COMPLICATIONS - INFECTIONS

R. Hattori¹, T. Kinukawa², Y. Funahashi¹, H. Kimura^3
1Urology, Nagoya University School of Medicine, Nagoya/JAPAN, ², Chukyo Hospital, Nagoya/JAPAN, ³Virology, Nagoya University School of Medicine, Nagoya/JAPAN

Body: (Introduction) For the early diagnosis and treatment of BK polyomavirus (BKPyV), urine cytology is useful for screening high-risk patients in kidney transplant recipients. The virus-infected urothelial cells called “decoy cells,” identified by their typical ground glass intranuclear inclusions on cellular smear stained by the Papanicolaou method, are observed in most patients with BKPyV nephropathy. However, decoy cells are not specific for the presence of BKPyV in urine and can be found in JC polyomavirus (JCPyV ) and adenovirus infection. We developed a multiplex real-time PCR assay for the simultaneous quantification of BKPyV, JCPyV, and adenovirus and validated it for the quantitative determination of viral loads. (Materials and methods) Urine samples from 124 renal transplant recipients were obtained to test the accuracy of the assay. The mean patients ages were 47(range, 8 to 75 years). The periods after transplantation were 78 (2 to 275) months. The immunosuppresunt included cyclosporine(n=53), tacrolimus (n=62), and mycophenolate mofetil (n=75). Multiplex and single real-time PCR were performed by using a QuantiTect multiplex PCR kit (Qiagen). (Results) BKPyV was detected in 28(22.6%)samples, JCPyV in 51 (41.1%) samples, and adenovirus in 2 (1.6%) samples from 124 urine specimens. BKPyV and JCPyV were detected together in five urine specimens, BKPyV and adenovirus were detected together in one, and all three viruses were amplified in one. The maximum amounts of each virus were 2.7×10⁹ ,8.7×10⁸ ,1.2 ×10² copies/ml for BKPyV, JCPyV, and adenovirus, respectively. Decoy cells were observed in 31 patients. Of these patients, only BKPyV was detected in 5 patients, only JCPyV was detected in 21 patients, and both viruses were detected in 5 patients. Limited to the cases with only BKPyV, the quantity of BKPyV DNA was higher in samples with decoy cells than in those without (mean, 10^7.0 versus 10^3.5 copies/ml; p=0.001), and all samples positive for decoy cells had a viral load of >10⁵ copies/ml. Conversely, decoy cells were observed in five of seven (71%) samples with>10⁵ copies of BKPyV DNA/ml. (Conclusion) We have developed a multiplex real-time PCR assay for the simultaneous detection of BKPyV, JCPyV, and adenovirus DNA. Thesavings in cost, time, and labor associated with these multiples real-time PCR would be useful in the management of post-transplant recipients.

Disclosure: All authors have declared no conflicts of interest.