Lilli Gard1, Hubert G M Niesters2, Annelies Riezebos-Brilman2. 1. Department of Medical Microbiology, Division of Clinical Virology, The University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9713 GZ Groningen, The Netherlands. Electronic address: l.gard@umcg.nl. 2. Department of Medical Microbiology, Division of Clinical Virology, The University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9713 GZ Groningen, The Netherlands.
Abstract
BACKGROUND: Polyomavirus BK (BKV) may cause nephropathy in renal transplant recipients and hemorrhagic cystitis in bone marrow recipients. We developed real-time PCRs (RT-PCR) to determine easily and rapidly the different BKV genotypes (BKGT) (I-IV). METHODS: On the VP1 gene a duplex of RT-PCRs was developed and validated to differentiate the four main BKGT. 212 BKV positive samples (21 plasma, 191 urine) were tested with these specific PCRs. Of these 212 samples, 55 PCR results were additionally confirmed by sequencing a VP1 gene fragment (nucleotide 1630-1956). RESULTS: For every genotype, a highly specific, precise and internally controlled assay was developed with a limit of detection of log 3 copies per ml. In 18 (8.5%) of these samples genotyping was not successful due to a low viral load. By sequence analysis, the genotype of 46 out of 55 and 2 out of 4 samples with double infection could be confirmed. CONCLUSIONS: This study describes RT-PCRs for detection of the main BKGT. It proved to be rapid, cheap and sensitive compared to sequencing. Double infections can also be detected. This method will be of value to investigate the role of BKV infection in relation to the genotype.
BACKGROUND:Polyomavirus BK (BKV) may cause nephropathy in renal transplant recipients and hemorrhagic cystitis in bone marrow recipients. We developed real-time PCRs (RT-PCR) to determine easily and rapidly the different BKV genotypes (BKGT) (I-IV). METHODS: On the VP1 gene a duplex of RT-PCRs was developed and validated to differentiate the four main BKGT. 212 BKV positive samples (21 plasma, 191 urine) were tested with these specific PCRs. Of these 212 samples, 55 PCR results were additionally confirmed by sequencing a VP1 gene fragment (nucleotide 1630-1956). RESULTS: For every genotype, a highly specific, precise and internally controlled assay was developed with a limit of detection of log 3 copies per ml. In 18 (8.5%) of these samples genotyping was not successful due to a low viral load. By sequence analysis, the genotype of 46 out of 55 and 2 out of 4 samples with double infection could be confirmed. CONCLUSIONS: This study describes RT-PCRs for detection of the main BKGT. It proved to be rapid, cheap and sensitive compared to sequencing. Double infections can also be detected. This method will be of value to investigate the role of BKV infection in relation to the genotype.
Authors: Herman F Wunderink; Caroline S de Brouwer; Els van der Meijden; Diana V Pastrana; Aloysius C M Kroes; Christopher B Buck; Mariet C W Feltkamp Journal: J Clin Virol Date: 2018-12-01 Impact factor: 3.168
Authors: Lilli Gard; Willem van Doesum; Hubert G M Niesters; Willem J van Son; Arjan Diepstra; Coen A Stegeman; Henk Groen; Annelies Riezebos-Brilman; Jan Stephan Sanders Journal: PLoS One Date: 2017-06-13 Impact factor: 3.240
Authors: H F Wunderink; C S De Brouwer; L Gard; J W De Fijter; A C M Kroes; J I Rotmans; M C W Feltkamp Journal: Open Forum Infect Dis Date: 2019-02-19 Impact factor: 3.835
Authors: Sergio Kamminga; Igor A Sidorov; Michaël Tadesse; Els van der Meijden; Caroline de Brouwer; Hans L Zaaijer; Mariet C W Feltkamp; Alexander E Gorbalenya Journal: iScience Date: 2021-12-11
Authors: Darlene Vigil; Nikifor K Konstantinov; Marc Barry; Antonia M Harford; Karen S Servilla; Young Ho Kim; Yijuan Sun; Kavitha Ganta; Antonios H Tzamaloukas Journal: World J Transplant Date: 2016-09-24