection, death-censored allograft failure (eGFR < 15 mL/min), patient mortality, delayed graft function, and diabetes mellitus. Results: A total of 2056 patients were in each cohort after matching (mean age: 50.7–51 years, 17.9–20.0% African American, 60–60.6% male.) The SCI cohort had a significantly higher cumulative incidence of composite viremia (18 vs. 28.1%, ESW vs. SCI, p < 0.01) driven by CMV, EBV, and BK virus. Post-transplant diabetes mellitus was significantly higher in the SCI cohort (3.21% vs. 5.49%, ESW vs. SCI, p < 0.01). Delayed graft function was also higher in the SCI cohort (19.55% vs. 22.79%, ESW vs. SCI, p < 0.01). Pyelonephritis (2.3 vs. 4.91%, ESW vs. SCI, p < 0.01) and sepsis (2.15 vs. 5.95%, ESW vs. SCI, p < 0.01) were higher in the SCI cohort. Rejection rates were similar between ESW and SCI (29 vs. 31%, ESW vs. SCI, p = 0.41). There were significantly higher incidences of graft failure (4.9 vs. 9.9%, ESW vs. SCI, p < 0.01) and mortality (0.8 vs. 2.1%, ESW vs. SCI, p < 0.01) in the SCI cohort. Conclusions: This well-matched case-control study suggests that ESW is associated with lower infectious outcomes, mortality, and graft failure without increasing rejection risk, supporting the potential benefits of ESW in kidney transplant patients. Full article

Despite the high prevalence of BK polyomavirus (BKPyV) and the associated risk for BKPyV-associated nephropathy (BKPyVAN) in kidney transplant (KTX) recipients, many details on viral processes such as replication, maturation, assembly and virion release from host cells have not been fully elucidated. VP1 is a polyomavirus-specific protein that is expressed in the late phase of its replicative cycle with important functions in virion assembly and infectious particle release. This study investigated the localization and time-dependent changes in the distribution of VP1-positive viral particles and their association within the spectrum of differing cell morphologies that are observed in the urine of KTX patients upon active BKPyV infection. We found highly differing recognition patterns of two anti-VP1 antibodies with respect to intracellular and extracellular VP1 localization, pointing towards independent binding sites that were seemingly associated with differing stages of virion maturation. Cells originating from single clones were stably cultured out of the urine sediment of KTX recipients with suspected BKPyVAN. The cell morphology, polyploidy, virus replication and protein production were investigated by confocal microscopy using both a monoclonal (mAb 4942) and a polyclonal rabbit anti-VP1-specific antibody (RantiVP1 Ab). Immunoblotting was performed to investigate changes in the VP1 protein. Both antibodies visualized VP1 and the mAb 4942 recognized VP1 in cytoplasmic vesicles exhibiting idiomorphic sizes when released from the cells. In contrast, the polyclonal antibody detected VP1 within the nucleus and in cytoplasm in colocalization with the endoplasmic reticulum marker CNX. At the nuclear rim, VP1 was recognized by both antibodies. Immunoblotting revealed two smaller versions of VP1 in urinary decoy cell extracts, potentially from different translation start sites as evaluated by in silico analysis. Oxford Nanopore sequencing showed integration of BKPyV DNA in chromosomes 3, 4 and 7 in one of the five tested primary cell lines which produced high viral copies throughout four passages before transcending into senescence. The different staining with two VP1-specific antibodies emphasizes the modification of VP1 during the process of virus maturation and cellular exit. The integration of BKPyV into the human genome leads to high virus production; however, this alone does not transform the cell line into a permanently cycling and indefinitely replicating one. Full article

The BK polyomavirus (BKPyV) is a small DNA non-enveloped virus whose infection is asymptomatic in most of the world’s adult population. However, in cases of immunosuppression, the reactivation of the virus can cause various complications, and in particular, nephropathies in kidney transplant recipients or hemorrhagic cystitis in bone marrow transplant recipients. Recently, it was demonstrated that BKPyV virions can use extracellular vesicles to collectively traffic in and out of cells, thus exiting producing cells without cell lysis and entering target cells by diversified entry routes. By a comparison to other naked viruses, we investigated the possibility that BKPyV virions recruit the Endosomal-Sorting Complexes Required for Transport (ESCRT) machinery through late domains in order to hijack extracellular vesicles. We identified a single potential late domain in the BKPyV structural proteins, a YPX₃L motif in the VP1 protein, and used pseudovirions to study the effect of point mutations found in a BKPyV clinical isolate or known to ablate the interaction of such a domain with the ESCRT machinery. Our results suggest that this domain is not involved in BKPyV association with extracellular vesicles but is crucial for capsomere interaction and thus viral particle assembly. Full article

Since the significance of viral infections in children and adolescents with nephrotic syndrome (NS) is yet to be defined, this study intended to estimate the occurrence, pattern, and outcomes of some DNA viral infections in children with NS. Methods: A prospective study was conducted to determine the genome identification of the viruses Epstein-Barr (EBV), human cytomegalovirus (HCMV), human herpesvirus 6 (HHV-6 type A and type B) and 7 (HHV-7), polyomavirus (BKV), and human adenovirus (HAdV) in plasma and urine samples of pediatric patients with NS. Results: A total of 35 patients aged 1 to 18 years with NS and under immunosuppressant drugs participated in the study. Plasma and urine samples were collected at regular intervals during a median follow-up of 266 days (range 133–595), and DNA was analyzed to detect the selected DNA viruses. Eleven patients (31.4%) had active virus infections, and patterns were classified as coinfection, recurrent, and consecutive. Of these, six patients (54.5%) presented viral coinfection, six (54.5%) viral recurrence, and seven patients (63.3%) had viral consecutive infection. Ten of the eleven patients with active infection had a proteinuria relapse (91%) and eight (72.7%) were hospitalized (p = 0.0022). Active HCMV infection was the most frequent infection and was observed in six patients (54.5%), three of the eleven patients (27.2%) had suspected HCMV disease in the gastrointestinal tract, and one had HHV-7 coinfection. The frequency of other infections was: 9% for HHV-6, 45.5% for BKV, 27.3% for HHV-7, 18.2% for EBV, and 18.2% for HAdV. Conclusion: viral infections, especially HCMV, can be an important cause of morbidity and nephrotic syndrome relapse in children. Full article