title: Polyomavirus BK DNA quantification assay to evaluate viral load in renal transplant recipients.
abstract: BACKGROUND
  Several studies have disclosed a correlation between polyomavirus BK (BKV) and interstitial nephritis in renal transplant recipients and its quantification in urine and serum is therefore required to assess the role of BKV infection in nephropathy.
  
  
  OBJECTIVE
  This paper describes a urine and serum BKV-DNA quantification protocol devised to evaluate the viral load.
  
  
  METHODS
  Screening of samples containing > or =10(3)/ml viral genome copies by a semi-quantitative polymerase chain reaction (PCR) assay is followed by precise quantification of the samples containing a high number of viral genomes in a quantitative-competitive (QC)-PCR assay. Generation of the competitor construct relied on the different sizes of wild-type and competitor amplicons.
  
  
  CONCLUSIONS
  Screening by semi-quantitative PCR selects samples with a high number of viral genomes for use in the more labor-intensive and -expensive QC-PCR assay and thus provides a handy means for quantitative DNA analysis of large numbers of samples. The results obtained in BKV-DNA quantification in urine and serum samples from 51 renal transplant recipients (22 on treatment with tacrolimus (FK506) and 29 on cyclosporine A (Cy A)) are interesting: BKV-DNA findings (43.1%) in urine samples are in agreement with the BKV urinary shedding reported in literature (5-45%). With regard to immunosuppressive treatment, the percentage of activation of the infection (revealed by BKV-DNA detection in urine samples) in the two groups of therapy is similar (40.9% vs 44.8%). The observation that the viral load in urine is dissociated with that of serum suggests that both parameters should be investigated in evaluation of the pathogenetic role of BKV reactivation in renal transplant recipients. Moreover, our BKV-DNA quantification protocol could be used to monitor viral load in urine and serum samples from renal transplant recipients so as to detect those at risk of nephropathy and monitor their response to immunosuppression reduction therapy if it occurs.
authors.affiliation: Department of Public Health and Microbiology, Virology Unit, University of Turin, Via Santena 9-10126, Turin, Italy. chiara.merlino@unito.it
authors.forename: Chiara
authors.initials: C
authors.lastname: Merlino
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Bergallo
authors.forename: Giorgio
authors.initials: G
authors.lastname: Gribaudo
authors.forename: Gabriella
authors.initials: G
authors.lastname: Gregori
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Paolo Segoloni
authors.forename: Franca
authors.initials: F
authors.lastname: Giacchino
authors.forename: Alessandro Negro
authors.initials: AN
authors.lastname: Ponzi
authors.forename: Rossana
authors.initials: R
authors.lastname: Cavallo
mesh_terms: D000328:Adult; D001739:BK Virus / Q000235:genetics / Q000302:isolation & purification*; D004279:DNA, Viral / Q000097:blood* / Q000652:urine*; D005260:Female; D006801:Humans; D016030:Kidney Transplantation / Q000009:adverse effects*; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods*; D027601:Polyomavirus Infections / Q000821:virology; D014412:Tumor Virus Infections / Q000821:virology; D019562:Viral Load
chemical_list: D004279:DNA, Viral