1887

Abstract

BK polyomavirus (BKPyV) quantification is useful for monitoring renal transplant patient response to therapy. The Abbott 2000 RealTie System employed by some clinical laboratories to perform US Food and Drug Administration-approved assays can also be used to develop in-house assays such as the one presented here. This study aimed to validate an in-house quantitative real-time PCR assay targeting the BKPyV major capsid VP1 gene for assessment of viral load using the Abbott 2000 RealTie System. BKPyV load was measured in 95 urine and plasma samples previously tested for BKPyV by one of three laboratories (46 BKPyV-positive samples consisting of 35 plasma and 11 urine samples; 49 samples negative for BKPyV consisting of 47 plasma and two urine samples). Two additional plasma specimens from the College of American Pathologists proficiency testing survey were also analysed. Precision studies were performed by diluting a high-viral-titre patient sample into BKPyV-negative pooled plasma to create high-positive (6.16 log copies ml) and low-positive (3.16 log copies ml) samples. For precision studies of inter-assay variability, a high-positive (7.0 log copies ml) and a low-positive (3.0 log copies ml) sample were measured in 20 separate runs. The assay’s limit of quantification and limit of detection were 2.70 and 2.25 log copies ml, respectively. The assay was linear from 2.70 to 9.26 log copies ml. Of the 48 known positives, 43 were detected as positive, with three reported by the reference laboratory as values lower than the limit of detection. Two known positives at 3.27 and 3.80 log copies ml tested negative by the 2000 BKPyV assay. Of the 49 known negative samples, 48 were negative by the 2000 BKPyV load assay, with one sample confirmed positive by a reference laboratory. Qualitative analysis prior to discrepancy testing demonstrated a sensitivity of 89.58 % and a specificity of 97.96 %. Precision studies demonstrated inter-assay coefficients of variation of 0.63 % (high positive) and 4.38 % (low positive). Genotyping was performed on 22 patient samples, of which 21 (95.45 %) were type I and one (4.55 %) was type II. In conclusion, the 2000 BKPyV viral load assay sensitivity, specificity, linear range, precision and cost effectiveness make it an attractive methodology for clinical laboratories using the Abbott 2000 RealTie System.

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2013-11-01
2019-10-20
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References

  1. Adeyi O. A., Belloni D. R., Dufresne S. D., Schned A. R., Tsongalis G. J.. ( 2005;). Real-time polymerase chain reaction and laser capture microdissection for the diagnosis of BK virus infection in renal allografts. . Am J Clin Pathol 124:, 537–542. [CrossRef][PubMed]
    [Google Scholar]
  2. Amendola A., Coen S., Belladonna S., Pulvirenti F. R., Clemens J. M., Capobianchi M. R.. ( 2011;). Improving clinical laboratory efficiency: a time-motion evaluation of the Abbott m2000 RealTime and Roche COBAS AmpliPrep/COBAS TaqMan PCR systems for the simultaneous quantitation of HIV-1 RNA and HCV RNA. . Clin Chem Lab Med 49:, 1283–1288. [CrossRef][PubMed]
    [Google Scholar]
  3. Bechert C. J., Schnadig V. J., Payne D. A., Dong J.. ( 2010;). Monitoring of BK viral load in renal allograft recipients by real-time PCR assays. . Am J Clin Pathol 133:, 242–250. [CrossRef][PubMed]
    [Google Scholar]
  4. Biel S. S., Held T. K., Landt O., Niedrig M., Gelderblom H. R., Siegert W., Nitsche A.. ( 2000;). Rapid quantification and differentiation of human polyomavirus DNA in undiluted urine from patients after bone marrow transplantation. . J Clin Microbiol 38:, 3689–3695.[PubMed]
    [Google Scholar]
  5. Bourlet T., Signori-Schmuck A., Roche L., Icard V., Saoudin H., Trabaud M.-A., Tardy J.-C., Morand P., Pozzetto B.. & other authors ( 2011;). HIV-1 load comparison using four commercial real-time assays. . J Clin Microbiol 49:, 292–297. [CrossRef][PubMed]
    [Google Scholar]
  6. Cheng A., Qian Q., Kirby J. E.. ( 2011;). Evaluation of the Abbott RealTime CT/NG assay in comparison to the Roche Cobas Amplicor CT/NG assay. . J Clin Microbiol 49:, 1294–1300. [CrossRef][PubMed]
    [Google Scholar]
  7. Chung B. H., Hong Y. A., Kim H. G., Sun I. O., Choi S. R., Park H. S., Lee S. H., Choi B. S., Park C. W.. & other authors ( 2012;). Clinical usefulness of BK virus plasma quantitative PCR to prevent BK virus associated nephropathy. . Transpl Int 25:, 687–695. [CrossRef][PubMed]
    [Google Scholar]
  8. Cimbaluk D., Pitelka L., Kluskens L., Gattuso P.. ( 2009;). Update on human polyomavirus BK nephropathy. . Diagn Cytopathol 37:, 773–779. [CrossRef][PubMed]
    [Google Scholar]
  9. College of American Pathologists ( 2012;). Viral load survey VLS-B. Northfield, IL: College of American Pathologists. . [PubMed]
    [Google Scholar]
  10. Costa C., Bergallo M., Sidoti F., Astegiano S., Terlizzi M. E., Mazzucco G., Segoloni G. P., Cavallo R.. ( 2009;). Polyomaviruses BK- and JC-DNA quantitation in kidney allograft biopsies. . J Clin Virol 44:, 20–23. [CrossRef][PubMed]
    [Google Scholar]
  11. Decker D. B., Karam J. A., Wilcox D. T.. ( 2009;). Pediatric hemorrhagic cystitis. . J Pediatr Urol 5:, 254–264. [CrossRef][PubMed]
    [Google Scholar]
  12. Dingle T., Sedlak R. H., Cook L., Jerome K. R.. ( 2013;). Comparison of the tolerance of real-time quantitative PCR versus droplet digital PCR to inhibitors. . In 29th Clinical Virology Symposium, 26 April–1 May, Daytona Beach, Florida, USA:.
    [Google Scholar]
  13. Funahashi Y., Iwata S., Ito Y., Kojima S., Yoshikawa T., Hattori R., Gotoh M., Nishiyama Y., Kimura H.. ( 2010;). Multiplex real-time PCR assay for simultaneous quantification of BK polyomavirus, JC polyomavirus, and adenovirus DNA. . J Clin Microbiol 48:, 825–830. [CrossRef][PubMed]
    [Google Scholar]
  14. Höcker B., Tönshoff B.. ( 2009;). Treatment strategies to minimize or prevent chronic allograft dysfunction in pediatric renal transplant recipients: an overview. . Paediatr Drugs 11:, 381–396. [CrossRef][PubMed]
    [Google Scholar]
  15. Hoffman N. G., Cook L., Atienza E. E., Limaye A. P., Jerome K. R.. ( 2008;). Marked variability of BK virus load measurement using quantitative real-time PCR among commonly used assays. . J Clin Microbiol 46:, 2671–2680. [CrossRef][PubMed]
    [Google Scholar]
  16. Iwaki K. K., Qazi S. H., Garcia-Gomez J., Zeng D., Matsuda Y., Matsuda K., Martinez M. E., Toyoda M., Kore A.. & other authors ( 2010;). Development of a real-time quantitative PCR assay for detection of a stable genomic region of BK virus. . Virol J 7:, 295. [CrossRef][PubMed]
    [Google Scholar]
  17. Kipp B. R., Sebo T. J., Griffin M. D., Ihrke J. M., Halling K. C.. ( 2005;). Analysis of polyomavirus-infected renal transplant recipients’ urine specimens: correlation of routine urine cytology, fluorescence in situ hybridization, and digital image analysis. . Am J Clin Pathol 124:, 854–861. [CrossRef][PubMed]
    [Google Scholar]
  18. Korup S., Rietscher J., Calvignac-Spencer S., Trusch F., Hofmann J., Moens U., Sauer I., Voigt S., Schmuck R., Ehlers B.. ( 2013;). Identification of a novel human polyomavirus in organs of the gastrointestinal tract. . PLoS ONE 8:, e58021. [CrossRef][PubMed]
    [Google Scholar]
  19. Krumbholz A., Zell R., Egerer R., Sauerbrei A., Helming A., Gruhn B., Wutzler P.. ( 2006;). Prevalence of BK virus subtype I in Germany. . J Med Virol 78:, 1588–1598. [CrossRef][PubMed]
    [Google Scholar]
  20. Limaye A. P., Jerome K. R., Kuhr C. S., Ferrenberg J., Huang M. L., Davis C. L., Corey L., Marsh C. L.. ( 2001;). Quantitation of BK virus load in serum for the diagnosis of BK virus-associated nephropathy in renal transplant recipients. . J Infect Dis 183:, 1669–1672. [CrossRef][PubMed]
    [Google Scholar]
  21. Luo C., Bueno M., Kant J., Randhawa P.. ( 2008;). Biologic diversity of polyomavirus BK genomic sequences: Implications for molecular diagnostic laboratories. . J Med Virol 80:, 1850–1857. [CrossRef][PubMed]
    [Google Scholar]
  22. Marcén R.. ( 2009;). Immunosuppressive drugs in kidney transplantation: impact on patient survival, and incidence of cardiovascular disease, malignancy and infection. . Drugs 69:, 2227–2243. [CrossRef][PubMed]
    [Google Scholar]
  23. Marchetti S., Graffeo R., Siddu A., Santangelo R., Ciotti M., Picardi A., Favalli C., Fadda G., Cattani P.. ( 2007;). BK virus DNA detection by real-time polymerase chain reaction in clinical specimens. . New Microbiol 30:, 119–126.[PubMed]
    [Google Scholar]
  24. Miller S., Liverman C. S., Post L., Khan Y., Wright C.. ( 2012;). Analytical and clinical performance characteristics of the Simplexa BK virus quantitative PCR assay for the diagnosis of polyomavirus-associated nephropathy in renal transplant recipients using plasma and urine specimens. . J Clin Virol 55:, 310–316. [CrossRef][PubMed]
    [Google Scholar]
  25. Møller J. K., Pedersen L. N., Persson K.. ( 2010;). Comparison of the Abbott RealTime CT new formulation assay with two other commercial assays for detection of wild-type and new variant strains of Chlamydia trachomatis. . J Clin Microbiol 48:, 440–443. [CrossRef][PubMed]
    [Google Scholar]
  26. Moret H., Brodard V., Barranger C., Jovenin N., Joannes M., Andréoletti L.. ( 2006;). New commercially available PCR and microplate hybridization assay for detection and differentiation of human polyomaviruses JC and BK in cerebrospinal fluid, serum, and urine samples. . J Clin Microbiol 44:, 1305–1309. [CrossRef][PubMed]
    [Google Scholar]
  27. Pang X. L., Doucette K., LeBlanc B., Cockfield S. M., Preiksaitis J. K.. ( 2007;). Monitoring of polyomavirus BK virus viruria and viremia in renal allograft recipients by use of a quantitative real-time PCR assay: one-year prospective study. . J Clin Microbiol 45:, 3568–3573. [CrossRef][PubMed]
    [Google Scholar]
  28. Pang X. L., Fox J. D., Fenton J. M., Miller G. G., Caliendo A. M., Preiksaitis J. K..American Society of Transplantation Infectious Diseases Community of PracticeCanadian Society of Transplantation ( 2009;). Interlaboratory comparison of cytomegalovirus viral load assays. . Am J Transplant 9:, 258–268. [CrossRef][PubMed]
    [Google Scholar]
  29. Pastrana D. V., Brennan D. C., Çuburu N., Storch G. A., Viscidi R. P., Randhawa P. S., Buck C. B.. ( 2012;). Neutralization serotyping of BK polyomavirus infection in kidney transplant recipients. . PLoS Pathog 8:, e1002650. [CrossRef][PubMed]
    [Google Scholar]
  30. Preiksaitis J. K., Pang X. L., Fox J. D., Fenton J. M., Caliendo A. M., Miller G. G..American Society of Transplantation Infectious Diseases Community of Practice ( 2009;). Interlaboratory comparison of Epstein–Barr virus viral load assays. . Am J Transplant 9:, 269–279. [CrossRef][PubMed]
    [Google Scholar]
  31. Ramos E., Drachenberg C. B., Wali R., Hirsch H. H.. ( 2009;). The decade of polyomavirus BK-associated nephropathy: state of affairs. . Transplantation 87:, 621–630. [CrossRef][PubMed]
    [Google Scholar]
  32. Randhawa P., Ho A., Shapiro R., Vats A., Swalsky P., Finkelstein S., Uhrmacher J., Weck K.. ( 2004;). Correlates of quantitative measurement of BK polyomavirus (BKV) DNA with clinical course of BKV infection in renal transplant patients. . J Clin Microbiol 42:, 1176–1180. [CrossRef][PubMed]
    [Google Scholar]
  33. Randhawa P., Kant J., Shapiro R., Tan H., Basu A., Luo C.. ( 2011;). Impact of genomic sequence variability on quantitative PCR assays for diagnosis of polyomavirus BK infection. . J Clin Microbiol 49:, 4072–4076. [CrossRef][PubMed]
    [Google Scholar]
  34. Singh H. K., Andreoni K. A., Madden V., True K., Detwiler R., Weck K., Nickeleit V.. ( 2009;). Presence of urinary Haufen accurately predicts polyomavirus nephropathy. . J Am Soc Nephrol 20:, 416–427. [CrossRef][PubMed]
    [Google Scholar]
  35. Swanson P., Holzmayer V., Huang S., Hay P., Adebiyi A., Rice P., Abravaya K., Thamm S., Devare S. G., Hackett J. Jr. ( 2006;). Performance of the automated Abbott RealTime HIV-1 assay on a genetically diverse panel of specimens from London: comparison to VERSANT HIV-1 RNA 3.0, AMPLICOR HIV-1 MONITOR v1.5, and LCx HIV RNA Quantitative assays. . J Virol Methods 137:, 184–192. [CrossRef][PubMed]
    [Google Scholar]
  36. Swanson P., Huang S., Abravaya K., de Mendoza C., Soriano V., Devare S. G., Hackett J. Jr. ( 2007;). Evaluation of performance across the dynamic range of the Abbott RealTime HIV-1 assay as compared to VERSANT HIV-1 RNA 3.0 and AMPLICOR HIV-1 MONITOR v1.5 using serial dilutions of 39 group M and O viruses. . J Virol Methods 141:, 49–57. [CrossRef][PubMed]
    [Google Scholar]
  37. Trofe-Clark J., Sparkes T., Gentile C., Van Deerlin V., Sawinski D., Bloom R. D.. ( 2013;). BK virus genotype variance and discordant BK viremia PCR assay results. . Am J Transplant 13:, 1112–1113. [CrossRef][PubMed]
    [Google Scholar]
  38. Wiseman A. C.. ( 2009;). Polyomavirus nephropathy: a current perspective and clinical considerations. . Am J Kidney Dis 54:, 131–142. [CrossRef][PubMed]
    [Google Scholar]
  39. Wu J. K., Harris M. T.. ( 2008;). Use of leflunomide in the treatment of polyomavirus BK-associated nephropathy. . Ann Pharmacother 42:, 1679–1685. [CrossRef][PubMed]
    [Google Scholar]
  40. Yamaguchi H., Kobayashi S., Ishii A., Ogawa H., Nakamura I., Moonga L., Hang’ombe B. M., Mweene A. S., Thomas Y.. & other authors ( 2013;). Identification of a novel polyomavirus from vervet monkeys in Zambia. . J Gen Virol 94:, 1357–1364. [CrossRef][PubMed]
    [Google Scholar]
  41. Yu G., Greninger A. L., Isa P., Phan T. G., Martínez M. A., de la Luz Sanchez M., Contreras J. F., Santos-Preciado J. I., Parsonnet J.. & other authors ( 2012;). Discovery of a novel polyomavirus in acute diarrheal samples from children. . PLoS ONE 7:, e49449. [CrossRef][PubMed]
    [Google Scholar]
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