Progress in Quantitative Viral Load Testing: Variability and Impact of the WHO Quantitative International Standards

Abstract

It has been hoped that the recent availability of WHO quantitative standards would improve interlaboratory agreement for viral load testing; however, insufficient data are available to evaluate whether this has been the case. Results from 554 laboratories participating in proficiency testing surveys for quantitative PCR assays of cytomegalovirus (CMV), Epstein-Barr virus (EBV), BK virus (BKV), adenovirus (ADV), and human herpesvirus 6 (HHV6) were evaluated to determine overall result variability and then were stratified by assay manufacturer. The impact of calibration to international units/ml (CMV and EBV) on variability was also determined. Viral loads showed a high degree of interlaboratory variability for all tested viruses, with interquartile ranges as high as 1.46 log 10 copies/ml and the overall range for a given sample up to 5.66 log 10 copies/ml. Some improvement in result variability was seen when international units were adopted. This was particularly the case for EBV viral load results. Variability in viral load results remains a challenge across all viruses tested here; introduction of international quantitative standards may help reduce variability and does so more or less markedly for certain viruses.

Copy number heterogeneity, large origin tandem repeats, and interspecies recombination in HHV-6A and HHV-6B reference strains

“…Recently several quantitative cut-offs have been proposed that are associated with end-organ disease or iciHHV-6 status (5–7). There have been only a few limited studies comparing PCR methods between clinical labs (8–10). In addition, newer studies often utilize commercial reference laboratories or commercial reagents that in general do not make their primer/probe locations known.…”

Calibration of BK Virus Nucleic Acid Amplification Testing to the 1st WHO International Standard for BK Virus

“…In fact, such discrepancies may be appropriate realities for institutions using different assays that produce different quantitative results for a given sample. As seen with CMV and EBV (16), the use of an internationally accepted common calibrator offers promise for improving the harmonization of BKV NAAT results and the opportunity to define broadly applicable interinstitutional thresholds needed for optimizing the primary and secondary prevention of BKV-related end-organ disease in transplantation.…”

New advances in the management of cytomegalovirus in allogeneic haemopoietic stem cell transplantation

“…Both qualitative and quantitative assays are performed using standardised commercial assays with viral loads now reported in standardised form as international units per millilitre (IU/mL) against a World Health Organisation (WHO) standard . Laboratories should be enrolled in quality assurance programmes with results calibrated to the WHO international standards in order to improve the interpretation of quantitative results from different testing laboratories . NAT testing on blood/plasma is generally performed weekly from early post‐transplant to approximately day 100 or later post‐HSCT in patients with active GVHD on immunosuppression.…”

“…Nucleic acid tests are the most commonly used method to diagnose CMV infection and can be performed on blood (whole blood and plasma), fluids (cerebrospinal and vitreous) and tissue samples (including colonic tissue, bronchoalveolar lavage (BAL) and brain tissue). Both qualitative and quantitative assays are performed using standardised commercial assays with viral loads now reported in standardised form as 12 Laboratories should be enrolled in quality assurance programmes with results calibrated to the WHO international standards in order to improve the interpretation of quantitative results from different testing laboratories. 12 NAT testing on blood/plasma is generally performed weekly from early post-transplant to approximately day 100 or later post-HSCT in patients with active GVHD on immunosuppression.…”

“…Both qualitative and quantitative assays are performed using standardised commercial assays with viral loads now reported in standardised form as 12 Laboratories should be enrolled in quality assurance programmes with results calibrated to the WHO international standards in order to improve the interpretation of quantitative results from different testing laboratories. 12 NAT testing on blood/plasma is generally performed weekly from early post-transplant to approximately day 100 or later post-HSCT in patients with active GVHD on immunosuppression. Definitive diagnosis of CMV organ disease remains dependent on proven histological evidence of viral tissue invasive effects such as presence of viral inclusion bodies or positive CMV immunohistochemistry staining.…”