A 18-year-old patient with a history of Matched-Unrelated-Donor Peripheral Blood Stem Cell Transplantation (MUD-PBSCT) was admitted to the hospital because of renal failure. In November 2007 the patient was first diagnosed with acute lymphoblastic leukemia of T cells (T-ALL) at another institution, where he was treated with induction therapy according to Hyper-CVAD Protocol (Cyclophosphamide 300 mg/m2 intravenously (IV) over 3 hours every 12 hours for six doses on days 1 through 3, with mesna given by continuous infusion, vincristine 2 mg IV days 4 and 11, doxorubicin 50 mg/m2 IV day 4, and dexamethasone 40 mg daily on days 1 through 4 and 11 through 14) and was refractory when he presented in our department of hematology/oncology. The diagnosis of T-ALL has been confirmed, immunophenotype precursor T-ALL (CD7, CD3, CD2, CD4, and CD8 pos), cytogenetic 46, XY, Fluorescein in sito hybridisation (FISH) analysis 87% deletion of p16 on the chromosome 9 region p21. He was then treated with induction therapy according to the German multicenter ALL protocol (GMALL) achieving only reduction of blasts after Induction II. Briefly, the induction in GMALL protocol was composed of eight cytotoxic drugs administered sequentially in two phases over an 8-week period. During the first 4 weeks (phase I), the patient received 60 mg/m2 prednisolone PO daily (days 1 through 28) plus 45 mg/m2 daunorubicin and 2 mg vincristine IV weekly (days 1, 8, 15, and 22). L-asparaginase (5 000 U/m2) was administered IV daily (days 15 through 28). In the second 4 weeks (phase II), the patient received two doses of 650 mg/m2 cyclophosphamide IV (days 29, 43, and 57) together with 60 mg/m2 6-mercaptopurine PO daily (days 29 through 57) and four courses of 75 mg/m2 cytarabine IV (days 31 through 34, 38 through 41, 45 through 48, and 52 through 55). He was then treated with nelarabine and achieved complete remission (CR1) following MUD-PBSCT. The conditioning consisted of total body irradiation (TBI), 12 Gy, given fractionated 2 times daily for three days, from day -6 to day -4, and cyclophosphamide 60 mg/kg once daily i.v. on days -3 and -2 (total dose 120 mg/kg b.w.). A Human Leukocyte Antigen (HLA) identical unrelated donor peripheral blood stem cells were infused on day 0. Graft-versus-Host Disease (GvHD) prophylaxis consisted of thymoglobulin (2 mg/kg b.w. daily, from day -3 to -1, total dose of 6 mg/kg b.w.), cyclosporine 5 mg/kg starting on day -1, and methotrexate at days +1, +3, +6, and +11 intravenously 15 mg absolute. The bone marrow puncture on day +28 showed a complete hematological remission with 100% donor chimerism. Along with the regeneration of white blood cells the patient developed a methylprednisolone-dependent GvHD of the skin and gastrointestinal, grade II. After 10 days the therapy with corticosteroids could be discontinued by rapid improvement of the GvHD-related symptoms. Under the therapy with corticosteroids the patient developed a herpes simplex related oesophagitis on day 21 post transplant which was treated with 30 g daily over 5 days i.v. immunoglobulin and foscarnet 90 mg/kg b.w. divided in 2 doses. On day +28 after PBSCT the patient developed a hemorrhagic cystitis with high urine titers of BK polyoma virus and CMV infection. A therapy with ganciclovir was initiated for ten days. Under treatment with ganciclovir, CMV infection improved, and the treatment with cidofovir was started (1 mg/kg b.w. for two weeks). The BK Virus viruria and viremia improved, and the patient received ganciclovir treatment for additional two weeks (Figure 1). The cyclosporine level was from 200 to 150 from day 0 to day 30, followed with level of 100 from day 30 to day 40 post transplant, and level of 50 from day 40 to day 50. The cyclosporine has been discontinued on day +70 after MUD-PBSCT in absence of acute GvHD and as recommended in GMALL protocol. On day +48 after PBSCT a cystoscopy with a continuous intravesical saline irrigation was started and discontinued 2 weeks later. The patient symptoms improved, and creatinine levels returned to normal. He was discharged from hospital; his medications at the time of discharge included ciprofloxacin (250 mg twice daily), valacyclovir (500 mg twice daily), fluconazole (200 mg daily), folate, and cotrimoxazole twice weekly for prophylaxis of pneumocystis carinii infection.
Over the next year the levels of urea nitrogen and creatinine gradually rose (Table 1), which was attributed to drug toxicity. cotrimoxazole prophylaxis was stopped. Cyclosporine was discontinued since no signs of GvHD were present 6 months after PBSCT. The serum creatinine level was 153 mumol/l (Table 1). Routine urinary diagnostic measures were carried out and showed inconsistent microhematuria. BK viruria was negative in this period. 
On admission, blood pressure was 130/80 mmHg with 90 heart beats per minute, a temperature of 36.4 C, and an oxygen saturation of 99% when the patient was breathing ambient air. The lungs were clear, the results of the remainder of the examination were normal. The complete blood count and levels of serum electrolytes, liver function tests, and levels of total protein and albumin were normal. The urine sediment was not nephritic but contained a small amount of leukocytes indicating an inflammatory process such as cystitis. Results of other laboratory tests are shown in Table 1. 
Ultrasonographic examination of the kidneys showed bilateral hydronephrosis stage III-IV (Figure 2(a)). In addition, decreased echotexture bladder tumours were seen (Figure 2(b)). The patient was presented to the urologist. Cystoscopy with urethrography has shown exophytic bladder tumours in both ostii. The extirpation of the tumours was performed and sent off for pathological examination. In addition a double-J catheter was placed. The hydronephrosis of the left kidney improved immediately and improved to grade II in the right kidney. For histological examination tissues were formalin-fixed overnight and embedded in paraffin. Hematoxylin & Eosin staining of 5 micrometer sections was performed following standard procedures. 
The pathohistology of the specimen showed a chronic cystical urocystitis with bleedings (Figure 3(a)). Tissue probes of both ureters were stained for SV40 large T antigen (clone pAb416, Oncogene Science, Boston, Mass, USA; dilution 1 : 100, antigen retrieval and amplification with catalyzed system amplification, Dako, Carpinteria, CA). The reactions were detected by means of the streptavidin-biotin method and were revealed using diaminobenzidine as a chromogen. In all tissue probes immunohistochemistry for large T antigen was negative (Figure 3(b)). 
The patient has undergone double-J catheter controls and changes every two months and is doing well 2 years after PBSCT and 1 year after operative extirpation of tumours. The last control was in April 2010 (Table 1).
Two different regions of the BKV genome were amplified by PCR, VP2 in serum and urine, and large T antigen from tissue biopsies.
Nucleic acids were extracted from 200 microliter blood or urine using magnetic beads (MagNA Pure LC total NA isolation kit, Roche, Mannheim, Germany) according to the manufacturer's instructions while from formalin-fixed, paraffin-embedded material nucleic acids were extracted using the QIAamp DNA FFPE Tissue Kit (Qiagen, Hilden, Germany). VP2 primers lead to amplification of a 131 bp fragment. PCR reaction was carried out in 20 microliter capillaries containing master mix (6 mM MgCl2, 0.1 mg/ml BSA (Sigma, St. Louis, USA)) and 10 picoM of each primer (Metabion, Martinsried, Germany), 3 picoM of each hybridization probe (5'-GGCTGCTGCTGCCACAGGATTTTC-FL (probe 1) and 5'LC Red640-GTAGCTGAAATTGCTGCTGGAGAG-ph (probe 2), 0.25 mM dNTP's (Roche)), and 1U Platinum Taq Polymerase (Invitrogen, Karlsruhe, Germany). Target DNA was added in a volume of 5 microliters. Two negative and 4 positive controls in different concentrations were included into each run performed on a LightCycler 1.0 instrument (Roche, Mannheim, Germany) under the following conditions: initial denaturation and activation of hot-start enzyme at 95 C for 30 s, followed by 45 cycles of denaturation at 95 C for 0 s, annealing of primers at 58 C for 10 s, and primer extension at 72 C for 20 s. Fluorescence was monitored by single acquisition during the annealing phase of each PCR cycle and channel setting F2/F1 (640 nm/530 nm). Differentiation of BK virus amplicons from JC virus depends on the binding of the probe 2, which perfectly matches JC virus but differs at the two underlined positions from BK virus. Thus binding to BK virus results in a shifted melting curve (61.5 C for BK versus 63.8 C for JC).
For detection of the viral large T-antigen, a 128 bp long fragment was amplified using modified primers from that match BKV genotypes 1 to 4, forward primer 5'-CAGGCAAGGGTTCTATTACTAAAT-3', reverse primer 5'-GCAACAGCAGATTCYCAACA-3', probe 5'-6FAM-AAACTGGTGTAGATCAGARGGAAAGTCTTTAGGGTCTT-BBQ. 
PCR conditions were identical to the one described above except TaqMan probe that was used at 4 pico molar, primer annealing was 56 C for 10 s, and fluorescence acquisition was at the end of each primer extension phase at 530 nm.